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Microinjection of angiotensin II in the caudal ventrolateral medulla induces hyperalgesia
Abstract
Nociceptive transmission from the spinal cord is controlled by supraspinal pain modulating systems that include the caudal ventrolateral medulla (CVLM). The neuropeptide angiotensin II (Ang II) has multiple effects in the CNS and at the medulla oblongata. Here we evaluated the expression of angiotensin type 1 (AT(1)) receptors in spinally-projecting CVLM neurons, and tested the effect of direct application of exogenous Ang II in the CVLM on nociceptive behaviors. Although AT(1)-immunoreactive neurons occurred in the CVLM, only 3% of AT(1)-positive neurons were found to project to the dorsal horn, using double-immunodetection of the retrograde tracer cholera toxin subunit B. In behavioral studies, administration of Ang II (100 pmol) in the CVLM gave rise to hyperalgesia in both the tail-flick and formalin tests. This hyperalgesia was significantly attenuated by local administration of the AT(1) antagonist losartan. The present study demonstrates that Ang II can act on AT(1) receptors in the CVLM to modulate nociception. The effect on spinal nociceptive processing is likely indirect, since few AT(1)-expressing CVLM neurons were found to project to the spinal cord. The renin-angiotensin system may also play a role in other supraspinal areas implicated in pain modulation.
... Evidence indicates that Ang II is involved in the modulation of nociceptive transmission. Namely, Ang II causes hyperalgesia in the caudal ventrolateral medulla (CVLM) [11] and hypoalgesia in the periaqueductal gray (PAG) and the rostral ventromedial medulla (RVM) [12][13][14]. However, the role of spinal Ang II in the modulation of nociceptive transmission remains unclear. ...
... It has been demonstrated that the administration of Ang II either i.c.v. or directly in key components of the supraspinal pain modulatory system, namely the PAG or RVM (for review, see Ref [30]), induces antinociceptive effects, which are reversed by losartan [12,13,31,32]. On the other hand, Marques-Lopes et al. [11] have recently reported that the microinjection of Ang II into the CVLM induces hyperalgesia through AT 1 receptors, and that the effect of Ang II on spinal nociceptive processing is likely indirect, since few AT 1 receptor-expressing CVLM neurons were found to project to the spinal cord. These reports lead us to suggest that supraspinal Ang II may participate in both inhibition and facilitation of the nociceptive transmission and its effect is region-dependent. ...
It has been demonstrated that angiotensin II (Ang II) participates in either the inhibition or the facilitation of nociceptive transmission depending on the brain area. Neuronal Ang II is locally synthesized not only in the brain, but also in the spinal cord. Though the spinal cord is an important area for the modulation of nociception, the role of spinal Ang II in nociceptive transmission remains unclear. Therefore, in order to elucidate the role of Ang II in nociceptive transmission in the spinal cord, we examined the effect of intrathecal (i.t.) administration of Ang II into mice.
I.t. administration of Ang II produced a behavioral response in mice mainly consisting of biting and/or licking of the hindpaw and the tail along with slight hindlimb scratching directed toward the flank. The behavior induced by Ang II (3 pmol) was dose-dependently inhibited by intraperitoneal injection of morphine (0.1-0.3 mg/kg), suggesting that the behavioral response is related to nociception. The nociceptive behavior was also inhibited dose-dependently by i.t. co-administration of losartan (0.3-3 nmol), an Ang II type 1 (AT1) receptor antagonist, and SB203580 (0.1-1 nmol), a p38 MAPK inhibitor. However, the Ang II type 2 (AT2) receptor antagonist PD123319, the upstream inhibitor of ERK1/2 phosphorylation U0126, and the JNK inhibitor SP600125 had no effect on Ang II-induced nociceptive behavior. Western blot analysis showed that the i.t. injection of Ang II induced phosphorylation of p38 MAPK in the lumbar dorsal spinal cord, which was inhibited by losartan, without affecting ERK1/2 and JNK. Furthermore, we found that AT1 receptor expression was relatively high in the lumbar superficial dorsal horn.
Our data show that i.t. administration of Ang II induces nociceptive behavior accompanied by the activation of p38 MAPK signaling mediated through AT1 receptors. This observation indicates that Ang II may act as a neurotransmitter and/or neuromodulator in the spinal transmission of nociceptive information.
... AngII may act as a sensory neurotransmitter and on up-regulated AT2R in sensory neurons by autocrine, paracrine and systemic mechanisms, particularly in relation to vascular innervation, via peripheral and central nerve terminals (including pre-synaptic nerve terminals), and DRG neuronal cell bodies. In the CNS, AngII and its metabolite AngIII both act at the AT1R and the AT2R (Martens et al., 1996;Zini et al., 1996;Wright et al., 2003;Pelegrini-da-Silva et al., 2005), playing a role in pain mechanisms (see Georgieva and Georgiev, 1999, Pelegrini-Da-Silva et al., 2005, Sakagawa et al., 2000Marques-Lopes et al., 2009, 2010, by the distinct expression of AT1R in neurons involved in descending pain modulation (Marques-Lopes et al., 2009), and the conversion of AngII to AngIII (Pelegrini-da-Silva et al., 2009). However, our study relates to the effects of EMA401 on peripheral mechanisms, since it does not show significant CNS distribution. ...
... AngII may act as a sensory neurotransmitter and on up-regulated AT2R in sensory neurons by autocrine, paracrine and systemic mechanisms, particularly in relation to vascular innervation, via peripheral and central nerve terminals (including pre-synaptic nerve terminals), and DRG neuronal cell bodies. In the CNS, AngII and its metabolite AngIII both act at the AT1R and the AT2R (Martens et al., 1996;Zini et al., 1996;Wright et al., 2003;Pelegrini-da-Silva et al., 2005), playing a role in pain mechanisms (see Georgieva and Georgiev, 1999, Pelegrini-Da-Silva et al., 2005, Sakagawa et al., 2000Marques-Lopes et al., 2009, 2010, by the distinct expression of AT1R in neurons involved in descending pain modulation (Marques-Lopes et al., 2009), and the conversion of AngII to AngIII (Pelegrini-da-Silva et al., 2009). However, our study relates to the effects of EMA401 on peripheral mechanisms, since it does not show significant CNS distribution. ...
Background
The angiotensin II (AngII) receptor subtype 2 (AT2R) is expressed in sensory neurons and may play a role in nociception and neuronal regeneration.
Methods
We used immunostaining with characterized antibodies to study the localization of AT2R in cultured human and rat dorsal root ganglion (DRG) neurons and a range of human tissues. The effects of AngII and AT2R antagonist EMA401 on capsaicin responses in cultured human and rat (DRG) neurons were measured with calcium imaging, on neurite length and density with Gap43 immunostaining, and on cyclic adenosine monophosphate (cAMP) expression using immunofluorescence.
Results
AT2R expression was localized in small-/medium-sized cultured neurons of human and rat DRG. Treatment with the AT2R antagonist EMA401 resulted in dose-related functional inhibition of capsaicin responses (IC50 = 10 nmol/L), which was reversed by 8-bromo-cAMP, and reduced neurite length and density; AngII treatment significantly enhanced capsaicin responses, cAMP levels and neurite outgrowth. The AT1R antagonist losartan had no effect on capsaicin responses. AT2R was localized in sensory neurons of human DRG, and nerve fibres in peripheral nerves, skin, urinary bladder and bowel. A majority sub-population (60%) of small-/medium-diameter neuronal cells were immunopositive in both control post-mortem and avulsion-injured human DRG; some very small neurons appeared to be intensely immunoreactive, with TRPV1 co-localization. While AT2R levels were reduced in human limb peripheral nerve segments proximal to injury, they were preserved in painful neuromas.
Conclusions
AT2R antagonists could be particularly useful in the treatment of chronic pain and hypersensitivity associated with abnormal nerve sprouting.
... Stereotaxy was performed as described above (n 5 5 per group) and to place the guide cannula. Control and antisense vector-injected animals were allowed to recover for 7 days, and BP was determined as described previously (Marques-Lopes et al., 2009). Briefly, with animals under i.p. anesthesia with pentobarbital (30 mg/kg body weight) and after tracheostomy for ventilation, a catheter was inserted into the abdominal aorta through the femoral artery and connected to a pressure transducer (B. ...
... Rectal temperature was maintained at 37–388C. NTS injections were performed as described above, but the needle remained in the same position for at least 20 min before microinjections to allow BP stabilization (Marques-Lopes et al., 2009). One nanomole of NMDA was slowly injected into the NTS using the volume described above. ...
N-methyl-D-aspartate receptors (NMDAR) have a role in cardiovascular control at the nucleus tractus solitarii (NTS), eliciting increases or decreases in blood pressure (BP), depending on the area injected with the agonists. In spite of the association between cardiovascular control and pain modulation, the effects of manipulating NMDAR in pain responses have never been evaluated. In this study, we decreased the expression of NMDAR in the NTS using gene transfer to target receptor subunits and evaluate long-term effects. Seven days after the injection of lentiviral vectors containing the NR1a subunit cDNA of NMDAR, in antisense orientation, into the intermediate NTS of Wistar rats, BP was measured, and the formalin test of nociception was performed. The antisense vector induced a decrease of NR1 expression in the NTS and elicited BP rises and hypoalgesia. Antisense vectors inhibited formalin-evoked c-Fos expression in the spinal cord, indicating decreased nociceptive activity of spinal neurons. Using a time-course approach, we verified that the onset of both the increases in BP and the hypoalgesia was at 4 days after vector injection into the NTS. The injection of NMDA into the NTS reversed the effects of antisense vectors in pain behavioral responses and spinal neuronal activation and decreased BP and heart rate. The present study shows that the NR1 subunit of the NMDAR at the NTS is critical in the regulation of tonic cardiovascular and nociceptive control and shows an involvement of the nucleus in the modulation of sustained pain.
... In addition to the spinal cord, Ang II increases pain sensitivity in the caudal ventrolateral medulla [41] and decreases pain sensitivity in the rostral ventromedial medulla and periaqueductal gray and [42][43][44]. Although these results indicate that Ang II contributes to pain modulation at the supraspinal level in the central nervous system, further studies are required to identify the underlying mechanisms. ...
The present study examined the underlying mechanisms of mechanical allodynia and thermal hyperalgesia induced by the intracisternal injection of angiotensin (Ang) II. Intracisternal Ang II injection decreased the air puff threshold and head withdrawal latency. To determine the operative receptors for each distinct type of pain behavior, we intracisternally injected Ang II receptor antagonists 2 h after Ang II injection. Losartan, an Ang II type 1 receptor (AT1R) antagonist, alleviated mechanical allodynia. Conversely, PD123319, an Ang II type 1 receptor (AT2R) antagonist, blocked only thermal hyperalgesia. Immunofluorescence analyses revealed the co-localization of AT1R with the astrocyte marker GFAP in the trigeminal subnucleus caudalis and co-localization of AT2R with CGRP-positive neurons in the trigeminal ganglion. Intracisternal pretreatment with minocycline, a microglial inhibitor, did not affect Ang II-induced mechanical allodynia, whereas L-α-aminoadipate, an astrocyte inhibitor, significantly inhibited Ang II-induced mechanical allodynia. Furthermore, subcutaneous pretreatment with botulinum toxin type A significantly alleviated Ang II-induced thermal hyperalgesia, but not Ang II-induced mechanical allodynia. These results indicate that central Ang II-induced nociception is differentially regulated by AT1R and AT2R. Thus, distinct therapeutic targets must be regulated to overcome pain symptoms caused by multiple underlying mechanisms.
... Ang-related peptide-mediated AT1 receptor activation is known to have opposing effects on pain depending on the brain region. For instance, microinjection of Ang II into the caudal ventrolateral medulla (CVLM), a supraspinal region that regulates pain, induces hyperalgesia via AT1 receptors [14]. In the CVLM, only 3% of AT1 receptor-positive neurons projected into the spinal cord, indicating that this pain response was not a direct neural pathway between the CVLM and spinal cord. ...
Simple Summary
Evidence indicates that angiotensin (Ang)-related peptides, also known as hypertensive peptides, are involved in pain regulation. Ang-related peptides exhibit various physiological effects via their receptors expressed throughout the body, and changes in the expression of Ang system components at the site of a lesion affect the local inflammatory response and pain transmission. Therefore, an extensive understanding of the pain modulatory mechanisms of Ang-related peptides and their receptors in several tissues and organs will aid in the development of drug therapies targeting the Ang system. This review article focuses on the current evidence regarding the mechanisms of pain regulation by Ang-related peptides in the central and peripheral regions involved in pain transmission.
Abstract
Angiotensin (Ang)-generating system has been confirmed to play an important role in the regulation of fluid balance and blood pressure and is essential for the maintenance of biological functions. Ang-related peptides and their receptors are found throughout the body and exhibit diverse physiological effects. Accordingly, elucidating novel physiological roles of Ang-generating system has attracted considerable research attention worldwide. Ang-generating system consists of the classical Ang-converting enzyme (ACE)/Ang II/AT1 or AT2 receptor axis and the ACE2/Ang (1–7)/MAS1 receptor axis, which negatively regulates AT1 receptor-mediated responses. These Ang system components are expressed in various tissues and organs, forming a local Ang-generating system. Recent findings indicate that changes in the expression of Ang system components under pathological conditions are involved in the development of neuropathy, inflammation, and their associated pain. Here, we summarized the effects of changes in the Ang system on pain transmission in various organs and tissues involved in pain development process.
... This hyperalgesia was significantly attenuated by local administration of losartan (AT1R antagonist; Tables 1 and 2). 79 These results suggest that Ang II may exert pronociceptive effects in components of the supraspinal pain control system. However, Parlo et al. 101 showed that Ang I, II, or III injected into rat periaqueductal gray induced dose-dependent antinociception in the tail-flick test. ...
Growing evidence implicates the renin–angiotensin system (RAS) in multiple facets of neuropathic pain (NP). This narrative review focuses primarily on the major bioactive RAS peptide, Angiotensin II (Ang II), and its receptors, namely type 1 (AT1R) and type 2 (AT2R). Both receptors are involved in the development of NP and represent potential therapeutic targets. We first discuss the potential role of Ang II receptors in modulation of NP in the central nervous system. Ang II receptor expression is widespread in circuits associated with the perception and modulation of pain, but more studies are required to fully characterize receptor distribution, downstream signaling, and therapeutic potential of targeting the central nervous system RAS in NP. We then describe the peripheral neuronal and nonneuronal distribution of the RAS, and its contribution to NP. Other RAS modulators (such as Ang (1-7)) are briefly reviewed as well. AT1R antagonists are analgesic across different pain models, including NP. Several studies show neuronal protection and outgrowth downstream of AT2R activation, which may lead to the use of AT2R agonists in NP. However, blockade of AT2R results in analgesia. Furthermore, expression of the RAS in the immune system and a growing appreciation of neuroimmune crosstalk in NP add another layer of complexity and therapeutic potential of targeting this pathway. A growing number of human studies also hint at the analgesic potential of targeting Ang II signaling. Altogether, Ang II receptor signaling represents a promising, far-reaching, and novel strategy to treat NP.
... 63 In addition, sensory information, learning, memory and emotional responses have been linked to different components of RAAS. 63 Angiotensin II mediated hyperalgesia has been experimentally documented 64 while the pain attenuating properties of RAAS via AT-II receptors 65 and endogenous opioids 66 have also been described. This might be one reason for myalgia reported by a third of COVID-19 patients. ...
... 76 In addition, sensory information, learning, memory and emotional responses have been linked to different components of RAAS. 76 Angiotensin II mediated hyperalgesia has been experimentally documented 77 while the pain attenuating properties of RAAS via AT-II receptors 78 and endogenous opioids 79 have also been described. This might be one reason for myalgia reported by a third of COVID-19 patients. ...
Coronavirus 2019 disease (COVID-19) was first recognised as an acute respiratory syndrome. Soon, it was realised that acute kidney injury, myocarditis, hyperimmune activation and prothrombotic disseminated intravascular coagulation are also part of the disease complex. Currently, it is being recognised as causing microthrombi as well as thrombo-embolic complications. The SARS-CoV-2 coronavirus enters cells via the ACE2 (Angiotensin converting enzyme-2) receptor and leads to its downregulation. The loss of ACE2 is postulated to cause activation of the Renin Angiotensin Aldosterone System (RAAS) that contributes to cardiac and renal pathology. Supporting the hypothesis that ACE2 loss can explain the entire pathology, we have reviewed the available literature to determine mechanistic associations between RAAS and the various manifestations of COVID-19.
ACE2 dependent protective pathways include the Angiotensin(1-7)/Mas receptor and Alamandine/MrgD receptor pathways. These are anti-hypertensive, anti-inflammatory, and anti-thrombotic. Loss of ACE2 leads to bradykinin and Angiotensin II accumulation that causes thrombosis via release of tissue Plasminogen Activator(tPA). Angiotensin II, with altered ACE/ACE2 balance, can activate immune cells. Animal models of various viral pneumonia have previously shown that ACE2 loss is associated with severe lung injury. Angiotensin II and aldosterone are known causes of cardiomyopathy and also potentiate acute kidney injury. There is some emerging evidence that inhibitors of canonical RAAS such as ACE inhibitors, Angiotensin receptor blockers, and statins may have mortality benefit in COVID-19. Heparin inhibits both thrombosis and RAAS, and has definite mortality benefit at least in a subset of COVID-19 patients. We strongly advocate that treatment strategies for COVID-19 should include synergistic targeting of the RAAS system.
... There is evidence indicating that Ang II is involved in the nociception. It was found that Ang II injected in the caudal ventrolateral medulla causes hyperalgesia [ 5 ], administered in the periaqueductal gray and the rostral ventromedial medulla produced hypoalgesia [ 6 ]. ...
The effects of angiotensin II (Ang II) and losartan (antagonist of Ang II type I receptors-AT1) infused unilaterally or bilaterally into the central nucleus of the amygdala (CeA) on nociception of male Wistar rats with a model of depression (bilateral olfactory bulbectomy-OBX) were investigated. Nociception was examined by applying mechanical pressure on the paw of the rat (analgesy-meter). The OBX rats were divided into three groups: 1st group-bilaterally microinjected into CeA with Ang II (0.5 µg), losartan (100 µg,) or saline; 2nd group-unilaterally (left-or right-side) microinjected into CeA with Ang II (0.5 µg), losartan (100 µg,) or saline, and 3rd group-sham-operated rats. It was found that Ang II microinjected bilaterally and into right-side CeA decreased the pain threshold (nociceptive effect). The inhibition of AT1 receptors by losartan microinjected uni-or bilaterally into CeA of bulbectomized rats, increased pain threshold (antinociceptive effect) compared to the respective OBX controls; the antinociceptive effect was more pronounced in the right-side. This study for the first time provides information on a lateralized nociceptive effect of Ang II and antinociceptive effect of losartan and involvement of AT1 receptors in nociception of rats with a model of depression. These findings point to the predominant involvement of the right CeA in nociceptive responses of OBX rats, suggesting an asymmetry of the Ang II-connected behaviour responses of the left or right CeA and a different distribution of AT1 receptors in the two hemispheres. Introduction. The octapeptide angiotensin II (Ang II) is a main bioactive component of the renin-angiotensin system (RAS). Ang II mediates its biological effects through Ang II type 1 (AT1) receptors and Ang II type 2 (AT2) receptors, which are seven transmembrane G-protein coupled receptors [ 1 ]. The brain RAS is characterised by the presence of all RAS components, including angiotensinogen, renin, ACE, Ang I, Ang II, and Ang II receptors [ 2 ]. The AT1 and AT2 receptors have a wide distribution in different brain structures, including the amygdala, hippocampus, lateral septum, and frontal cortex [ 3 ]. It is known that angiotensin modulates functions such as stress, exploratory behaviour, anxiety, learning and memory [ 4 ]. There is evidence indicating that Ang II is involved in the nociception. It was found that Ang II injected in the caudal ventrolateral medulla causes hyperalgesia [ 5 ], administered in the periaque-ductal gray and the rostral ventromedial medulla produced hypoalgesia [ 6 ]. Previously, we have found that Ang II microinjected bilaterally and into the left hippocampal CA1 area exerted a nociceptive effect. In contrast, the inhibition of AT1 receptors by losartan, microinjected bilaterally and into the left hip-pocampal CA1 area, produced an antinociceptive effect [ 7 ]. Takai et al. [ 8 ] have shown that AT1 receptor antagonists and ACE inhibitors administrated orally exert antinociceptive effect in a hot-plate test. The bilateral olfactory bulbectomy (OBX) is an animal model of depression. Removal of the olfactory bulbs in rats causes dysfunction of the cortical-hippocampal-amygdala circuit and produces a complex constellation of behaviou-ral, neurochemical, neuroendocrine, and neuroimmune alterations, many of which reflect symptoms reported in patients with major depression. Behavioural abnormalities of OBX rats include exploratory hyperactivity in response to novel environmental stress, a decrease of libido, memory deficits, deficits in appetite-motivated behaviours, anxiety symptom-resembling behaviour in social interaction and plus-maze tests, etc. [ 9 ]. An analgesic-like effect (decreased pain threshold) has been demonstrated in bulbectomized rats (analgesy-meter test) [ 10 ]. The amygdala is a brain structure classically related to emotional states. Central nucleus of the amygdala (CeA) is one of the most important regulatory centres for the behavioural responses to stress. In the last years increasing anatomical, behavioural, and physiological studies support the role of the CeA as a neural modulator of pain perception [ 11 ]. The CeA receives multiple nociceptive information from the brainstem, as well as highly processed polymodal information from the thalamus and the cerebral cortex. It also possesses the connections that allow influencing most of the descending pain control systems as well as higher centres involved in emotional, affective and cognitive functions. Among the different neurotransmitter systems present in this nucleus, the CeA also contains Ang II, angiotensin converting enzyme, and AT1 receptors [ 12 ] but their role in nociception is not studied.
... The inherent pharmacological consequences of such an integrated mechanism of action are perhaps most clearly seen with Ang-II associated analgesia. Chronic administration of AT1R antagonists has been shown to ameliorate migraine and neuropathic and nociceptive pain (Marques-Lopes et al. 2009;Bali et al. 2014), whereas selective AT2R antagonists have been proposed as a treatment for chronic inflammatory pain and peripheral neuropathy Smith and Muralidharan 2015;Shepherd and Mohapatra 2018). It remains unclear whether the beneficial effects of RAS blockers in models of neuropathic pain are mediated directly through action on sensory pathways, or indirectly through their attenuating effects on pro-inflammatory mediators. ...
Neuropathic and inflammatory pain results from cellular and molecular changes in dorsal root ganglion (DRG) neurons. The type‐2 receptor for Angiotensin‐II (AT2R) has been involved in this type of pain. However, the underlying mechanisms are poorly understood, including the role of the type‐1 receptor for Angiotensin‐II (AT1R). Here, we used a combination of immunohistochemistry and immunocytochemistry, RT‐PCR and in vitro and in vivo pharmacological manipulation to examine how cutaneous inflammation affected the expression of AT1R and AT2R in subpopulations of rat DRG neurons and studied their impact on inflammation‐induced neuritogenesis. We demonstrated that AT2R‐neurons express C‐ or A‐neuron markers, primarily IB4, trkA, and substance‐P. AT1R expression was highest in small neurons and co‐localized significantly with AT2R. In vitro, an inflammatory soup caused significant elevation of AT2R mRNA, whereas AT1R mRNA levels remained unchanged. In vivo, we found a unique pattern of change in the expression of AT1R and AT2R after cutaneous inflammation. AT2R increased in small neurons at 1 day and in medium size neurons at 4 days. Interestingly, cutaneous inflammation increased AT1R levels only in large neurons at 4 days. We found that in vitro and in vivo AT1R and AT2R acted co‐operatively to regulate DRG neurite outgrowth. In vivo, AT2R inhibition impacted more on non‐peptidergic C‐neurons neuritogenesis, whereas AT1R blockade affected primarily peptidergic nerve terminals. Thus, cutaneous‐induced inflammation regulated AT1R and AT2R expression and function in different DRG neuronal subpopulations at different times. These findings must be considered when targeting AT1R and AT2R to treat chronic inflammatory pain.
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... Angiotensin II, in fact, can modulate neuronal transmission within the central nervous system; type 1 receptors have been demonstrated in spinal cord, dorsal root ganglia and sciatic nerve of the rat [15]. Moreover, other works have shown that angiotensin II can play different roles in the modulation of pain transmission within the central nervous system; in fact, it can have hyperalgesic effects when injected into the caudal ventrolateral medulla or intrathecal [16,17], or it can have hypoalgesic effects when injected into the periacqueductal grey or the rostral ventrolateral medulla [6,7,18,19]. Interestingly, an AT1RA can block the intrathecal-hyperalgesic effect of angiotensin II; moreover, it has also been shown that in mice the oral treatment with several ACE inhibitors or AT1RA resulted in hypoalgesia [7]. ...
Background: Auto-algometry has been previously proposed as a toll to evaluate hypertension-associated hypoalgesia. The aim of the present work was to confirm the robustness of this method and to evaluate other possible variables associated with hypertension and hypoalgesia.
Methods: All routinary ambulatory data were collected from 111 hypertensive subjects along with the evaluation of pain threshold assessed with the auto-algometer.
Results: The results confirmed the sensibility of the auto-algometer in revealing the hypertension-associated hypoalgesia and revealed unexpected higher pain thresholds in patients consuming angiotensin receptor 1 antagonists compared to patients consuming other medications.
Conclusion: The results encourage the use of the auto-algometer to study unexplored mechanisms involved in hypertension and in hypoalgesia.
... Decreased activation of angiotensin type 1 receptors in the caudal ventrolateral medulla reduces hyperalgesia and likely serves a role in descending pain modulation and nociception [14]. The renin angiotensin system plays a significant role in inflammation and oxidative stress via the production of nitric oxide. ...
Opinion statement:
Angiotensin converting enzyme (ACE) inhibitors and angiotensin receptor blockers (ARBs) are antihypertensive agents that can be considered for migraine preventative therapy. Although the exact mechanisms by which ACE inhibitors and ARBs may work for migraine prophylaxis are unknown, there are several plausible hypotheses as to why modulating the activity of the renin angiotensin system could result in migraine prevention. Clinical trials of ACE inhibitors and ARBs provide evidence that they are effective and generally well tolerated when used for migraine prophylaxis. Based upon biologic plausibility, the quality of evidence for efficacy from clinical trials, and recommendations in published guidelines, we consider ACE inhibitors and ARBs as second- or third-line options for migraine prophylaxis.
... The involvement of the RAS in pain modulation in the supra-spinal areas is mainly due to AT 1 -expressing CVLM neurons projecting to the spinal cord. 31 The same group of scientists demonstrated that AngII-induced hyperalgesia, elicited from the CVLM, is mediated to the spinal cord by an indirect pathway relayed at the pontine noradrenergic A(5)group. 32 A very recent study has demonstrated that the systemic continuous delivery of sub-pressor doses of AngII (150 ng/kg/min) induces pain in terms of tactile and cold hyperalgesia in chronic constriction injury (CCI)-subjected animals which was shown to be attenuated with losartan. ...
Apart from the well-documented role of the renin-angiotensin-aldosterone system (RAAS) in regulating the blood pressure and other related parameters, its role in modulating different physiological/pathological functions, including pain, has also been described. Like its dual role in regulating stress-related anxiety and cognitive functions, its dual role has also been documented in pain modulation in different disease states. Drugs blocking the RAAS activation, viz., renin inhibitors, angiotensin converting enzyme (ACE) inhibitors, AT1 receptor antagonists and aldosterone antagonists, have been shown to produce beneficial effects in migraine and neuropathic and nociceptive pain. Their beneficial effects have been mainly attributed to inhibition of the inflammatory cascade of reactions by inhibiting the generation of key cytokines, including tumor necrosis factor (TNF)-α. On the contrary, clinical as well as preclinical studies have also shown the pain-inducing actions of renin-angiotensin system (RAS) blocking drugs. Furthermore, the pain-relieving actions of angiotensin II (AngII) and pain-inducing actions of AT1 blockers have also been described. The pain-inducing actions of ACE inhibitors have been mainly attributed to interference with metabolism of bradykinin and substance P, while the analgesic actions of AngII have been mainly related to activation of brain localized AT2 receptors and release of endogenous opioids. The present review describes the dual role of the RAAS in different states of pain.
© The Author(s) 2013.
... Angiotensin related peptides have been posited as neurotransmitters in the periaqueductal gray 55 and other pain relevant brain areas, where they modulate pro-and anti-nociceptive pathways. 10,45 In addition to angiotensin-I, endogenous peptide substrates of ACE include bradykinin, substance P, and opioids such as dynorphin and enkephalin, 20,72 suggesting the angiotensin system plays a pivotal role underlying the relationship between blood pressure and pain sensitivity. Pharmacological inhibition of ACE has been associated with increase in nociceptive thresholds and tolerance 30 and risk of complex regional pain syndrome (CRPS), 13 suggesting lower levels of ACE activity due to genetic variation might underlie altered somatic sensitivity. ...
Genetic risk factors are believed to combine with environmental exposures and contribute to the risk of developing temporomandibular disorder (TMD). In this prospective cohort study, 2,737 people without TMD were assessed for common genetic variation in 358 genes known to contribute to nociceptive pathways, inflammation, and affective distress. During a median follow-up period of 2.8 years, 260 people developed first-onset TMD. Hazard ratios were computed as measures of association between 2,924 single-nucleotide polymorphisms and TMD incidence. After correction for multiple testing, no single single-nucleotide polymorphism was significantly associated with risk of onset TMD. However, several single-nucleotide polymorphisms exceeded Bonferroni correction for multiple comparison or false discovery rate thresholds (.05, .1, or .2) for association with intermediate phenotypes shown to be predictive of TMD onset. Nonspecific orofacial symptoms were associated with voltage-gated sodium channel, type I, alpha subunit (SCN1A, rs6432860, P = 2.77 × 10−5) and angiotensin I–converting enzyme 2 (ACE2, rs1514280, P = 4.86 × 10−5); global psychological symptoms with prostaglandin-endoperoxide synthase 1 (PTGS1, rs3842803, P = 2.79 × 10−6); stress and negative affectivity with amyloid-β (A4) precursor protein (APP, rs466448, P = 4.29 × 10−5); and heat pain temporal summation with multiple PDZ domain protein (MPDZ, rs10809907, P = 3.05 × 10−5). The use of intermediate phenotypes for complex pain diseases revealed new genetic pathways influencing risk of TMD.
Perspective
This article reports the findings of a large candidate gene association study of first-onset TMD and related intermediate phenotypes in the OPPERA Study. Although no genetic markers predicted TMD onset, several genetic risk factors for clinical, psychological, and sensory phenotypes associated with TMD onset were observed.
... Our approach, the determination of pain threshold in combination with measurement of withdrawal latencies at pain threshold temperature, seems to be more reliable method for study of thermonociception in conscious animals. Recently, it was shown that activation of AT 1 receptors by local Ang II administration into components of the supraspinal pain modulatory system elicited hyperalgesia, and this effect was significantly attenuated by local administration of the AT 1 receptor antagonist, losartan (Marques-Lopes et al. 2009). Similarly, our findings showed a marked elevation of pain perception (hyperalgesia) after systemic Ang II administration, demonstrated by von Frey paw withdrawal threshold, and by measurements of withdrawal latencies at noxious heat and cold temperatures. ...
To clarify the role of angiotensin II (Ang II) in the regulation of sensory signaling, we studied the effect of subpressor dose (150 ng/kg/min) of Ang II on pain-related behavior in relation with neuronal injury and activation of satellite glial cells (SGCs) in the dorsal root ganglia (DRGs) after chronic constriction injury (CCI). Systemic continuous delivery of Ang II induced the tactile, heat and cold hyperlagesia, when measured at 7 days ofpost-injury. Blockade of the AT1 receptor with losartan (2.5 mg/kg/day) prevented tactile hyperalgesia and attenuated cold hyperalgesia, but did not affect the response to noxious heat stimulus. A marked increase of large-sized injured primary afferent neurons, detected by ATF3 immunolabeling, was seen in lower lumbar DRGs on ipsilateral side after Ang II treatment. Subpressor dose of Ang II induced an increase of activated SGCs (detected by GFAP immunolabeling) enveloping large-diameter neurons. Our results suggested that Ang II through the AT1 receptor activation is an important regulatory factor in neuropathic pain perception and plays an important role in the injury of large-sized primary afferent neurons and activation of SGCs elicited by the CCI.
... This network connected 12 of our identified genes into a causal network (Figure 3). Angiotensin II has been already been implicated in central pain: it has been shown to facilitate pain-related behaviours in experimental animals [31] including responses to thermal stimuli similar to those employed in the current studies. The mechanism appears to be via the modulation of descending brainstem pathways. ...
Sensitivity to pain varies considerably between individuals and is known to be heritable. Increased sensitivity to experimental pain is a risk factor for developing chronic pain, a common and debilitating but poorly understood symptom. To understand mechanisms underlying pain sensitivity and to search for rare gene variants (MAF<5%) influencing pain sensitivity, we explored the genetic variation in individuals' responses to experimental pain. Quantitative sensory testing to heat pain was performed in 2,500 volunteers from TwinsUK (TUK): exome sequencing to a depth of 70× was carried out on DNA from singletons at the high and low ends of the heat pain sensitivity distribution in two separate subsamples. Thus in TUK1, 101 pain-sensitive and 102 pain-insensitive were examined, while in TUK2 there were 114 and 96 individuals respectively. A combination of methods was used to test the association between rare variants and pain sensitivity, and the function of the genes identified was explored using network analysis. Using causal reasoning analysis on the genes with different patterns of SNVs by pain sensitivity status, we observed a significant enrichment of variants in genes of the angiotensin pathway (Bonferroni corrected p = 3.8×10(-4)). This pathway is already implicated in animal models and human studies of pain, supporting the notion that it may provide fruitful new targets in pain management. The approach of sequencing extreme exome variation in normal individuals has provided important insights into gene networks mediating pain sensitivity in humans and will be applicable to other common complex traits.
... A significant number of CPA neurons were activated and immunolabeled with Fos after underwater submersion compared to control and swimming rats in the present study. Neurons in and around the CPA have been implicated in cardiovascular regulation, in the processing of noxious information, as well as important for the exercise pressure response Panneton, 2002, 2005;Panneton et al., 2008Panneton et al., , 2011Marques-Lopes et al., 2009;Takakura et al., 2011a,b). Although we have shown previously that the area of the MDH activated by voluntary diving projects to the CPA (Panneton et al., 2006), it plays no role in the cardiorespiratory adjustments seen with nasal stimulation (Panneton et al., 2008). ...
The mammalian diving response is a powerful autonomic adjustment to underwater submersion greatly affecting heart rate, arterial blood pressure, and ventilation. The bradycardia is mediated by the parasympathetic nervous system, arterial blood pressure is mediated via the sympathetic system and still other circuits mediate the respiratory changes. In the present study we investigate the cardiorespiratory responses and the brainstem neurons activated by voluntary diving of trained rats, and, compare them to control and swimming animals which did not dive. We show that the bradycardia and increase in arterial blood pressure induced by diving were significantly different than that induced by swimming. Neuronal activation was calculated after immunohistochemical processing of brainstem sections for Fos protein. Labeled neurons were counted in the caudal pressor area, the medullary dorsal horn, subnuclei of the nucleus tractus solitarii (NTS), the nucleus raphe pallidus (RPa), the rostroventrolateral medulla, the A5 area, the nucleus locus coeruleus, the Kölliker–Fuse area, and the external lateral and superior lateral subnuclei of the parabrachial nucleus. All these areas showed significant increases in Fos labeling when data from voluntary diving rats were compared to control rats and all but the commissural subnucleus of the NTS, A5 area, and RPa were significantly different from swimming rats. These data provide a substrate for more precise experiments to determine the role of these nuclei in the reflex circuits driving the diving response.
... Although the functions of Ang II in the DRG system have not been extensively studied and therefore are not fully clarified, our results are not without clinical interest. Ang II has been proposed to be involved in the regulation of sensory information, and in particular nociception [24,36,48,49,61]. Hence, we attempted to determine whether Ang II was associated with other nociceptive-controlling neuropeptides in the DRG. ...
To elucidate the local formation of angiotensin II (Ang II) in the neurons of sensory dorsal root ganglia (DRG), we studied the expression of angiotensinogen (Ang-N)-, renin-, angiotensin converting enzyme (ACE)- and cathepsin D-mRNA, and the presence of protein renin, Ang II, Substance P and calcitonin gene-related peptide (CGRP) in the rat and human thoracic DRG. Quantitative real time PCR (qRT-PCR) studies revealed that rat DRG expressed substantial amounts of Ang-N- and ACE mRNA, while renin mRNA as well as the protein renin were untraceable. Cathepsin D-mRNA and cathepsin D-protein were detected in the rat DRG indicating the possibility of existence of pathways alternative to renin for Ang I formation. Angiotensin peptides were successfully detected with high performance liquid chromatography and radioimmunoassay in human DRG extracts. In situ hybridization in rat DRG confirmed additionally expression of Ang-N mRNA in the cytoplasm of numerous neurons. Intracellular Ang II staining could be shown in number of neurons and their processes in both the rat and human DRG. Interestingly we observed neuronal processes with angiotensinergic synapses en passant, colocalized with synaptophysin, within the DRG. In the DRG, we also identified by qRT-PCR, expression of Ang II receptor AT(1A) and AT(2)-mRNA while AT(1B)-mRNA was not traceable. In some neurons Substance P and CGRP were found colocalized with Ang II. The intracellular localization and colocalization of Ang II with Substance P and CGRP in the DRG neurons may indicate a participation and function of Ang II in the regulation of nociception. In conclusion, these results suggest that Ang II may be produced locally in the neurons of rat and human DRG and act as a neurotransmitter.
Enteric glial cells (EGCs) play an important role in visceral hypersensitivity associated with irritable bowel syndrome (IBS). Losartan (Los) is known to reduce pain; however, its function in IBS is unclear. The present study aimed to investigate Los's therapeutic effect on visceral hypersensitivity in IBS rats. Thirty rats were randomly divided into control, acetic acid enema (AA), AA + Los low, medium and high dose groups in vivo. EGCs were treated with lipopolysaccharide (LPS) and Los in vitro. The molecular mechanisms were explored by assessing the expression of EGC activation markers, pain mediators, inflammatory factors and angiotensin-converting enzyme 1(ACE1)/angiotensin II (Ang II)/Ang II type 1 (AT1) receptor axis molecules in colon tissue and EGCs. The results showed that the rats in the AA group showed significantly higher visceral hypersensitivity than the control rats, which was alleviated by different doses of Los. The expression of GFAP, S100β, substance P (SP), calcitonin gene-related peptide (CGRP), transient receptor potential vanilloid 1 (TRPV1), tumor necrosis factor (TNF), interleukin-1β (IL-1β) and interleukin-6 (IL-6) was considerably increased in colonic tissues of AA group rats and LPS-treated EGCs compared with control rats and EGCs, and reduced by Los. In addition, Los reversed ACE1/Ang II/AT1 receptor axis upregulation in AA colon tissues and LPS-treated EGCs. These results show that Los inhibits ACE1/Ang II/AT1 receptor axis upregulation by suppressing EGC activation, resulting in reduced expression of pain mediators and inflammatory factors, thereby alleviating visceral hypersensitivity.
In many instances, the perception of pain is disproportionate to the strength of the algesic stimulus. Excessive or inadequate pain sensation is frequently observed in cardiovascular diseases, especially in coronary ischemia. The mechanisms responsible for individual differences in the perception of cardiovascular pain are not well recognized. Cardiovascular disorders may provoke pain in multiple ways engaging molecules released locally in the heart due to tissue ischemia, inflammation or cellular stress, and through neurogenic and endocrine mechanisms brought into action by hemodynamic disturbances. Cardiovascular neuropeptides, namely angiotensin II (Ang II), angiotensin-(1-7) [Ang-(1-7)], vasopressin, oxytocin, and orexins belong to this group. Although participation of these peptides in the regulation of circulation and pain has been firmly established, their mutual interaction in the regulation of pain in cardiovascular diseases has not been profoundly analyzed. In the present review we discuss the regulation of the release, and mechanisms of the central and systemic actions of these peptides on the cardiovascular system in the context of their central and peripheral nociceptive (Ang II) and antinociceptive [Ang-(1-7), vasopressin, oxytocin, orexins] properties. We also consider the possibility that they may play a significant role in the modulation of pain in cardiovascular diseases. The rationale for focusing attention on these very compounds was based on the following premises (1) cardiovascular disturbances influence the release of these peptides (2) they regulate vascular tone and cardiac function and can influence the intensity of ischemia – the factor initiating pain signals in the cardiovascular system, (3) they differentially modulate nociception through peripheral and central mechanisms, and their effect strongly depends on specific receptors and site of action. Accordingly, an altered release of these peptides and/or pharmacological blockade of their receptors may have a significant but different impact on individual sensation of pain and comfort of an individual patient.
Migraine is a common neurologic condition marked by recurrent episodes of headache. Its pathophysiology is highly complex involving neuronal, inflammatory and vascular mechanisms. The Renin-Angiotensin System (RAS) can modulate all these mechanism, being a potential pharmacological target for migraine treatment. We carried out a systematic review of the studies evaluating the involvement of RAS in patients with migraine. There is evidence from genetic studies exploring the relation between migraine and RAS-related genes and from clinical trials evaluating the efficacy of Angiotensin II Type 1 (AT1) receptor antagonists and angiotensin converting enzyme inhibitors in migraine prophylaxis. RAS seems to play a role in the pathophysiology of migraine, but more direct evidence is still missing.
Angiotensin (Ang) II, the main bioactive component of the renin-angiotensin system, is reported to participate in either the inhibition or the facilitation of pain transmission depending on brain area. Although Ang II is known to exist in the superficial dorsal horn of the spinal cord, the role of Ang II in spinal pain transmission remains unclear. In order to elucidate that role, we examined the effect of intrathecal administration of Ang II to mice. We found that Ang II produced pain-related behavior accompanied by the phosphorylation of p38 mitogen-activated protein kinase (MAPK) via Ang II type 1 (AT1) receptors, which are highly expressed in the superficial dorsal horn (laminae I and II). In addition, pain-related behavior was caused by acting on spinal neurons and astrocytes that express AT1 receptors. We next examined whether the spinal Ang II system is responsible for diabetic neuropathic pain. In streptozotocin-induced diabetic mice, neuronal Ang-converting enzyme was upregulated in the superficial dorsal horn, which led to an increase in spinal Ang II levels. Furthermore, this increase in Ang II caused mechanical hypersensitivity accompanied by the activation of p38 MAPK via AT1 receptors. In conclusion, our findings suggest that Ang II may act as a neurotransmitter and/or neuromodulator in spinal pain transmission. This review describes our recent efforts regarding the role of spinal Ang II in spinal pain transmission.
Elevated blood pressure (BP) and headache have long been linked in the medical literature. Headache associated with arterial hypertension is a main concern in emergency department. It is believed that headache may be a symptom attributed to arterial hypertension only if the BP values are very high or rise quickly. Many studies support the hypothesis that migraine patients have an increased risk of developing hypertension, while hypertensive subjects do not seem to have an increased risk of migraine or other types of headache. Conversely many studies found an inverse association. Hypertension has been identified as one of the most important factors of chronic transformation of episodic migraine and increases the cerebrovascular and cardiovascular risk of migraine patients. Migraine and arterial hypertension may share common mechanisms like endothelial dysfunction, deficiency of autonomic cardiovascular regulation and renin angiotensin system involvement. Preventive effects of migraine were described by several antihypertensive agents traditionally beta-blockers, and more recently angiotensin-converting-enzyme inhibitors and angiotensin II receptor blockers.
Lomerizine, calcium channel blocker, is the most used medication for migraine prophylaxis in Japan. The effectiveness of this drug is reported as 50–75%. Telmisartan is angiotensin II receptor blockers which plasma half-life is 24 h. We examined whether telmisartan has preventative benefits in lomerizine non-responsive migraineurs. Lomerizine non-responders received telmisartan (20 mg/day) for 3 months after the investigation period of 3 months. Blood pressure, frequency of headache days/month, headache severity, and doses of triptans and analgesics were analyzed by Wilcoxon signed rank test. Thirty-three migraineurs (25 women and 8 men) participated in this study. Seven patients had migraine with aura and 26 patients had migraine without aura. Mean age (SD) was 46.6 (10.3) years. Mean duration (SD) of migraine was 20.4 (12.5) years. Headache severity exhibited mild degree in 5 patients, moderate degree in 9 patients and severe degree in 19 patients. Mean frequency (SD) of headache days was 10.9 (8.5) days/month. Mean usage (SD) of triptans was 4.8 (5.1) tablets/month and that of analgesics was 15.2 (22.2) tablets/month. Five patients (15%) had hypertension. Telmisartan administration had benefits in 30 patients (90%). This medication significantly decreased frequency of headache days (P < 0.01) and headache severity (P < 0.01). Doses of triptans were reduced at one-third (P < 0.05) and those of analgesia at one-fifth after telmisartan treatment (P < 0.01). After telmisartan, mean (SD) of systolic blood pressure was significantly decreased (P < 0.05). The present study supported that telmisartan treatment had preventive effects in 90% of lomerizine non-responders. Telmisartan non-responders (10%) exhibited chronic migraine and long migraine duration.
Background and Objectives: To investigate long term use of ramipril, and losartan on pain. Methods: Vehicle (1% CMC), ramipril (0.125mg/kg and 0.25mg/kg) and losartan (12.5mg/kg and 25mg/kg) were given orally for 21 days. The tail flick test on Wistar rats and hot plate test on swiss albino mice were conducted on day 7, 14 and 21 whereas writhing test was performed on swiss albino mice at the end of study period. Results: Losartan (25mg/kg) but not ramipril significantly increased the tail flick latency likewise it significantly increased the time of onset of writhing and also reduced the number of writhes in the mice. losartan (25mg/kg) showed a tendency to increase the basal reaction time at 21st day of treatment in the hot plate test in mice. Interpretation & Conclusion: Long term administration of losartan demonstrated significant analgesic effects in the tail flick test and writhing test. losartan can be used as adjuvant for pain relief.
Background Headache is the most frequent and typical ma-nifestation of arterial hypertension (HTN) but the prevalence of pain in other locations and its detailed characteristics in patients with HTN remain unknown. Furthermore, the influence of pain on the effectiveness of the antihypertensive treatment has never been comprehensively studied. The aim of the study was to analyze the prevalence and characteristics of pain and relation to blood pressure control and the effect of antihypertensive treatment on pain in patients with HTN. Material and methods 122 patients with the diagnosis of primary HTN (70 women and 57 men; age 55.7 ± 15.6 years) filled the modified McGill Melzack Pain Questionnaire. Mean time from HTN diagnosis was 12.9 ± 10.9 years. All patients were treated for hypertension receiving a mean number of 2.5 ± 1.5 blood pressure-lowering medications. 27% of the patients fulfilled the criteria of resistant hypertension. Mean arterial blood pressure from the last three measurements was 141/84 mm Hg . Results As many as 87% of the patients with HTN reported pain in any location. The most common pain locations were lower limbs - shanks, knees, feet (60%) and the head (52%). The most prevalent pain characteristics were neuromuscular (36%), dull (25%) and paroxysmal (47%). Pain was associated with body movements in 64% of the patients. Mean pain intensity in a 10-point scale was 5.9 ± 2.2. The intensity of pain did not correlate with the number of antihypertensive drugs. In a group of patients with well-controlled blood pressure (<140/90 mm Hg ) the intensity of the pain tended to be lower than in patients with poorly controlled HTN (5.6 ± 2.5 vs. 6.3 ± 2.3; respectively; p = 0.02). Only 56% of the patients declaring pain were using pain-relief drugs. The number of pain localizations was higher in patients treated with renin-angiotensin- aldosterone system inhibitors (2.7 ± 1.7 vs. 2.2 ± 1.1) and in patients with diabetes (2.89 ± 1.64 vs. 2.35 ± 1.42; p = 0,09). Conclusions The prevalence of pain in patients with HTN is exceedingly high and seems to be related to the degree of blood pressure control.
The objective is to suggest a possible role of different angiotensin receptor blockers in the treatment of chronic migraine, especially in hypertensive subjects. Chronic migraine is a highly disabling disorder affecting between 1.4 and 2.2 % of the general population. Despite many pharmacological and non-pharmacological treatments proposed, the results are rather discouraging. Therefore, we believe that should be highlighted all the possible therapies that may lead to an improvement of the symptomatology. Particularly, data available on efficacy of ARBs in preventing chronic migraine are poor. Methods include case reports, literature review and discussion. We report three cases recently treated with angiotensin II receptor blockers that showed a significant improvement, never previously presented with more conventional treatments, including beta blockers. In all three cases, we obtained the reversibility from a chronic migraine to an episodic. Taking a cue from this observation, we consider desirable large controlled, randomized trials to assess the effectiveness of ARBs both in CM hypertensive patients and in patients who do not require anti-hypertensive therapy; furthermore are desirable comparative studies between the various ARB inhibitors to assay any intermolecular differences in efficacy.
We have recently reported that an intrathecal (i.t.) administration of angiotensin II (Ang II) into mice induces a nociceptive behavior accompanied by the activation of p38 MAPK signaling via AT1 receptors (Nemoto et al., 2013, Mol. Pain 9, 38). These results suggested that Ang II participates in the facilitation of nociceptive transmission in the spinal cord. In the present study, we used formalin test to examine the effect of i.t.-administered losartan, an AT1 receptor antagonist, and determine whether Ang II acts as a neurotransmitter and/or neuromodulator in the spinal transmission of nociceptive information. When administered i.t. 5 min before the injection of a 2% formalin solution into the plantar surface of the hindpaw, losartan (30–100 nmol) produced a dose-dependent and significant antinociceptive effect during both the first and second phases of the test. In the superficial dorsal horn of the spinal cord (laminae I and II), the fluorescence intensities for Ang II and phospho-p38 MAPK were both significantly increased on the ipsilateral side 3 min after the injection of formalin compared to saline-treated controls. Moreover, the increase of phospho-p38 MAPK fluorescence intensity was significantly inhibited by the i.t. administration of losartan (54.8 nmol) 5 min prior to formalin. These results indicate that losartan produces an antinociceptive effect through the inhibition of p38 MAPK phosphorylation in the mouse formalin test and that Ang II may act as a neurotransmitter and/or neuromodulator in the spinal transmission of nociceptive information.
Pharmacological, surgical, psychological, and alternative medicine approaches for the treatment of chronic pain, including neuropathic pain, provide only partial relief for most patients, with the efficacy of existing medications often blunted by dose-limiting side effects arising from drug actions on cells outside the pain-signaling axis. The development of more effective treatments for pain-particularly chronic pain states such as neuropathic pain-has been hampered by lack of predictive animal models and biomarkers, variation in pain characteristics between patients or on a day-to-day basis for single patients, patient stratification on the basis of symptoms rather than mechanism, and a high rate of placebo responses. We discuss genetic and genomic approaches to translational pain research. We review examples of the identification and validation of human pain targets through rodent genome-wide association studies (GWAS) and global mRNA expression studies, functional screening in flies and mice, human GWAS and whole-exome sequencing studies, and the targeted candidate gene approach. These and other emerging genetic and genomic strategies are likely to facilitate the development of new, more effective pain therapeutics.
The presence of a tissue-based renin-angiotensin system, independent of the systemic one, has been identified in several organs including the brain. Experimental models have suggested the involvement of the renin-angiotensin system in neurogenic inflammation, susceptibility to oxidative stress, endothelial dysfunction, and neuromodulation of nociceptive transmission, thus potentially contributing to the pathogenesis of migraine. Genetic factors that increase susceptibility to migraine may include angiotensin-converting enzyme polymorphism, although available data are controversial. Clinical studies have suggested that angiotensin-converting enzyme inhibitors and angiotensin receptor blockers may be effective in migraine prophylaxis. However, further research should clarify whether the postulated preventive effect is attributable to a pharmacological action over and above the antihypertensive effect and should test their tolerability in subjects with normal blood pressure values. In patients with contraindications or not responding to conventional prophylactic drugs and in patients with comorbid arterial hypertension, angiotensin-converting enzyme inhibitors and angiotensin receptor blockers may be used for migraine prophylaxis.
Background
We have recently demonstrated that intrathecal (i.t.) administration of angiotensin II (Ang II) induces nociceptive behaviour in mice accompanied by a phosphorylation of p38 mitogen-activated protein kinase (MAPK) mediated through Ang II type 1 (AT1) receptors. The N-terminal fragment of Ang II, Ang (1–7), plays a pivotal role in counterbalancing many of the well-established actions induced by Ang II. However, the role of Ang (1–7) in spinal nociceptive transmission remains unclear. Therefore, we examined whether i.t. administration of Ang (1–7) can inhibit the Ang II-induced nociceptive behaviour in mice.Methods
In the behavioural experiments, the accumulated response time of nociceptive behaviour consisting of scratching, biting and licking in conscious mice was determined during a 25-min period starting after i.t. injection. The distribution and localization of AT1 or Mas receptors were analysed using a MapAnalyzer and confocal microscope, respectively. Phosphorylation of p38 MAPK in the dorsal spinal cord was measured by Western blotting.ResultsThe nociceptive behaviour induced by Ang II was dose-dependently inhibited by the co-administration of Ang (1–7). The inhibitory effect of Ang (1–7) was reversed by the co-administration of A779, a Mas receptor antagonist. Western blot analysis showed that the increase in spinal p38 MAPK phosphorylation following the i.t. administration of Ang II was also inhibited by Ang (1–7), and the Ang (1–7) induced-inhibition was prevented by A779.Conclusions
Our data show that the i.t. administration of Ang (1–7) attenuates an Ang II-induced nociceptive behaviour and is accompanied by the inhibition of p38 MAPK phosphorylation mediated through Mas receptors.
The renin-angiotensin system (RAS) is a major regulatory system controlling many different homeostatic mechanisms both within the brain and in the periphery. While it is primarily associated with blood pressure and salt/water regulation, increasing evidence points to the involvement of the RAS in both headache disorders specifically and pain regulation in general. Several publications have indicated that drugs blocking various elements of the renin-angiotensin system lead to a reduction in migraine. Additionally, interventions on different angiotensin peptides or their receptors have been shown to both reduce and increase pain in animal models. As such, modulation of the renin-angiotensin system is a promising approach to the treatment of headaches and other pain conditions.
Angiotensins (Angs) modulate blood pressure, hydro-electrolyte composition, and antinociception. Although Ang (5–8) has generally been considered to be inactive, we show here that Ang (5–8) was the smallest Ang to elicit dose-dependent responses and receptor-mediated antinociception in the rat ventrolateral periaqueductal gray matter (vlPAG). Ang (5–8) antinociception seems to be selective, because it did not alter blood pressure or act on vascular or intestinal smooth muscle cells. The non-selective Ang-receptor (Ang-R) antagonist saralasin blocked Ang (5–8) antinociception, but selective antagonists of Ang-R types I, II, IV, and Mas did not, suggesting that Ang (5–8) may act via an unknown receptor. Endopeptidase EP 24.11 and amastatin-sensitive aminopeptidase from the vlPAG catalyzed the synthesis (from Ang II or Ang III) and inactivation of Ang (5–8), respectively. Selective inhibitors of neuronal-nitric oxide (NO) synthase, soluble guanylyl cyclase (sGC) and a non-selective opioid receptor (opioid-R) inhibitor blocked Ang (5–8)-induced antinociception. In conclusion, Ang (5–8) is a new member of the Ang family that selectively and strongly modulates antinociception via NO–sGC and endogenous opioid in the vlPAG.
We characterized response properties of neurons in the caudal ventrolateral medulla (CVLM), a structure involved in pain modulation. Electrophysiological recordings were performed in pentobarbitone-anesthetized control and monoarthritic rats. Noxious pinch, heat, cold and colorectal distension were used for peripheral test stimulation. To study central modulation of CVLM neurons and role in mediating descending pain regulation from the hypothalamus, glutamate was administered into the paraventricular nucleus of the hypothalamus (PVN). CVLM neurons gave excitatory, inhibitory or no response to noxious test stimulation. Response patterns for part of the neurons varied with submodality of test stimulation; e.g., a cell with an excitatory response to heat could give no or an inhibitory response to cold. Arthritis induced cell type-dependent changes in the spontaneous activity, most prominent of which was increased discharge rate of CVLM cells with an excitatory response to noxious stimulation. Significant arthritis-induced changes were also observed in the magnitudes of the nociceptive responses, among which was an increase in the pinch-induced excitatory response. Glutamate in the PVN of arthritic but not control animals suppressed the excitatory responses of CVLM cells, independent of the submodality of test stimulation and without influence on their spontaneous discharge rates. The results indicate that CVLM neurons give differential responses to noxious stimulation. Arthritis induces changes in response properties of CVLM neurons and in their central regulation by the PVN. These findings are in line with the evidence indicating that the CVLM plays a role in processing of nociceptive signals under inflammatory as well as control conditions.
Targeting supraspinal pain control centers by gene transfer is known to induce sustained analgesia. In this study, we evaluated the effects of injecting a Herpes Simplex Virus type 1 vector which expresses enkephalin (HSV-ENK vector) in the lateralmost part of the caudal ventrolateral medulla (VLMlat), a pain control center that exerts mainly descending inhibitory effects on pain modulation. Overexpression of enkephalin at the VLMlat reduced the number of flinches during the early and delayed phases of the formalin test and decreased c-fos expression in the spinal cord. These antinociceptive effects were detected at 2 and 10days after injection of HSV-ENK in the VLMlat and were completely reversed by local administration of naloxone. Virally driven-enkephalin was expressed from transduced neurons located in the VLMlat and, at lower extent, in the rostral ventromedial medulla. Our results show that HSV-mediated expression of enkephalin in the VLMlat induced antinociceptive effects, likely due to an enhancement of the opioidergic input to the VLMlat which accounted for descending inhibition of the nociceptive transmission at the spinal cord. This study also demonstrates the value of HSV-1 derived vectors to manipulate, in a sustained and directed manner, pain modulatory pathways in the brain, which is important in the study of supraspinal pain control circuits.
The caudal ventrolateral medulla (CVLM) is a key component of the supraspinal pain modulatory system. Pain modulation from the CVLM is partially relayed by spinally projecting noradrenergic neurons of the pontine A(5) cell group, which leave collateral fibres at the CVLM. The injection of angiotensin II (Ang II) into the CVLM was recently shown to induce hyperalgesia mediated by angiotensin type 1 (AT(1)) receptors, expressed by CVLM neurons that do not project to the spinal cord. The present study evaluates the effects of lesioning the noradrenergic pontine A(5) cell group by the retrograde transport of the selective toxin anti-dopamine beta-hydroxylase-saporin (anti-DBH-SAP) from the CVLM in pain behavioural responses elicited by Ang II injection into the CVLM. The injection of anti-DBH-SAP induced neurodegeneration, identified by the marker Fluoro-Jade B, restricted to the A(5) noradrenergic cell group. These results were confirmed by the decrease in the number of noradrenergic neurons only in the A(5) group. Pain behavioural evaluation using the formalin test showed that Ang II injection into the CVLM induced hyperalgesia, which was partially prevented by lesion of the A(5) noradrenergic cell group with anti-DBH-SAP. Immunostaining of AT(1) receptors in CVLM neurons retrogradely labelled from the A(5) noradrenergic cell group showed that CVLM neurons that project to the A(5) express AT(1) receptors, indicating that Ang II can modulate directly the CVLM-A(5) connection. The results show that Ang II-induced hyperalgesia elicited from the CVLM is mediated by an indirect pathway relayed at the pontine noradrenergic A(5) group.
Investigators have demonstrated pressor areas in the medullas of various species. The present study precisely localized the pressor area in the caudal medulla of the rat and determined its projections to the caudal and rostral ventrolateral medulla. The caudal medulla first was mapped grossly in rats with injections (30 nl) of glutamate (30-, 15-, and 7.5-nmol doses) placed 0.5, 1.0, and 1.5 mm caudal to the calamus scriptorius, 1.0, 1.5, and 2.0 mm lateral to the midline, and 1.8, 1.7, and 1.6 mm ventral to the dorsal medullary surface, respectively, and their arterial pressures were recorded. One of these nine injections showed significant increases in arterial pressure. We micromapped this area with a total of 27 injections of glutamate (10 nl; 5 nmol) placed 300 μm apart at 3 different dorsoventral levels. This micromapping study pinpointed the precise location of caudal pressor area (CPA) neurons in a restricted region lateral to the caudal end of the lateral reticular nucleus and ventromedial to the medullary dorsal horn near the level of the pyramidal decussation. Injections of glutamate into this spot, 1.0 mm caudal to the calamus scriptorius, 2.0 mm lateral to the midline, and 1.7 mm ventral from the dorsal surface of the medulla, induced significant increases in arterial pressure. The neuroanatomic connections of neurons in the CPA to the ventrolateral medulla were then investigated with iontophoretic injections of either the anterograde tracer biotinylated dextran amine (BDA) made into the CPA or the retrograde tracer Fluoro-Gold (FG) injected into either the caudal or rostral ventrolateral medulla. BDA injections resulted in bouton-laden fibers throughout both caudal and rostral portions of the ventrolateral medulla. Either of the F G injections resulted in numerous spindle-shaped neurons interspersed between the longitudinal fiber bundles running through the CPA area. The proximity of the CPA neurons to the A1 catecholaminergic cell group is discussed.
An overview of the role of ventrolateral medulla (VLM) in regulation of cardiovascular activity is presented. A summary of VLM anatomy and its functional relation to other areas in the central nervous system is described. Over the past few years, various studies have investigated the VLM and its involvement in cardiovascular regulation during static muscle contraction, a type of static exercise as seen, for example, during knee extension or hand-grip exercise. Understanding the neural mechanisms that are responsible for regulation of cardiovascular activity during static muscle contraction is of particular interest since it helps understand circulatory adjustments in response to an increase in physical activity. This review surveys the role of several receptors and neurotransmitters in the VLM that are associated with changes in mean arterial pressure and heart rate during static muscle contraction in anesthetized animals. Possible mechanisms in the VLM that modulate cardiovascular changes during static muscle contraction are summarized and discussed. Localized administration of an excitatory amino-acid antagonist into the rostral portion of the VLM (RVLM) attenuates increases in blood pressure and heart rate during static muscle contraction, whereas its administration into the caudal part of the VLM (CVLM) augments these responses. Opioid or 5-HT1A receptor stimulation in the RVLM, but not in the CVLM, attenuates cardiovascular responses to muscle contraction. Furthermore, intravenous, intracerebroventricular or intracisternal injection of an α2-adrenoceptor agonist or a cholinesterase inhibitor attenuates increases in blood pressure and heart rate during static muscle contraction. Finally, the possible involvement of endogenous neurotransmitters in the RVLM and the CVLM associated with cardiovascular responses during static muscle contraction is discussed. An overview of the role of the VLM in the overall cardiovascular control network in the brain is presented and critically reviewed.
Significant advances have been made in our understanding of nociceptive modulation from RVM. Among the most useful conceptually has been the discovery that there are two classes of modulatory neurons in the RVM that are likely to have opposing actions on nociception: on-cells, which may facilitate nociceptive transmission, and off-cells, which probably have a net inhibitory effect on nociception. The similarity in response properties among the members of each class, their large, somatic "receptive fields," and the wide distribution of the terminal fields of axons of individual neurons to the trigeminal sensory complex and to multiple spinal segments indicate that these neurons exert a global influence over nociceptive responsiveness. Drug microinjections into the RVM presumably shift the balance between states of on- or off-cell firing and also produce measurable changes in the threshold for nocifensor reflexes. The meaningful unit of function in the RVM nociceptive modulatory system therefore probably consists of large ensembles of physiologically and pharmacologically similar neurons. The strong coordination of activity of the two classes of RVM neuron may depend largely upon intranuclear projections from RVM off-cells that excite other off-cells and inhibit on-cells. The off-cell pause is GABA-mediated, and it is likely that there is a subset of GABA-containing RVM on-cells that directly inhibit off-cells. Furthermore, the available evidence indicates that exogenous opiates activate off-cells by inhibiting GABAergic release. Presumably, enkephalinergic cells in the RVM disinhibit off-cells in a similar way. Although non-serotonin-containing off-cells certainly exist, we propose that some off-cells contain serotonin. Other possible connections are based on more limited data; however, ACh, neurotensin, NE, and EAAs are present in neurons that project to the RVM, and each of these compounds, when microinjected into the RVM, has a modulating effect on nociceptive transmission. The local circuits in the RVM that underlie these actions remain to be elucidated. At the level of the dorsal horn, there is good evidence for each of three inhibitory mechanisms: direct inhibition of nociceptive projection neurons, inhibition of excitatory relay interneurons, and excitation of an inhibitory interneuron. The relative contribution made by each of these circuits is unknown.(ABSTRACT TRUNCATED AT 400 WORDS)
Inhibition of the spinal nociceptive tail-flick (TF) reflex by focal electrical stimulation in the caudal medulla was examined and characterized in lightly pentobarbital-anesthetized rats. Systematic mapping studies revealed that inhibition of the TF reflex was produced at low intensities of stimulation (12.5-25 microA) only from the lateral reticular nucleus (LRN). Areas dorsal and medial to the LRN required higher intensities of stimulation to produce descending inhibition of the TF reflex, likely reflecting spread of current to the LRN at these higher intensities of stimulation (50-100 microA). At threshold inhibitory intensities of stimulation in the LRN, changes in blood pressure were not produced. Strength-duration characterization of stimulation and the microinjection of glutamate into the LRN at the same site where focal electrical stimulation was effective suggest that the descending inhibition produced arises from activation of cell bodies in the LRN. The intrathecal administration of a variety of pharmacological antagonists revealed the descending inhibition produced by stimulation in the LRN to be mediated at least in part by spinal alpha 2-adrenoceptors. These findings, together with previous observations, suggest a role for the LRN in the centrifugal modulation of spinal nociceptive transmission.
It has been previously established that a bulbar relay plays an important role in descending inhibition of spinal dorsal horn nociceptive neurons and nociceptive reflexes produced by stimulation in the midbrain periaqueductal gray (PAG). In the present study, selected receptor antagonists were microinjected into the medial medullary nucleus raphe magnus (NRM) to determine whether descending inhibition of the tail flick (TF) reflex in the rat produced by focal electrical stimulation in the midbrain PAG was mediated by serotonin, opioid, or glutamate receptors on bulbospinal neurons in the NRM. It was determined in initial experiments that the serotonin receptor antagonist methysergide, the opioid receptor antagonist naloxone, the local anesthetic lidocaine, and the glutamate receptor antagonists gamma-D-glutamylglycine (DGG) and DL-2-amino-5-phosphonovalerate (APV) microinjected into the medulla all significantly increased the threshold of focal electrical stimulation in the medulla required to inhibit the TF reflex. The antinociceptive efficacy of agonists at opioid, serotonin, and glutamate receptors was also tested in other experiments. The microinjection of morphine (2.5-10 micrograms) into the NRM increased significantly TF latencies in a dose-dependent manner in rats in the awake or lightly anesthetized state; morphine was more potent in awake rats. Inhibition of the TF reflex produced by the microinjection of morphine was reversed by a subsequent microinjection of naloxone into the same site in the medulla. The microinjection of serotonin (5 and 10 micrograms), however, did not affect the latency of the TF reflex in either awake or lightly anesthetized rats. Glutamate (100 microM, 0.5 microliter) microinjected into the rostral ventral medulla produced an inhibition of the TF reflex of short duration that could be blocked or attenuated significantly by the glutamate receptor antagonists DGG or APV microinjected into the same site. In subsequent experiments, a nonspecific functional block was introduced adjacent to the NRM bilaterally in the medullary reticular formations (MRFs) by the microinjection of the local anesthetic lidocaine; receptor antagonists were then microinjected into the NRM and their effect on the threshold of focal electrical stimulation in the PAG to inhibit the TF reflex determined. No increase was seen in stimulation thresholds in the PAG following the microinjection of either methysergide or naloxone into the NRM. Following the microinjection of lidocaine, DGG or APV into the NRM, the stimulation threshold in the PAG for inhibition of the TF reflex was increased significantly.(ABSTRACT TRUNCATED AT 400 WORDS)
Focal electrical stimulation and microinjection of the excitatory amino acid glutamate in the lateral reticular nucleus (LRN) both inhibit the heat-evoked tail flick (TF) reflex in rats. The stimulation-produced inhibition from the LRN has previously been demonstrated to be mediated by spinal monoaminergic receptors. In the present study, inhibition of responses to noxious thermal stimuli by glutamate microinjected into the LRN was examined and characterized; this study is the first to examine the spinal receptors mediating inhibition produced by selective activation of cell bodies in the LRN. Microinjection of glutamate (100 mM) into the LRN in rats lightly anesthetized with pentobarbital produced a transient (less than 5 min) inhibition of the heat-evoked TF reflex, the magnitude of which increased with the volume of glutamate injected (100, 200, or 400 nl). This glutamate-produced inhibition of the TF reflex was antagonized by the intrathecal administration of phentolamine (30 micrograms), yohimbine (15 and 30 micrograms), or methysergide (15 and 30 micrograms) to the level of the lumbar spinal cord, but was not antagonized by prazosin (30 micrograms) or naloxone (20 micrograms). Yohimbine (15 and 30 micrograms) administered to the level of the cervical spinal enlargement did not significantly alter inhibition of the TF reflex produced by glutamate microinjected into the LRN. Microinjection of glutamate (100 mM, 400 nl) into the LRN elevated TF latencies and hindpaw lick latencies in the hot plate test performed on conscious rats. This inhibition of responses to noxious thermal stimuli in conscious rats was short-lasting (less than 5 min), and was also attenuated by intrathecal administration of yohimbine (30 micrograms) or methysergide (30 micrograms), but not by prazosin (30 micrograms) or naloxone (20 micrograms). While it has previously been established that cell bodies in the LRN mediate descending inhibition of spinal nociceptive reflexes, the present results establish that spinal alpha 2-adrenoceptors and serotonin receptors mediate LRN-produced antinociception and extend our understanding of LRN-mediated modulation of nociceptive responses integrated spinally and supraspinally.
Despite the wealth of information on the anatomy of descending pathways controlling cardiovascular SPNs, there is still a surprising lack of knowledge concerning the precise role of these pathways in cardiovascular regulation. The exception to this is the descending pathway from the RVLM, which has been the most thoroughly studied. Of all the sympathetic premotor nuclei, it is the only one so far identified that is critical for the operation of cardiovascular reflexes and that has been shown to be a major source of tonic excitatory input to cardiovascular SPNs. This does not necessarily mean that other sympathetic premotor nuclei are not important components of the central mechanisms subserving cardiovascular reflexes; rather, the experimental methods so far employed have not yet provided enough information on the functions of these other premotor nuclei to allow their functional roles to be defined.
Opioid-produced antinociception in mammals seems to be mediated in part by pathways originating in the periaqueductal gray (PAG) and the rostroventral medulla (RVM), and these pathways may include serotonergic neurons. In the present study, we examined the relationship of the cloned mu- and delta-receptors (MOR1 and DOR1, respectively) to PAG neurons projecting to the RVM, and RVM neurons projecting to the dorsal spinal cord. This was carried out by combining immunocytochemical staining for MOR1, DOR1, and serotonin with fluorescent retrograde tract-tracing. Of 133 retrogradely labeled cells in the RVM, 31% were immunoreactive for MOR1. Of the double-labeled cells, 41% also were immunoreactive for 5HT. Fifty-three percent of retrogradely labeled cells were apposed by DOR1-ir varicosities; 29% of the apposed cells were immunoreactive for 5HT. In the mesencephalon, cells retrogradely labeled from the RVM were usually surrounded by MOR1-ir structures; however, retrogradely labeled cells were never observed to be immunoreactive for MOR1. Similarly, retrogradely labeled cells in the caudal midbrain were seldom, if ever, labeled for DOR1; however, they frequently were apposed by DOR1-ir varicosities. Of 156 retrogradely labeled profiles from three rats, 52 (33%) were apposed by DOR1-ir varicosities. We conclude that both mu- and delta-opioid receptors could be involved in the antinociception mediated by the PAG-RVM-spinal cord circuit. In addition, opioids seem likely to have both direct and indirect effects on spinally projecting RVM cells in general, and on serotonergic RVM cells in particular.
The effects of angiotensin II (AngII), the AngII analogues saralasin--[Sar1, Ala8]AngII, sarmesin--[Sar1Tyr(Me)4]AngII, the nonpeptide AngII receptor antagonists DuP753 (losartan) (for AT1 receptor subtype) and PD123319 (for AT2 receptor subtype), as well as combinations of AngII and each of its analogues and receptor antagonists, administered intracerebroventricularly (ICV), were studied on mice using the acetic acid-induced abdominal constrictions test (acetic acid 1% intraperitoneally, IP). The abdominal constrictions were counted at 5-min intervals for 30 min. AngII at doses of 0.05, 0.1, and 1 microg exerted a dose-dependent antinociceptive effect. Saralasin, sarmesin, losartan, and PD123319 exhibited a dose-dependent effect on nociception: they either increased or decreased it. PD123319 antagonized the antinociceptive effect of AnglI while losartan was ineffective. The importance of AT2 receptor subtype for the nociception reducing effect of AngII is considered.
Many studies indicate that blood pressure control systems can attenuate pain (hypoalgesia) of short duration; however, we recently found exaggerated nociceptive responses (hyperalgesia) of persistent duration in the spontaneously hypertensive rat (SHR). Here, we used SHR, Dahl Salt-Sensitive (SS), and normotensive control rats to evaluate the contribution of sustained elevations in arterial pressure to nociceptive responses. Compared with Sprague-Dawley and/or Wistar-Kyoto controls, SHR were 1) hypoalgesic in the hot plate test and 2) hyperalgesic in longer latency tail and paw-withdrawal tests and in two models of inflammatory nociception. These differences were not observed between SS and salt-resistant controls fed a high-salt diet. Inflammatory hyperalgesia in SHR was correlated with neither paw edema nor the number of Fos-positive spinal cord neurons. Our results indicate that "pain" phenotype of the SHR is not restricted to hypoalgesia. This phenotype is related to genetic factors or to the autonomic systems that control blood pressure and not to sustained elevations in blood pressure, differences in spinal neuron activity, or inflammatory edema.
The objective of the present study was to determine the contribution of the autonomic nervous system and nitric oxide to the depressor effect produced by unilateral microinjection of ANG-(1-7) and ANG II into the caudal ventrolateral medulla (CVLM). Unilateral microinjection of ANG-(1-7), ANG II (40 pmol), or saline (100 nl) was made into the CVLM of male Wistar rats anesthetized with urethane before and after intravenous injection of 1) methyl-atropine, 2.5 mg/kg; 2) prazosin, 25 microg/kg; 3) the nitric oxide synthase (NOS) inhibitor, N(G)-nitro-L-arginine methyl ester (L-NAME), 5 mg/kg; or 4) the specific inhibitor of neuronal NOS, 7-nitroindazole (7-NI), 45 mg/kg. Arterial pressure and heart rate (HR) were continuously monitored. Microinjection of ANG-(1-7) or ANG II into the CVLM produced a significant decrease in mean arterial pressure (MAP; -11 +/- 1 mmHg, n = 12 and -10 +/- 1 mmHg, n = 10, respectively) that was not accompanied by consistent changes in HR or in cardiac output. The effect of ANG-(1-7) was abolished after treatment with methyl-atropine (-3 +/- 0.6 mmHg, n = 9) or L-NAME (-2.3 +/- 0.5 mmHg, n = 8) or 7-NI (-2.8 +/- 0.6 mmHg, n = 5). In contrast, these treatments did not significantly interfere with the ANG II effect (-10 +/- 2.6 mmHg, n = 8; -8 +/- 1.5 mmHg, n = 8; and -12 +/- 3.6 mmHg, n = 6; respectively). Peripheral treatment with prazosin abolished the hypotensive effect of ANG-(1-7) and ANG II. Microinjection of saline did not produce any significant change in MAP or in HR. These results suggest that the hypotensive effect produced by ANG II at the CVLM depends on changes in adrenergic vascular tonus and, more importantly, the hypotensive effect produced by ANG-(1-7) also involves a nitric oxide-related mechanism.
EthoLog is a tool that aids in the transcription and timing of behavior observation sessions—experimental or naturalistic, from video/audio tapes or registering real time. It was created with Visual Basic and runs on Windows (3.x / 9x). The user types the key codes for the predefined behavioral categories, and EthoLog registers their sequence and timing and saves the resulting data in ASCII output files. A sequential analysis matrix can be generated from the sequential data. The output files may be edited, converted to plain text files for printing, or exported to a spreadsheet program, such as MS Excel, for further analyses.
The caudal ventrolateral medulla (CVLM) plays a critical role in cardiovascular regulation. Convincing data now support the hypothesis that inhibition of sympathoexcitatory neurons in the rostral ventrolateral medulla (RVLM) by CVLM neurons constitutes the necessary inhibitory link in baroreceptor reflex mediated control of sympathetic vasomotor outflow. Inhibition or destruction of the CVLM produces severe acute hypertension, consistent with blockade of baroreceptor reflexes and withdrawal of inhibition of RVLM sympathoexcitatory neurons. However, other data indicate that the CVLM also tonically inhibits RVLM sympathoexcitatory neurons in a manner not driven by baroreceptor input. In some studies, inhibition of the CVLM results in an increase in arterial pressure (AP) without inhibiting baroreceptor reflexes, possibly reflecting baroreceptor-independent and baroreceptor-dependent sub-regions of the CVLM. Furthermore, in baroreceptor-denervated rats, inhibition of the CVLM still leads to large increases in AP. In addition, in spontaneously hypertensive rats (SHR) central processing of baroreceptor reflexes appears normal but CVLM-mediated inhibition of the RVLM seems to be attenuated, suggesting that it is specifically a baroreceptor-independent mechanism of cardiovascular regulation in SHR that is altered. Taken together, these findings support an important, tonic, baroreceptor-independent inhibition of RVLM sympathoexcitatory neurons exerted by the CVLM.
We evaluated the participation of nucleus reticularis gigantocellularis (NRGC), a medullary nucleus that plays an important role in the regulation of nociceptive processes, in the antinociceptive effect of angiotensin III (AIII), a biologically active peptide of the renin-angiotensin system. Adult, male Sprague-Dawley rats anesthetized with pentobarbital sodium (40 mg/kg, i.p., with 10 mg/kg/h i.v. infusion supplement) were used. Bilateral, site-specific microinjection of AIII (80 or 160 pmol) into the NRGC produced a dose-related increase in the latency of tail-flick response to noxious thermal stimuli (50°C hot water). Such an antinociceptive action of AIII was blocked by concomitant administration of the AIII receptor antagonist, Ile7-angiotensin III (Ile7-AIII, 10 nmol). At the neuronal level, microiontophoretic application of AIII suppressed, Ile7-AIII reversibly, the responsiveness of nociception-related neurons in the NRGC to tail-clamping. These results demonstrated that central AIII may elicit antinociception via a process that may at least take place at the NRGC.
A review of pharmacological, neuroanatomical, electrophysiological, and behavioral data indicates that systems controlling cardiovascular function are closely coupled to systems modulating the perception of pain. This view is directly supported by experiments from our laboratory showing that activation of either the cardiopulmonary baroreceptor reflex arc or the sinoaortic baroreceptor reflex arc induces antinociception. The outcomes of studies using pharmacological treatments, peripheral nerve stimulation, peripheral nerve resection, and CNS lesions are also presented as a preliminary means of characterizing cardiovascular input to pain regulatory systems. The network formed by these systems is proposed to participate in the elaboration of adaptive responses to physical and psychological stressors at various levels of the neuroaxis, and possibly to participate in “diseases of adaptation.” In particular, the present analysis suggests that the inhibition of pain brought about by elevations in either arterial or venous blood pressure may provide a form of psychophysiological relief under situations of stress and contribute to the development of essential hypertension in humans.
A method for assessing pain and analgesia in rats and cats is described. The procedure involves subcutaneous injection of dilute formalin into the forepaw, after which the animal's responses are rated according to objective behavioral criteria. The formalin test is a statistically valid technique which has two advantages over other pain tests: (1) little or no restraint is necessary, permitting unhindered observation of the complete range of behavioral responses; and (2) the pain stimulus is continuous rather than transient, thus bearing greater resemblance to most clinical pain. The analgesic effects of morphine, meperidine, and stimulation of the periaqueductal grey matter are evaluated using this test.
The formalin test for nociception, which is predominantly used with rats and mice, involves moderate, continuous pain generated by injured tissue. In this way it differs from most traditional tests of nociception which rely upon brief stimuli of threshold intensity. In this article we describe the main features of the formalin test, including the characteristics of the stimulus and how changes in nociceptive behaviour may be measured and interpreted. The response to formalin shows an early and a late phase. The early phase seems to be caused predominantly by C-fibre activation due to the peripheral stimulus, while the late phase appears to be dependent on the combination of an inflammatory reaction in the peripheral tissue and functional changes in the dorsal horn of the spinal cord. These functional changes seem to be initiated by the C-fibre barrage during the early phase. In mice, the behavioural response in the late phase depends on the ambient temperature. We argue that the peripheral tissue temperature as well as other factors influencing the peripheral inflammation may affect the response, possibly confounding the results obtained with the test. Furthermore, we discuss the methods of recording the response and the value of observing more than one aspect of behaviour. Scoring of several behavioural variables provides a means of assessing motor or sensorimotor function as possible causes for changes in behaviour. In conclusion, the formalin test is a valuable addition to the battery of methods available to study nociception.
We examined the vasomotor and respiratory effects of angiotensin II microinjection into the rabbit ventrolateral medulla (VLM). Angiotensin II in the rostral and caudal VLM increased and decreased arterial pressure, respectively, but had no effect on phrenic nerve activity. In contrast, L-glutamate injections in the same areas altered both arterial pressure and phrenic nerve activity. The results suggest that angiotensin II may activate specifically vasomotor neurons but not respiratory neurons in the VLM.
Multiple separate and distinct supraspinally organized descending inhibitory systems have been identified which are capable of powerfully modulating spinal nociceptive transmission. Until recently, brainstem sites known to be involved in the centrifugal modulation of spinal nociceptive transmission were few in number, being limited to midline structures in the midbrain and medulla (e.g., periaqueductal gray and nucleus raphe magnus). However, with continued investigation, that number has increased and brainstem sites previously thought to be primarily involved in cardiovascular function and autonomic regulation (e.g., nucleus tractus solitarius; locus coeruleus/subcoeruleus (LC/SC); A5 cell group; lateral reticular nucleus) also have been demonstrated to play a role in the modulation of spinal nociceptive transmission. Spinal monoamines (norepinephrine (NE) and serotonin) have been shown to mediate stimulation-produced descending inhibition of nociceptive transmission from these brainstem sites. The majority of NE-containing fibers and terminations in the spinal cord arise from supraspinal sources; thus, the LC/SC, the parabrachial nuclei, the Kölliker-Fuse nucleus and the A5 cell group have all been suggested as possible sources of the spinal noradrenergic (NA) innervation involved in the centrifugal modulation of spinal nociceptive transmission. Several lines of evidence suggest that the LC/SC plays a significant role in a functionally important descending inhibitory NA system. Focal electrical stimulation in the LC produces an antinociception and increases significantly the spinal content of NA metabolites. The inhibition of the nociceptive tail-flick withdrawal reflex produced by electrical stimulation in the LC/SC has been demonstrated to be mediated by postsynaptic alpha 2-adrenoceptors in the lumbar spinal cord. Similarly, electrical or chemical stimulation given in the LC/SC inhibits noxious-evoked dorsal horn neuronal activity. Thus, results reported in electrophysiological experiments confirm those reported in functional studies and the NA coeruleospinal system appears to play a significant role in spinal nociceptive processing.
Angiotensin II receptor subtypes (AT1 and AT2) were characterized in rat brain by displacement with the specific angiotensin antagonists Du Pont 753 and CGP 42112A, respectively, and quantitative autoradiography. Young (2-wk-old) rats expressed AT1 receptors in selected limbic system areas, structures involved in cardiovascular and fluid regulation, parts of the hippocampal formation, and the choroid plexus. In young rats, AT2 receptors were concentrated in areas involved in control and learning of motor activity, sensory areas, and selected limbic system structures. The cingulate cortex, the molecular layer of the cerebellar cortex, and the superior colliculus contained both AT1 and AT2 receptors. The number of AT1 receptors in most areas of adult (8-wk-old) rats was similar to or even higher than that present in young rats. Conversely, AT2 receptors were always much lower in number in adult animals, and in some areas they were undetectable in adults. Their differential localization and development suggest different functions for the specific angiotensin II receptor subtypes.
Intrathecal administration of the neurotoxins 5,6-dihydroxytryptamine (5,6-DHT) and 6-hydroxydopamine (6-OHDA) in rats selectively lesioned the descending spinal serotonergic and noradrenergic pathways, respectively. Four days after neurotoxin administration the behaviour was evaluated in the formalin test. Several behavioural variables were recorded. The statistical analysis of the results was supplemented using a multivariate statistical method (partial least squares projection to latent variables, PLS) in addition to traditional analysis of variance. The described methods for recording and statistical analysis of behaviour appear to be useful in describing drug-induced differences in behavioural patterns in the formalin test. Both types of lesion reduced the pain-related behaviour in the formalin test (protection of the paw, biting and licking). The results indicate that the descending monoaminergic pathways are parts of a network which maintains adequate nociceptive responses to a chemical stimulus, or to stimuli lasting several minutes, as in the formalin test.
Two physiologically and pharmacologically distinct classes of putative nociceptive modulatory neurons have been identified in the rostral ventral medulla (RVM) of the lightly anesthetized rat: on-cells and off-cells. We have previously shown that administration of morphine either systemically or by microinjection into the periaqueductal gray (PAG) produces an increase in the activity of all off-cells and a depression of the activity of all on-cells concomitant with inhibition of the tail flick reflex. We now demonstrate that morphine applied intrathecally has effects on RVM neurons that are indistinguishable from those of systemic or PAG administration. This may contribute to the known multiplicative effects of concurrent administration of opioids at spinal and supraspinal sites.
This study used postembedding immunocytochemistry to examine the organization of GABA‐immunoreactive synapses in the rostral ventral medulla (RVM) of the rat. To determine whether the outflow neurons of the RVM are under GABAergic control, we examined the distribution of GABA‐immunoreactive synapses upon bulbospinal projection neurons that were labelled by retrograde transport of wheatgerm agglutinin‐HRP from the cervical spinal cord. To study the possible convergence of GABAergic and periaqueductal gray (PAG) synaptic inputs to RVM neurons, we also made lesions in the PAG and examined the relationship between degenerating PAG axons and GABA‐immunoreactive terminals.
Approximately 45% of all synapses in the RVM, which includes the midline nucleus raphe magnus and the nucleus reticularis paragigantocellularis lateralis, were GABA‐immunoreactive. The vast majority of GABA‐immunoreactive terminals contained round, clear, and pleomorphic vesicles and made symmetrical axodendritic synapses; axoaxonic synapses were not found. Almost 50% of the retrogradely labeled dendrites in the NRM were postsynaptic to GABA‐immunoreactive terminals. Several examples of convergence of degenerating PAG terminals and GABAergic terminals onto the same unlabelled dendrite were also found. These data indicate that the projection neurons of the RVM are urider profound GABAergic inhibitory control. The results are discussed with regard to the hypothesis that the analgesic action of narcotics and electrical stimulation of the midbrain PAG involves the regulation of tonic GABAergic inhibitory controls that are exerted upon spinally‐projecting neurons of the nucleus raphe magnus.
The respiratory modulation of the lumbar sympathetic nerve discharge (LSND) was examined in halothane-anesthetized, paralyzed, and vagotomized rats by means of phrenic nerve discharge (PND)-triggered histograms. The respiratory modulation was 1) proportional to PND amplitude during chemoreceptor activation with CO2 and 2) reduced at elevated arterial pressure. Bilateral injections of bicuculline [gamma-aminobutyric acid (GABA)A receptor antagonist, n = 5] into the rostral ventrolateral medulla (RVLM), but not into medullary raphe, reversibly increased mean arterial pressure (MAP) and resting LSND, decreased the baroreflex, reduced PND amplitude and central respiratory rate, and greatly magnified the respiratory modulation of LSND. Injections of strychnine (glycine receptor antagonist, n = 5) or phaclofen (GABAB receptor antagonist, n = 2) into RVLM were without effect. Injections of kynurenic acid (excitatory amino acid receptor antagonist) into RVLM (n = 8), but not raphe (n = 3), reduced PND amplitude, increased central respiratory rate, reduced MAP, elevated resting LSND slightly, and greatly reduced the respiratory modulation of LSND. These data suggest that the rostral tip of the ventrolateral medulla represents a critical link between the central respiratory rhythm generator and the vasomotor outflow. Also, it indicates that the respiratory modulation of SND does not involve a gating of the activity of the medullary neurons that convey baroreceptor information to RVLM sympathoexcitatory neurons.
Various angiotensins, bradykinins, and related peptides were examined for their inhibitory activity against several enkephalin-degrading enzymes, including an aminopeptidase and a dipeptidyl aminopeptidase, purified from a membrane-bound fraction of monkey brain, and an endopeptidase, purified from the rabbit kidney membrane fraction. Angiotensin derivatives having a basic or neutral amino acid at the N-terminus showed strong inhibition of the aminopeptidase. Dipeptidyl aminopeptidase was inhibited by angiotensins II and III and their derivatives, whereas the endopeptidase was inhibited by angiotensin I and its derivatives. The most potent inhibitor of aminopeptidase and dipeptidyl aminopeptidase was angiotensin III, which completely inhibited the degradation of enkephalin by enzymes in monkey brain or human CSF. The Ki values for angiotensin III against aminopeptidase, dipeptidyl aminopeptidase, endopeptidase, and angiotensin-converting enzyme, which degraded enkephalin, were 0.66 X 10(-6), 1.03 X 10(-6), 2.3 X 10(-4), and 1.65 X 10(-6) M, respectively. Angiotensin III potentiated the analgesic activity of Met-enkephalin after intracerebroventricular coadministration to mice in the hot plate test. Angiotensin III itself also displayed analgesic activity in that test. These actions were blocked by the specific opiate antagonist naloxone.
An increasing amount of anatomical, physiological, and pharmacological evidence suggest that pain inhibitory circuitry is linked with cardiovascular regulatory systems in man and laboratory animals. Induction of hypertension in rats by different methods (mineralocorticoid treatment, stenosis of renal artery, or social deprivation) is associated with reduced responsiveness to noxious thermal stimuli (hot-plate) or to noxious mechanical stimuli (paw pressure). Genetically hypertension-prone rats derived from the SABRA strain and spontaneously hypertensive rats derived from Wistar/Kyoto strain also display a similar hypoalgesia. Acute increases in blood pressure are associated with reduced sensitivity to painful stimuli. Additionally, the interaction between blood pressure and pain perception has also been supported by the demonstration that various experimental interventions that diminish the magnitude of hypertension also attenuate the hypoalgesia. Recent clinical findings are also in agreement with the laboratory animal findings since sensory and pain thresholds have been shown to be significantly higher in unmedicated essential hypertensive subjects compared to normotensive controls. Thus, the human data corroborate animal data and suggest that a relation between blood pressure and pain sensitivity is likely to be a general phenomenon. It is unlikely that damage to peripheral pain fibers caused by a change in blood pressure contributes to the observed hypoalgesia. Naloxone, which has no effect on blood pressure, returns the pain sensitivity to normal levels. Behavioral tests (open field and motor activity cage) of normotensive and of renal and genetically (SBH and SHR) hypertensive rats exclude the possibility of a general motor deficit in hypertensive rats. Endogenous opioid peptides in central and peripheral nervous systems as well as in endocrine organs are implicated, although non-opioid mechanisms are also evident. Activation of baroreceptor afferents by acute or chronic increases in arterial or venous blood pressure may play an important role in the somatosensory responses associated with the increase in blood pressure. Coordinated cardiovascular-pain regulatory responses may be part of an adaptive mechanism that helps the body to face stressful events.
Studies concerning variations of the central renin-angiotensin system (RAS) during immobilization stress in rats have shown a significant increase in renin-like activity in the hypothalamus and fronto-parietal cortex, together with a definite decrease in the hypophysis and pineal gland. The resultant stress analgesia is blocked by the previous administration of naloxone and saralasin (angiotensin II antagonist). The intracerebral administration of renin and angiotensin II produces an increase in latencies to thermoalgesic stimuli; this is reduced, as is immobilization stress, by naloxone and saralasin. Both chemical hypophysectomy obtained by dexamethasone pretreatment as well as surgical epiphysectomy block the stress-induced analgesia. The experimental data obtained argue in favour of the participation of the cerebral RAS in stress analgesia through the indirect mechanism of release of opioid peptides.
In this review, the emerging functional roles of the brain angiotensin system have been considered. The major effects of Ang II can be classified into three groups, which imply three possible functions: The first, and largest, group is actions associated with the regulation of body fluid volume in response to hypovolemia. These include thirst, blood pressure increase, vasopressin release, sodium appetite and excretion, and ACTH and aldosterone release. This function alone has important implications for the control of blood pressure and the disease of hypertension. Another possible function is a role for angiotensin in the activity of gonadotropic hormone releasing hormones and pituitary hormones during the reproductive cycle and pregnancy. A third group of functions is the synaptic, neurotransmitter interactions of Ang II with catecholamines, serotonin, prostaglandins, and other peptides, not all of which could be reviewed here due to space limitations. This interaction is significant for all functions mentioned and leads to alterations in motivation (thirst, pain), memory (and possibly learning), and motor control. The amount of data available, however, is so limited that to claim angiotensin plays any major role in the latter functions would be premature. Throughout this review, we compared the central and peripheral effects of Ang II. We suggest that normally, a blood-CVO barrier prevents diffusion of peripheral Ang II to brain receptors inside the BBB. Because of this mechanism, the responses to the two routes of administration are distinctly different. When systemic peptide levels are low, Ang II activates only receptors in the CVOs; however, when these levels are high, the peptide diffuses to receptors that are normally activated only by brain Ang II.
The need for rapid histological feedback on neural tissue is ever present. Although there are several stains which can be readily used for staining either cell bodies or fiber tracts, adequate contrasting stains which are both rapid and easy to apply are not generally available. In 1936 Chang presented a technique for whole brains utilizing the metachromatic properties of thionin. Unfortunately this procedure was very time consuming. For the last several years we have worked with several variations of this stain and have found that thionin can be reliably used as a polychrome stain for sections of neural tissue obtained from a freezing microtome.
Thanks largely to the study of the brainstem nuclei that mediate stimulation analgesia, the involvement of the monoamines in the descending control of pain is now well established. The periaqueductal grey, the raphe nuclei (NRM and DRN) and the locus coeruleus are all key brainstem sites for the control of nociceptive transmission in the spinal cord. Although the initial emphasis was on 5-HT as the transmitter mediating this control at spinal levels, it is clear from more recent work that NA has an equally important part to play. How (or even if) the two amines differ in their roles and actions in analgesia is, however, still an open question. The small size and complexity of the brainstem areas from which analgesia may be elicited by electrical stimulation complicates the interpretation of the data. Stimulating currents may spread to surrounding regions mediating opposite effects to that of the main region stimulated. Opiates and GABA are clearly involved in descending control at both brainstem and spinal levels, although the relative roles of the different types of amino-acid and opiate receptors is still hotly debated. Despite the fact that the first report on stimulation analgesia appeared more than a quarter of a century ago in 1969, the precise connections and cord synaptology are still the basis of ongoing research. It is perhaps ironic, in an issue dedicated to new molecules and mechanisms, that those transmitters most involved in descending inhibition should be such old and familiar friends.
The formalin test is increasingly used as a model of injury-produced pain but there is no generally accepted method of pain rating. To examine the properties of various pain rating methods we established dose-response relations for formalin injected in the plantar surface of one hind paw, and the analgesic effects of morphine and amphetamine using the most frequently reported behavioural measures of pain (favouring, lifting, licking and flinching/shaking of the injured paw) and combinations of these. Licking, elevation and favouring of the injected paw showed a biphasic response at all formalin doses. Flinching varied in form across the time course of formalin, and the biphasic nature of the behaviour was not as apparent. In untreated rats all these behaviours were infrequent. Flinching and favouring were increased after injection of local anaesthetic into the paw but remained negligible relative to the effect of formalin. Grooming other than that directed to the injected paw was elevated in a dose-dependent manner by formalin. Intercorrelations between the behaviours were different for the initial response and the second phase. Correlational analysis indicated that no single behavioural measure was a strong predictor of formalin, morphine and amphetamine dose. A simple sum of time spent licking plus elevating the paw, or the weighted pain score of Dubuisson and Dennis (1977), were superior to any single measure (r ranging from 0.75 to 0.86). Addition of flinching and favouring to the combined pain score using multiple regression did not increase variance explained. Depending on the measure used, a sedative dose of pentobarbital produced apparent analgesia, hyperalgesia or no effect. The interphase depression of pain, as well as the analgesic effects of morphine and amphetamine, were all associated with increased motor activation. Power analysis indicated that using a moderate dose of formalin and a combined pain score gave the greatest power to detect differences in pain. It was also found that pain scores increase with ambient temperature and that rat strains may differ in formalin pain sensitivity.
The location of neurons in the caudal medulla oblongata that project to the superficial or deep dorsal horn was studied in the rat, by means of retrograde labelling from confined spinal injection sites. The tracer cholera toxin subunit B was injected into laminae I-III (fuve rats) or I-V (three rats) at C4-7 spinal segments. Neurons projecting to the superficial dorsal horn were located in the dorsomedial part of the dorsal reticular nucleus ipsilaterally, the subnucleus commissuralis of the nucleus tractus solitarius bilaterally, and a region occupying the lateralmost part of the ventrolateral reticular formation between the lateral reticular nucleus and the caudal pole of the spinal trigeminal nucleus, pars caudalis, bilaterally. Neurons projecting to the deep dorsal horn, which were only labelled when laminae I-V were filled by the tracer, occurred in the dorsomedial and ventrolateral parts of the dorsal reticular nucleus and in the ventral reticular nucleus bilaterally. A few cells were located in the above described lateralmost portion of the ventrolateral reticular formation bilaterally and in the ventral portion of the ipsilateral cuneate nucleus. In the light of previous data demonstrating that dorsal horn neurons project to the dorsal reticular nucleus, the ventrolateral reticular formation, and the nucleus tractus solitarius, and that neurons in these three medullary regions are involved in pain inhibition at the spinal level, the descending projections demonstrated here suggest the occurrence of spino-medullary-spinal loops mediating the analgesic actions elicited in each nucleus upon the arrival of nociceptive input from the dorsal horn.
The aim of the current study was to investigate the role of the rostroventromedial medulla (RVM) in alpha 2-adrenoceptor-mediated antinociception. Medetomidine or clonidine, selective alpha 2-adrenoceptor agonists were microinjected into the RVM in unanesthetized rats with a chronic guide cannula. The antinociceptive effects were evaluated using the tail-flick and hot-plate tests. For comparison, medetomidine was microinjected into the cerebellum or the periaqueductal gray (PAG). To study the role of medullospinal pathways, the tail-flick latencies were also measured in spinalized rats. The reversal of the antinociception induced by intracerebral microinjections of medetomidine was attempted by s.c. atipamezole, a selective alpha 2-adrenoceptor antagonist. The reversal of the antinociception induced by systemic administration of medetomidine was attempted by microinjections of 5% lidocaine or atipamezole into the RVM. When administered into the RVM, medetomidine produced a dose-dependent (1-30 micrograms) antinociception in the tail-flick and hot-plate tests, which antinociceptive effect was completely reversed by atipamezole (1 mg/kg, s.c.). Also clonidine produced a dose-dependent (3-30 micrograms) antinociception following microinjection into the RVM. Microinjections of medetomidine into the cerebellum or the PAG produced an identical dose-response curve in the tail-flick test as that obtained following microinjection into the RVM. In spinalized rats the antinociceptive effect (tail-flick test) induced by medetomidine microinjected into the RVM was not less effective than in intact rats. Lidocaine (5%) or atipamezole (5 micrograms) microinjected into the RVM did not attenuate the antinociception induced by systemically administered medetomidine (100 micrograms/kg, s.c.).(ABSTRACT TRUNCATED AT 250 WORDS)
A behavioral hypalgesia (increased response threshold to noxious stimuli) has been consistently, although not invariably, reported in spontaneous and experimental acute and chronic hypertension in the rat. Studies in human hypertension have also demonstrated a diminished perception of pain, assessed as pain thresholds or ratings. The sensitivity to painful stimuli correlated inversely with blood pressure levels, and this relationship extended into the normotensive range. Evidence in humans and rats points to a role of the baroreflex system in modulating nociception. In the rat, blood pressure-related antinociception may be due to attenuated transmission of noxious stimuli at the spinal level secondary to descending inhibitory influences that are projected from brain stem sites involved in cardiovascular regulation and that may depend on baroreceptor activation and/ or on a central "drive." Both endorphinergic and noradrenergic central neurons (the latter acting through postsynaptic alpha 2-receptors) have been shown to be involved, and other mediators probably also play a role. Functionally, blood pressure-related antinociception may represent an aspect of a more-complex coordinated adaptive response of the body to "stressful" situations. It is still uncertain whether in human essential hypertension hypalgesia is secondary to elevated blood pressure or whether both depend on some common mechanism. Studies on the effect of hypotensive treatment are too few to allow conclusions. According to one hypothesis, the reduction in pain perception caused by baroreceptor activation secondary to blood pressure elevation may represent a rewarding mechanism that may be reinforced with repeated stress and may be involved in the development of hypertension in some individuals. Hypertension-associated hypalgesia may have clinically relevant consequences, especially in silent myocardial ischemia and unrecognized myocardial infarction, both of which are more prevalent in hypertensive individuals.
The pathway conveying the descending inhibitory noradrenergic input elicited from the caudal ventrolateral medulla (VLM) onto the spinal cord dorsal horn was studied in the rat. Retrograde labeling with cholera toxin subunit B (CTb) injected into the dorsal horn was combined with immunostaining for dopamine-beta-hydroxylase (DBH) in the VLM and other brainstem nuclei containing noradrenergic cells. CTb-labeled neurons occurred in the lateral part of the VLM (VLMlat), located ventrolaterally to the DBH-immunoreactive cells of the A1 noradrenergic cell group. Neuronal profiles stained for CTb and DBH (double labeled) occurred in the A5 (31%), A6 (57%), and A7 (12%) noradrenergic cell groups. To ascertain whether noradrenergic cells targeting the spinal cord in those groups received projections from the VLMlat, this area was injected with the anterograde tracer biotinylated dextran amine (BDA). Labeled terminal fibers with boutons en passant were apposed to numerous double-stained neurons in the A5 cell group. Similar appositions occurred in small amounts in the ventral subcoerulear component of the A6. Correlated light and electron microscopic analyses of the labeled appositions revealed that the BDA-labeled axonal boutons contained spherical vesicles and were presynaptic at asymmetrical contacts to somata and dendritic profiles of the double-stained A5 neurons. These data indicate the occurrence of an indirect dysynaptic pathway connecting the VLM to the spinal cord, with a relay in the A5 cells. This pathway may convey the antinociceptive effects mediated by alpha 2-adrenoreceptors, which have been previously observed in the spinal cord following VLM stimulation.
A polyclonal antibody has been prepared against a synthetic peptide derived from the C-terminal tail of the cloned rat AT2 angiotensin receptor, corresponding to amino acid residue 341-351. The antibody was of high titer and displayed monospecific activity toward the synthetic peptide in the ELISA assay. Western blot analysis indicated that the antiserum recognised only a single protein band with a mean apparent molecular mass of 75.4 kDa in the rat adrenals. Immunohistochemical studies with affinity purified antibody localised immunoreactive AT2 angiotensin receptor in medulla cells of the adrenals. Immunoreactivity was also observed in pyramidal tract, but no specific immunoreactivity can be detected in regions of rat brain that are known to express AT2 angiotensin receptors, including inferior olive, locus coeruleus and cerebellum.
A disynaptic pathway linking the caudal ventrolateral medulla (VLM) to the spinal cord via the A5 noradrenergic cell group of the pons has recently been described in the rat. In the present work, the projections of the A5 to the VLM and to the spinal dorsal horn were studied with double-tracing techniques combined with immunostaining of the noradrenaline-synthesizing enzyme dopamine-beta-hydroxylase. Cholera toxin subunit B (CTb) injected into the VLM and fluoro-gold injected into the spinal dorsal horn produced double retrograde labelling of A5 neurons immunoreactive for dopamine-beta-hydroxylase, which received appositions of fibre varicosities labelled anterogradely with CTb injected into the VLM. After injecting CTb into the A5, retrogradely labelled neurons occurred in the VLM. These neurons were contacted by anterogradely labelled fibres from the A5 group. These observations indicate that the VLM cells acting upon the A5 spinally projecting neurons, which are likely to exert an alpha2-adrenoreceptor-mediated inhibition on the spinal cord, are targeted by collaterals of the A5 spinal cord-bound axons. The A5-VLM pathway may be the anatomical substrate of a negative feedback circuit whereby the modulatory action of the VLM on the spinal cord is self-inhibited through activation of the A5.
The influence of the dorsal reticular nucleus (DRt) on pain behaviour during the formalin test was studied in the rat by lesioning the nucleus through local application of electrical current or quinolinic acid. Animals in which the DRt was lesioned ipsilaterally to the paw injected with formalin spent less time in focused (licking, biting or scratching the injected paw) and total (focused pain behaviour plus protection of the injected paw during movements) pain behaviour, and showed paw-jerks less frequently than non-lesioned animals in both phases 1 and 2 of the test. Animals in which the DRt was lesioned contralaterally to the injected paw presented a decrease in total pain behaviour and number of paw-jerks only during phase 2. The number of superficial (laminae I–II) and deep (laminae III–VI) spinal dorsal horn cells expressing the c-fos proto-oncogene 2 h after subcutaneous injection of formalin was reduced by 34% and 50%, respectively, in animals with an ipsilateral DRt lesion as compared to non-lesioned rats. No differences in c-fos expression were observed after lesioning the DRt contralateral to the formalin injection.
The results indicate that the DRt is involved in the facilitation of nociception during the formalin test by enhancing the response capacity of dorsal horn neurons to noxious stimulation. It is suggested that the pronociceptive action of the DRt is mediated by the reciprocal connections it establishes with the spinal dorsal horn [Almeida et al. (1993) Neuroscience, 55, 1093].
Physiological stress is known to produce analgesia and memory disruption. Brain renin angiotensin system (RAS) has been reported to participate in stress response and plays a role in the processing of sensory information. Angiotensin receptors (AT), particularly AT1 subtypes have been reported to be distributed in brain areas that are intimately associated with stress response. The purpose of present study was to examine the modulation of AT1 receptor in the immobilization stress and angiotensin II (AngII)-induced analgesia and impaired retention, and to determine whether resultant behavioral changes involve common sensory signals. Result of present experiments showed that immobilization stress in mice and rats, and intracerebroventricular (ICV) administration of AngII (10 and 20 ng) in rats produced an increase in tail-flick latency. Similarly, post training administration of AngII or immobilization stress produced impairment of retention tested on plus-maze learning and on passive avoidance step-down task. Both these responses were sensitive to reversal by prior treatment with losartan (10 and 20 mg/kg), an AT1 AngII receptor antagonist. On the other hand, naloxone, an opiate antagonist preferentially attenuated the stress and AngII-induced analgesia and retention deficit induced by immobilization stress, but failed to reverse the AngII induced retention deficit. These results suggest immobilization stress-induced analgesia and impaired retention involves the participation of brain RAS. Further, failure of naloxone to reverse AngII-induced retention impairment shows. AngII-induced behavioral changes are under control of different sensory inputs.
The aim of this paper is to study the synergistic anti-analgesic effect of angiotensin II (Ang II) plus cholecystokinin octapeptide (CCK-8). Our previous studies have shown that both CCK-8 and Ang II are potent anti-opioid substances. Intracerebroventricular (i.c.v.) injection of CCK-8 or Ang II dose-dependently antagonizes morphine-induced analgesia (MIA). In the present study, we observed the combined effect of CCK-8 and Ang II in antagonizing MIA. CCK-8 and Ang II were injected intracerebroventricularly to rats in various proportions and doses. The results were analyzed with isobolographic analysis. Combined injection of CCK-8 and Ang II in a ratio of 1 ng: 2.5 microg or 1 ng: 5 microg produced significantly greater effect in antagonizing MIA. The ED(50) of the two ratios are only 18.5% and 27.5%, respectively, of the theoretical dose of simple addition. We conclude that CCK-8 and Ang II used in such dose ratios may antagonize MIA synergistically.
The endogenous pain control system is composed of multiple functionally distinct brain regions, which are thought to integrate nociceptive information with various brain functions. The clear involvement of some pain control centres in cardiovascular modulation has been claimed as a strong indication of their role in nociceptive-cardiovascular integration. Particular emphasis has been given to their putative function in triggering cardiovascular reactions to pain. However, the possibility of their participation in the less-studied influence of cardiovascular conditions in pain perception has been largely ignored. We have recently addressed this issue by investigating the involvement of the caudal ventrolateral medullary reticular formation (cVLM) in hypertension-induced hypoalgesia. Circuits capable of conveying cVLM-elicited antinociception include a direct reciprocal cVLM-spinal loop, and two disynaptic spinal pathways relaying in rostroventromedial medullary (RVM) neurones and A(5) noradrenergic neurones. In the three pathways, the cVLM neurones involved are circumscribed to a small area of reticular formation located laterally to the lateral reticular nucleus, the VLMlat. The VLMlat has a vasodepressor effect similar to that obtained from the cVLM. In the spinal cord dorsal horn, c-fos expression evoked by noxious stimuli is decreased in hypertensive animals, as compared to normotensive animals. In hypertensive animals following lesion of the VLMlat, spinal c-fos expression is identical to that observed in normotensive animals. The collected data point to a role for the VLMlat in the depression of spinal nociceptive processing in response to rises in blood pressure. Since hypertension-induced hypoalgesia is mediated by spinal alpha(2)-adrenoreceptors, this effect could be conveyed by the cVLM-A(5)-spinal pathway.
1. Cardiovascular effects of microinjections of nociceptin and endomorphin-2 into the following medullary areas were studied in urethane-anaesthetized rats: chemoreceptor projection site (CPS), intermediate portion of the nucleus tractus solitarius (I-NTS), caudal ventrolateral medullary depressor area (CVLM) and rostral ventrolateral medullary pressor area (RVLM).
2. Microinjections of nociceptin or endomorphin-2 (0.6 mmol/L each) into the CPS and RVLM elicited depressor and bradycardic responses, whereas similar injections into the I-NTS and CVLM elicited pressor and tachycardic responses.
3. The mechanism of cardiovascular responses to microinjections of these opioid peptides into different medullary areas involved in cardiovascular function can be postulated as follows: the direct effect of nociceptin and endomorphin-2 on neurons is usually inhibitory. Because the activation of CPS and RVLM by microinjections of L-glutamate results in pressor and tachycardic responses, inhibition of these areas by nociceptin and endomorphin-2 elicits depressor and bradycardic responses. Similarly, activation of neurons in the I-NTS and CVLM by microinjections of L-glutamate elicits depressor and bradycardic responses. Therefore, inhibition of these areas by microinjections of these opioid peptides elicits an increase in blood pressure and heart rate.
Upon receipt in the dorsal horn (DH) of the spinal cord, nociceptive (pain-signalling) information from the viscera, skin and other organs is subject to extensive processing by a diversity of mechanisms, certain of which enhance, and certain of which inhibit, its transfer to higher centres. In this regard, a network of descending pathways projecting from cerebral structures to the DH plays a complex and crucial role. Specific centrifugal pathways either suppress (descending inhibition) or potentiate (descending facilitation) passage of nociceptive messages to the brain. Engagement of descending inhibition by the opioid analgesic, morphine, fulfils an important role in its pain-relieving properties, while induction of analgesia by the adrenergic agonist, clonidine, reflects actions at alpha(2)-adrenoceptors (alpha(2)-ARs) in the DH normally recruited by descending pathways. However, opioids and adrenergic agents exploit but a tiny fraction of the vast panoply of mechanisms now known to be involved in the induction and/or expression of descending controls. For example, no drug interfering with descending facilitation is currently available for clinical use. The present review focuses on: (1) the organisation of descending pathways and their pathophysiological significance; (2) the role of individual transmitters and specific receptor types in the modulation and expression of mechanisms of descending inhibition and facilitation and (3) the advantages and limitations of established and innovative analgesic strategies which act by manipulation of descending controls. Knowledge of descending pathways has increased exponentially in recent years, so this is an opportune moment to survey their operation and therapeutic relevance to the improved management of pain.
Angiotensinogen, the precursor molecule for angiotensins I, II and III, and the enzymes renin, angiotensin-converting enzyme (ACE), and aminopeptidases A and N may all be synthesised within the brain. Angiotensin (Ang) AT(1), AT(2) and AT(4) receptors are also plentiful in the brain. AT(1) receptors are found in several brain regions, such as the hypothalamic paraventricular and supraoptic nuclei, the lamina terminalis, lateral parabrachial nucleus, ventrolateral medulla and nucleus of the solitary tract (NTS), which are known to have roles in the regulation of the cardiovascular system and/or body fluid and electrolyte balance. Immunohistochemical and neuropharmacological studies suggest that angiotensinergic neural pathways utilise Ang II and/or Ang III as a neurotransmitter or neuromodulator in the aforementioned brain regions. Angiotensinogen is synthesised predominantly in astrocytes, but the processes by which Ang II is generated or incorporated in neurons for utilisation as a neurotransmitter is unknown. Centrally administered AT(1) receptor antagonists or angiotensinogen antisense oligonucleotides inhibit sympathetic activity and reduce arterial blood pressure in certain physiological or pathophysiological conditions, as well as disrupting water drinking and sodium appetite, vasopressin secretion, sodium excretion, renin release and thermoregulation. The AT(4) receptor is identical to insulin-regulated aminopeptidase (IRAP) and plays a role in memory mechanisms. In conclusion, angiotensinergic neural pathways and angiotensin peptides are important in neural function and may have important homeostatic roles, particularly related to cardiovascular function, osmoregulation and thermoregulation.
The intracerebroventricular administration of renin substrate or angiotensin II evokes antinociception in rodents, but the brain sites where most of the renin-angiotensin system peptides act are not yet known. This study describes the antinociceptive effects of microinjecting porcine renin substrate tetradecapeptide (RS) or angiotensins I (AI), II (AII) or III (AIII) into different regions of the periaqueductal gray matter (PAG), using the rat tail flick test. All the above peptides were effective following administration into several PAG regions. Their antinociceptive effects were strongly evoked from the caudal ventrolateral and ventral PAG, including the dorsal raphe nucleus. A dose-dependent antinociception following administration into the ventrolateral PAG was demonstrated for all peptides studied. The effect of AII from the ventrolateral PAG was inhibited by the previous local administration of saralasin, a non-selective angiotensin receptor antagonist. Moreover, the peak effects of RS and AI occurred later than those of AII and AIII. The time-course of antinociception suggests that longer-chain peptides are locally processed to biologically active smaller-chain peptides. This study shows for the first time the antinociceptive effect of RS, AI, AII and III in well-defined PAG regions, an effect that is receptor mediated for AII.
Studies using an AT(1) receptor antagonist, losartan, demonstrated that depressor and bradycardic responses to angiotensin II (Ang II) injection into the nucleus tractus solitarii (NTS) are mediated via those receptors. We further characterized Ang II-evoked cardiovascular responses in this nucleus in spontaneously hypertensive rats (SHR) using a new, selective AT(1) receptor antagonist, valsartan. In alpha-chloralose-anesthetized Sprague-Dawley (S-D) rats, Wistar-Kyoto (WKY) rats, and SHR, unilateral injection of Ang II into the NTS decreased arterial pressure (AP) and heart rate (HR). This response was eliminated by preinjection of valsartan. Depressor responses were much greater in SHR than in WKY rats. In normotensive rats, bilateral valsartan injection did not alter baseline AP or HR, or baroreceptor reflex index (BRI) calculated as the maximal change in HR (bpm) divided by phenylephrine- or nitroprusside-induced maximal change in mean AP (mmHg). In SHR, this treatment did not alter baseline HR and BRI, but significantly increased AP. Preinjection of valsartan did not alter injected glutamate effects in any strain. Thus, stimulation of AT(1) receptors within the NTS contributes to cardiovascular regulation independently of the baroreceptor reflex and the glutamatergic system. This angiotensinergic system in SHR acts tonically to reduce AP.
The caudal ventrolateral medulla (VLM) has emerged during the last decade as one of the main components of the endogenous pain control system. Profound and long-lasting analgesia is produced by mild stimulation of the VLM. The VLMlat, the reticular formation located between the spinal trigeminal nucleus and the lateral reticular nucleus (LRt), appears to play a major role in that antinociceptive action. The projections to spinal cord laminae involved in nociceptive transmission originate exclusively in the VLMlat. The VLMlat participates in a disynaptic pathway involving spinally projecting pontine A5 noradrenergic neurons, which appears to convey alpha(2)-adrenoreceptor-mediated analgesia produced from the VLM. Neurons in the VLMlat and in lamina I are reciprocally connected by a closed loop that is likely to mediate feedback control of supraspinal nociceptive transmission. On the other hand, the LRt, which is targeted by ventral (lamina VII) and deep dorsal (laminae IV to V) horn inputs, projects to the premotor lamina VII. Nociceptive input ascending from the cord and increases in blood pressure are discussed as possible physiologic triggers of the analgesia produced by the VLM. The overall role of the VLM as a center for integration of nociceptive, cardiovascular, and motor functions is discussed. The putative therapeutic benefits of manipulating the VLM for the control of chronic pain are envisaged.