N-acetylaspartylglutamate and beta-NAAG protect against injury induced by NMDA and hypoxia in primary spinal cord cultures

The acidic dipeptide N-acetylaspartylglutamate (NAAG) is the most prevalent peptide in the central nervous system. NAAG is a low potency agonist at the NMDA receptor, and hydrolysis of NAAG yields the more potent excitatory amino acid neurotransmitter glutamate. beta-NAAG is a competitive inhibitor of the NAAG hydrolyzing enzyme N-acetylated alpha-linked acidic dipeptidase (NAAG peptidase activity) or glutamate carboxypeptidase II, and may also act as a NAAG-mimetic at some of the sites of NAAG pharmacological activity. Since NAAG has been shown to have neuroprotective characteristics in a number of experimental preparations, it is the purpose of the present study to specifically evaluate the possible efficacy of NAAG and beta-NAAG against NMDA- and hypoxia-induced injury to spinal cord mixed neuronal and glial cell cultures. NAAG (500-1000 microM) protected against NMDA- or hypoxia-induced injuries to spinal cord cultures, and the nonhydrolyzable analog beta-NAAG (250-1000 microM) completely eliminated the loss of viability caused by either insult. Both peptides also attenuated NMDA-induced increases in intraneuronal Ca(2+). Nonspecific mGluR antagonists, pertussis toxin, a stable cAMP analog, and manipulation of NAAG peptidase activity did not by themselves alter cell damage and did not influence the neuroprotective effects of NAAG. NAAG was not protective against kainate- or AMPA-induced cellular injury, while beta-NAAG was partially neuroprotective against both insults. At 2 mM, NAAG and beta-NAAG reduced neuronal survival and increased intraneuronal Ca(2+); these effects were only marginally attenuated by dizocilpine and APV. The results indicate that NAAG and beta-NAAG protect against excitotoxic and hypoxic injury to spinal cord neurons, and do so predominantly by interactions with NMDA and not mGluR receptors.     

Increased adenine nucleotide translocator 1 in reactive astrocytes facilitates glutamate transport

A hallmark of central nervous system (CNS) pathology is reactive astrocyte production of the chronic glial scar that is inhibitory to neuronal regeneration. The reactive astrocyte response is complex; these cells also produce neurotrophic factors and are responsible for removal of extracellular glutamate, the excitatory neurotransmitter that rises to neurotoxic levels in injury and disease. To identify genes expressed by reactive astrocytes, we employed an in vivo model of the glial scar and differential display PCR and found an increase in the level of Ant1, a mitochondrial ATP/ADP exchanger that facilitates the flux of ATP out of the mitochondria. Ant1 expression in reactive astrocytes is regulated by transforming growth factor-beta1, a pluripotent CNS injury-induced cytokine. The significance of increased Ant1 is evident from the observation that glutamate uptake is significantly decreased in astrocytes from Ant1 null mutant mice while a specific Ant inhibitor reduces glutamate uptake in wild-type astrocytes. Thus, the astrocytic response to CNS injury includes an apparent increase in energy mobilization capacity by Ant1 that contributes to neuroprotective, energy-dependent glutamate uptake.     

Neuroprotective effects of (+/-)-huprine Y on in vitro and in vivo models of excitoxicity damage

We have investigated the neuroprotective effects of (+/-)-huprine Y on excitotoxic lesions in rat cerebellar granule cells (CGCs). (+/-)-Huprine Y prevented cell death induced by 100 microM glutamate, as well as, 10 microM MK-801, a NMDA receptor antagonist, in a significant manner. On the other hand, intracellular calcium increase induced by NMDA (200 microM), measured by fura-2 fluorescence, was prevented by (+/-)-huprine Y with an EC(50) of 12.44 microM, which evidences the modulatory action of this compound on NMDA-induced calcium currents. In vivo, we have studied (+/-)-huprine Y neuroprotective effects on striatal lesions induced by the subacute administration of the mitochondrial toxin 3-nitropropionic acid (3-NP, 30 mg/kg, ip, for 10 days). We have assessed that both the behavioral and the morphological consequences of the lesion were prevented by pretreatment with (+/-)-huprine Y (2.5 mg/kg/twice a day, ip). Striatal gliosis induced by 3-NP treatment was prevented by (+/-)-huprine Y pretreatment, as demonstrated by the attenuation of both the increase in [(3)H]PK 11195 specific binding indicative of microgliosis and the expression of hsp27 kDa, a chaperone expressed mainly in astrocytes. In conclusion, (+/-)-huprine Y attenuated excitotoxic-induced lesions, both in vitro and in vivo, and further evidence is provided for the potential use of this compound in the prevention of neurodegenerative disorders.     

Ovarian steroid modulation of seizure severity and hippocampal cell death after kainic acid treatment

To determine whether maintained estrogen or progesterone levels affect kainic acid (KA) seizure patterns or the susceptibility of hippocampal neurons to death from seizures, ovariectomized Sprague-Dawley rats were implanted with estrogen pellets, 0.1 or 0.5 mg, that generated serum levels of 42.4 +/- 6.6 (mean +/- SEM) and 242.4 +/- 32.6 pg/ml or one to six capsules of progesterone that generated serum levels of 11.00 +/-.72 to 48.62 +/- 9.4 ng/ml. Seven days later, the rats were administered KA (8.5mg/kg, ip) and scored for seizure activity; 96 h later, the rats were killed and their brains processed for localization of neuron nuclear antigen (NeuN), a general neuronal marker. The hippocampus was scored for spread (the number of separate regions showing cell loss), and the area within the CA fields occupied by NeuN immunoreactivity was measured (indicating surviving neurons). Administration of estrogen or progesterone (independent of dose) significantly reduced mortality from KA seizures. Progesterone reduced seizure severity in animals that received one to four implants; compared with controls, no difference in seizure severity was noted for animals with six progesterone implants. The reduced seizures in progesterone-treated animals were accompanied by a reduction in the spread of hippocampal damage (r(2) = 0.87; P < 0.05). Likewise, in progesterone-treated rats, neuron survival and reduction in seizure scores were correlated (r(2) = 0.76; P < 0.0001). Estrogen had no effect on seizure severity (P > 0.05), but reduced both the spread (P < 0.05) and degree of neuronal loss (P < 0.05). Indeed, in the estrogen-treated rats, neuronal death was significantly lower than that observed in progesterone-treated animals with equally severe seizures (P < 0.05). These data are consistent with the hypothesis that progesterone produces its effects by reducing seizures, whereas estrogen has little beneficial effect on seizure behavior but protects the hippocampus from the damage seizures produce.     

Recombinant adeno-associated viral vector (rAAV) delivery of GDNF provides protection against 6-OHDA lesion in the common marmoset monkey (Callithrix jacchus)

Glial cell line-derived neurotrophic factor (GDNF) has shown potential as a treatment for Parkinson's disease. Recombinant adeno-associated viral vectors expressing the GDNF protein (rAAV-GDNF) have been used in rodent models of Parkinson's disease to promote functional regeneration after 6-OHDA lesions of the nigrostriatal system. The goal of the present study was to assess the anatomical and functional efficacy of rAAV-GDNF in the common marmoset monkey (Callithrix jacchus). rAAV-GDNF was injected into the striatum and substantia nigra 4 weeks prior to a unilateral 6-OHDA lesion of the nigrostriatal bundle. Forty percent of the dopamine cells in the lesioned substantia nigra of the rAAV-GDNF-treated monkeys survived, compared with 21% in the untreated monkeys. Fine dopaminergic fibres were observed microscopically in the injected striatum of some rAAV-GDNF-treated monkeys, suggesting that rAAV-GDNF treatment may have prevented, at least in part, the loss of dopaminergic innervation of the striatum. Protection of dopamine cells and striatal fibre innervation was associated with amelioration of the lesion-induced behavioural deficits. rAAV-GDNF-treated monkeys showed partial or complete protection not only in the amphetamine and apomorphine rotation but also in head position and the parkinsonian disability rating scale. Therefore, our study provides evidence for the behavioural and anatomical efficacy of GDNF delivered via an rAAV vector as a possible treatment for Parkinson's disease.     

2-OH-estradiol, an endogenous hormone with neuroprotective functions

We compared the neuroprotective effects of the catecholestrogen 2-hydroxy-estradiol (2-OH-E₂) to the actions of 17-β-estradiol (E₂), since catecholestrogens have been clinically implicated in the pathophysiology of major depression and other psychiatric diseases. Using the hippocampal HT22 cell line as a well-established in vitro model system, we here show that the extent of the neuroprotective effects of 2-OH-E₂ was significantly increased compared to the physiological estrogen E₂ at equimolar concentrations after a toxic challenge with hydrogen peroxide. Statistically significant effects of neuroprotection as measured by survival of HT22 cells were detectable at concentrations of 1 and 10 μM of 2-OH-E₂ or E₂. Studies on the time-dependence of the evoked reactions showed that a pre-incubation and a post-incubation up to 30 min with a dose of 10 μM of 2-OH-E₂ resulted in a significant decrease in cell death after incubation with hydrogen peroxide if compared to E₂. Further characterization of the effects in rat brain homogenates with an assay for the induction of cellular lipid peroxidation (LPO) revealed, that 2-OH-E₂ was more effective in the reduction of LPO than E₂ in equimolar concentrations. This indicates a pharmacologically relevant effect of this hormone metabolite and a mechanism of action, which does not involve the classical estrogen receptor. In conclusion, the catecholestrogen 2-OH-E₂ induces increased neuroprotective actions in comparison to the major physiological estrogen E₂, suggesting a clinically relevant physiological function of catecholestrogens during health and disease.     

血管内皮生长因子在脑缺血损伤的血管和神经保护作用

血管内皮生长因子是目前所知唯一作用于血管内皮的生长因子,在血管发生中处枢纽地位。它选择性地作用于内皮细胞、促进新生血管形成;它参与脑缺血损伤的病理生理过程。本文就VEGF血管和神经保护作用进展作一综述。     

Efficacy and tolerability of 0.2% brimonidine tartrate for the treatment of acute non-arteritic anterior ischemic optic neuropathy (NAION): a 3-month, double-masked, randomised, placebo-controlled trial

INTRODUCTION: Neuroprotection may be an option in ischemic optic nerve disease. There have been promising reports about the neuroprotective ability of topical brimonidine in animal studies. Therefore, we tested whether 0.2% brimonidine tartrate could improve the outcome of patients with non-arteritic anterior ischemic optic neuropathy (NAION). The study was stopped after an interim analysis, having not proven its feasibility within practicable time frame. METHODS: A 3-month, double-masked, placebo-controlled, randomised European multicenter trial conducted according to good clinical practice rules. Thirty-six patients (22 m, 14 f), older than 40 years with first eye involvement and typical signs of NAION were included in the study within the 1st week after visual loss (VA 0.05-1.0) and were randomized to treatment with either brimonidine 0.2% (Alphagan) or placebo. Visual acuity (VA, primary endpoint), visual field (VF, Humphrey 30-2 and Goldmann, quantified by a modified Esterman grid) and automated swinging flashlight test (SWIFT) were performed at baseline, 2 weeks, 4 weeks and 12 weeks after first visit. Primary analysis aimed at intention-to-treat group (ITT, n=29), secondary analysis to the per protocol population (PP, n=25). Tolerability and safety were tested in the safety group (n=36). A two-sample two-sided t-test was used for statistical analysis (alpha level at 0.05). RESULTS: VA did not show statistically significant difference by treatment. There were non-significant trends for better visual field results in the brimonidine group. Adverse events consisting of local irritation were observed six times in the verum and three times in the placebo group. No serious adverse events occurred. CONCLUSION: In contradiction to an open-labeled, retrospective study published by Fazzone et al., the results of this trial did not indicate any harmful effect of brimonidine in patients suffering from NAION. However, a statistically significant advantage for the patients receiving brimonidine tartrate could not be shown.     

Dose-dependent protective effects of apomorphine against methamphetamine-induced nigrostriatal damage

(R)-apomorphine is a non-selective dopamine (DA) agonist which is used in the treatment of Parkinson's disease. In addition to symptomatic effects, apomorphine exerts a neuroprotective activity in specific experimental models. For instance, apomorphine prevents experimental parkinsonism induced by the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine. Neuroprotection obtained with apomorphine does not seem to be related to its dopamine (DA) agonist properties, instead it appears to be grounded on the antioxidant and the free radical scavenging effects of the compound. In this study, we sought to determine whether apomorphine protects against methamphetamine toxicity. We found that apomorphine (1; 5 and 10 mg/kg) dose-dependently protects against methamphetamine- (5 mg/kg X3, 2 h apart) induced striatal DA loss and reduction of tyrosine hydroxylase (TH) activity in the rat striatum. These protective effects are neither due to a decrease in the amount of striatal methamphetamine nor to hypothermia as indicated by measurement of striatal methamphetamine and body temperature at different time intervals after drug administration. The effects of apomorphine were neither opposite to, nor reversed by the DA antagonist haloperidol despite no decrease in body temperature was observed when apomorphine was given in combination with haloperidol. The present data are in line with recent studies suggesting a DA receptor-independent neuroprotective effect of apomorphine on DA neurons and call for further studies aimed at evaluating potential neuroprotective effects of apomorphine in Parkinson's disease.     

SKF-38393, a dopamine receptor agonist, attenuates 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced neurotoxicity

Parkinson's disease (PD) is characterized by progressive degeneration of nigrostriatal dopaminergic neurons. Several factors such as inhibition of the mitochondrial respiration, generation of hydroxyl radicals and reduced free radical defense mechanisms causing oxidative stress, have been postulated to contribute to the degeneration of dopaminergic neurons. 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) treated animals is a useful experimental model of PD, exhibiting most of the clinical features, as well as the main biochemical and pathologic symptoms of the disease. In the present study, we have examined a dopaminergic (D1) receptor agonist, SKF-38393 HCl (SKF) for its possible neuroprotective action against MPTP-induced insults on dopaminergic neurons. MPTP is converted by monoamine oxidase-B (MAO-B) to its neurotoxic metabolite 1-methyl-4-phenyl-pyridinium (MPP+), which is then taken up into the dopaminergic neurons. SKF-38393 had no effects either on total or monoamine oxidase B in the striatum. SKF-38393 blocked the MPTP-induced depletion of glutathione and attenuated MPTP-induced depletion of dopamine. Furthermore, it enhanced the activity of superoxide dismutase and hence mimicked the action of selegiline. The results of these studies are interpreted to suggest that SKF-38393 may prove a valuable drug in the treatment of Parkinson's disease.