Marti Santiago

THE ROLE OF GABA RECEPTORS IN THE CONTROL OF NIGROSTRIATAL DOPAMINERGIC-NEURONS - DUAL-PROBE MICRODIALYSIS STUDY IN AWAKE RATS

A microdialysis probe implanted into the substantia nigra was used to infuse gamma-aminobutyric acid-ergic (GABAergic) compounds onto cell bodies/dendrites of dopaminergic neurons, while a second microdialysis probe was used to record the extracellular concentrations of dopamine and 3,4-dihydroxy-phenylacetic acid (DOPAC) in the ipsilateral striatum. The GABAA receptor agonist muscimol (10 mumol/l) increased the release of dopamine in the ipsilateral striatum to 120% of the control values. The GABAB receptor agonist, (Z)-3[(aminoiminomethyl)-thiol]-prop-2- enoic acid (500 mumol/l), was without effect. Infusion of the GABAA receptor antagonists, bicuculline (50 mumol/l) and picrotoxin (50 mumol/l), stimulated the release of dopamine in the ipsilateral striatum to 160 and 130% of the controls, respectively. The GABAB receptor agonist, baclofen (10 and 50 mumol/l), strongly inhibited the release of striatal dopamine, whereas infusion of the GABAB receptor antagonist, 2-hydroxy-saclofen (100 mumol/l), was without effect. The results indicate that, in the substantia nigra, GABAA as well as GABAB receptors participate in controlling the activity of dopaminergic neurons.

Regulation of the prefrontal cortical dopamine release by GABAA and GABAB receptor agonists and antagonists

The gamma-aminobutyric acid-dopamine (GABA-DA) relationship was studied by intracerebral microdialysis in the prefrontal cortex. Nomifensine (5 microM) was included in the Ringer solution during all the dialysis experiments. Muscimol, a GABAA receptor agonist (50 and 500 microM) did not affect the extracellular output of DA and 3,4-dihydroxyphenylacetic acid (DOPAC). Baclofen, a GABAB receptor agonist (50 microM) significantly decreased the extracellular output of DA and DOPAC. On the other hand, picrotoxin and phaclofen, GABAA and GABAB receptor antagonists respectively, at a concentration of 50 microM, both significantly increased the release of DA. While the DOPAC level was affected only by picrotoxin perfusion. The present study indicates that GABA could control the release of DA in the prefrontal cortex.

Group II metabotropic glutamate receptor activation protects striatal dopaminergic nerve terminals against MPP+‐induced neurotoxicity along with brain‐derived neurotrophic factor induction

We have studied the in vivo effect of the selective agonist for group II metabotropic glutamate receptors (2S, 2′R, 3′R)‐2‐(2′3′‐dicarboxycyclopropyl)glycine (DCG‐IV) against MPP+‐induced toxicity on rat striatal dopaminergic nerve terminals by using both microdialysis and immunohistochemical techniques. Perfusion of 1 mm DCG‐IV during 1 h protected dopaminergic nerve terminals against the degeneration induced by a 15‐minute perfusion of 1 mm MPP+. In addition, the microglial cell population was markedly activated 24 h after DCG‐IV perfusion. The astroglial cell population was only markedly activated around the microdialysis probe. This protective effect seems to be dependent on protein synthesis since 1 mm cycloheximide, an inhibitor of protein synthesis, abolished the neuroprotective effect of 1 mm DCG‐IV against MPP+ toxicity. Perfusion of DCG‐IV induced an upregulation of striatal brain‐derived neurotrophic factor (BDNF) mRNA expressing cells which were confined precisely around the microdialysis probe. Taken together, our results suggest that the induction and release of brain‐derived neurotrophic factor (BDNF) by activated glial cells induced by DCG‐IV perfusion may account for its protective action against MPP+‐induced dopaminergic terminal degeneration.

Simvastatin prevents the inflammatory process and the dopaminergic degeneration induced by the intranigral injection of lipopolysaccharide.

Anti‐inflammatory strategies have attracted much interest for their potential to prevent further deterioration of Parkinson’s disease. Recent experimental and clinical evidence indicate that statins – extensively used in medical practice as effective lipid‐lowering agents – have also anti‐inflammatory effects. In this study, we investigated the influence of simvastatin on the degenerative process of the dopaminergic neurons of the rat following intranigral injection of lipopolysaccharide (LPS), a potent inductor of inflammation that we have previously used as an animal model of Parkinson’s disease. We evaluated TH positive neurons, astroglial, and microglial populations and found that simvastatin prevented the inflammatory processes, as the induction of interleukin‐1β, tumor necrosis factor‐α, and iNOS and the consequent dopaminergic degeneration induced by LPS. Moreover, simvastatin produced the activation of the neurotrophic factor BDNF, along with the prevention of the oxidative damage to proteins. Moreover, it also prevents the main changes produced by LPS on different mitogen‐activated protein kinases, featured as increases of P‐c‐Jun N‐terminal protein kinase, P‐extracellular signal‐regulated kinase, p‐38, and P‐glycogen synthase kinase and the decrease of the promotion of cell survival signals such as cAMP response element‐binding protein and Akt. Our results suggest that statins could delay the progression of dopaminergic degeneration in disorders involving inflammatory processes.

Histamine Infusion Induces a Selective Dopaminergic Neuronal Death Along with an Inflammatory Reaction in Rat Substantia Nigra

We have evaluated the effects of a direct infusion of histamine, as mediator of inflammatory response, in substantia nigra, striatum, medial septum, and medial lemniscus. Injection of 100 and 250 nmol of histamine in substantia nigra produced a selective damage in dopaminergic neurons evidenced by the loss of tyrosine hydroxylase mRNA‐expressing cells, tyrosine hydroxylase‐immunolabeled‐positive cell bodies, and dopamine and 3,4‐dihydroxyphenylacetic acid levels. In parallel we found an acute inflammatory response manifested by a loss of glial fibrillary acidic protein‐immunolabeled astrocytes and, at precisely the same area, an activation of microglia. In the striatum, only high doses (500 nmol) produced an evident terminal degeneration. The selective neurotoxicity of histamine for dopaminergic cells was demonstrated by the unaltered transcription of glutamic acid decarboxylase mRNA in substantia nigra. Moreover, intraseptal injection of 100 nmol of histamine failed to alter the pattern of choline acetyltransferase mRNA‐expressing cells, and intraparenchymal injection of histamine in medial lemniscus failed to alter the pattern of serotonin‐immunolabeled cells. We conclude that the substantia nigra is highly sensitive to histamine‐derived neurotoxicity, where inflammatory processes mediated by histamine could be important in the pathological changes that lead to dopaminergic neuronal damage after histamine infusion.

Regulation of prefrontal cortical dopamine release by dopamine receptor agonists and antagonists

The effect of dopamine D1 and D2 receptors agonists and antagonists on extracellular dopamine release was evaluated by microdialysis in the prefrontal cortex. Nomifensine (5 microM) was included in the Ringer solution during the experiments. Cortical dopamine release was tetrodotoxin- and calcium-dependent and was stimulated by high potassium (60 mM) Ringer solution. 1-Methyl-4-phenylpyridinium ion (MPP+) (10 mM) increased the extracellular output of dopamine. SKF-38393 decreased the release of dopamine in a dose-related manner to about 80, 40 and 0% of the control values at 0.1, 1 and 10 microM, respectively. The decrease produced by SKF-38393 (10 microM) was partially antagonized by SCH-23390 at a concentration of 1 microM. Perfusion of CY-208243 (10 microM) produced a decrease in the release of dopamine to about 70% of controls. Quinpirole, at a concentration of 10 microM, produced a decrease in the release of dopamine to about 65% of controls. SCH-23390 and sulpiride, at 10 microM, increased the extracellular output of dopamine to about 150% of controls. These results indicate that dopamine D1 and D2 receptors are implicated in the autoregulation of dopamine release in the prefrontal cortex.

Peripheral inflammation increases the deleterious effect of CNS inflammation on the nigrostriatal dopaminergic system

Evidence supports the role of inflammation in the development of neurodegenerative diseases. In this work, we are interested in inflammation as a risk factor by itself and not only as a factor contributing to neurodegeneration. We tested the influence of a mild to moderate peripheral inflammation (injection of carrageenan into the paws of rats) on the degeneration of dopaminergic neurons in an animal model based on the intranigral injection of lipopolysaccharide (LPS), a potent inflammatory agent. Overall, the treatment with carrageenan increased the effect of the intranigral injection of LPS on the loss of dopaminergic neurons in the SN along with all the other parameters studied, including: serum levels of the inflammatory markers TNF-α, IL-1β, IL-6 and C-reactive protein; activation of microglia, expression of proinflammatory cytokines, the adhesion molecule ICAM and the enzyme iNOS, loss of astrocytes and damage to the blood brain barrier (BBB). The possible implication of BBB rupture in the increased loss of dopaminergic neurons has been studied using another Parkinsons disease animal model based on the intraperitoneal injection of rotenone. In this experiment, loss of dopaminergic neurons was also strengthened by carrageenan, without affecting the BBB. In conclusion, our data show that a mild to moderate peripheral inflammation can exacerbate the degeneration of dopaminergic neurons caused by a harmful stimulus.

In vivo release of dopamine from rat striatum, substantia nigra and prefrontal cortex: differential modulation by baclofen

1 The effect of baclofen, a GABAB receptor agonist, on the release of dopamine from the striatum (ST), substantia nigra (SN) and prefrontal cortex (PFC) of the rat was examined by intracerebral microdialysis. 2 Perfusion of baclofen 50 μm did not affect the striatal release of dopamine. However, dopamine release was markedly reduced in the SN and PFC. 3 3,4‐Dihydroxyphenylacetic acid and homovanillic acid output increased in the ST and decreased in the SN and PFC when baclofen was perfused through the microdialysis probe. 5‐Hydroxyindoleacetic acid levels were not affected in any experimental condition by baclofen perfusion. 4 The results suggest that GABAB receptors modulate the release of dopamine in the SN and PFC, but do not affect the striatal release of dopamine, which indicates that the role of GABA receptor activation is different in the dopaminergic terminals of the ST and PFC.

In vivo protection of striatum from MPP+ neurotoxicity by N-methyl-D-aspartate antagonists

The present study was designed to assess by microdialysis whether N-methyl-D-aspartate (NMDA) non-competitive receptor antagonist, MK-801, is able to protect dopaminergic neurons against 1-methyl-4-phenylpyridinium ion (MPP+) neurotoxicity. An intraperitoneal injection of MK-801, 10 mumol/kg, half an hour before striatal MPP+, 10 mM, perfusion did not protect against its neurotoxicity. Afterwards, rats received an intraperitoneal injection of MK-801 every 4 h, during 24 h. Under these conditions, one day after MPP+ perfusion, DA basal extracellular levels were close to the detection limit of our HPLC equipment in both control and MK-801 treated rats. 3,4-Dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) basal output were lower in the former than in the latter rats. A second MPP+ perfusion statistically increased extracellular levels of DA and decreased DOPAC and HVA output in both groups of rats. However, the increase in extracellular DA overflow was higher in MK-801 treated rats than in the control group, indicating a higher number of surviving dopaminergic terminals. These results suggest that MK-801 is not able to protect against the primary direct neurotoxic action of MPP+, but a second MPP+ neurotoxic action mediated by excitatory amino acids could be partially prevented by MK-801.

Neuroprotective Effect of the Iron Chelator Desferrioxamine Against MPP+ Toxicity on Striatal Dopaminergic Terminals

Abstract: Microdialysis was used to evaluate the effect of desferrioxamine (DES) against 1‐methyl‐4‐phenylpyridinium (MPP+) toxicity. The presence of DES (40 fmol‐40 nmol/15 min for a total of 90 min) in the Ringer solution, coperfused with MPP+ (40 nmol/15 min) on day 1, produced on day 2 a higher extracellular dopamine output after perfusion of MPP+ than in control MPP+ perfusion experiments, in which no DES was administered on day 1. Both Ringer perfusion alone (control Ringer) and coperfusion of 40 nmol DES with 40 nmol MPP+ on day 1 produced on day 2 similar increases in extracellular dopamine output after a second MPP+ perfusion. In the control Ringer experiment, note that the MPP+ on day 2 is the first MPP+ perfusion. Perfusion of 800 fmol FeCl3/15 min along with 40 nmol MPP+ and 400 fmol DES on day 1 completely abolished on day 2 the neuroprotective effect found with 40 nmol MPP+ and 400 fmol DES; 800 fmol FeCl3 did not increase the neurotoxic effect of 40 nmol MPP+ perfusion. The ability of DES to protect against MPP+ toxicity may indicate a therapeutic strategy in the treatment of diseases when iron is implicated.