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Pain and analgesia: The dual effect of nitric oxide in the nociceptive system


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Nitric oxide (NO) is involved in many physiological processes and several lines of evidence have indicated that NO plays a complex and diverse role in the modulation of pain. Nitric oxide is an important neurotransmitter involved in the nociceptive process and, in the dorsal horn of the spinal cord, it contributes to the development of central sensitization. On the other hand, experimental data have also demonstrated that NO inhibits nociception in the peripheral and also in the central nervous system. In addition, it has been shown that nitric oxide mediates the analgesic effect of opioids and other analgesic substances. The information included in the present review aims to present and analyze data about the dual effect of NO on pain transmission and control, the molecular mechanisms involved in these effects and also the potential use of nitric oxide in pain therapy.
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Pain and analgesia: The dual effect of nitric oxide in the nociceptive system
Yara Cury
, Gisele Picolo
, Vanessa Pacciari Gutierrez
, Sergio Henrique Ferreira
Special Laboratory of Pain and Signaling, Butantan Institute, Vital Brazil Avenida, 1500, 05503-900 São Paulo, SP, Brazil
Department of Pharmacology, Faculty of Medicine, Ribeirão Preto, University of São Paulo, Ribeirão Preto, 14049-900 São Paulo, Brazil
article info
Article history:
Received 29 June 2010
Revised 17 February 2011
Available online 24 June 2011
Nitric oxide
Dual effect
NO–cGMP pathway
Nitric oxide (NO) is involved in many physiological processes and several lines of evidence have indicated
that NO plays a complex and diverse role in the modulation of pain. Nitric oxide is an important neuro-
transmitter involved in the nociceptive process and, in the dorsal horn of the spinal cord, it contributes to
the development of central sensitization. On the other hand, experimental data have also demonstrated
that NO inhibits nociception in the peripheral and also in the central nervous system. In addition, it has
been shown that nitric oxide mediates the analgesic effect of opioids and other analgesic substances. The
information included in the present review aims to present and analyze data about the dual effect of NO
on pain transmission and control, the molecular mechanisms involved in these effects and also the poten-
tial use of nitric oxide in pain therapy.
Ó2011 Elsevier Inc. All rights reserved.
Introduction. . . . . ...................................................................................................... 244
Nitric oxide and pain . . . . . . . . . . . . . . . . ................................................................................... 244
Nitric oxide as a mediator of pain at central level . . . . . . . . . . . . . . . . . . ................... ................................... 244
Nitric oxide as a mediator of pain in the periphery . . . . . . . . . . . . . . . . . .......... ............................................ 246
Nitric oxide and analgesia . . . . . . . . . . . . ................................................................................... 246
Evidences for the involvement of nitric oxide in the analgesic action of opioids, non-steroidal antiinflammatory drugs, natural products
and other analgesic drugs . . ................................................ ......................................... 247
Molecular mechanisms involved in the analgesic action of nitric oxide . ...................................................... 248
Understanding the dual role of nitric oxide on the nociceptive system . . . . . . . . . . . . . . ............................................. 248
Perspectives on the clinical use of NO as an analgesic drug . . . . ................................................................ 250
Concluding remarks . . . . . . . . . . . . . . . . . ................................................................................... 251
Acknowledgment . . . . . . . . . . . . . . . . . . ................................................................................... 251
References . . . . ...................................................................................................... 251
1089-8603/$ - see front matter Ó2011 Elsevier Inc. All rights reserved.
Abbreviations: 7-NI, 7-nitroindazole; AA, arachidonic acid; Ach, acetylcholine; AG, aminoguanidine hemisulfate; AKT, protein kinase B; AMPA,
methyl-4-isoxazolepropionic acid; ATP, adenosine triphosphate; CFA, complete Freund’s adjuvant; cGMP, cyclic guanosine 3
-monophosphate; CINODS, cyclooxygenase
inhibitor nitric oxide donors; CNGs, cyclic nucleotide gated cation channels; CNS, central nervous system; COX, cyclooxygenase; cPKG, cGMP-dependent protein kinase;
DbcGMP, dibutyrylguanosine 3:5
-cyclic monophosphate; GABA, gamma-aminobutyric acid;, intraplantar; IL-1b, interleukin-1 beta; IL-10, interleukin-10;
-arginine methyl Ester;
-NMMA, NG-monomithyl-
-arginina; NMDA, N-methyl-
-aspartate; NO, nitric oxide; NO
, nitrite;
, nitrate; NOS, nitric oxide synthase; eNOS, endothelial NOS; iNOS, inducible NOS; nNOS, neuronal NOS; NSAID, non-steroidal anti-inflammatory drugs; ODQ, 1H-
[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one; PAG, periaquedutal gray; PDE, phosphodiesterases; PG, prostaglandin; PGE
, prostaglandin E
; PGH, prostaglandin endoperoxide;
, prostacyclin; PI3K
, phosphoinositide-3-kinase
isoform; PKA, protein kinase A; PKC, protein kinase C; PKG, protein kinase G; PPAR-
, peroxisome proliferator
activated receptors; sGC, soluble guanylyl cyclase; SIN-1, 3-morpholinosydnonimine; SNAP, S-nitroso-N-acetylpenicillamine; SP, P substance; TNF
, tumor necrosis factor-
Corresponding author. Fax: +55 11 3726 1505.
E-mail address: (Y. Cury).
These two authors contributed equally to the review article.
Nitric Oxide 25 (2011) 243–254
Contents lists available at ScienceDirect
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The nitric oxide (NO) is derived from
-arginine by the action of
specific neuronal and non-neuronal forms of NO synthase (nNOS
and endothelial/eNOS or inducible/iNOS, respectively) [1–3]. Nitric
oxide and its associated enzymes are involved in many physiolog-
ical [1] and pathophysiological processes [2,3].
Several lines of evidence have indicated that both neuronal and
non-neuronal pools of NO play a complex and diverse role in the
modulation of nociception (see the sections ‘Nitric oxide and pain’,
‘Nitric oxide and analgesia’ and ‘Understanding the dual role of ni-
tric oxide on the nociceptive system’, for details).
Pain is described as an unpleasant sensation produced by pro-
cesses that either damage or are capable of damaging the tissues
(review in [4–6]). Such noxious stimuli are detected by high
threshold primary sensory nerve fibers called nociceptors (Ad
and C fibers). Nociceptors are free nerve endings that terminate
in the superficial layers of the dorsal horn of the spinal cord. In
the spinal cord, these primary sensory nerve fibers release neuro-
transmitters such as amino acids (glutamate) and neuropeptides
(such as substance P and calcitonin gene related peptide), which
activate second-order neurons. The second-order neurons convey
information via specific tracts that reach the thalamus where the
sensation of pain occurs. In the thalamus, a third-order neuron is
activated, traveling from the thalamus to the somatosensory cor-
tex, which allows for the perception of pain (review in [4–6]).
Clinical pain can arise either from damage to the nervous sys-
tem (neuropathic pain) or inflammatory states (inflammatory
pain) [7]. This type of pain is characterized by the presence of
spontaneous pain, hyperalgesia (an increase in the response to
noxious stimuli) and allodynia (the presence of pain in response
to normally innocuous stimuli). Multiple mechanisms are involved
in the generation of clinical pain. Inflammation or tissue lesion
causes the release of a variety of agents (bradykinin, cytokines,
eicosanoids, serotonin, histamine, cations), which, by acting on
their own receptors, contribute to alter the firing pattern of the pri-
mary sensory neurons, leading to nociception (review in [8]). The
binding of these substances to receptors results in activation of
intracellular signaling pathways, such as the cyclic adenosine
-monophosphate/protein kinase A or protein kinase C
ways, which phosphorylates voltage-gated ion channels [9–12].
Some of these substances also activate non-selective cationic chan-
nels present in the nociceptors. These channels include, for exam-
ple, the vanilloid-type receptors, the transient receptor potential
ankyrin type 1, the ATP-gated ion channels and acid-gated ion
channels. Once activated, these channels produce a net inward
electrical current that depolarizes the neuronal membrane and in-
creases the probability of action potential generation. At the Na
threshold, voltage gated sodium channels are activated, initiating
a burst of action potentials and propagating the conduction of sen-
sory input from the peripheral nerve ending to the dorsal horn of
the spinal cord. In addition to sodium channels, transmission of
the afferent pain signal to the central nervous system (CNS) is
mediated by voltage-gated calcium channels. It is generally
accepted that the increased intracellular calcium concentration
controls the release of neurotransmitters from the pre-synaptic
terminals in the dorsal horn upon arrival of the action potential,
being involved in neuronal excitability (review in [8,13–17]).
The dorsal horn of the spinal cord is the first site of synaptic
transfer in the nociceptive pathway and it is a major site for the
integration and modulation of the peripheral afferent signals.
Following peripheral lesion, the prolonged activity of the C fibers
increases the synaptic conduction in dorsal root neurons resulting
in central sensitization. This phenomenon is mediated by neuro-
transmitters such as glutamate, substance P and neurokinin. One
of the mechanisms involved in central sensitization is the
activation of the NMDA (N-methyl-
-aspartic acid) receptors by
glutamate. Activation of these receptors generates NO, which dif-
fuses out of the neuron to act on nerve endings and astrocyte pro-
cesses, acting as a neurotransmitter (review in [7,18,19]). The
central sensitization plays a major role in the generation of hyper-
sensitivity, resulting in the already described phenomena of hyper-
algesia and allodynia. The dorsal horn of the spinal cord is also
subject to local and descending mechanisms of pain modulation
(review in [6,18]). This is a physiological process that occurs in
the CNS and involves the release of opioid peptides, biogenic
amines and other transmitters. The endogenous pain modulation
systems consist of intermediate neurons within the dorsal horn
of the spinal cord and descending neural tracts that inhibit trans-
mission of the pain signal. These systems are activated by opioid
and GABAergic mechanisms in and around the periaqueductal gray
(PAG) region. Neurons from PAG project themselves, via glutama-
tergic descending pathways, to sites in the medullary reticular for-
mation and the locus ceruleus. The descending fibers from both
regions project themselves to the dorsal horn of the spinal cord
to synapse with the primary afferent neurons, the second-order
neurons, or interneurons. In the spinal cord, the descending
neurons release neurotransmitters (such as serotonin and norepi-
nephrine) and also activate small interneurons to release opioid
peptides, which modulate the incoming pain information (review
in [6,18]).
The pharmacological control of inflammatory pain is based
on two strategies. The first one involves the use of drugs that inhi-
bit nociceptor sensitization, and therefore, the development of
hypernociception. This is the main mechanism of action of aspirin
and aspirin-like drugs that, by inhibiting cyclooxygenases, prevent
nociceptor sensitization [20]. The second strategy involves the use
of drugs that directly block ongoing pain, resulting in antinocicep-
tion. This can be achieved by the use of morphine, dipyrone, nitric
oxide or NO donors, and also by other anti-inflammatory drugs (via
activation of the NO–cGMP pathway) (see references in the section
‘Nitric oxide and analgesia’).
As described above, nitric oxide is an important mediator of
nociception and it is unequivocally involved in central sensitiza-
tion; however, experimental and clinical evidences have demon-
strated that NO is also capable of inducing analgesia. Taken
together, these data indicate that NO plays a complex and diverse
role in the modulation of nociceptive processing. The information
included in the present review aims to present and analyze data
about the dual effect of NO on pain transmission and control, the
molecular mechanisms involved in these effects and also the
potential use of nitric oxide in pain therapy.
Nitric oxide and pain
Nitric oxide is a pivotal signaling molecule that plays an impor-
tant role in acute [21] and chronic [22] pain states at both central
[23] and peripheral [24] levels (Table 1).
Nitric oxide as a mediator of pain at central level
In the central nervous system (CNS), NO is concentrated in the
dorsal horn of the spinal cord, wherein it is derived from diverse
sources (including glial cells). Neuronal NO synthase (nNOS) is
the predominant form of NOS in the dorsal horn and has a definite
role in spinal cord circuits. Besides nNOS, the eNOS is also found in
some neuronal populations and blood vessels [23,25]. Concerning
iNOS, there is some disagreement whether this isoform also partic-
ipates in the central transmission of pain or if it is expressed only
244 Y. Cury et al. / Nitric Oxide 25 (2011) 243–254
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peripherally during inflammatory or neuropathic pain processes
The production of nitric oxide by nNOS in the CNS requires the
influx of Ca
. This influx occurs through, and is dependent on, acti-
vation of N-methyl-
-aspartate (NMDA) receptors [26,27]. The in-
crease in the intracellular levels of Ca
triggers a cascade of
events that include activation of nNOS, followed by the increase
in NO production [25].
The major intracellular receptor for NO is a soluble guanylyl cy-
clase (sGC). Its activation by NO, results in conversion of guanosine
triphosphate to the second messenger cGMP. The NO–cGMP sig-
naling pathway is present in neurons of the spinal cord and has
been implicated in synaptic plasticity such as central sensitization
[28]. The stimulation of cGMP is one of the numerous direct biolog-
ical actions of NO and the activity of this intracellular signaling
molecule modulates the activity of many cell targets, including
cGMP-dependent protein kinase (cPKG), ion channels and phos-
phodiesterases [25].
As pointed out above, the NO generated by activation of NMDA
receptors, has been implicated in synaptic plasticity and multiple
mechanisms are involved in central sensitization caused by nitric
oxide (review in [29]). NO can diffuse out of the neuron to act on
nerve endings and astrocyte processes, acting as a neurotransmit-
ter. NO can also enhance the release of SP and calcitonin-gene-
related peptide from C-fiber terminals [25], contributing to the
development of secondary hyperalgesia (Fig. 1). Furthermore, it
has been suggested that spinal NO participates in glutamatergic
mechanisms of descending facilitation triggered by illness and
inflammation, weakening the influence of descending inhibition
upon dorsal horn neurons. This weakening occurs, at least partially,
by interfering with GABAergic and glycinergic inhibitory tone upon
projection neurons [30,31].
The involvement of NO in nociceptive processes is supported by
experiments in which inhibitors of NOS were used in order to re-
duce NO production. By using the nitric oxide synthase inhibitor
-arginine methyl ester (
-NAME), it was demonstrated
that NO is involved in the maintenance of behavioral signs of neu-
ropathic pain induced in rats, by constriction of both L5 and L6
spinal nerves [32] and sciatic nerve [33]. In the sciatic nerve con-
striction model, the thermal hyperalgesia, one of the phenomena
that characterizes neuropathic pain, is blocked by intrathecal
administration of either an alternate substrate for nitric oxide syn-
thase (NW-nitro-
-arginine methyl ester), or the soluble guanylate
cyclase inhibitor, methylene blue, for a period of 2 and 4 h, respec-
tively. This suggests that thermal hyperalgesia produced by liga-
tion of sciatic nerve is mediated by the production of nitric oxide
and subsequent activation of soluble guanylate cyclase in the lum-
bar spinal cord [34]. The NO involved in hyperalgesia and allodynia
detected in rats undergone sciatic nerve transaction is derived
from the neuronal isoform of NOS, since these phenomena are
inhibited by intrathecal administration of 7-NI, a selective inhibitor
of nNOS [33].
Table 1
‘‘In vivo’’ evidences for NO as analgesic and/or hyperalgesic mediator at central and
peripheral levels.
Effect in the nociceptive system Site of action Reference
Thermal hyperalgesia CNS Meller et al. [34]
Thermal hyperalgesia Periphery Chen et al. [22]
Mechanical allodynia, cold allodynia
and cold-stress exacerbated
ongoing pain
CNS Yoon et al. [32]
Mechanical hyperalgesia
and allodynia
CNS Chacur et al. [33]
Antinociception CNS Chung et al. [68]
Antinociception Periphery Duarte et al. [67]
Dual effect CNS Tegeder et al. [145]
Dual effect CNS Sousa and Prado [166]
Dual effect CNS Li and Qi [59]
Dual effect Periphery Kawabata et al. [162]
Dual effect Periphery Prado et al. [163]
Dual effect Periphery Rocha et al. [161]
This provides a summary of anti- or pro-nociceptive or dual effect induced by NO at
peripheral and central levels. CNS, central nervous system.
Fig. 1. Involvement of nitric oxide in central sensitization. The prolonged activity of C fibers releases, in the dorsal horn of the spinal cord, a variety of neurotransmitters, such
as substance P, calcitonin-gene related peptide and glutamate that binds to their own receptors. Activation of NMDA receptors by glutamate induces an influx of Ca
triggers a cascade of events, including activation of nNOS, followed by increased production of NO. This NO not only activates the guanyl cyclase on the postsynaptic neuron
but can diffuse across the synaptic cleft back into the synapse that originally released glutamate. This retrograde action of NO reinforces the glutamatergic signaling. NO may
also diffuse out of the neuron, acting on astrocytes and microglia. NO, acting on these cells, can induce the formation of NO from iNOS, which can act again on the pre- and/or
post-synaptic neurons. NKA, neurokinin A; SP, substance P; NK-1, substance P receptor; NK-2, neurokinin A receptor; CGRP, calcitonin-gene related peptide; CGRPr,
calcitonin-gene related peptide receptor; NMDA, ionotropic receptor by glutamate; AMPA, ionotropic receptor by glutamate; mGlu, metabotropic receptor; NO, nitric oxide;
nNOS, neuronal nitric oxide synthase.
Y. Cury et al. / Nitric Oxide 25 (2011) 243–254 245
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Corroborating those behavioral data, a marked increase of nNOS
mRNA expression in the lumbar spinal cord tissue and in dorsal
root ganglia has been observed in rats, few days after sciatic nerve
transaction, persisting for more than 2 months [33,35]. Also, an in-
crease in the number of neurons that express nNOS immunoreac-
tivity has been detected, being this immunoreactivity located in
neurons present at the dorsal horn (lamina II–III) and around the
central canal (lamina X) of the lumbar spinal cord [33]. Interest-
ingly, recent data demonstrated that the presence of iNOS is re-
quired to increase the spinal cord expression of nNOS caused by
sciatic nerve injury [36]. The increase in the number of nNOS-posi-
tive neurons observed at the spinal level after peripheral injury is
detected either in the ipsilateral and contralateral side of the le-
sion, being the increase detected in the ipsilateral side higher than
that of the contralateral one [37].
These results indicate that NO may play a role in the central
mechanisms involved in the development of nociceptive phenom-
ena after peripheral inflammation/tissue lesion.
Nitric oxide as a mediator of pain in the periphery
The participation of NO, derived from different isoforms of NOS,
as a key mediator of nociceptive phenomena has also been demon-
strated in the periphery, by different experimental approaches
Chen et al. [22] recently demonstrated that pretreatment with
-NAME (non-selective NOS inhibitor), 7-nitroindazole (selective
nNOS inhibitor), aminoguanidine hydrochloride (selective iNOS
inhibitor), but not with
-N(5)-(1-iminoethyl)-ornithine (
selective eNOS inhibitor), significantly attenuated thermal hyper-
algesia induced by intraplantar ( injection of complete Fre-
und’s adjuvant (CFA) in mice. Using a real-time reverse
transcription-polymerase chain reaction, these authors observed
an increase in nNOS, iNOS, and eNOS gene expression in plantar
skin tissue of mice after intraplantar ( injection of CFA. An in-
crease in tumor necrosis factor-alpha (TNF
), interleukin-1 beta
(IL-1b), and interleukin-10 (IL-10) gene expression was also
observed. Pretreatment with the NOS inhibitors prevented the
CFA-induced increase in the pro-inflammatory cytokines TNF
and IL-1bbut, interestingly, these inhibitors increased even more
the anti-inflammatory cytokine IL-10 gene expression. In addition,
these authors also observed that nNOS-, iNOS- or eNOS-knockout
(KO) mice had lower gene expression of TNF
, IL-1b, and IL-10
following CFA. These results demonstrated a great interaction be-
tween NO and cytokines in the periphery, suggesting a feedback
loop between NO and cytokines on the modulation of inflamma-
tory pain.
The involvement of NO in peripheral nociception is also corrob-
orated by data demonstrating the local release of NO by inflamma-
tory stimulus. Omote et al. [38] and Toriyabe et al. [21] observed an
increase of the total amount of nitrite and nitrate ðNO
lected by a microdialysis probe implanted subcutaneously into the
glabrous skin of hindpaws of Sprague–Dawley rats, after injection
of carrageenin. The increase in the dialysate concentrations of
induced by carrageenin was suppressed by
-NMMA in
a period of 8 h of collection. When using aminoguanidine hemisul-
fate (AG), an iNOS inhibitor, this suppression occurs from 2.5 h
after carrageenin injection. These results indicate that the increase
in the production of NO induced by carrageenin is mediated by
nNOS in the early phase (3 h) and by both nNOS and iNOS in
the late phase (3.5 h). These data also suggest that both nNOS,
in peripheral nerves, and iNOS, in inflammatory cells, contribute
to the production of peripherally released NO during inflammation
Toriyabe et al. [21] also observed that, in addition to NO, periph-
eral concentrations of prostaglandin E
), and prostacyclin
) were increased after carrageenin injection into the rat
paw. Interestingly, the increase in the prostanoids concentrations
was completely inhibited by the NOS inhibitor
-NMMA. Several
studies have suggested an interaction between NO and PG, being
the PGE
production increased in the presence of NO and inhibited
in the presence of NOS inhibitors [41–43]. It has been proposed
that the NO-induced increase in PGs is due to the ability of NO to
activate cyclooxygenases (COX), demonstrated in in vitro assays
[44–46]. Furthermore, Toriyabe et al. [21] observed that the
peripheral release of PGE
and PGI
induced by carrageenan is
due to activation of COX-1 in the early phase, and up-regulation
of COX-2 expression in the late phase of the inflammatory re-
sponse, and that both phenomena are facilitated by NO. Taken to-
gether, these results indicate that NO can induce peripheral
hyperalgesia by regulation of the expression and/or activity of
cyclooxygenases, resulting in an increase of PGs release. It is
important to stress that the molecular mechanisms by which NO
activates COX remain undetermined as yet. Some possible mecha-
nisms have been proposed: (1) COX is a heme containing enzyme,
and heme has very high affinity for NO. Then, it is possible that NO
stimulates COX-1 and/or COX-2 increasing the catalytic activity of
COX manifold, by combining with heme moiety or by nitrosylating
the tyrosine residues of COX [47]; (2) peroxynitrite generated by
NO is able to react with the peroxidase of prostaglandin endoper-
oxide (PGH) synthase to induce its cyclooxygenase activity. In-
creased activation of PGH synthase results in enhanced
conversion of arachidonic acid (AA) to prostaglandins [48]; (3)
NO nitrosylates cysteine residues in the catalytic domain of COX
enzymes, leading to the formation of nitrosothiols, which can pro-
duce changes in the structure of the enzyme, resulting in an in-
creased catalytic efficiency (review in [21,39]). NO interacts with
, decreasing the amount of the radical in the tissue. Since this
radical has been postulated to be involved in the auto-inactivation
of the COX enzyme, it is therefore possible that NO augments COX
activity preventing its inactivation (review in [39]).
Nitric oxide and analgesia
Besides the role of nitric oxide in nociceptive pathways, several
lines of evidence have indicated that NO induces analgesia (Table
1) and also that it mediates the peripheral and central antinocicep-
tive effect of analgesic compounds, such as opioids, non-steroidal
anti-inflammatory drugs and natural products [49–62] (Table 2).
The first evidence that NO induces analgesia and is also in-
volved in the peripheral antinociceptive effect of analgesic drugs
was provided by Professor Sergio Henrique Ferreira and his group,
in the early 1990s. Based on previous data from the literature
showing that acetylcholine (ACh) exerts analgesic action [63,64]
and that ACh induces nitric oxide (NO) release from endothelial
cells [65,66], Prof. Ferreira investigated if the analgesic action of
ACh is mediated by the release of nitric oxide. By using the rat
paw hyperalgesia test [67], it was observed that intraplantar injec-
tion of sodium nitroprusside, a substance which nonenzymatically
releases nitric oxide (NO), causes analgesia. Antinociception in-
duced by acetylcholine, but not by sodium nitroprusside, was
blocked by NG-monomethyl-
-arginine (
-NMMA), an inhibitor of
the formation of NO from
-arginine. Furthermore, the analgesic
effects of acetylcholine (ACh) and sodium nitroprusside were
enhanced by intraplantar injection of My5445 (inhibitor of cyclic
GMP phosphodiesterase) and inhibited by methylene blue (inhibi-
tor of guanylate cyclase) [67]. Taken together, these results
indicated that NO mediates the peripheral analgesic action of
acetylcholine and also that NO induces, per se, analgesic effect.
Moreover, these data suggested that the analgesic action of NO
depends on an intracellular signaling pathway involving the
formation of cyclic GMP [67].
246 Y. Cury et al. / Nitric Oxide 25 (2011) 243–254
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In addition to data showing that nitric oxide donor induces
peripheral antinociception, the role of NO in pain control was also
shown at central levels.
-Arginine and the nitric oxide donor 3-
morpholinosydnoimine (SIN-1), intracerebroventricularly admin-
istered to mice, produce antinociception. This effect seems to be
mediated by dynorphin and dependent on nitric oxide. This sug-
gestion was based on the observation that naloxone and also the
intracerebroventricular administration of a rabbit antiserum
against rat dynorphin block the analgesic effect of both substances.
Moreover, the effect induced by
-arginine was also antagonized by
an inhibitor of nNOS [68].
Several lines of evidence have also indicated that, in the CNS, ni-
tric oxide is involved in the descending inhibitory control of noci-
ception (review in [31]). NO interacts with noradrenergic and
cholinergic mechanisms involved in the descending inhibition
pathways, and a reciprocal interplay between NO,
and ACh in the induction of antinociception in the dorsal horn of
the spinal cord has been observed [69–79].
Evidences for the involvement of nitric oxide in the analgesic action of
opioids, non-steroidal antiinflammatory drugs, natural products and
other analgesic drugs
Since the pioneering observations of Duarte et al. [67] indicat-
ing that NO induces analgesia and also mediates the analgesic
effect of ACh, several experimental works have been carried out
indicating NO as a mediator of the peripheral and central antinoci-
ceptive effect of analgesic compounds, such as opioids, non-
steroidal antiinflammatory drugs and natural products. Based on
(a) the demonstration that NO mediates the peripheral analgesic
action of Ach, (b) the similarity of the local action of ACh and opi-
oids, and (c) data from the literature showing that opioids induce
the production of cGMP in neurons [80], Prof. Ferreira and his
group investigated whether agents which affect the
NO–cGMP pathway also interfere with morphine-induced periph-
eral analgesia. By using, again, the rat paw hyperalgesia test and
pharmacological approaches, Ferreira et al. [81] evidenced that
peripheral morphine analgesia is subsequent to activation of the
-arginine/NO–cGMP pathway.
It is interesting to point out that, when evaluating the involve-
ment of the NO–cGMP pathway in the effect of morphine in the
central nervous system, Ferreira et al. [81] did not detect a role
for nitric oxide, despite evidencing the involvement of cGMP in this
effect. As discussed earlier in the section ‘Nitric oxide and pain’,
experimental evidence has indicated that endogenous nitric oxide,
produced at the central nervous system, mainly at spinal and
supraspinal levels, acts as a nociceptive mediator. Furthermore,
several data have also shown that the increase in NO levels, at cen-
tral sites, attenuates morphine antinociception, whereas the de-
crease in its levels enhances the analgesic effect of the opioid
[82–95]. Moreover, the involvement of NO in the adverse effects
of opioids has also been evidenced by the demonstration that tol-
erance to and dependence on morphine or its withdrawal syn-
drome are prevented by nitric oxide synthase inhibition (review
in [96]). Despite the conflicting data, the role of NO as a mediator
of the central analgesic effect of morphine has been consistently
confirmed and several data have indicated that it also potentiates
the actions of systemically administered morphine [97–102].In
addition, it has been shown that morphine, via activation of opioid
receptors, induces an overall stimulation of NOS biosynthesis,
increasing the levels of NOS mRNA and nNOS expression (review
in [96]). Data from the literature also suggest that supraspinal ni-
tric oxide production modulates the transmission of opioid pain-
inhibitory signals in the central nervous system [100].
In addition to morphine, the contribution of the
NO–cGMP pathway to the analgesic action of other non-selective
and selective
-, d-, and
-opioid receptor agonists has also been
evidenced [53,84,85,103–115]. Furthermore, the involvement of
this pathway has also been demonstrated for a number of other
substances capable of inducing analgesia, such as dipyrone and
other non-steroidal anti-inflammatory drugs, statins, cannabi-
noids, the antiepileptic gabapentin, adenosine, PPAR-
Table 2
NO as a mediator of the analgesic effect of drugs or alternative analgesic therapies.
Drug or drug class or alternative therapy Reference
Ach Duarte et al. [67]
Morphine Ferreira et al. [81]
Morphine Kolesnikov et al. [97], Pataki and Telegdy [98], Song et al. [99], Abacioglu et al. [101], Chen and Pan [102] and Javanmardi
et al. [100]
Opioids Javanmardi et al. [100], Brignola et al. [84], Dambisya and Lee [85], Granados-Soto et al. [53], Nozaki-Taguchi and
Yamamoto [103], Mixcoatl-Zexuatl et al. [115], Amarante and Duarte [104], Jain et al. [114], Hayashida et al. [113],
Tasatargil and Sadan [105], Oritz et al. [106], Pacheco et al. [107], Yalcin and Aksu [108], Chena et al. [109], Menendez
et al. [110], Clemente-Napimoga et al. [112] and Hervera et al. [111]
Loperamide Menendez et al. [110]
Dipyrone and other non-steroidal anti-
inflammatory drugs
Lozano-Cuenca et al. [178] and Ventura-Martínez et al. [57]
Statins Santodomingo-Garzón et al. [123]
Cannabinoids Reis et al. [179]
Gabapentin Ortiz et al. [152]
agonists Pena-dos-Santos et al. [150]
-Adrenoceptor agonist xylazine Romero and Duarte [180]
Hormones such as estradiol and melatonin Fávaro-Moreira et al. [122] and Hernández-Pacheco [119]
Bovine lactoferrin Hayashida et al. [113]
Anesthetic gas nitrous oxide Li et al. [139], Emmanouil et al. [181] and Cope et al. [138]
Isosorbide dinitrate spray Yuen et al. [117]
Hydrogen sulfide releasing drugs Distrutti et al. [151]
Phosphodiesterase inhibitors Mixcoatl-Zecuatl et al. [115], Jain et al. [114] and Vale et al. [182]
Electroacupuncture Xu et al. [72] and Almeida and Duarte [127]
Exercise Galdino et al. [183]
Hyperbaric oxygen therapy Ohgami et al. [129] and Zelinski et al. [137]
Natural products, including plants extracts
and animal toxins
Pu et al. [125], Picolo et al. [126], Luiz et al. [131], Guginski et al. [130] and Gutierrez et al. [61]
This provides a summary of drugs or class of drugs or alternative therapies that induces antinociception mediated by NO. Ach, acetylcholine; PPAR-
, peroxisome proliferator
activated receptors.
Y. Cury et al. / Nitric Oxide 25 (2011) 243–254 247
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-adrenoceptor agonist xylazine, hormones such as estradiol
and melatonin, bovine lactoferrin, the anesthetic gas nitrous oxide,
isosorbide dinitrate spray, hydrogen sulfide releasing drugs and
phosphodiesterase inhibitors [50,51,54–58,116–124]. In most of
the studies referred above, the involvement of the NO–cGMP path-
way in the antinociceptive effect of the analgesics was demon-
strated by the use of NO synthase inhibitors, as well as of
guanylyl cyclase inhibitors, administered in doses that display only
peripheral (local) effects. It is important to stress that all these
analgesic drugs, including morphine, are capable of blocking ongo-
ing hypernociception. Taken together, these data indicate that NO
is probably the common denominator for the mode of action of
peripheral analgesic drugs that directly block ongoing nociceptor
In addition to the analgesic drugs, the importance of nitric oxide
for alternative analgesic therapies such as hyperbaric oxygen
therapy, electroacupuncture, natural products (including plants
extracts and animal venoms/toxins), and for the analgesic action
of exercise, has also been demonstrated [58,61,125–134]. Alto-
gether, these literature findings confirm that NO displays an
important role in pain control.
Despite the available data on the role of NO as a mediator of the
analgesic activity of various drugs, the type of NO synthase (NOS)
isoform involved in NO generation is not well determined yet. By
using pharmacological or genetic approaches (knock out animals
or antisense oligodeoxynucleotide directed against nNOS), several
authors have undoubtedly demonstrated that peripheral and also
central nNOS are involved in the analgesic effect induced by
substances that activate the NO–cGMP pathway [60–62,97,99,
127,129,135–141]. A contribution of eNOs for the analgesic activity
of opioids was also observed. Menendez et al. [110] showed that
the peripheral antihyperalgesic effect of loperamide on osteosar-
coma-induced thermal hyperalgesia in mice is mediated by activa-
tion of the NO/cGMP and that eNOS is involved in the activation of
this pathway. Despite being well demonstrated that iNOS displays
pro-inflammatory and nociceptive effects, Almeida and Duarte
[127] evidenced in rats, that orofacial antinociception induced by
electroacupuncture is mediated by activation of the nitric oxide/
cGMP pathway and that, in addition to nNOS, the activity of iNOS
is important for the generation of NO.
Molecular mechanisms involved in the analgesic action of nitric oxide
As pointed out above, the antinociceptive effect of nitric oxide
involves the generation of cGMP. Several lines of evidence have
suggested a link between activation of the NO–cGMP pathway
and the opening of ATP-sensitive K
channels. This suggestion is
based on experimental data demonstrating that the peripheral
antinociceptive effect of nitric oxide donors as well as of dibu-
tyrylguanosine 3:5
-cyclic monophosphate (DbcGMP), a mem-
brane permeable analog of cyclic GMP, is mediated through
specific opening of ATP-sensitive K
channels [142,143]. Moreover,
an additive antinociceptive effect of the combination of diazoxide
(activator of ATP-sensitive K
channels), sodium nitroprusside
and dibutyryl-cGMP was also observed [144]. Furthermore, other
works have shown that the peripheral antinociceptive effect of
various drugs or natural products that stimulate the
NO–cGMP pathway, is mediated [145] by activation of ATP-sensi-
tive K
channels [56,58,61,107,108,110,119,121,132,133,146–153].
These findings were further supported by electrophysiological
studies indicating that morphine induces, in nociceptive neurons,
an increase in ATP-sensitive K
channels followed by hyperpolar-
ization of the neuron [60]. These data indicate that NO, or drugs
capable of activating the NO–cGMP pathway, causes antinocicep-
tion via opening ATP-sensitive K
channels with consequent
increase in the K
current, restoring the normal high nociceptor
threshold. It is important to stress that the role of ATP-sensitive
channels in modulating the enhanced excitability that results
from inflammatory or injury conditions was also confirmed by
electrophysiological studies [154].
It is well demonstrated that cGMP can directly or indirectly (via
PKG stimulation), modulate the activity of ion channels [155–160].
PKG is a protein kinase that is stimulated selectively, but not exclu-
sively, by cyclic GMP. Once stimulated, PKG induces inhibition of
phospholipase C activity, stimulation of Ca
-ATPase activity, inhi-
bition of inositol 1,4,5-triphosphate, inhibition of Ca
and/or stimulation of K
channels activity. The understanding of
how the increased concentration of cGMP promotes the opening
of ATP-sensitive K
channels and the consequent nociceptor desen-
sitization was achieved by Sachs et al. [160]. These authors demon-
strated, in rat models of acute and persistent pain, that the specific
PKG inhibitor, KT5823, inhibited, in a dose-dependent manner, the
peripheral antinociceptive effect of the cGMP analog, 8-bromo-
cGMP, morphine, dipyrone, and SNAP. Since PKG is capable to
phosphorylate ion channels [156–158], it is plausible to suggest
that cGMP, at least in part, by activating PKG, may promote the
opening of the ATP-sensitive K
channels, restoring the nociceptor
All the data presented in this section demonstrate that the anal-
gesic action of nitric oxide depends on an intracellular signaling
pathway. The first step in this pathway involves the formation of
cyclic GMP, activation of PKG and the consequent opening of K
channels. Data herein presented also indicate that the NO–
cGMP–PKG pathway mediates the analgesic effect of various drugs,
including opioids, dipyrone and some antiinflammatory agents.
The mechanisms by which these drugs stimulate the production
of NO were not well characterized yet; however, Cunha et al.
[60] have recently demonstrated, through in vivo and in vitro stud-
ies that, at least for morphine, activation of the nitric-oxide path-
way is dependent on an initial stimulation of PI3K
/AKT protein
kinase B (AKT) that occurs after activation of
opioid receptors
by the agonist. In Fig. 2, a schematic representation of the molecu-
lar basis of peripheral NO-induced analgesia is proposed.
Understanding the dual role of nitric oxide on the nociceptive
As evidenced by data presented in the sections ‘Nitric oxide and
pain’ and ‘Nitric oxide and analgesia’ (Table 1), NO has a dual role
in the regulation of pain processes, i.e., it can mediate nociception
or induce an antinociceptive effect. This dual effect occurs at cen-
tral and/or peripheral levels and many studies have been carried
out in order to understand the dual effect of NO (Table 3).
Rocha et al. [161] investigating the contribution of nitric oxide
to the nociceptive phenomena induced by intraarticular injection
of zymosan in rats, have shown that inhibitors of NOS decrease
articular inflammatory pain; however this inhibitory effect was ob-
served only when the inhibitors were given before the induction of
arthritis. These authors also observed that administration of NO
donors induces antinociceptive effect, being this effect observed
for the drugs administered after the injection of zymosan. These re-
sults indicate that the endogenous NO is an important mediator in-
volved in the development of zymosan-induced arthritis, whereas
pharmacological administration of NO can inhibit the ongoing
nociceptive phenomena.
Kawabata et al. [162] also observed that different doses of NO
can induce nociception or antinociceptive effect. These authors
demonstrated that intraplantar injection of low doses (0.1–1
paw) of
-arginine enhances the second phase response of the for-
malin-induced nociception in mice, whereas the administration of
a high dose (10
g/paw) suppresses this nociceptive effect. Both
248 Y. Cury et al. / Nitric Oxide 25 (2011) 243–254
Author's personal copy
actions of
-arginine were reversed by injection of
demonstrating that both the pro- and antinociceptive effects of
arginine are mediated by peripheral (local) NO formation.
Prado et al. [163] also observed that different doses of NO can
induced a dual effect on the nociceptive system. These authors
showed that local application of a cream containing a nitric oxide
donor S-nitroso-N-acetylpenicillamine (SNAP 1% and 5%), signifi-
cantly reduced the tactile allodynia in an incision model of pain.
The antinociceptive effect of the NO donor was detected 2 h after
drug application and persisted for at least 24 h. In contrast, higher
concentrations of NO donors produced a smaller or no effect. More-
over, a cream containing 30% SNAP caused a significant pro-allo-
dynic effect, detected 24 h after drug administration. It is
important to point out that the antinociceptive effect of SNAP
was due to a local effect of NO, since the incision allodynia was sig-
nificantly inhibited only in the paw treated with the SNAP cream,
but remained unchanged in the contralateral one.
Some nitric oxide synthase inhibitors, such as
-NAME, may act
as a partial agonist, stimulating, instead of inhibiting, iNOS and
guanylyl cyclase, which may contribute to observed dual effect of
NO in nociceptive system [164,165]. In agreement with this
hypothesis, it was demonstrated that chronic administration of
-NAME to guinea pigs and rats does not result in a sustained
suppression of nitric oxide synthesis, because of the compensatory
expression of the inducible form of nitric oxide synthase [165].
The dual role of NO in nociception has also been demonstrated
in the CNS. Sousa and Prado [166] showed that intrathecal
administration of 3-morpholinosydnonimine SIN-1, a NO donor,
produces a dual dose-dependent effect in a model of neuropathic
pain in rats. Low intrathecal doses (0.1–2.0
l) reduced
(i.e., caused antinociception), whereas higher doses enhanced
(i.e., caused pronociceptive effect – 10 and 20
l) or had no
effect (5 or 100
l) on the mechanical allodynia evoked by
chronic ligature of the sciatic nerve. In contrast, the same dose
range of SIN-1 (10 and 100
l) produced a dose-dependent
increase in the latency for the tail-flick reflex (i.e., caused antinoci-
ception to thermal noxious stimulation). The inhibitor of guanylate
cyclase, 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) (4
l; i.t.), abolished the antinociceptive effects of SIN-1 in both
tests and reduced the pronociceptive effect of high doses of SIN-
1 in neuropathic rats, indicating that both effects (nociception
and antinociception) occur through a spinal mechanism that in-
volves activation of guanylate cyclase. Similar results were re-
cently observed by Li and Qi [59]. These authors demonstrated
that, in rats, intrathecal administration of low doses of
inhibited the nociceptive responses induced by the intraplantar
Fig. 2. Schematic representation of the molecular basis of peripheral NO-induced analgesia. The analgesic action of nitric oxide depends on an intracellular signaling pathway
that includes the formation of cyclic GMP, activation of PKG and the consequent opening of K
channels. The opening of these channels increases the K
current causing
hyperpolarization of nociceptive neurons. NO–cGMP–PKG pathway mediates the analgesic effect of various substances, including opioids, dipyrone and animal venoms. The
stimulation of nitric-oxide pathway by morphine is dependent on an initial stimulation of PI3K
/AKT. Ca
, calcium channel; Na
, sodium channel; K ATP, ATP-sensitive K
channels; nNOS, neuronal nitric oxide synthase; NO, nitric oxide; GTP, guanosine triphosphate; Gc, guanylate cyclase; cGMP, cyclic guanosine monophosphate; PKG, protein
kinase G; PI3K
/AKT, phosphoinositide-3-kinase
isoform/protein kinase B.
Table 3
Dual effect of NO on nociceptive system.
Variables Experimental protocol Effect References
Time Post-treatment with NO donors No effect Rocha et al. [161]
Pre-treatment with NO donors Antinociception
Dose (NO donors or precursor) Low dose Antinociception Sousa et al. [166], Li and Qi et al. [59],
Prado et al. [163] and Tegeder et al. [145]High dose Nociception
High dose Antinociception Kawabata et al. [162]
Low dose Nociception
Model Thermal noxious stimulation Antinociception Sousa et al. [166]
Mechanical allodynia Nociception
Administration route of NO donors Subcutaneous Antinociception Vivancos et al. [168]
Intradermal Nociception
Y. Cury et al. / Nitric Oxide 25 (2011) 243–254 249
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administration of formalin, whereas the injection of high doses of
the NO precursor increases this response. Taken together, these
data suggest that the dual effect of NO depends not only on the
doses of the NO donors, but also on the experimental model used
for pain evaluation.
Based on data demonstrating that NO displays a dual effect in
pain transmission, a possible mirrored dual effect of cGMP was also
investigated. The rat formalin assay was used to assess the effects
of the cyclic guanosine mono-phosphate (cGMP) analog, 8-bromo-
cGMP on nociception and on the expression of cGMP dependent
protein kinase I (protein kinase G; PKG-I) in lumbar spinal cord.
Intrathecal (i.t.) delivery of low doses of 8-bromo-cGMP (0.1–
0.25 mmol) reduced nociceptive behavior and formalin induced
upregulation of PKG-I in the spinal cord while medium doses
(0.5–1 mmol i.t.) had no effect. On the other hand, high doses of
8-bromo-cGMP (2.5 mmol i.t.) caused hyperalgesia associated with
a further increase of PKG-I expression [145]. Trying to explain
these dose dependent contrary effects, these authors investigated
the potential involvement of various cGMP targets in both
nociceptive and antinociceptive effects induced by 8-bromo
c-GMP. The obtained data indicate that cGMP-induced hyperalge-
sia apparently involves protein kinase G (PKG-I) activation and
upregulation, whereas cGMP-induced antinociception is PKG,
cyclic nucleotide gated cation channels (CNGs) and phosphodies-
terases (PDE2 and PDE3) independent. In addition, these authors
observed that a reduction of AMPA (
methyl-4-isoxazolepropionic acid) receptor currents is important
for to the antinociceptive effect of 8-bomo-cGMP [145]. These
results suggest that the dual effect of NO may result from distinct
intracellular signaling pathways.
All together, these data indicate that: (a) the pro- or anti-
nociceptive effect of NO is dependent on the type and the phase
of the nociceptive process and also on the nociceptive stimuli
(mechanical or thermal) used [53,161,166,167] and (b) NO donors
or inhibitors, depending on the dose, could cause nociception or
antinociception [163,166]. In addition, it has been observed that
depending on the site of activation, the
-arginine/NO/cGMP path-
way could induce opposite effects, i.e., nociception or antinocicep-
tion. Intradermal administration in the rat paw, of drugs capable of
activating the NO/cGMP pathway, induces nociceptive phenomena,
whereas the subcutaneous injection of these drugs results in
antinociception. These findings suggest the existence of different
subsets of nociceptive primary sensory neurons in which NO plays
opposing roles [168].
Concerning the role of NO as a mediator of the action of analge-
sic drugs, it has also been observed that NO display a dual effect. As
pointed out in the section ‘Evidences for the involvement of nitric
oxide in the analgesic action of opioids, non-steroidal antiinflam-
matory drugs, natural products and other analgesic drugs’, it is well
demonstrated that NO mediates the peripheral analgesic effect of
morphine, but decreases the analgesic potency of this opioid in
the CNS (review in [96]). The mechanisms involved in this dual role
of NO in modulating morphine activities are still not well under-
stood and the discrepancies that are to be found in the Literature
might be related to differences in (a) animal species, (b) nocicep-
tive stimuli, (c) site of opioid administration and/or (d) doses of
the inhibitors of NO or NOS substrate used in the studies. In fact,
the importance of the site of administration of opioids and/or the
doses of NOS substrate to the distinct results described in the liter-
ature could be evidenced by data from Bhargava and Bian [82,88]
and Bian and Bhargava [89]. These authors showed that acute or
chronic administration to mice of
-arginine (a substrate for NOS
that forms NO) decreases morphine analgesia. The reduced effect
of the opioid is related to the decrease in the amount of morphine
that enters the central sites (midbrain, pons and medulla, hippo-
campus, corpus striatum and spinal cord). Interestingly, the
inhibitory effect of
-arginine on morphine antinociception and
distribution in the CNS is dependent on the dose of the NOS sub-
strate and observed only for morphine administered by s.c. route,
but not detected with intracerebroventricularly-administered
morphine. These observations suggest that the action of
might be related to the decreased levels of the opioid in the brain.
The inhibitory effect of
-arginine on morphine-analgesia and entry
into the CNS is blocked by the NOS inhibitor
-NNA, suggesting that
the NO–NOS system may play a role in the regulation of blood–
brain barrier to morphine. The complexity of the role of NO in
modulating the action of opioids could also be evidenced by
experimental data showing that, depending on the isoform of neu-
ronal nitric oxide synthase (nNOS-1 or nNOS-2), different roles of
nitric oxide in morphine analgesia could be observed. Kolesnikov
et al. [169] demonstrated that the nNOS-2 isoform mediates the
analgesic action of morphine, whereas the nNOS-1 isoform is
involved in the development of tolerance to the analgesic action
of the opioid. Furthermore, these authors also observed that the
facilitating nNOS-2 system predominates at the spinal level, while
the inhibitory nNOS-1 system is the major supraspinal nNOS sys-
tem, indicating that, at the functional level, the two isoforms of
nNOS can display opposing effects on morphine actions.
The distinct effects of NO in the nociceptive system and in the
antinociceptive action of opioids may be also correlated to gender.
Fatehi-Hassanabad et al. (2005) [170] demonstrated that adminis-
tration of
-NAME to male mice increases the reaction time to ther-
mal nociceptive stimulus in the hot plate test, but does not alter
the response in female animals. These results indicate that the
endogenous control of nociception by nitric oxide is related to
sex hormones, being NO pro-nociceptive only in male animals.
On the other hand,
-NAME increases the antinociceptive effect of
morphine in the tail-flick test only in female rats, suggesting that
the decreased efficacy of opioids in females is dependent on the
NO signaling systems [171].
Perspectives on the clinical use of NO as an analgesic drug
Despite the controversial data, the antinociceptive effect of NO
has been consistently demonstrated. Therefore, the clinical use of
NO should be considered as an important strategy for pain therapy.
In fact, modification of pre-existing analgesic and anti-inflamma-
tory drugs by addition of NO-releasing moieties has been shown
to improve the analgesic efficacy of these drugs and also to reduce
the expression of their side effects. The cyclooxygenase inhibitor
nitric oxide donors (CINODs) are an example of a new class of
anti-inflammatory/analgesic drugs generated by addition of a NO
generating moiety to the parent non-steroidal anti-inflammatory
drugs (NSAID) (review in [172]). This strategy reduces the gastro-
intestinal toxicity of NSAID and confers a potent anti-inflammatory
activity. NCX-701 or nitroparacetamol is also a new drug resulting
from the combination of paracetamol and a nitrooxybutyroyl moi-
ety, which releases nitric oxide at a low, but steady, level [173].
NCX-701 has been shown to be effective in acute nociception
and in neuropathic pain, with a better outcome when compared
to paracetamol alone, since the combination of these molecules
also results in an enhancement of the analgesic activity of paracet-
amol. In addition, whereas paracetamol lacks antiinflammatory
activity, NCX-701 might reduce inflammation [173]. Another strat-
egy that has been used for pain control is the delivery of nitric
oxide donors (usually as a nitroglycerin patch) together with opi-
oids in cancer pain management (review in [96]). This strategy en-
hances the analgesic efficacy of morphine in patients with cancer
pain, delays morphine tolerance and decreases the incidence of
the adverse effects of opioids [174,175]. The use of NO donors to
reduce pain in patients undergoing surgery has also been
evaluated. It was demonstrated by Sen et al. [176] that the addition
250 Y. Cury et al. / Nitric Oxide 25 (2011) 243–254
Author's personal copy
of nitroglycerin to lidocaine for IV regional anesthesia improves
sensory and motor block, tourniquet pain, and postoperative anal-
gesia without side effects. Furthermore, the analgesic efficacy of
epidural ketamine in patients undergoing orthopedic surgery is en-
hanced by transdermal nitroglycerin [177]. Although clinical trials
with some of these drugs have provided results consistent with
pre-clinical data, there is only limited information available to sup-
port the beneficial use of nitric oxide as an analgesic in patients
and further studies are needed to support their clinical use
Concluding remarks
Nitric oxide plays a complex and diverse role in the modulation
of nociceptive transmission in both the peripheral and central ner-
vous system. The mechanisms involved in the nociceptive as well
antinociceptive effects of NO are not fully characterized yet and
considerable work is necessary to elucidate its role in nociceptive
transmission. The nociceptive effect of NO involves the stimulation
of soluble guanylyl cyclase resulting in conversion of guanosine tri-
phosphate to cGMP, which, in turn, modulates the activity of many
targets in the cells, including PKG, ion channels and phosphodies-
terases. NO may interact with many systems, such as NMDA
receptors and COXs, to induce hyperalgesic effect. Concerning the
analgesic effect of NO, experimental data have indicated that this
effect also depends on activation of an intracellular signaling
pathway, involving the cGMP–PKG–ATP-sensitive K
pathway. The experimental evidences demonstrating that nitric
oxide undoubtedly contributes to the mechanisms underlying the
analgesic action of opioids, anti-inflammatory and other analgesic
drugs could foster the development of strategies for appropriate
analgesic drug therapy. Furthermore, the demonstration that addi-
tion of NO-releasing moieties increases the analgesic efficacy of the
parent drug could represent a useful strategy for an improved and
more efficient pain treatment.
Authors would like to thank the Instituto Nacional de Ciencia e
Tecnologia em Toxinologia (INCTTOX PROGRAM) of Conselho Nac-
ional de Desenvolvimento Científico e Tecnológico (CNPq)/FAPESP
(Grant number 2008/57898-0).
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... Los investigadores buscan nuevas respuestas y los estudios se dirigen hacia los mecanismos moleculares que repercuten en los sistemas moduladores del dolor y que aporten nuevos elementos a los mecanismos de acción de estas técnicas para facilitar su validación científica. Sobre la base de la importancia de los sistemas de señalización celular en los diferentes sistemas biológicos, se presta atención en particular a una molécula tan controversial en del área de la nocicepción e hiperalgesia como el óxido nítrico (NO, por sus siglas en inglés) (Ferreira et al., 2005;Latremoliere y Woolf, 2009;Cury et al. 2011). ...
... Posteriormente, en las décadas sucesivas, se producen nuevos descubrimientos que constituyen anomalías para el nuevo paradigma establecido, lo que conduce a la revisión y modificación de la teoría por los mismos autores. Pero las alteraciones del procesamiento nociceptivo, de la conectividad central, la desinhibición y los cambios fenotípicos en las propias fibras gruesas Aß, que en estas condiciones pueden ser mediadoras del dolor; han revolucionado nuevamente la ciencia nociceptiva y la necesidad de búsqueda de otras explicaciones (Millares et al., 1997;Ferreira et al., 2005;Latremoliere y Woolf, 2009;Cury et al., 2011). Se está, entonces, ante el nacimiento de otro paradigma y se debe ir más allá en la búsqueda de los blancos moleculares de la acupuntura y sus técnicas afines a la par con el nuevo patrón, pero sobre la base de muchos conceptos conocidos. ...
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Despite the diffusion of acupuncture and its related techniques in Cuba and the World, its mechanism of action is still controversial, being considered by the most sceptics as placebo or some kind oriental myth, and it only should by related to this subjects as a matter of cultural-historical elements and not to science. The purpose of this revision is to characterize the pain sensation, after a critical analysis of the different philosophical streams related to the human knowledge and its expression in the historical evolution of the algology. On the other hand, to emphasize the importance of electroacupuncture-induced neuro-modulation in the field of experimental pain researches. In this content will be analyzed the concept of Khun paradigm and his ideas about the structure of scientific revolution in the theory of gates control and the explosion of pain researches in the last decades. It will related the introduction to acupuncture and its techniques in pain clinics, with scientific context of the historical moment. In addition, a space will be dedicated to the topic of complementary and alternative medicine on the century of evidence based medicine, given its scientific needs of validation in ours times.
... This activation results in the production of cGMP and modulation of various cellular targets, including cPKG and ion channels. NO acts as a neurotransmitter, enhancing the release of substance P and calcitonin-gene related peptide from C-fiber terminals, contributing to the development of secondary hyperalgesia [59]. The inhibition of eNOS function has been observed to upregulate the expression of APP (amyloid precursor protein) and BACE1 (β-site APP-cleaving enzyme 1) in human brain endothelial cells. ...
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Purpose of Review Individuals with dementia and Alzheimer’s disease (AD) often receive inadequate analgesia compared to those without cognitive impairments [24-26]. However, undertreated pain contributes more to cognitive decline than analgesics. A comprehensive evaluation of available therapies considering administration modes, dosages, potential interactions, and side effects is necessary. Our review provides an extensive report on current methods of pain assessment, strategies for dealing with postoperative pain, and the challenges anesthesiologists face, and points out the inadequacy of current literature to formulate a comprehensive strategy. Recent Findings Trials have shown that for demented patients, there is a significant decrease in the dose of analgesia. Moreover, the ineffective assessment of the degree of pain leads to under-treatment. Studies are being conducted to assess validity of pain scales, current limitations, and suggestions to enhance their sensitivity. Individualizing the opioid dose is vital for effective pain management in older adults. Multimodal analgesia can enhance pain relief and reduce opioid consumption and related adverse events. Acetaminophen is a safe non-opioid option, while NSAIDs and centrally acting anticholinergics should be avoided due to potential toxicity. Among opioids, fentanyl, morphine, and oxycodone are considered safe, while pethidine and tramadol should be avoided due to their association with postoperative delirium. Summary Effective postoperative pain management in dementia patients requires individualized approaches, considering specific needs and risks associated with different drugs and administration methods. Certain medications may interact with analgesics and cause serious side effects in demented patients. Further research is necessary to establish guidelines for managing postoperative pain in Alzheimer’s disease.
... It is recognized the role of NO as a critical mediator of synaptic plasticity, longterm potentiation, and consolidation of long-term memory (Yamada and Nabeshima, 1998). Also, it has been suggested that NO mediates the effects of estradiol in many organs (Cury et al., 2011). Estradiol affects the release of NO and the activity of nitric oxide synthase isoforms in the brain (Lopez-Jaramillo and Teran, 1999;Gotti et al., 2010). ...
Context: Ovarian hormone deficiency observed in menopausal women increases the production of reactive oxygen species, which could be implicated in central sensitization subjacent in chronic functional pain syndromes. Aims: To examine the hyperalgesic state induced by ovariectomy in adult rats and its relation to some oxidative stress outcomes. Methods: The female Wistar rats were divided into normal, sham ovariectomized (OVX) and OVX groups, which were tested for mechanical and thermal hypernociception during 6 weeks and a single acetic acid-induced test 6 weeks after surgery. Redox biomarkers determinations of superoxide dismutase (SOD) enzyme activity, glutathione (GSH) and nitrates/nitrites as an indicator of nitric oxide (NO) concentrations were determined in the brain and cerebellum of 6 animals of each group. Results: Exclusivity OVX rats developed a robust state of mechanical hypernociception and allodynia in the abdomen, hindlimbs and proximal tail. Besides, thermal pain thresholds (hot plate) decreased. That was established 3-4 weeks after OVX and lasted for the 6 weeks of the experiment. Increases in visceral sensitivity were also observed in OVX rats. SOD enzyme activity decreased in OVX rats, which showed major deficit for this enzymatic defense under visceral inflammatory injury. However GSH concentrations were increased in brain of OVX animals that allow the balance during acute inflammation. NO concentrations were raised only in OVX rats exposure to chemical inflammatory injury. Conclusions: OVX in rats provide a useful model, which mimics the functional pain in females that could be related with brain oxidative stress.
... In our hands, we demonstrated that FCH metabolites limited the rise of secondary pro-inflammatory mediators by chondrocytes upon rhIL-1β incubation with a trendy but not significant blunt of PGE2 and a very significant reduction of NO production. Interestingly, nitric oxide is a well acknowledged mediator of the nociceptive system in the periphery [50]. Consistently, the reduction of NO may explain, at least partly, the positive influence of FCH on pain as observed in the multicentric clinical trial (exploratory clinical study). ...
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Background Aging is frequently associated with impairments of the musculoskeletal system and many elderly people experience joint discomfort or pain which might reduce their ability to move and consequently alter their quality of life. A beneficial effect of fish cartilage hydrolysate (FCH) on pain and joint function has recently been shown in an ACLT/pMMx osteoarthritis rat model. Methods We therefore performed an exploratory, non-comparative, multi-centric clinical trial including 33 subjects with moderate knee joint discomfort and loss of functionality to investigate the efficacy of FCH on their algo-functional status. We further determined the potential health benefit of FCH in an original clinical ex vivo study investigating the role of FCH human metabolites on primary human chondrocytes. Results FCH significantly improved knee pain and function, as assessed by the Knee injury and Osteoarthritis Outcome Score (KOOS). Moreover, FCH significantly reduced pain at rest and while walking, and patient global assessment (PGA), as assessed by the Visual Analogue Scale (VAS), and improved patients’ quality of life (SF-36). FCH metabolites decreased the synthesis of catabolic factors (MMP-13) and pro-inflammatory mediators (NO, PGE2) and limited the inhibitory effect of IL-1β on the synthesis of cartilage matrix components (GAG and collagen). Conclusions Thus, these data provide insights on the mode of action of FCH in humans and contribute to explain how FCH may relieve pain and improve joint function in subjects with knee discomfort. Although these preliminary data need to be confirmed in a randomized controlled trial, they strongly support the potential health benefit of such an active ingredient. Trial registration: The study was registered on with the identifier NCT04420091 (09/06/2020).
... NO is a gasotransmitter involved in the pathogenesis of migraine; it can be produced in the meninges by eNOS, expressed in the endothelium of cerebral vessels (including the MMA), and by nNOS, localized in the trigeminal afferents [13][14][15]33]. To analyze the effects of NO on the activity of the trigeminal nerve afferents, we used a rat hemiskull preparation that allows us to record the nociceptive activity directly at the site of its occurrence [34]. ...
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Nitric oxide is one of the endogenous molecules that play a key role in migraine. However, the interaction between NO and the main players in the nociceptive activity of the meningeal trigeminal afferents—TRPV1 and P2X3 receptors—remains unstudied. In the current project, the effects of acute and chronic NO administration on the activity of TRPV1 and P2X3 receptors in the peripheral afferents were studied using electrophysiological recording of action potentials of the trigeminal nerve in the rat hemiskull preparations. The data obtained indicate that exogenous and endogenous NO increased the activity of the trigeminal nerve independent on the inhibition of the TRPV1 and P2X3 receptors. The activity of the trigeminal nerve triggered by ATP changed neither in acute incubation in the NO donor—sodium nitroprusside (SNP) nor in the chronic nitroglycerine (NG)-induced migraine model. Moreover, the chronic NG administration did not increase in the number of degranulated mast cells in the rat meninges. At the same time, the capsaicin-induced activity of the trigeminal nerve was higher with chronic NO administration or after acute NO application, and these effects were prevented by N-ethylmaleimide. In conclusion, we suggested that NO positively modulates the activity of TRPV1 receptors by S-nitrosylation, which may contribute to the pro-nociceptive action of NO and underlie the sensitization of meningeal afferents in chronic migraine.
... In addition, NO has neuroprotective effects by facilitating regeneration and preventing the loss of DRG neurons [148]. Moreover, NO itself can induce analgesia and mediates the anti-nociceptive effect of analgesics via activation of the cGMP-PKG-ATP-sensitive potassium channel pathway [149]. Paradoxically, NO synthesis in mouse DRG is elevated in response to peripheral inflammation and consequently, neuronal hyperexcitability which amplifies nociceptive sensitization [150,151]. ...
The pentameric nicotinic acetylcholine receptors (nAChRs) are typically classed as muscle- or neuronal-type, however, the latter has also been reported in non-neuronal cells. Given their broad distribution, nAChRs mediate numerous physiological and pathological processes including synaptic transmission, presynaptic modulation of transmitter release, neuropathic pain, inflammation, and cancer. There are 17 different nAChR subunits and combinations of these subunits produce subtypes with diverse pharmacological properties. The expression and role of some nAChR subtypes have been extensively deciphered with the aid of knock-out models. Many nAChR subtypes expressed in heterologous systems are selectively targeted by the disulfide-rich α-conotoxins. α-Conotoxins are small peptides isolated from the venom of cone snails, and a number of them have potential pharmaceutical value.
The purpose of the narrative review is to present the current knowledge on the mechanisms of action of vitaminD in the processes of nociception. The role of vitamin D in calcium homeostasis and bone metabolism is wellknown and documented, and there is a strong correlation between hypovitaminosis D and the occurrence of bonepain in humans. However, the mechanisms by which vitamin D may influence nociception processes are poorlyunderstood. The review was based on selected publications from 1973October 2022, relating to the mechanismsand analgesic efficacy of vitamin D in animal and human studies. The potential interaction between the vitamin Dpathway and nociception processes, demonstrated in the above studies, is discussed. The analysis of the results ofexperimental and clinical studies indicates a significant correlation and participation of vitamin D in the processesof nociception through its influence on receptors, numerous ion channels, modulation of the expression of pro- andanti-inflammatory cytokines, neurotrophic factors, prostaglandin levels, oxidative processes, the opioid system andthe activity of immunocompetent cells. Indirectly, vitamin D also affects the risk and course of autoimmune diseases, cardiovascular diseases and cancer, which may be associated with chronic pain in humans.
Background: With a huge impact on the quality of life and productivity of young women, dysmenorrhea remains a major gynecological disorder. Although abnormal myometrial activity and vasoconstriction are believed to play a major role, the pathophysiology of dysmenorrhea is not fully understood. Our aim was therefore to examine a possible relationship between the Glu298Asp polymorphism (rs1799983) of the endothelial nitric oxide synthase (eNOS) gene and dysmenorrhea. Methods: We sought the participation of women of reproductive age (n = 239). Each participant completed a questionnaire to obtain data on their menstrual cycles and a Numeric Pain Rating Scale (NPRC) to rate their pain experience during menstrual periods. Genotyping was done by PCR-RFLP method while a possible association between the presence and severity of menstrual pain and genotypes was determined. Results: Among the participants, pain was reported by 167 (89.9%) while 72 (30.1%) reported no pain. Seventy-one (29.7%) and 93 (38.9%) participants reported moderate and severe pains respectively while others reported no or mild pain. The frequencies of G and T alleles were 316 and 162 while frequencies of GG, GT, and TT were 119, 78, and 42 respectively. The population deviated from Hardy-Weinberg equilibrium (χ² = 17.6, df = 1, p < 0.01). Participants with a T allele were more than three times as likely as those without a T allele to report pain (OR = 3.3, p < 0.01 95%CI = 1.8–5.9). Conclusion: Carrying a T (Asp298) allele of Glu298Asp polymorphism of the eNOS gene may predispose to dysmenorrhea indicating that nitric oxide might play a role in dysmenorrhea.
Pregnancy is character-ized by hemodynamic and body fluid alterations. Increased nitric oxide (NO) production has been suggested to play a role in the hemodynamic alterations of pregnancy and has also been reported to increase arginine vasopressin (AVP) release. We therefore hypothesized that gestation could increase both NO synthase (NOS) constitutive isoforms, neuronal NOS and endothelial NOS, and thereby contribute to the hyposmolality and peripheral arterial vasodilation of pregnancy, respectively. The present study was therefore undertaken to examine the constitutive NOS isoforms in aortas, mesenteric arteries, and hypothalami of pregnant rats on day 20 of gestation compared with age-matched nonpregnant rats. Plasma AVP was determined by radioimmunoassay and hypothalamic mRNA AVP by solution hybridization assay. Hypothalamic neuronal NOS was assessed by Northern blot and Western blot; endothelial NOS was assessed by Western blot in arteries and hypothalamus. The results demonstrated that 1) plasma AVP and hypothalamic AVP mRNA are increased in pregnant rats (n = 8), 2) neuronal NOS protein and mRNA are increased in hypothalamus of pregnant rats (n = 4), and 3) endothelial NOS expression, as assessed by Western blot analysis, is increased in both conductance (aorta) as well as resistance (mesenteric) arteries of pregnant rats (n = 4). We conclude that both of the constitutive NOS isoforms are increased in pregnant rats, suggesting that the peripheral arterial vasodilation and hyposmolality of pregnancy could be mediated by these isoforms.
The retina is a thin layer of neural tissue that lines the back of the eye. It is part of the central nervous system displaced into the eye during development. In addition to the light-sensitive photoreceptor cells, the retina contains five basic classes of neurons and one principal type of glial cell, the Müller cell. The neurons are organized into three cellular (nuclear) layers which are separated by two synaptic (plexiform) layers. Virtually all of the junctions (synapses) between the retinal neurons are made in the two synaptic layers, and all visual information passes across at least two synapses, one in the outer plexiform layer and another in the inner plexiform layer, before it leaves the eye. Processing of visual information occurs in both plexiform layers. The outer plexiform layer separates visual information into on- and off-channels and carries out a spatial type of analysis on the visual input. The output neurons of this layer, the on- and off-bipolar cells, demonstrate a center-surround antagonistic receptive field organization. The inner plexiform layer, on the other hand, is concerned more with the temporal aspects of light stimuli.
Polycrystalline Nd(Mn1−xFex)2Ge2 (0≤x≤1) compounds have been prepared by arc-melting. X-ray powder diffraction analysis shows that all samples crystallize in the ThCr2Si2-type structure with the space group I4/mmm. The substitution of Fe for Mn results in decreases of the lattice constants a, c and the unit-cell volume V. Magnetic properties and magnetocaloric effect of the compounds Nd(Mn0.9Fe0.1)2Ge2 and Nd(Mn0.8Fe0.2)2Ge2 have been studied by magnetic measurements. A spin reorientation transition at about 188K is observed for Nd(Mn0.9Fe0.1)2Ge2. The Nd(Mn0.8Fe0.2)2Ge2 compound shows more complicated magnetic behavior, which is characterized by four magnetic ordering states below room temperature. The maximum values of the magnetic entropy change are 2.35 and 1.84Jkg−1K−1 for x=0.1 and 0.2, respectively, under the applied field changing from 0 to 5T. The relative cooling powers of Nd(Mn0.9Fe0.1)2Ge2 and Nd(Mn0.8Fe0.2)2Ge2 are 145.9 and 133.8Jkg−1 under an applied field change of 5T.
In the present study, it was found that intraperitoneal (i.p.) pre-injection of N-G-nitro-L-arginine methyl ester (LNAME) significantly influenced the endomorphin-1 (EM-1) and endomorphin-2 (EM-2) induced antinociception. These effects could be inhibited or reversed by L-Arg or naloxone. Our results suggest that the modulatory effect of NO system on the mu-receptor evoked analgesia is different between the two mu receptor subtypes.
The potent vasodilator factor released from endothelial cells upon activation of muscarinic acetylcholine receptors has recently been identified as nitric oxide (NO). This discovery has sparked intensive research efforts aiming at understanding the functional role of this short-lived, highly reactive free radical. As a result, it has been shown that NO is a very important second messenger involved in a wide spectrum of physiological functions. One important aspect that differentiates NO from other second messengers is that NO is a ‘traveling’ messenger that diffuses out of the cells of its origin to produce marked effects in neighboring cells. This paper provides a synopsis of the diverse biological roles of NO and the mechanisms of its generation upon activation of muscarinic acetylcholine receptors. Drug Dev. Res. 40:205–214, 1997. © 1997 Wiley-Liss, Inc.