[Show abstract][Hide abstract] ABSTRACT: Although the renin-angiotensin system (RAS) was classically considered as a circulating system that regulates blood pressure, many tissues are now known to have a local RAS. Angiotensin, via type 1 receptors, is a major activator of the NADPH-oxidase complex, which mediates several key events in oxidative stress (OS) and inflammatory processes involved in the pathogenesis of major aging-related diseases. Several studies have demonstrated the presence of RAS components in the basal ganglia, and particularly in the nigrostriatal system. In the nigrostriatal system, RAS hyperactivation, via NADPH-oxidase complex activation, exacerbates OS and the microglial inflammatory response and contributes to progression of dopaminergic degeneration, which is inhibited by angiotensin receptor blockers and angiotensin converting enzyme (ACE) inhibitors. Several factors may induce an increase in RAS activity in the dopaminergic system. A decrease in dopaminergic activity induces compensatory upregulation of local RAS function in both dopaminergic neurons and glia. In addition to its role as an essential neurotransmitter, dopamine may also modulate microglial inflammatory responses and neuronal OS via RAS. Important counterregulatory interactions between angiotensin and dopamine have also been observed in several peripheral tissues. Neurotoxins and proinflammatory factors may also act on astrocytes to induce an increase in RAS activity, either independently of or before the loss of dopamine. Consistent with a major role of RAS in dopaminergic vulnerability, increased RAS activity has been observed in the nigra of animal models of aging, menopause and chronic cerebral hypoperfusion, which also showed higher dopaminergic vulnerability. Manipulation of the brain RAS may constitute an effective neuroprotective strategy against dopaminergic vulnerability and progression of Parkinson's disease.
Frontiers in Neuroanatomy 01/2014; 8:67. · 4.06 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Dysfunction of iron homeostasis has been shown to be involved in aging, Parkinson's disease and other neurodegenerative diseases. Increased levels of labile iron result in increased reactive oxygen species and oxidative stress. Angiotensin II, via type-1 receptors, exacerbates oxidative stress, the microglial inflammatory response and progression of dopaminergic degeneration. Angiotensin activates the NADPH-oxidase complex, which produces superoxide. However, it is not known whether angiotensin affects iron homeostasis. In the present study, administration of angiotensin to primary mesencephalic cultures, the dopaminergic cell line MES23.5 and to young adult rats, significantly increased levels of transferrin receptors, divalent transporter-1 and ferroportin, which suggests an increase in iron uptake and export. In primary neuron-glia cultures and young rats, angiotensin did not induce significant changes in levels of ferritin or labile iron, both of which increased in neurons in the absence of glia (neuron-enriched cultures, dopaminergic cell line) and in the N9 microglial cell line. In aged rats, which are known to display high levels of angiotensin activity, ferritin levels and iron deposits in microglial cells were enhanced. Angiotensin-induced changes were inhibited by angiotensin type-1 receptor antagonists, NADPH-oxidase inhibitors, antioxidants and NFK-β inhibitors. The results demonstrate that angiotensin, via type-1 receptors, modulates iron homeostasis in dopaminergic neurons and microglial cells, and that glial cells play a major role in efficient regulation of iron homeostasis in dopaminergic neurons.
[Show abstract][Hide abstract] ABSTRACT: A major limiting factor for cell therapy in Parkinson's disease is that the survival of grafted dopaminergic neurons is very poor, which may be improved by administration of GDNF, for which the carotid body is a good source.
Rats with total unilateral dopaminergic denervation were grafted with a cell suspension of rat dopaminergic neuroblasts with or without cell aggregates from the rat carotid body. At 1, 2 and 3 months after grafting, the rats were tested in the cylinder and the rotometer and killed 4 months after grafting.
We observed that the survival of dopaminergic neurons and graft-derived dopaminergic innervation were higher in rats that received mixed grafts. Both grafted groups showed complete recovery in the amphetamine-induced rotation test. However, rats with cografts performed significantly better in the cylinder test.
Cografting of carotid body cells may constitute a useful strategy for cell therapy in Parkinson's disease.
Regenerative Medicine 05/2012; 7(3):309-22. · 3.87 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: A number of C-3 spirocyclic 2-benzazepine analogs of α-phenyl-N-tert-butyl nitrone (PBN) were synthesized and tested for their activity in protecting rat brain mitochondria and dopaminergic (DA) neurons against 6-hydroxydopamine (6-OHDA), a toxin inducing destruction of the DA nigro-striatal pathway in rodent models of Parkinson's disease. The newly synthesized nitrone derivatives were firstly investigated for their activity in decreasing the level of hydroxyl radicals generated during 6-OHDA oxidation, and inhibit lipid peroxidation (TBARS assay) and protein carbonyl content (PCC) in rat brain mitochondria. Most of the studied 2-benzazepine nitrones showed inhibitory potencies in both TBARS and PCC assays at least two magnitude orders higher than that of PBN. The data obtained usefully complemented the known structure-activity relationships. In particular, 5 and 10, bearing C-3 spiro cyclopentyl and tetrahydropyranyl moieties, respectively, at 8 µM concentration proved to be significantly more effective than PBN in protecting cultured DA neurons exposed to 6-OHDA, which alone causes about 45% cell loss in 24 h. In addition, we found that 5 inhibited butyrylcholinesterase with an IC(50) value of 16.8 µM, which would enhance its potential as neuroprotective agent in Alzheimer's neurodegeneration. These findings extend the utility of benzazepine-based PBN analogs in the treatment of age-related free radical-mediated disorders.
Archiv der Pharmazie 04/2012; 345(8):598-609. · 1.54 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The pathogenic mechanism of Parkinson's disease (PD) appears to be multifactorial. However, oxidative stress and neuroinflammation, including activation of NADPH-dependent oxidases, play a major role in the progression of dopaminergic cell death. The renin-angiotensin system (RAS) was described as a circulating humoral system that regulates blood pressure and water homeostasis. However, there exist local RAS in many tissues, and locally formed angiotensin activates NADPH-dependent oxidases, which are a major source of superoxide and are upregulated in major aging-related diseases such as hypertension, diabetes and atherosclerosis. Furthermore, an intracellular or intracrine RAS, with still unknown functions, has been identified in several cell types. The brain has an independent local RAS, which has been involved in several brain disorders, including neurodegenerative diseases. It is particularly interesting for PD the important interaction observed between angiotensin and dopamine, which counterregulate each other in renal cells and also in the striatum and substantia nigra. In recent studies, we have observed both a local and an intracellular RAS in the rodent, monkey and human substantia nigra, and that dopamine depletion induced RAS upregulation possibly as a compensatory mechanism. However, RAS hyperactivation also exacerbated oxidative stress and neuroinflammation, which contributed to progression of dopaminergic degeneration. In addition, we observed increased RAS activity in the nigra of animals with higher vulnerability of dopaminergic neurons to degeneration, such as aged males, menopausal females and rats subjected to chronic brain hypoperfusion. RAS activity and dopaminergic vulnerability were significantly reduced by treatment with angiotensin type I receptor antagonists. Manipulation of the brain RAS may constitute an effective neuroprotective strategy against dopaminergic degeneration in PD.
American journal of neurodegenerative disease. 01/2012; 1(3):226-44.
[Show abstract][Hide abstract] ABSTRACT: Recent studies have shown that renin-angiotensin system overactivation is involved in the aging process in several tissues as well as in longevity and aging-related degenerative diseases by increasing oxidative damage and inflammation. We have recently shown that angiotensin II enhances dopaminergic degeneration by increasing levels of reactive oxygen species and neuroinflammation, and that there is an aging-related increase in angiotensin II activity in the substantia nigra in rats, which may constitute a major factor in the increased risk of Parkinson's disease with aging. The mechanisms involved in the above mentioned effects and particularly a potential angiotensin-mitochondria interaction have not been clarified. The present study revealed that activation of mitochondrial ATP-sensitive potassium channels [mitoK(ATP)] may play a major role in the angiotensin II-induced effects on aging and neurodegeneration. Inhibition of mitoK(ATP) channels with 5-hydroxydecanoic acid inhibited the increase in dopaminergic cell death induced by angiotensin II, as well as the increase in superoxide/superoxide-derived reactive oxygen species levels and the angiotensin II-induced decrease in the mitochondrial inner membrane potential in cultured dopaminergic neurons. The present study provides data for considering brain renin-angiotensin system and mitoK(ATP) channels as potential targets for protective therapy in aging-associated diseases such as Parkinson's disease.
[Show abstract][Hide abstract] ABSTRACT: For years, the renin-angiotensin system (RAS) was described as a circulating humoral system that regulates blood pressure and water homeostasis. Angiotensin II (AII) is the most important effector peptide. However, in addition to the "classical" humoral RAS there exist local RAS in many tissues and locally formed AII activates NADPH-dependent oxidases, which are a major source of superoxide and are upregulated in major aging-related diseases such as hypertension, diabetes and atherosclerosis. Accordingly, disruption of AII receptors promotes longevity in mice. The brain has an independent local RAS, which was also initially associated with the central control of blood pressure. However, more recent studies have involved brain RAS in brain disorders, including neurodegenerative diseases. The interaction between AII and dopamine is particularly interesting. Recent evidence suggests that dopamine and AII systems directly counterregulate each other in renal cells as well as in the striatum and substantia nigra. Dopamine depletion may induce RAS upregulation as a potential compensatory mechanism. However, RAS hyperactivation also exacerbates NADPH-oxidase activity, oxidative stress and the microglial inflammatory response and contribute to progression of dopaminergic neuron loss, as observed in recent studies with animal models of Parkinson's disease (PD). Aging is the most prominent risk factor for PD and other neurodegenerative diseases. Interestingly, we observed increased activation of the NADPH oxidase complex and increased levels of the pro-inflammatory cytokines in the nigra of aged male rats, which was associated with increased RAS activity and was reduced by treatment with AII antagonists. We also observed that the lack of oestrogen may act as an additional factor for increasing RAS activity in the nigra in aged females, which was significantly reduced by treatment with AII antagonists. Manipulation of the brain RAS may constitute an effective neuroprotective strategy against the aging-related risk of dopaminergic degeneration.
[Show abstract][Hide abstract] ABSTRACT: Reactive oxygen species signaling has been suggested to regulate stem cell development. In the present study, we treated neurospheres of rat mesencephalic precursors with inhibitors of the NADPH oxidase complex and mitochondrial ATP-sensitive potassium (mitoKATP) channel blockers during the proliferation and/or the differentiation periods to study the effects on generation of dopaminergic neurons. Treatment with low doses (100 or 250 μM) of the NADPH inhibitor apocynin during the proliferation period increased the generation of dopaminergic neurons. However, higher doses (1 mM) were necessary during the differentiation period to induce the same effect. Treatment with general (glibenclamide) or mitochondrial (5-hydroxydecanoate) KATP channel blockers during the proliferation and differentiation periods increased the number of dopaminergic neurons. Furthermore, neither increased proliferation rate nor apoptosis had a major role in the observed increase in generation of dopaminergic neurons, which suggests that the redox state is able to regulate differentiation of precursors into dopaminergic neurons.
[Show abstract][Hide abstract] ABSTRACT: The basal ganglia have a local renin-angiotensin system and it has been shown that the loss of dopaminergic neurons induced by neurotoxins is amplified by local angiotensin II (AII) via angiotensin type 1 receptors (AT1) and nicotinamide adenine dinucleotide phosphate (NADPH) complex activation. Recent studies have revealed a high degree of counter-regulatory interactions between dopamine and AII receptors in non-neural cells such as renal proximal tubule cells. However, it is not known if this occurs in the basal ganglia. In the striatum and nigra, depletion of dopamine with reserpine induced a significant increase in the expression of AT1, angiotensin type 2 receptors (AT2) and the NADPH subunit p47(phox) , which decreased as dopamine function was restored. Similarly, 6-hydroxydopamine-induced chronic dopaminergic denervation induced a significant increase in expression of AT1, AT2 and p47(phox) , which decreased with L-dopa administration. A significant reduction in expression of AT1 mRNA was also observed after administration of dopamine to cultures of microglial cells. Transgenic rats with very low levels of brain AII showed increased AT1, decreased p47 (phox) and no changes in AT2 expression, whereas mice deficient in AT1 exhibited a decrease in the expression of p47 (phox) and AT2. The administration of relatively high doses of AII (100 nm) decreased the expression of AT1, and the increased expression of AT2 and p47(phox) in primary mesencephalic cultures. The results reveal an important interaction between the dopaminergic and local renin-angiotensin system in the basal ganglia, which may be a major factor in the progression of Parkinson's disease.
European Journal of Neuroscience 10/2010; 32(10):1695-706. · 3.75 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The neurotoxin MPTP reproduces most of the biochemical and pathological hallmarks of Parkinson's disease. In addition to reactive oxygen species (ROS) generated as a consequence of mitochondrial complex I inhibition, microglial NADPH-derived ROS play major roles in the toxicity of MPTP. However, the exact mechanism regulating this microglial response remains to be clarified. The peptide angiotensin II (AII), via type 1 receptors (AT1), is one of the most important inflammation and oxidative stress inducers, and produces ROS by activation of the NADPH-oxidase complex. Brain possesses a local angiotensin system, which modulates striatal dopamine (DA) release. However, it is not known if AII plays a major role in microglia-derived oxidative stress and DA degeneration. The present study indicates that in primary mesencephalic cultures, DA degeneration induced by the neurotoxin MPTP/MPP(+) is amplified by AII and inhibited by AT1 receptor antagonists, and that protein kinase C, NADPH-complex activation and microglial activation are involved in this effect. In mice, AT1 receptor antagonists inhibited both DA degeneration and early microglial and NADPH activation. The brain angiotensin system may play a key role in the self-propelling mechanism of Parkinson's disease and constitutes an unexplored target for neuroprotection, as previously reported for vascular diseases.
Journal of Neurochemistry 03/2009; 109(2):656-69. · 3.97 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The neurotoxin 6-hydroxydopamine is commonly used in models of Parkinson's disease, and a potential factor in the pathogenesis of the disease. However, the mechanisms responsible for 6-hydroxydopamine-induced dopaminergic degeneration have not been totally clarified. Reactive oxygen species (ROS) derived from 6-OHDA uptake and intraneuronal autooxidation, extracellular 6-OHDA autooxidation, and microglial activation have been involved. The mitochondrial implication is controversial. Mitochondrial ATP-sensitive K (mitoK(ATP)) channels may provide a convergent target that could integrate these different mechanisms. We observed that in primary mesencephalic cultures and neuron-enriched cultures, treatment with the mitoK(ATP) channel blocker 5-hydroxydecanoate, inhibits the dopaminergic degeneration induced by low doses of 6-OHDA. Furthermore, 5-hydroxydecanoate blocks the 6-OHDA-induced decrease in mitochondrial inner membrane potential and inhibits 6-OHDA-induced generation of superoxide-derived ROS in dopaminergic neurons. The results suggest that low doses of 6-OHDA may generate low levels of ROS through several mechanisms, which may be insufficient to induce neuron death. However, they could act as a trigger to activate mitoK(ATP) channels, thereby enhancing ROS production and the subsequent dopaminergic degeneration. Furthermore, the present study provides additional data for considering mitoK(ATP) channels as a potential target for neuroprotection.
Neurotoxicity Research 02/2009; 15(1):82-95. · 2.87 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: It is not known if aging induces changes in nigral vascularization and nigral vascular endothelial growth factor (VEGF) levels similar to those previously reported for Parkinson's disease (PD). In this study nonexercised rats displayed age-dependent decreases in the density of nigral microvessels and VEGF mRNA expression, which were reversed by physical exercise. Such changes may enhance the vulnerability of dopaminergic neurons and the risk of developing PD, and may be reduced by exercise. Furthermore, the observed pattern is the opposite of that previously observed in PD, suggesting that the process underlying PD is not an accelerated age-dependent decline in the dopaminergic system.
Journal of cerebral blood flow and metabolism: official journal of the International Society of Cerebral Blood Flow and Metabolism 10/2008; 29(2):230-4. · 5.46 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Angiotensin II (AII) plays a major role in the progression of inflammation and NADPH-derived oxidative stress (OS) in several tissues. The brain possesses a local angiotensin system, and OS and inflammation are key factors in the progression of Parkinson's disease. In rat mesencephalic cultures, AII increased 6-OHDA-induced dopaminergic (DA) cell death, generation of superoxide in DA neurons and microglial cells, the expression of NADPH-oxidase mRNA, and the number of reactive microglial cells. These effects were blocked by AII type-1 (AT1) antagonists, NADPH inhibitors, or elimination of glial cells. DA degeneration increased angiotensin converting enzyme activity and AII levels. In rats, 6-OHDA-induced dopaminergic cell loss and microglial activation were reduced by treatment with AT1 antagonists. The present data suggest that AII, via AT1 receptors, increases the dopaminergic degeneration process by amplifying the inflammatory response and intraneuronal levels of OS, and that glial cells play a major role in this process.
Neurobiology of Disease 08/2008; 31(1):58-73. · 5.62 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Production of dopaminergic neurons from stem/precursor cells for transplantation in Parkinson's disease has become a major focus of research. However, the inductive signals mediating this process have not been clarified. Reported data on the effects of Sonic hedgehog on differentiation of dopaminergic and serotonergic neurons from cultures of neural precursors are controversial. In the present study, cultures of proliferating neurospheres of mesencephalic precursors treated with anti-sonic hedgehog antibodies showed significantly less serotonergic and GABAergic cells and a markedly higher number of dopaminergic neurons generated from the neurospheres than control cultures. Treatment of the neurospheres with cyclopamine, which selectively inhibits sonic hedgehog signaling by preventing Smoothened activation, did not induce significant changes in generation of serotonergic and dopaminergic neurons. This suggests that Smoothened activation is not significantly involved in the above-mentioned effects and that sonic hedgehog may exert effects on the mesencephalic precursors that do not involve the canonical Patched-Smoothened-Gli signaling.
[Show abstract][Hide abstract] ABSTRACT: Neurotransmitters have been shown to control CNS neurogenesis, and GABA-mediated signaling is thought to be involved in the regulation of nearly all key developmental stages. Generation of dopaminergic (DA) neurons from stem/precursor cells for cell therapy in Parkinson's disease has become a major focus of research. However, the possible effects of GABA on generation of DA neurons from proliferating neurospheres of mesencephalic precursors have not been studied. In the present study, GABA(A), and GABA(B) receptors were found to be located in DA cells. Treatment of cultures with GABA did not cause significant changes in generation of DA cells from precursors. However, treatment with the GABA(A) receptor antagonist bicuculline (10(-5) M) led to a significant increase in the number DA cells, and treatment with the GABA(B) receptor antagonist CGP 55845 (10(-5) M) to a significant decrease. Simultaneous treatment with bicuculline and CGP 55845 did not induce significant changes. Apoptotic cell death studies and bromodeoxyuridine immunohistochemistry indicated that the aforementioned differences in generation of DA neurons are not due to changes in survival or proliferation of DA cells, but rather to increased or decreased differentiation of mesencephalic precursors towards the DA phenotype. The results suggest that these effects are exerted via GABA receptors located on DA precursors, and are not an indirect consequence of effects on the serotonergic or glial cell population. Administration of GABA(A) receptor antagonists in the differentiation medium may help to obtain higher rates of DA neurons for potential use in cell therapy for Parkinson's disease.
[Show abstract][Hide abstract] ABSTRACT: Cell death induced by 6-hydroxydopamine (6-OHDA) is thought to be caused by reactive oxygen species (ROS) derived from 6-OHDA autooxidation and by a possible direct effect of 6-OHDA on the mitochondrial respiratory chain. However, the process has not been totally clarified. In rat primary mesencephalic cultures, we observed a significant increase in dopaminergic (DA) cell loss 24 h after administration of 6-OHDA (40 micromol/L) and a significant increase in NADPH subunit expression, microglial activation and superoxide anion/superoxide-derived ROS in DA cells that were decreased by the NADPH inhibitor apocynin. Low doses of 6-OHDA (10 micromol/L) did not induce a significant loss of DA cells or a significant increase in NADPH subunit expression, microglial activation or superoxide-derived ROS. However, treatment with the NADPH complex activator angiotensin II caused a significant increase in all the latter. Forty-eight hours after intrastriatal 6-OHDA injection in rats, there was still no loss of DA neurons although there was an increase in NADPH subunit expression and NADPH oxidase activity. The results suggest that in addition to the autooxidation-derived ROS and the inhibition of the mitochondrial respiratory chain, early microglial activation and NADPH oxidase-derived ROS act synergistically with 6-OHDA and constitute a relevant and early component of the 6-OHDA-induced cell death.
Journal of Neurochemistry 11/2007; 103(1):145-56. · 3.97 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Inductive signals mediating the differentiation of neural precursors into serotonergic (5-HT) or dopaminergic neurons have not been clarified. We have recently shown that in cell aggregates obtained from rat mesencephalic precursors, reduction of serotonin levels induces a marked increase in generation of dopaminergic neurons. In the present study we treated rat neurospheres with antagonists of the main subtypes of 5-HT receptors, 5-HT transport inhibitors, or 5-HT receptor agonists, and studied the effects on generation of dopaminergic neurons. Cultures treated with Methiothepin (5-HT(1,2,5,6,7) receptor antagonist), the 5-HT(4) receptor antagonist GR113808;67:00-.or the 5-HT(7) receptor antagonist SB 269970 showed a significant increase in generation of dopaminergic cells. Treatment with the 5-HT(1B/1D) antagonist GR 127935, the 5-HT(2) antagonist Ritanserin, the 5-HT transporter inhibitor Fluoxetine, the dopamine and norepinephrine transport inhibitor GBR 12935, or with both inhibitors together, or 5-HT(4) or 5-HT(7) receptor agonists induced significant decreases in generation of dopaminergic cells. Cultures treated with WAY100635 (5-HT(1A) receptor antagonist), the 5-HT(3) receptor antagonist Ondasetron, or the 5-HT(6) receptor antagonist SB 258585 did not show any significant changes. Therefore, 5-HT(4) and 5-HT(7) receptors are involved in the observed serotonin-induced decrease in generation of dopaminergic neurons from proliferating neurospheres of mesencephalic precursors. 5-HT(4) and 5-HT(7) receptors were found in astrocytes and serotonergic cells using double immunolabeling and laser confocal microscopy, and the glial receptors appeared to play a major role.
[Show abstract][Hide abstract] ABSTRACT: The search for alternative sources of dopaminergic cells, other than primary fetal tissue for transplantation in Parkinson's disease has become a major focus of research. Different methodological approaches have led to generation in vitro of cells expressing DA-cell markers, although these cells are frequently unable to survive for a long time in vivo after transplantation and/or induce functional effects in the host brain. In the present study, we grafted cell aggregates treated with antibodies against fibroblast growth factor 4 into dopaminergic-denervated striata in rats. Furthermore, we grafted cell suspensions from primary mesencephalic fetal tissue. Grafts from expanded precursors were able to survive (at least 3 months postgrafting) and most decreased the lesion-induced ipsiversive rotation. In addition, immunolabeling for tyrosine hydroxylase and/or Fos showed that the grafts reinnervated the surrounding striatal tissue with dopaminergic terminals, and induced the expression of Fos in the striatal neurons of the reinnervated area after administration of amphetamine to the host rat. The number of dopaminergic cells in grafts from expanded precursors inducing rotational recovery was usually lower (1,226+/-314) than that in grafts from primary fetal tissue (1,671+/-122), but they were more densely packed in grafts that were of smaller volume and did not have the characteristic central nondopaminergic area observed in grafts from primary fetal tissue. The results suggest that long-term survival and functional integration into the DA-denervated striatum can be achieved with grafts of expanded mesencephalic precursors.
[Show abstract][Hide abstract] ABSTRACT: Cultures of rat mesencephalic precursors treated with interleukin-1beta or angiotensin II contained significantly more dopaminergic neurons than controls. However, simultaneous treatment with angiotensin II and interleukin-1beta did not induce any further increase. Treatment with the angiotensin type-2 receptor antagonist PD 123319 precluded both the angiotensin- and the interleukin-induced increase. The present results indicate that angiotensin type-2 receptors and interleukin-1 cooperate to induce the dopaminergic phenotype.
Developmental Brain Research 09/2005; 158(1-2):120-2. · 1.78 Impact Factor