dizocilpine-maleate and 3-nitropropionic-acid

The striatum is particularly vulnerable to mitochondrial dysfunction and this problem is linked to pathology created by environmental neurotoxins, stimulants like amphetamine, and metabolic disease and ischemia. We studied the course of recovery following a single systemic injection of the mitochondrial complex II inhibitor 3-nitropropionic acid (3-NP) and found 3-NP caused lasting changes in motor behavior that were associated with altered activity-dependent plasticity at corticostriatal synapses in Fischer 344 rats. The changes in synapse behavior varied with the time after exposure to the 3-NP injection. The earliest time point studied, 24h after 3-NP, revealed 3-NP-induced an exaggeration of D1 Dopamine (DA) receptor dependent long-term potentiation (LTP) that reversed to normal by 48 h post-3-NP exposure. Thereafter, the likelihood and degree of inducing D2 DA receptor dependent long-term depression (LTD) gradually increased, relative to saline controls, peaking at 1 month after the 3-NP exposure. NMDA receptor binding did not change over the same post 3-NP time points. These data indicate even brief exposure to 3-NP can have lasting behavioral effects mediated by changes in the way DA and glutamate synapses interact.

Topics: Adrenergic Agents; Analysis of Variance; Animals; Benzazepines; Biophysics; Cerebral Cortex; Corpus Striatum; Dizocilpine Maleate; Dopamine Antagonists; Dose-Response Relationship, Drug; Electric Stimulation; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Excitatory Postsynaptic Potentials; Functional Laterality; In Vitro Techniques; Movement; Nitro Compounds; Oxidopamine; Propionates; Protein Binding; Rats; Rats, Inbred F344; Substantia Nigra; Sulpiride; Synapses; Time Factors; Tritium; Tyrosine 3-Monooxygenase

Mice heterozygous for the homeobox gene Engrailed-1 (En1) display progressive loss of mesencephalic dopaminergic (mDA) neurons. We report that exogenous Engrailed-1 and Engrailed-2 (collectively Engrailed) protect mDA neurons from 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), a mitochondrial complex I toxin used to model Parkinson's disease in animals. Engrailed enhances the translation of nuclearly encoded mRNAs for two key complex I subunits, Ndufs1 and Ndufs3, and increases complex I activity. Accordingly, in vivo protection against MPTP by Engrailed is antagonized by Ndufs1 small interfering RNA. An association between Engrailed and complex I is further confirmed by the reduced expression of Ndufs1 and Ndufs3 in the substantia nigra pars compacta of En1 heterozygous mice. Engrailed also confers in vivo protection against 6-hydroxydopamine and α-synuclein-A30P. Finally, the unilateral infusion of Engrailed into the midbrain increases striatal dopamine content, resulting in contralateral amphetamine-induced turning. Therefore, Engrailed is both a survival factor for adult mDA neurons and a regulator of their physiological activity.

Topics: alpha-Synuclein; Animals; Cell Count; Cells, Cultured; Chromatography, High Pressure Liquid; Dizocilpine Maleate; Dopamine; Dopamine Plasma Membrane Transport Proteins; Electron Transport Chain Complex Proteins; Embryo, Mammalian; Homeodomain Proteins; In Vitro Techniques; Male; Mesencephalon; Mice; Mice, Inbred C57BL; Mice, Transgenic; NADH Dehydrogenase; Nerve Tissue Proteins; Neurons; Neurotoxins; Nitro Compounds; Oxidopamine; Propionates; RNA, Small Interfering; Rotenone; Stereotyped Behavior; Tyrosine 3-Monooxygenase

There is accumulating evidence that excitotoxicity and oxidative stress resulting from excessive activation of glutamate (N-methyl-D-aspartate) NMDA receptors are major participants in striatal degeneration associated with 3-nitropropionic acid (3NP) administration. Although excitotoxic and oxidative mechanisms are implicated in 3NP toxicity, there are conflicting reports as to whether NMDA receptor antagonists attenuate or exacerbate the 3NP-induced neurodegeneration. In the present study, we investigated the involvement of NMDA receptors in striatal degeneration, protein oxidation and motor impairment following systemic 3NP administration. We examined whether NMDA receptor antagonists, memantine and ifenprodil, influence the neurotoxicity of 3NP. The development of striatal lesion and protein oxidation following 3NP administration is delayed by memantine but not affected by ifenprodil. However, in behavioral experiments, memantine failed to improve and ifenprodil exacerbated the motor deficits associated with 3NP toxicity. Together, these findings suggest caution in the application of NMDA receptor antagonists as a neuroprotective agent in neurodegenerative disorders associated with metabolic impairment.

Topics: Adenosine Diphosphate; Animals; Corpus Striatum; Dizocilpine Maleate; Drug Interactions; Male; Memantine; Motor Activity; Nerve Degeneration; Neuroprotective Agents; Nitro Compounds; Piperidines; Poly (ADP-Ribose) Polymerase-1; Poly(ADP-ribose) Polymerases; Propionates; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Tyrosine

Preconditioning by excitatory stimuli such as N-methyl-d-aspartate (NMDA) offers good neuroprotection against excitotoxic insults, but is potentially limited by the risk of damage associated with the treatment. We report here the potential of an alternative strategy, tested on rat neonatal cerebellar granule neurons, which involves a 48-hour preconditioning step using the potassium channel blocker 4-aminopyridine (4-AP), at a low (50 microM) and at a higher (2500 microM) concentration (in the presence or absence of the GABA(A) receptor antagonist, bicuculline). 4-Aminopyridine gave extensive protection against a number of stressors (glutamate, NMDA and 3-nitropropionic acid) applied 24 h following the end of the preconditioning period. Blockade of neuronal depolarisation by tetrodotoxin during preconditioning attenuated but did not eliminate protection, whilst co-application with the NMDA receptor blocker MK-801 increased protection. Western blot analysis showed that CREB phosphorylation was significantly increased by the 4-AP preconditioning, although bcl-2 expression was not stimulated. Glutamate induced cell death without significant activation of caspase-3, suggesting that 4-AP preconditioning is effective primarily against necrotic excitotoxicity. Since 4-AP preconditioning affords extensive protection against a range of neurotoxic insults we propose that it could provide the basis for a novel neuroprotective therapy worthy of further investigation.

Topics: 4-Aminopyridine; Animals; Bicuculline; Cell Death; Cells, Cultured; Cerebellum; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; GABA Antagonists; Glutamic Acid; N-Methylaspartate; Neurons; Neuroprotective Agents; Neurotoxins; Nitro Compounds; Propionates; Rats; Rats, Sprague-Dawley; Time Factors

3-nitropropionic acid (3-NP), a substance used for modelling Huntington's disease, was given to male Wistar rats in a single 20 mg/kg b.w. dose, and the resulting behavioral alterations in spontaneous locomotor activity were measured after 30 minutes. To detect the involvement of neurotransmitter systems in this immediate effect, the NMDA antagonist MK-801 (0.8 mg/kg); as well as an agonist, quinpirole (QP, 5 mg/kg) and an antagonist, sulpiride (SP, 80 mg/kg) of the dopamine D2 receptors, were given before 3-NP to separate groups of rats. Controls were given saline. All substances were injected ip. 3-NP decreased the rats' locomotor, especially vertical, activity, whereas local activity was increased. Based on the further changes of 3-NP effects in the combination groups it could be concluded that dopaminergic rather than glutamatergic mechanisms were possibly involved in the acute behavioral effect of 3-NP.

Topics: Animals; Behavior, Animal; Disease Models, Animal; Dizocilpine Maleate; Dopamine Agents; Dopamine D2 Receptor Antagonists; Excitatory Amino Acid Agents; Huntington Disease; Male; Motor Activity; N-Methylaspartate; Nitro Compounds; Propionates; Quinpirole; Rats; Rats, Wistar; Receptors, Dopamine; Receptors, Dopamine D2; Receptors, Glutamate; Sulpiride

3-Nitropropionic acid (3NP), an irreversible inhibitor of succinate dehydrogenase, induces both rapid necrotic and slow apoptotic death in rat hippocampal neurons. Low levels of extracellular glutamate (10 microM) shift the 3NP-induced cell death mechanism to necrosis, while NMDA receptor blockade results in predominantly apoptotic death. In this study, we examined the 3NP-induced alterations in free cytosolic and mitochondrial calcium levels, ATP levels, mitochondrial membrane potential, and calpain and caspase activity, under conditions resulting in the activation of apoptotic and necrotic pathways. In the presence of 10 microM glutamate, 3NP administration resulted in a massive elevation in [Ca(2+)](c) and [Ca(2+)](m), decreased ATP, rapid mitochondrial membrane depolarization, and a rapid activation of calpain but not caspase activity. In the presence of the NMDA receptor antagonist MK-801, 3NP did not induce a significant elevation of [Ca(2+)](c) within the 24h time period examined, nor increase [Ca(2+)](m) within 1h. ATP was maintained at control levels during the first hour of treatment, but declined 64% by 16h. Calpain and caspase activity were first evident at 24h following 3NP administration. 3NP treatment alone resulted in a more rapid decline in ATP, more rapid calpain activation (within 8h), and elevated [Ca(2+)](m) as compared to the results obtained with added MK-801. Together, the results demonstrate that 3NP-induced necrotic neuron death is associated with a massive calcium influx through NMDA receptors, resulting in mitochondrial depolarization and calpain activation; while 3NP-induced apoptotic neuron death is not associated with significant elevations in [Ca(2+)](c), nor with early changes in [Ca(2+)](m), mitochondrial membrane potential, ATP levels, or calpain activity.

Topics: Adenosine Triphosphate; Animals; Brain; Calcium; Cell Death; Cytosol; Dizocilpine Maleate; Female; Fetus; Glutamic Acid; In Vitro Techniques; Intracellular Membranes; Membrane Potentials; Mitochondria; Neurons; Neurotoxins; Nitro Compounds; Pregnancy; Propionates; Rats; Rats, Sprague-Dawley

3-Nitropropionic acid (3NP), an irreversible inhibitor of succinate dehydrogenase, has been used to model features of neurodegenerative disorders including Huntington disease, as well as acute neuronal insults such as cerebral ischemia. 3NP induces rapid necrosis and delayed apoptosis in primary cultures of rat hippocampal neurons. Low levels of extracellular glutamate shift the cell death mechanism to necrosis, whereas antagonism of NMDA receptors results in predominately apoptotic death. In the present study, the involvement of cysteine proteases in the morphologic and biochemical alterations accompanying 3NP-induced neuron death was investigated. Immunoblots of spectrin breakdown products indicated Ca(2+)-dependent cysteine protease (calpain) activation within the 8 hours of 3NP administration, whereas caspase-3 activation was not evident until 16 to 48 hours after treatment. The NMDA receptor antagonist MK-801 (dizocilpine) decreased 3NP-induced calpain activity, but did not alter caspase-3 activity. Similar to MK-801, calpain inhibitors (Z-Val-Phe.H and Z-Leu-Phe-CONHEt) shifted the cell death morphology towards apoptosis and delayed, but did not prevent, the 3NP-induced cell death. Together, the results indicate that following 3NP administration, increased calpain activity precedes caspase-3 activation, contributes to the necrotic morphology, and facilitates and accelerates the cell death.

Topics: Amino Acid Chloromethyl Ketones; Animals; Apoptosis; Calpain; Caspases; Cell Death; Cell Survival; Cells, Cultured; Cysteine Proteinase Inhibitors; Dizocilpine Maleate; Drug Synergism; Embryo, Mammalian; Excitatory Amino Acid Antagonists; Glutamic Acid; Hippocampus; Immunoblotting; Immunohistochemistry; Necrosis; Neurons; Neurotoxins; Nitro Compounds; Oligopeptides; Propionates; Rats; Spectrin; Time Factors

3-Nitropropionic acid (3-NP), an irreversible inhibitor of succinate dehydrogenase, induced ATP depletion and both necrosis and apoptosis in human NT2-N neurons. Necrosis occurred predominantly within the first two days, and increased in a dose-dependent fashion with the concentration of 3-NP, whereas apoptosis was observed after 24 h or later at a similar rate in 0.1 mM and 5 mM 3-NP. We focused our efforts on intracellular calcium homeostasis during the first 48 h in 1 mM 3-NP, a period during which 10% of the neurons died by necrosis and 3% by apoptosis. All NT2-N neurons showed a stereotyped [Ca(2+)](i) rise, from 48+/-2 to 140+/-12 nM (mean +/-S.E.M.), during the first 2 h in 3-NP. Despite severe ATP depletion, however, [Ca(2+)](i) remained above 100 nM in only 17% and 25% of the NT2-N neurons after 24 and 48 h in 3-NP, respectively, indicating that most neurons were able to recover from acute [Ca(2+)](i) rise, and suggesting that chronic [Ca(2+)](i) dysregulation is a better indicator of subsequent necrosis. Blockade of N-methyl-D-aspartate-glutamate receptor by MK-801 substantially ameliorated 3-NP-induced ATP depletion, subsequent chronic [Ca(2+)](i) elevation, and survival. Moreover, xestospongin C, an inhibitor of endoplasmic reticulum Ca(2+) release, enhanced the capacity of NT2-N neurons to maintain [Ca(2+)](i) homeostasis and resist necrosis while subjected to sustained energy deprivation. As far as we know, this report is the first to employ human neurons to study the pathophysiology of 3-NP neurotoxicity.

Topics: Adenosine Triphosphate; Apoptosis; Calcium; Calcium Channels; Dizocilpine Maleate; Dose-Response Relationship, Drug; Endoplasmic Reticulum; Enzyme Inhibitors; Humans; In Situ Nick-End Labeling; Macrocyclic Compounds; Mitochondria; Necrosis; Neurons; Neurotoxins; Nitro Compounds; Oxazoles; Propionates; Receptors, Glutamate; Receptors, N-Methyl-D-Aspartate; Succinate Dehydrogenase; Time Factors; Tumor Cells, Cultured

Senile plaques composed mainly by beta-amyloid (Abeta) protein are one of the pathological hallmarks of Alzheimer's disease (AD). In vitro, Abeta and its active fragment 25-35 have been shown either to be directly neurotoxic or to exacerbate the damaging effect of other neurotoxic insults. However, the attempts to replicate Abeta neurotoxicity in vivo have yielded conflicting results. One of the most consistent alterations in AD is a reduced resting glucose utilization. Important evidence suggests that impairment of brain energy metabolism can lead to neuronal damage or facilitate the deleterious effects of some neurotoxic agents. In the present study we have investigated the influence of glycolysis inhibition induced by iodoacetate, and mitochondrial impairment induced by 3-nitropropionic acid (3-NP), in the toxicity of Abeta. We have studied Abeta neurotoxicity during energy deficiency both in vivo in the dentate gyrus of the hippocampal formation and in presynaptic terminals isolated from neocortex and hippocampus. Results show that during metabolic inhibition an enhanced vulnerability of hippocampal neurons to Abeta peptide toxicity occurs, probably resulting from decreased glucose metabolism and mitochondrial ATP production. Synaptosomal response to energy impairment and Abeta toxicity was evaluated by the MTT assay. Results suggest that synapses may be particularly sensitive to metabolic perturbation, which in turn exacerbates Abeta toxicity. The present data provide experimental support to the hypothesis that certain risk factors such as metabolic dysfunction and amyloid accumulation may interact to exacerbate AD, and that metabolic substrates such as pyruvate may play a role as a therapeutic tool.

Topics: Adenosine Triphosphate; Alzheimer Disease; Amyloid beta-Peptides; Animals; Disease Models, Animal; Dizocilpine Maleate; Energy Metabolism; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Glucose; Glycolysis; Hippocampus; Injections, Intraperitoneal; Iodoacetates; Male; Microinjections; Mitochondria; Neocortex; Neuroprotective Agents; Nitro Compounds; Peptide Fragments; Presynaptic Terminals; Propionates; Pyruvic Acid; Rats; Rats, Wistar; Synaptosomes

We studied the effect of an acute infusion of quinolinic acid (QUIN) on in vivo hydroxyl radical (.OH) formation in the striatum of awake rats. Using the microdialysis technique, the generation of.OH was assessed through electrochemical detection of the salicylate hydroxylation product 2,3-dihydroxybenzoic acid (2,3-DHBA). The .OH extracellular levels increased up to 30 times over basal levels after QUIN infusion (240 nmol/microl), returning to the baseline 2 h later. This response was attenuated, but not abolished, by pretreatment with the NMDA receptor antagonist MK-801 (10 mg/kg, i.p.) 60 min before QUIN infusion. The mitochondrial toxin 3-nitropropionic acid (3-NPA, 500 nmol/microl) had stronger effects than QUIN on .OH generation, as well as on other markers of oxidative stress explored as potential consequences of .OH increased levels. These results support the hypothesis that early .OH generation contributes to the pattern of toxicity elicited by QUIN. The partial protection by MK-801 suggests that QUIN neurotoxicity is not completely explained through NMDA receptor overactivation, but it may also involve intrinsic QUIN oxidative properties.

Topics: Animals; Chromatography, High Pressure Liquid; Corpus Striatum; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Glutathione; Glutathione Peroxidase; Hydroxybenzoates; Hydroxyl Radical; Lipid Peroxidation; Male; Microdialysis; Microinjections; Neuroprotective Agents; Nitro Compounds; Oxidative Stress; Perfusion; Propionates; Quinolinic Acid; Rats; Rats, Wistar; Salicylic Acid; Wakefulness

Excitotoxicity is implicated in the pathogenesis of several neurologic diseases, such as chronic neurodegenerative diseases and stroke. Recently, it was reported that excitotoxicity has a relationship to apoptotic neuronal death, and that the mitochondrial toxin, 3-nitropropionic acid (3-NP), could induce apoptosis in the striatum. Although striatal lesions produced by 3-NP could develop through an excitotoxic mechanism, the exact relationship between apoptosis induction and excitotoxicity after 3-NP treatment is still not clear. The authors investigated the role of excitotoxicity and oxidative stress on apoptosis induction within the striatum after intraperitoneal injection of 3-NP. The authors demonstrated that removal of the corticostriatal glutamate pathway reduced superoxide production and apoptosis induction in the denervated striatum of decorticated mice after 3-NP treatment. Also, the N-methyl-D-aspartate (NMDA) receptor antagonist, MK-801, prevented apoptosis in the striatum after 3-NP treatment for 5 days, whereas the non-NMDA receptor antagonist, 2,3-dihydroxy-6-nitro-7-sulphamoyl-benzo(F)quinoxaline, was ineffective. The authors also evaluated the initial type of neuronal death by 3-NP treatment for different durations from 1 to 5 days. In early striatal damage, apoptotic neuronal death initially occurred after 3-NP treatment. Our data show that excitotoxicity related to oxidative stress initially induces apoptotic neuronal death in mouse striatum after treatment with 3-NP.

Topics: Animals; Apoptosis; Caspases; Corpus Striatum; Decerebrate State; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Female; In Situ Nick-End Labeling; Mice; Mice, Inbred C57BL; Mitochondria; Nervous System; Neurotoxins; Nitro Compounds; Oxidative Stress; Propionates; Quinoxalines; Receptors, N-Methyl-D-Aspartate; Superoxides

Magnetic resonance imaging and in vivo proton magnetic resonance spectroscopy were used to evaluate the therapeutic effect of lamotrigine and MK-801 on rat brain lesions induced by 3-nitropropionic acid. Systemic administration of 3-nitropropionic acid (15 mg/kg per day) to two-month-old Sprague-Dawley rats (n = 10 for each group) for five consecutive days induced selective striatal and hippocampal lesions and specific behavioral change. Pretreatment with lamotrigine (10 mg/kg or 20 mg/kg per day) or MK-801 (2 mg/kg per day) attenuated the lesions and behavioral change. There were no significant differences in T2 values of the striatum and hippocampus among rats pretreated with MK-801, lamotrigine (20 mg/kg) and sham controls. Significant elevations of succinate/creatine and lactate/creatine ratios and decreases of N-acetylaspartate/creatine and choline/creatine ratios were observed after 3-nitropropionic acid injections (P < 0.001). The changes were nearly prevented after pretreatment with lamotrigine (20 mg/kg). However, the N-acetylaspartate/creatine in rats pretreated with lamotrigine (10 mg/kg) (P < 0.01) and MK-801 (P < 0.05) still showed significant reduction as compared with sham controls. Thus we conclude that both lamotrigine and MK-801 are effective in attenuation of brain lesions induced by 3-nitropropionic acid. A higher dose of lamotrigine provides a better neuroprotective effect than MK-801. With a better therapeutic effect and fewer side effects, lamotrigine is more promising for potential clinical application.

Topics: Animals; Aspartic Acid; Behavior, Animal; Brain; Choline; Corpus Striatum; Creatine; Dizocilpine Maleate; Hippocampus; In Vitro Techniques; Lamotrigine; Magnetic Resonance Imaging; Magnetic Resonance Spectroscopy; Male; Neuroprotective Agents; Nitro Compounds; Propionates; Rats; Rats, Sprague-Dawley; Triazines

Glutamate promotes neuronal survival during brain development and destroys neurons after injuries in the mature brain. Glutamate antagonists are in human clinical trials aiming to demonstrate limitation of neuronal injury after head trauma, which consists of both rapid and slowly progressing neurodegeneration. Furthermore, glutamate antagonists are considered for neuroprotection in chronic neurodegenerative disorders with slowly progressing cell death only. Therefore, humans suffering from Huntington's disease, characterized by slowly progressing neurodegeneration of the basal ganglia, are subjected to trials with glutamate antagonists. Here we demonstrate that progressive neurodegeneration in the basal ganglia induced by the mitochondrial toxin 3-nitropropionate or in the hippocampus by traumatic brain injury is enhanced by N-methyl-d-aspartate antagonists but ameliorated by alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionate antagonists. These observations reveal that N-methyl-d-aspartate antagonists may increase neurodestruction in mature brain undergoing slowly progressing neurodegeneration, whereas blockade of the action of glutamate at alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionate receptors may be neuroprotective.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Brain Injuries; Cell Death; Dizocilpine Maleate; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Memantine; N-Methylaspartate; Neurons; Neuroprotective Agents; Neurotoxins; Nitro Compounds; Piperazines; Propionates; Quinoxalines; Rats; Rats, Wistar; Wounds and Injuries

Neuronal necrosis and apoptosis occur after traumatic brain injury (TBI) in animals and contribute to subsequent neurological deficits. In contrast, relatively little apoptosis is found after mechanical injury in vitro. Because in vivo trauma models and clinical head injury have associated cerebral ischemia and/or metabolic impairment, we transiently impaired cellular metabolism after mechanical trauma of neuronal-glial cultures by combining 3-nitropropionic acid treatment with concurrent glucose deprivation. This produced greater neuronal cell death than mechanical trauma alone. Such injury was attenuated by the NMDA receptor antagonist dizocilpine (MK801). In addition, this injury significantly increased the number of apoptotic cells over that accruing from mechanical injury alone. This apoptotic cell death was accompanied by DNA fragmentation, attenuated by cycloheximide, and associated with an increase in caspase-3-like but not caspase-1-like activity. Cell death was reduced by the pan-caspase inhibitor BAF or the caspase-3 selective inhibitor z-DEVD-fmk, whereas the caspase-1 selective inhibitor z-YVAD-fmk had no effect; z-DEVD-fmk also reduced the number of apoptotic cells after combined injury. Moreover, cotreatment with MK801 and BAF resulted in greater neuroprotection than either drug alone. Thus, in vitro trauma with concurrent metabolic inhibition parallels in vivo TBI, showing both NMDA-sensitive necrosis and caspase-3-dependent apoptosis.

Topics: Animals; Apoptosis; Brain Injuries; Caspase 3; Caspase Inhibitors; Caspases; Cell Death; Cells, Cultured; Cerebral Cortex; Coculture Techniques; Dizocilpine Maleate; Hypoglycemia; Necrosis; Neuroglia; Neurons; Nitro Compounds; Oligopeptides; Propionates; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Succinate Dehydrogenase

3-Nitropropionic acid (NPA), an inhibitor of succinate dehydrogenase, is dietary neurotoxin. It is not known if neurons and astrocytes differ in their vulnerability to NPA, therefore, we investigated its toxicity in primary cultures of cerebellar granule cells and astrocytes. NPA inhibited succinate dehydrogenase and tricarboxylic acid cycle activity to the same degree in neurons and astrocytes. Even so NPA acid was 16 times more toxic to neurons than to astrocytes (LC50: 0.7 and 11 mM, respectively). The neurotoxicity of NPA was mediated by NMDA-receptor activation, calcium influx, and formation of reactive oxygen species, as revealed by the protective effect of NMDA-receptor blockade, the accumulation of 45Ca, and the protective effect of N-t-butyl-alpha-phenylnitron (PBN), a scavenger of reactive oxygen species. Cytotoxic concentrations of NPA caused a reduction in the intracellular level of glutathione, which probably contributed to the oxidative damage in both neurons and astrocytes. The relative resistance of astrocytes to NPA appeared to be related to their low tricarboxylic acid cycle activity (5%-10% of that in neurons) and to the inability of NPA to cause astrocytic calcium overload. We conclude that NPA acid predominantly is an astrocyte-sparing neurotoxin.

Topics: Amino Acids; Animals; Astrocytes; Calcium; Cell Survival; Citric Acid Cycle; Cyclic N-Oxides; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Free Radical Scavengers; Glucose; Male; Nervous System Diseases; Neurons; Neurotoxins; Nitro Compounds; Nitrogen Oxides; Propionates; Rats; Rats, Wistar; Succinate Dehydrogenase

Fluoro-Jade is an anionic fluorochrome capable of selectively staining degenerating neurons in brain slices. The histochemical application of Fluoro-Jade results in a simple, sensitive and reliable method for staining degenerating neurons and their processes. The technique will detect neuronal degeneration resulting from exposure to a variety of neurotoxic insults. Fluoro-Jade can be combined with other fluorescent methodologies including immunofluorescence, fluorescent axonal tract tracing, and fluorescent Nissl counterstaining. Compared to conventional methodologies, Fluoro-Jade is a more sensitive and definitive marker of neuronal degeneration than hematoxylin and eosin (H&E) or Nissl type stains, while being comparably sensitive yet considerably simpler and more reliable than suppressed silver techniques.

Topics: 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine; Animals; Antihypertensive Agents; Axons; Dizocilpine Maleate; Dopamine Agents; Eosine Yellowish-(YS); Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Eye Enucleation; Fluorescent Antibody Technique; Fluorescent Dyes; Hallucinogens; Hematoxylin; Ibogaine; Kainic Acid; Male; Metals; Nerve Degeneration; Nissl Bodies; Nitro Compounds; Phencyclidine; Propionates; Rats; Rats, Sprague-Dawley; Sensitivity and Specificity; Silver Staining; Stereotaxic Techniques

Neuronal death in neurodegenerative diseases may involve energy impairment leading to secondary excitotoxicity, and free radical generation. Potential therapies for the treatment of neurodegenerative diseases therefore include glutamate release blockers, excitatory amino acid receptor antagonists, agents that improve mitochondrial function, and free radical scavengers. In the present study we examined whether these strategies either alone or in combination had neuroprotective effects against striatal lesions produced by mitochondrial toxins. The glutamate release blockers lamotrigine and BW1003C87 significantly attenuated lesions produced by intrastriatal administration of 1-methyl-4-phenylpyridinium. Lamotrigine significantly attenuated lesions produced by systemic administration of 3-nitropropionic acid. Memantine, an N-methyl-D-aspartate antagonist, protected against malonate induced striatal lesions. We previously found that coenzyme Q10 and nicotinamide, and the free radical spin trap n-tert-butyl-alpha-(2-sulfophenyl)-nitrone (S-PBN) dose-dependently protect against lesions produced by intrastriatal injection of malonate. In the present study we found that the combination of MK-801 (dizocipiline) with coenzyme Q10 exerted additive neuroprotective effects against malonate. Lamotrigine with coenzyme Q10 was more effective than coenzyme Q10 alone. The combination of nicotinamide with S-PBN was more effective than nicotinamide alone. These results provide further evidence that glutamate release inhibitors and N-acetyl-D-aspartate antagonists can protect against secondary excitotoxic lesions in vivo. Furthermore, they show that combinations of agents which act at sequential steps in the neurodegenerative process can produce additive neuroprotective effects. These findings suggest that combinations of therapies to improve mitochondrial function, to block excitotoxicity and to scavenge free radicals may be useful in treating neurodegenerative diseases.

Topics: 1-Methyl-4-phenylpyridinium; Animals; Anticonvulsants; Coenzymes; Cyclic N-Oxides; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Free Radicals; Lamotrigine; Male; Malonates; Memantine; Mitochondria; Nervous System Diseases; Neuroprotective Agents; Neurotoxins; Niacinamide; Nitro Compounds; Nitrogen Oxides; Propionates; Pyrimidines; Rats; Rats, Sprague-Dawley; Spin Labels; Thallium; Triazines; Ubiquinone

3-Nitropropionic acid (3-NPA) is a metabolic poison that produces lesions of striatal intrinsic neurones such as gamma-aminobutyric acid (GABA) neurones. This study was carried out to determine whether 3-NPA would impair the ability of striatal GABAergic neurones to withstand hypoglycaemic stress. 3-NPA (500 microM) did not affect [3H]GABA release from striatal slices under normal (11 mM) glucose concentrations. When the glucose concentration was lowered to 0.3 mM, however, 3-NPA greatly potentiated the leakage of [3H]GABA from the slices. Blockage of N-methyl-D-aspartate (NMDA) or alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors with 1 microM 5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine (MK-801) or 10 microM 2,3-dihydroxy-6-nitro-7-sulpha-moylbenzo[F]quinoxaline (NBQX), respectively, or a combination of both, had no effect. However, blockade of voltage-dependent sodium channels with tetrodotoxin totally antagonized the [3H]GABA overflow induced by the combination of 3-NPA and hypoglycaemis. Riluzole (10 to 100 microM), a neuroprotective agent that stabilizes the inactivated state of the voltage-dependent sodium channel, also dose-dependently antagonized the increase in [3H]GABA release induced by the combination of the two stresses.

Topics: Animals; Corpus Striatum; Dizocilpine Maleate; Dose-Response Relationship, Drug; Excitatory Amino Acid Antagonists; gamma-Aminobutyric Acid; Glucose; Hypoglycemia; Isotope Labeling; Male; Neurons; Neuroprotective Agents; Neurotoxins; Nitro Compounds; Propionates; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Regression Analysis; Sodium Channels; Tritium

Exposure of mesencephalic dopamine neurons to an irreversible inhibitor of succinate dehydrogenase (SDH), 3-nitropropionic acid (3-NPA), for 24 h on day 12 in vitro, produced a dose-dependent loss of high-affinity dopamine uptake when measured 48 h following 3-NPA removal. ATP concentrations in the cultures were reduced by 57% after 3 h of treatment with the highest concentration of 3-NPA tested (500 microM). To determine whether glutamate receptors mediated the dopamine toxicity by 3-NPA, cultures were examined for their sensitivity to excitatory amino acid-induced toxicity. Mesencephalic cultures exposed to either 100 microM NMDA or kainate, on day 12 for 24 h, showed complete loss of dopamine uptake following 48 h of recovery. The NMDA and non-NMDA antagonists, MK-801 (1 microM) or 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX; 15 microM), completely prevented the effects of NMDA or kainate, respectively, when present at the time of toxin exposure. In cultures treated with 3-NPA, MK-801, but not CNQX, significantly attenuated the loss of dopamine uptake. Direct measurement of the effect of 3-NPA on SDH activity showed that 3-NPA dose-dependently inhibited SDH in vitro in a manner commensurate with the loss of dopamine uptake by 3-NPA. MK-801 had no effect on basal SDH activity or on 3-NPA inhibition of SDH. These data are consistent with the interpretation that metabolic inhibition in dopamine neurons can trigger a secondary excitotoxicity that is mediated by NMDA receptors.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; Adenosine Triphosphate; Animals; Antihypertensive Agents; Cells, Cultured; Dizocilpine Maleate; Dopamine; Mesencephalon; N-Methylaspartate; Neurons; Nitro Compounds; Propionates; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Succinate Dehydrogenase

We examined the effects of chronic systemic administration of the mitochondrial toxin 3-nitropropionic acid (3-NP) in doses ranging from 12 to 16 mg/kg/day for 30 days on striatal cytoarchitecture in rats. Administration of 3-NP at a dose of 16 mg/kg/day resulted in large lesions with a central necrotic core that was depleted of both neurons and glia. Glial fibrillary acidic protein (GFAP) gene expression was decreased in the lesion core, whereas the tissue surrounding this area showed a massive increase in signal intensity. Enkephalin and substance P mRNA expression in the striatum showed dose-dependent decreases following administration of 3-NP. A substantial decrease occurred even in animals treated with 3-NP at a dose of 12 mg/kg/day, in which there was little discernible neuronal loss and no increase in GFAP gene expression. In contrast to the decrease in enkephalin and substance P mRNA expression, somatostatin mRNA-expressing neurons were largely preserved. There was no preferential loss of [3H]naloxone patches in the rat striatum following chronic administration of 3-NP. In animals treated with 12-15 mg/kg/day neither the area nor binding density of the patches was changed. To study the effect of 3-NP on N-methyl-D-aspartate (NMDA)-gated Ca2+ channels we used in vivo administration of [3H]MK-801. Three hours after a single injection of 3-NP at a dose of 30 mg/kg there was a three- to fivefold increase in [3H]MK-801 binding in cortex and striatum as compared with saline-treated animals, consistent with an activation of NMDA receptors.

Topics: Animals; Antihypertensive Agents; Base Sequence; Corpus Striatum; Dizocilpine Maleate; DNA; Dose-Response Relationship, Drug; Enkephalins; Gene Expression; Glial Fibrillary Acidic Protein; Histocytochemistry; In Situ Hybridization; Ligands; Male; Molecular Sequence Data; Naloxone; Neurons; Nitro Compounds; Propionates; Rats; Rats, Sprague-Dawley; RNA, Messenger; Somatostatin; Substance P; Tritium

Succinic dehydrogenase in mouse cortical explant cultures was inhibited by 3-nitropropionic acid (3-NPA). ATP concentrations declined upon application of 3-NPA. At 4 h, ATP levels of cultures treated with 3-NPA alone were no different from those in cultures treated additionally with MK-801 (20 microM), 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX; 10 microM) or a combination thereof. However, MK-801 and MK-801 plus CNQX mitigated morphological lesions caused by 3-NPA. CNQX alone did not influence the extent of morphological damage. In conclusion, MK-801, at concentrations which were neuroprotective against 3-NPA lesions in cortical explant cultures, did not modify 3-NPA dependent decreases in cellular ATP levels. These data indicate that the neuroprotective effects of glutamate receptor antagonists in this model are probably receptor mediated and do not involve effects on cellular metabolism.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; Adenosine Triphosphate; Animals; Culture Techniques; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Mice; Nerve Degeneration; Neurotoxins; Nitro Compounds; Oxidative Phosphorylation; Plant Extracts; Propionates

The ability of N-methyl-D-aspartate (NMDA) receptor agonists and antagonists to modify 3-nitropropionic acid toxicity was studied in cultured rat cerebellar granule neurons. Exposure of these neurons to 3-nitropropionic acid resulted in a concentration and time-dependent neurotoxicity. In contrast to glutamate toxicity, 3-nitropropionic acid toxicity was potentiated by preexposure to subtoxic concentrations of NMDA. Presumably, the 3-nitropropionic acid-induced energy depletion relieved the voltage-dependent Mg2+ block of the NMDA receptor and induced vulnerability to subtoxic concentrations of NMDA receptor agonists. MK-801 and 2-amino-5-phosphonovaleric acid (APV) delayed but did not prevent 3-nitropropionic acid toxicity, indicating that prolonged exposure to 3-nitropropionic acid ultimately resulted in histotoxic neuronal death. We conclude that there are at least two distinct mechanisms of 3-nitropropionic acid toxicity of cerebellar granule neurons, one indirectly involving NMDA receptor-mediated excitotoxicity and one that is NMDA receptor-independent.

Topics: 2-Amino-5-phosphonovalerate; Animals; Cells, Cultured; Cerebellum; Cytoplasmic Granules; Dizocilpine Maleate; Electrophysiology; Glutamates; Glutamic Acid; Neurons; Neurotoxins; Nitro Compounds; Propionates; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate