Glutamate-induced Excitation Research Articles

Rescue of Fmr1KO phenotypes with mGluR5 inhibitors: MRZ-8456 versus AFQ-056

Metabotropic glutamate receptor 5 (mGluR5) is a drug target for central nervous system disorders such as fragile X syndrome that involve excessive glutamate-induced excitation. We tested the efficacy of a novel negative allosteric modulator of mGluR5 developed by Merz Pharmaceuticals, MRZ-8456, in comparison to MPEP and AFQ-056 (Novartis, a.k.a. mavoglurant) in both in vivo and in vitro assays in a mouse model of fragile X syndrome, Fmr1KO mice. The in vivo assays included susceptibility to audiogenic-induced seizures and pharmacokinetic measurements of drug availability. The in vitro assays included dose response assessments of biomarker expression and dendritic spine length and density in cultured primary neurons. Both MRZ-8456 and AFQ-056 attenuated wild running and audiogenic-induced seizures in Fmr1KO mice with similar pharmacokinetic profiles. Both drugs significantly reduced dendritic expression of amyloid-beta protein precursor (APP) and rescued the ratio of mature to immature dendritic spines. These findings demonstrate that MRZ-8456, a drug being developed for the treatment of motor complications of L-DOPA in Parkinson’s disease and which completed a phase I clinical trial, is effective in attenuating both well-established (seizures and dendritic spine maturity) and exploratory biomarker (APP expression) phenotypes in a mouse model of fragile X syndrome.

Indicaxanthin from Opuntia ficus-indica Crosses the Blood-Brain Barrier and Modulates Neuronal Bioelectric Activity in Rat Hippocampus at Dietary-Consistent Amounts.

Indicaxanthin is a bioactive and bioavailable betalain pigment from the Opuntia ficus-indica fruits. In this in vivo study, kinetic measurements showed that indicaxanthin is revealed in the rat brain within 1 h from oral administration of 2 μmol/kg, an amount compatible with a dietary consumption of cactus pear fruits in humans. A peak (20 ± 2.4 ng of indicaxanthin per whole brain) was measured after 2.5 h; thereafter the molecule disappeared with first order kinetics within 4 h. The potential of indicaxanthin to affect neural activities was in vivo investigated by a microiontophoretic approach. Indicaxanthin, administered in a range between 0.085 ng and 0.34 ng per neuron, dose-dependently modulated the rate of discharge of spontaneously active neurons of the hippocampus, with reduction of the discharge and related changes of latency and duration of the effect. Indicaxanthin (0.34 ng/neuron) showed inhibitory effects on glutamate-induced excitation, indicating activity at the level of glutamatergic synapses. A molecular target of indicaxanthin is suggested by in silico molecular modeling of indicaxanthin with N-methyl-D-aspartate receptor (NMDAR), the most represented of the glutamate receptor family in hippocampus. Therefore, at nutritionally compatible amounts indicaxanthin (i) crosses the rat BBB and accumulates in brain; (ii) can affect the bioelectric activity of hippocampal neurons locally treated with amounts comparable with those measured in the brain; and (iii) modulates glutamate-induced neuronal excitation. The potential of dietary indicaxanthin as a natural neuromodulatory agent deserves further mechanistic and neurophysiologic investigation.

Lithium distinguishes between growth and circumnutation and augments glutamate-induced excitation of Helianthus annuus seedlings

An effect of lithium (Li+) on growth, circumnutation, and glutamate-induced excitation in sunflower (Helianthus annuus L.) seedlings was investigated using time-lapse photography and extracellular electrical potential measurements. The seedlings were treated with a micro and millimolar concentration of lithium chloride (LiCl) both persistently in a hydroponic medium and acutely through Li+ injection. The length of hypocotyls, fresh weight of seedlings, intensity and period of circumnutation, and the number of action potentials (APs) after glutamate (Glu) injection were determined. It was found that the circumnutation intensity and period did not depend on hypocotyl length and fresh weight of seedlings. Under persistent Li+ treatment, the circumnutation intensity was constant at a concentration between 0.2 and 20 mM although the hypocotyls were significantly shorter in relation to the control, whereas at a concentration of 40 mM circumnutation intensity decreased without any changes in the hypocotyl length. Under persistent treatment with 0.5 and 20 mM Li+, the period of circumnutation was significantly prolonged. The number of APs in a Glu-induced series significantly increased in the seedlings exhibiting an Li+-induced decrease in circumnutation intensity (in 40 and 60 mM Li+). Additionally Li+ injection before Glu injection also augmented the series of APs in seedlings growing without Li+ in hydroponic medium. These Li+-sensitive responses demonstrated that circumnutation and growth are partly independent processes and reveal a relationship between circumnutation intensity and excitability in Helianthus annuus seedlings.

Vitamin C neuroprotection against dose-dependent glutamate-induced neurodegeneration in the postnatal brain.

Glutamate-induced excitotoxicity due to over-activation of glutamate receptors and associated energy depletion (phosphorylation and activation of AMPK) results in neuronal cell death in various neurological disorders. Restoration of energy balance during an excitotoxic insult is critical for neuronal survival. Ascorbic acid (vitamin C), an essential nutrient with well-known antioxidant potential, protects the brain from oxidative damage in various models of neurodegeneration. In this study, we reported the therapeutic efficacy of vitamin C in response to glutamate-induced excitation, resulting in energy depletion and apoptosis in the hippocampus of the developing rat brain. A single subcutaneous injection of glutamate at two different concentrations (5 and 10 mg/kg) in postnatal day 7 rat pups increased brain glutamate levels and increased the protein expression of neuronal apoptotic markers. Both doses of glutamate upregulated the ratio of pro-apoptotic Bax to anti-apoptotic Bcl-2, cytochrome-c release, caspase-3 activation and the expression of PARP-1. However, co-treatment of vitamin C (250 mg/kg) with glutamate decreased brain glutamate levels and reversed the changes induced by glutamate in the developing hippocampus. Interestingly, only a high dose of glutamate caused the phosphorylation and activation of AMPK and induced neuronal cell death, whereas a low dose of glutamate failed to mediate these effects. Vitamin C supplementation reduced the glutamate-induced phosphorylation of AMPK and attenuated neuronal cell death, as assessed morphologically by Fluoro Jade B in the hippocampal CA1 region of the developing brain. Taken together, our results indicated that glutamate in both concentrations is toxic to the immature rat brain, whereas vitamin C is pharmacologically effective against glutamate-induced neurodegeneration.

Dopamine and Glutamate in Huntington's Disease: A Balancing Act

Huntington's disease (HD) is caused by a CAG repeat expansion in exon 1 of the HD gene resulting in a long polyglutamine tract in the N-terminus of the protein huntingtin. Patients carrying the mutation display chorea in early stages followed by akinesia and sometimes dystonia in late stages. Other major symptoms include depression, anxiety, irritability or aggressive behavior, and apathy. Although many neuronal systems are affected, dysfunction and subsequent neurodegeneration in the basal ganglia and cortex are the most apparent pathologies. In HD, the primary hypothesis has been that there is an initial overactivity of glutamate neurotransmission that produces excitotoxicity followed by a series of complex changes that are different in the striatum and in the cortex. This review will focus on evidence for alterations in dopamine (DA)-glutamate interactions in HD, concentrating on the striatum and cortex. The most recent evidence points to decreases in DA and glutamate neurotransmission as the HD phenotype develops. However, there is some evidence for increased DA and glutamate functions that could be responsible for some of the early HD phenotype. Significant evidence indicates that glutamate and dopamine neurotransmission is affected in HD, compromising the fine balance in which DA modulates glutamate-induced excitation in the basal ganglia and cortex. Restoring the balance between glutamate and dopamine could be helpful to treat HD symptoms.

Long-Term Treatment and Relapse Prevention of Alcohol and Benzodiazepine Dependence with Lamotrigine

Back to table of contents Previous article Next article LETTERFull AccessLong-Term Treatment and Relapse Prevention of Alcohol and Benzodiazepine Dependence with LamotrigineZoran M. Pavlovic M.D.,Zoran M. Pavlovic M.D.Search for more papers by this author,Published Online:1 Apr 2010https://doi.org/10.1176/jnp.2010.22.2.247.e25AboutSectionsPDF/EPUB ToolsAdd to favoritesDownload CitationsTrack Citations ShareShare onFacebookTwitterLinked InEmail To the Editor: Glutamate is the primary excitatory neurotransmitter in the CNS. The N -methyl- d- aspartate (NMDA) glutamate receptor subtype is also inhibited by ethanol and may be one of the primary molecular targets for its action in the brain. 1 – 4 Chronic inhibition of NMDA receptor function by repeated ethanol exposure induces an up-regulation of various NMDA receptor subunits. 5 When chronic alcohol administration ceases, NMDA receptors remain upregulated, and the resulting excessive glutamate-induced excitation may contribute to the hyperexcitatory symptoms of alcohol withdrawal. 6 Lamotrigine is an anticonvulsant that inhibits voltage-gated Na+ and Ca2+ channels and thereby prevents the release of various neurotransmitters, including glutamate. 7 , 8 A previous report by Krupitsky et al. 9 showed beneficial effects of lamotrigine as an aid in detoxification and as a treatment for dependence in bipolar patients with dual diagnosis (comorbid alcohol dependence). 10 Herein I report what is to my knowledge the first case report on the use of lamotrigine in alcohol and benzodiazepine withdrawal and dependence in a nonbipolar patient. Case Report Mrs. N is a 45-year-old woman who was diagnosed with alcohol and benzodiazepine dependence according to DSM-IV criteria and had a 5-year history of alcohol and benzodiazepine addiction at the time she was referred to our clinic. She experienced nightly insomnia and, during the previous 2 weeks, persistent vomiting. She presented with severe withdrawal symptoms, manifested as constant nausea and vomiting, extensive sweating, tactile disturbances on her scalp in the form of “electric cap,” and severe tremor. Her Clinical Institute Withdrawal Assessment of Alcohol Scale, Revised (CIWA) 11 score was 38, and her Severity of Dependence score was 9 for alcohol (cutoff for dependence is 3) and 12 for benzodiazepines (cutoff 7). 12 Her alcohol craving score according to the Penn Alcohol craving scale was 25 (maximum score 30). 13She agreed to undergo diazepam withdrawal to be followed by continuous topiramate treatment, with the target dose of 300 mg/day. She underwent 5-day diazepam detoxification which led to marked improvement in nausea and vomiting and a CIWA score of 20, making her very motivated to start on the new drug. After 2 days on topiramate, 100 mg/day, she experienced intensive dizziness. Both systolic and diastolic pressure had decreased by 20 mm/Hg and her resting pulse was 40 bpm (versus her usual 80 bpm); she also complained of new bruises which were 2–3 cm in diameter and mainly on her lower body. As it was judged that all these new symptoms and signs might be induced by topiramate according to the manufacturer’s instructions on safety (bradycardia and drop in arterial pressure) 14 and bruises as problems in coagulation and effect of topiramate on platelet function, 15 it was decided to slowly taper it down and to initiate lamotrigine, which was uptitrated according to manufacturer’s instructions to a final dose of 200 mg/day. During the next 2 weeks of full dosage treatment, she reported moderate improvements, which correlated with a decrease on her CIWA score. The symptoms continuously diminished during the following weeks, and after 16 weeks of active treatment, during the last assessment, she was only mildly anxious, with moist palms and moderate tremor (CIWA score <10) and Severity of Dependence score of 2 for alcohol and 4 for benzodiazepines. She also reported complete abstinence during the aforementioned period, while her Penn Alcohol craving score decreased to 12. DiscussionIn conclusion, I would like to suggest that lamotrigine might be a reasonable choice of antiepileptic drug for the long-term treatment of alcohol and benzodiazepine dependence. However, in my view it should not be used as detoxification agent, owing to the need to titrate it to therapeutic levels over several weeks. My suggestion would be to simultaneously use benzodiazepines as detoxification agent, and start long-term lamotrigine treatment in order to reduce remaining withdrawal symptoms following detoxification and prevent symptoms of relapse and dependence. My data suggest 200 mg/day as a reasonable target dose. Placebo-controlled trials are needed to test this hypothesis.Private Practice, Belgrade, Serbia

When and why amino acids?

This article reviews especially the early history of glutamate and GABA as neurotransmitters in vertebrates. The proposal that some amino acids could mediate synaptic transmission in the CNS initially met with much resistance. Both GABA and its parent glutamate are abundant in the brain; but, unlike glutamate, GABA had no obvious metabolic function. By the late 1950s, the switch of interest from electrical to chemical transmission invigorated the search for central transmitters. Its identification with Factor I, a brain extract that inhibited crustacean muscle, focused interest on GABA as a possible inhibitory transmitter. In the first microiontophoretic tests, though GABA strongly inhibited spinal neurons, these effects were considered 'non-specific'. Strong excitation by glutamate (and other acidic amino acids) led to the same conclusion. However, their great potency and rapid actions on cortical neurons convinced other authors that these endogenous amino acids are probably synaptic transmitters. This was partly confirmed by showing that both IPSPs and GABA greatly increased Cl() conductance, their effects having similar reversal potentials. Many anticonvulsants proving to be GABA antagonists, by the 1970s GABA became widely accepted as a mediator of IPSPs. Progress was much slower for glutamate. Being generated on distant dendrites, EPSPs could not be easily compared with glutamate-induced excitation, and the search for specific antagonists was long hampered by the lack of blockers and the variety of glutamate receptors. These difficulties were gradually overcome by the application of powerful techniques, such as single channel recording, cloning receptors, as well as new pharmacological tools.

Beyond Monoamines: Glutamatergic Function in Mood Disorders

The monoamine theory has implicated abnormalities in serotonin and norepinephrine in the pathophysiology of major depression and bipolar illness and contributed greatly to our understanding of mood disorders and their treatment. Nevertheless, some limitations of this model still exist that require researchers and clinicians to seek further explanation and develop novel interventions that reach beyond the confines of the monoaminergic systems. Recent studies have provided strong evidence that glutamate and other amino acid neurotransmitters are involved in the pathophysiology and treatment of mood disorders. Studies employing in vivo magnetic resonance spectroscopy have revealed altered cortical glutamate levels in depressed subjects. Consistent with a model of excessive glutamate-induced excitation in mood disorders, several antiglutamatergic agents, such as riluzole and lamotrigine, have demonstrated potential antidepressant efficacy. Glial cell abnormalities commonly associated with mood disorders may at least partly account for the impairment in glutamate action since glial cells play a primary role in synaptic glutamate removal. A hypothetical model of altered glutamatergic function in mood disorders is proposed in conjunction with potential antidepressant mechanisms of antiglutamatergic agents. Further studies elucidating the role of the glutamatergic system in the pathophysiology of mood and anxiety disorders and studies exploring the efficacy and mechanism of action of antiglutamatergic agents in these disorders, are likely to provide new targets for the development of novel antidepressant agents.

Inosine, Calcium Channels, and Neuroprotection Against Ischemic Brain Injury

To the Editor: We read with great interest the article by Shen et al,1 dealing with neuroprotective effects of inosine against ischemic brain injury. The results of their study demonstrated that intracerebroventricular administration of inosine before middle cerebral artery occlusion in rats resulted in a higher level of locomotor activity and less cerebral infarction. In addition, they indicated that coadministration of selective A3 receptor antagonist MRS1191 significantly attenuated inosine-mediated protection. In the electrophysiological study, it was shown that inosine antagonized glutamate-induced excitation in cerebral cortical neurons. The authors proposed that inosine may inhibit glutamate postsynaptic responses and reduce cerebral infarction via the activation of A3 receptors. Several studies have shown the mechanisms for glutamate-induced excitation of neural cells. In a study we presented earlier, …

Nitric oxide modulates striatal neuronal activity via soluble guanylyl cyclase: An in vivo microiontophoretic study in rats

It is now well established that nitric oxide (NO) acts as a neuromodulator in the central nervous system. To assess the role of NO in modulating striatal activity, single-unit recording was combined with iontophoresis to study presumed spiny projection neurons in urethane-anesthetized male rats. Striatal neurons recorded were essentially quiescent and were therefore activated to fire by the iontophoretic administration of glutamate, pulsed in cycles of 30 sec on and 40 sec off. In this study, iontophoresis of 3-morpholinosydnonimine hydrochloride (SIN 1), a nitric oxide donor, produced reproducible, current-dependent inhibition of glutamate-induced excitation in 12 of 15 striatal neurons, reaching its maximal inhibitory effect (76.2 +/- 5.6% below baseline) during the application of a 100 nA current. Conversely, microiontophoretic application of N-omega-nitro-L-arginine methyl ester (L-NAME), an inhibitor of nitric oxide synthase, produced clear and reproducible excitation of glutamate evoked firing in 7 of 10 cells (51.4 +/- 2.3%, at 100 nA). To evaluate the involvement of cyclic guanosine monophosphate (cGMP) in the electrophysiological effects produced by the NO donor, the effects of methylene blue, an inhibitor of guanylyl cyclase, on the responses of nine neurons to SIN 1 were tested. In six of nine neurons the effect of SIN 1 was significantly reduced during continuous iontophoretic administration (50 nA) of methylene blue. Taken together, these data show that NO modulates the striatal network and that inhibitory control of the output neurons is involved in this effect. These results also suggest that the effects of nitric oxide on striatal neurons are partially mediated via cGMP.

Local Application of Dopamine Inhibits Pyramidal Tract Neuron Activity in the Rodent Motor Cortex

Cortical neurons respond in a variety of ways to locally applied dopamine, perhaps because of the activation of different receptors within or among subpopulations of cells. This study was conducted to assess the effects of dopamine and the receptor subtypes that mediate the responses of a specific population of neurons, the pyramidal tract neurons (PTNs) in the rodent motor cortex. The specific subfamilies of dopamine receptors expressed by PTNs also were determined. PTNs were identified by antidromic stimulation in intact animals. Extracellular recordings of their spontaneous activity and glutamate-induced excitation were performed with multi-barrel pipettes to allow simultaneous recording and iontophoresis of several drugs. Prolonged (30 s) application of dopamine caused a progressive, nonlinear decrease in spontaneous firing rates for nearly all PTNs, with significant reductions from baseline spontaneous activity (71% of baseline levels) occurring between 20 and 30 s of iontophoresis. The D1 selective (SCH23390) and the D2 selective (eticlopride) antagonists were both effective in blocking dopamine-induced inhibition in nearly all PTNs. Mean firing levels were maintained within 3% of baseline levels during co-application of the D1 antagonist with dopamine and within 11% of baseline levels during co-application of the D2 antagonist and dopamine. SCH23390 was ineffective however, in 2 of 16 PTNs, and eticlopride was ineffective in 3 PTNs. The dopamine blockade by both antagonists in most neurons, along with the selective blockade by one, but not the other antagonist in a few neurons indicate that the overall population of PTNs exhibits a heterogeneous expression of dopamine receptors. The firing rate of PTNs was significantly enhanced by iontophoresis of glutamate (mean = 141% of baseline levels). These increases were attenuated significantly (mean= 98% of baseline) by co-application with dopamine in all PTNs, indicating dopaminergic interactions with glutamate transmission. The expression of dopamine receptors was studied with dual-labeling techniques. PTNs were identified by retrograde labeling with fast blue and the D1a, D2, or D5 receptor proteins were stained immunohistochemically. Some, but not all PTNs, showed labeling for D1a, D2, or D5 receptors. The D1a and D2 receptor immunoreactivity was observed primarily in the somata of PTNs, whereas D5 immunoreactivity extended well into the apical dendrites of PTNs. In accordance with findings of D1 and D2 receptor antagonism of dopamine's actions, the identification of three DA receptor subtypes on PTNs suggests that dopamine can directly modulate PTN activity through one or more receptor subtypes.

Involvement of glutamate in gastrointestinal vago-vagal reflexes initiated by gastrointestinal distention in the rat

Vago-vagal reflexes play an integral role in the regulation of gastrointestinal function. Although there have been a number of reports describing the effects of various stimuli on the firing rates of vagal afferent fibers and vagal motor neurons, little is known regarding the neurotransmitters that mediate the vago-vagal reflexes. In the present work, we investigated the role of glutamate in the vago-vagal reflex induced by gastrointestinal distention. Using single-cell recording techniques, we determined the effects of gastric and duodenal distention on the firing rates of gut-related neurons in the dorsal vagal complex, in the absence and presence of glutamate antagonists. Kynurenic acid, a competitive glutamate receptor antagonist, injected into the dorsal vagal complex, blocked the neuronal response of neurons in the dorsal motor nucleus of the vagus and the nucleus of the solitary tract to gastrointestinal distention. Injection of glutamate into the nucleus of the solitary tract produced inhibition of dorsal motor nucleus of the vagus neurons that were also inhibited by gastric and/or duodenal distention. Thus, the distention-induced inhibition of dorsal motor nucleus of the vagus neurons may be mediated by glutamate-induced excitation of gut-related nucleus of the solitary tract neurons. To investigate the role of the various glutamate receptor subtypes in the distention-induced events, we studied the effects of 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), a selective non-NMDA receptor antagonist, and dl-2-amino-5-phosphonopentanoic acid ( dl-AP5), a selective NMDA receptor antagonist. CNQX injected into the dorsal vagal complex either blocked or attenuated the inhibitory response of the neurons in the dorsal motor nucleus of the vagus and nucleus of the solitary tract neurons to gastric and duodenal distention. In contrast, dl-AP5 had less effect, especially in the vago-vagal reflex elicited by gastric distention. The results suggest (1) distention activates vagal afferents in the gastrointestinal tract; (2) the central branches of the vagal afferents from the gut terminate in the nucleus of the solitary tract and release glutamate that mainly act on non-NMDA receptors; (3) glutamate activates the inhibitory neurons in the nucleus of the solitary tract that project to the dorsal motor nucleus of the vagus; and (4) the inhibitory neurotransmitter suppresses the activity of the dorsal motor nucleus of the vagus neurons. For the excitatory neuronal responses of the dorsal motor nucleus of the vagus neurons to gastrointestinal distention, the possible circuit is that the vagal afferents containing glutamate directly activate the receptors on the dendrites of the dorsal motor nucleus of the vagus.

Influence of Serotonin on the Glutamate-Induced Excitations of Secondary Vestibular Neurons in the Rat

The excitatory responses evoked by glutamate and its agonists in secondary vestibular neurons of the rat were studied during microiontophoretic application of 5-hydroxytryptamine (5-HT). Ejection of 5-HT modified neuronal responsiveness to glutamate in 86% of the studied units, the effect being a depression of the excitatory responses in two-thirds of cases and an enhancement in the remaining third. 5-HT was also effective in modifying 94% of the responses evoked by N-methyl-d-aspartate (NMDA), inducing a depressive effect in 76% of cases and an enhancement in the remaining ones. Quisqualate-evoked effects were depressed and enhanced by 5-HT in about the same number of cases; in contrast, kainate-evoked responses were enhanced. The depressive action of 5-HT was mimicked by application of α-methyl-5-hydroxytryptamine (α-Me-5-HT), a 5-HT2 receptor agonist, whereas the enhancing effect could be evoked by application of 8-hydroxy-2(di-n-propylamino)tetralin (8-OH-DPAT), a selective 5-HT1A receptor agonist. The 5-HT2 receptor antagonist ketanserin was able to reduce, but not to block totally, the depressive action of 5-HT on glutamate- or NMDA-evoked responses. No significant difference was detected between neuronal responses in the lateral and the superior vestibular nucleus. These results indicate that 5-HT is able to modulate the responsiveness of secondary vestibular neurons to excitatory amino acids. Its action is mostly depressive, involves 5-HT2 receptors, and is exerted on NMDA receptors. A minor involvement of other 5-HT receptors (at least 5-HT1A) and other glutamate receptors (for quisqualate and kainate) in the modulatory action of 5-HT is plausible.

Glutamate-induced excitation and sensitization of nociceptors in rat glabrous skin

Anatomical studies demonstrate the presence of glutamate receptors on unmyelinated axons in peripheral cutaneous nerves. Pharmacological studies show that intraplantar injection of glutamate or glutamate agonists in the glabrous skin results in nociceptive behaviors. The present study describes a novel in vitro skin-nerve preparation using the glabrous skin from the rat hindpaw. In the first series of experiments, recordings were obtained from 141 fibers that responded to a strong mechanical search stimulus. Based on their conduction velocity they were classified as C (27%), Aδ (28%) and Aβ (45%) fibers. The C and Aδ fibers typically exhibited sustained firing during suprathreshold mechanical stimuli whereas both rapidly (66%) and slowly (34%) adapting responses were obtained from Aβ fibers. Noxious heat excited 46% of the C fibers but only 12% of the Aδ units. In another series of experiments application of an ascending series of glutamate concentrations (10, 100, 300, and 1000 μM) to Aδ ( n=14) and C ( n=19) nociceptors resulted in a significant excitation of 43% (6/14) Aδ fibers and 68% (13/19) C fibers. At these concentrations, there was no excitation of Aβ units ( n=13). Superfusion of the receptive fields of either mechanoheat-sensitive A (AMH, n=10) or C fibers (CMH, n=12) for 2 min with 300 μM glutamate resulted in sensitization of 90% (9/10) AMH and 92% (11/12) CMH fibers to subsequent thermal stimulation. This was evidenced by a significant (1) decrease in thermal threshold for activation, (2) increase in discharge rate, and (3) increase in peak instantaneous frequencies during the second heat trial. Glutamate-induced sensitization to heat occurred in the absence of either a glutamate-induced excitation or an initial heat response. Exposure of Aδ or C fibers to glutamate did not result in a decrease in von Frey thresholds. These data provide a physiological basis for the nociceptive behaviors that arise following intraplantar injection of glutamate or glutamate agonists. Furthermore, demonstration of glutamate-induced excitation and heat sensitization of nociceptors indicates that local or topical administration of glutamate receptor antagonists may have therapeutic potential for the treatment of pain.

Effects of intravenous cocaine administration on cerebellar Purkinje cell activity

The goal of the present study was to investigate the effects of intravenous cocaine administration on cerebellar Purkinje cell firing. Extracellular neuron activity was recorded and cells were locally excited with spaced microiontophoretic pulses of glutamate. Glutamate-evoked and spontaneous discharges were compared before and immediately following cocaine administration. Cocaine injections (1.0 and 0.25 mg/kg, i.v.) induced a reversible suppression of both spontaneous activity and glutamate-evoked excitation. Procaine was ineffective in producing similar actions. Cocaine only inhibited glutamate-induced excitation in animals pre-treated with reserpine (5 mg/kg, i.p.). Propranolol injections (10 mg/kg, i.p.) were ineffective in blocking cocaine-induced inhibitions. Yohimbine (5 mg/kg, i.p.) pre-treatment abolished cocaine-induced suppressions of either spontaneous or glutamate-evoked excitation. Therefore, cocaine administration decreases Purkinje cell spontaneous and glutamate-evoked discharges by a mechanism involving α 2-adrenoceptor activation. It is suggested that by changing the normal function of the cerebellum cocaine can produce drug-related alterations in overt behavior and cognition.

Ascorbate modulates glutamate-induced excitations of striatal neurons

To assess the role of ascorbate (AA), an antioxidant vitamin, in modulating striatal activity, single-unit recording was combined with iontophoresis in awake, unrestrained rats. Brief applications of AA (20 s, 5–80 nA) elicited few changes in either basal activity or activity evoked by continuous application of glutamate (GLU), but relatively high AA ejection currents (>40 nA) often inhibited fast-firing units. Comparable results were obtained with the antioxidant isomer, iso-AA, suggesting the AA-induced inhibition represents a high-dose, antioxidant effect. When applied for prolonged periods (2–4 min) at doses that failed to alter basal activity, AA either enhanced or attenuated the excitatory response to test pulses of GLU. The AA-induced enhancement occurred more frequently (16 vs. 6 applications) and was characterized by a more rapid (shorter onset and peak latencies) and more pronounced (greater peak magnitude) excitation to GLU without an evident change in offset latency. In most cases, further increases in AA ejection current attenuated the GLU response. Iso-AA, in contrast, had only inhibitory effects, which occurred at moderate- to high-dose applications. Collectively, these results suggest that AA, apart from its antioxidant effects, modulates phasic changes in striatal excitability induced by GLU. Because extracellular levels of striatal AA fluctuate in relation to behavioral activation, this neuromodulatory action of AA may contribute to behaviorally relevant changes in sensorimotor responsivity.

Heterogeneity of ventral tegmental area neurons: Single-unit recording and iontophoresis in awake, unrestrained rats

Single-unit recording combined with iontophoresis of dopamine, GABA, and glutamate was used in awake, unrestrained rats to characterize the electrophysiological and receptor properties of neurons in the ventral tegmental area under naturally occurring behavioural conditions. All isolated ventral tegmental area units ( n=90) were analysed and compared with cells ( n=58) recorded from dorsally adjacent areas of the pre-rubral area and red nucleus. Two distinct neuronal groups were identified in the ventral tegmental area: units with triphasic, long-duration spikes (78/90) and units with biphasic, short-duration spikes (12/90). Although all long-spike units discharged in an irregular, bursting pattern with varying degrees of within-burst decrements in spike amplitude, they could be further subdivided into at least three distinct subgroups. Type I long-spike units (36/78) discharged at a relatively slow and stable rate (mean: 6.03 imp/s; range: 0.42–15.78) with no evident fluctuations during movement. These cells were inhibited by dopamine and GABA and responded to glutamate with a low-magnitude excitation accompanied by a pronounced decrement in spike amplitude and a powerful rebound inhibition. Type II long-spike units (23/78) had relatively high and unstable discharge rates (mean: 22.82 imp/s; range: 4.42–59.67) and showed movement-related phasic activations frequently followed by partial or complete cessation of firing. Some Type II cells (4/9) were inhibited by dopamine, but all were excited by glutamate at very low currents (0–10 nA). With an increase in current, the glutamate-induced excitation often (18/22) progressed into a cessation of firing. All these cells were inhibited by GABA followed by a strong rebound excitation (8/9), which also frequently (6/8) resulted in cessation of firing. Type III long-spike units (19/78) had properties that differed from either Type I or Type II cells, including a lack of spontaneous firing (5/19). Short-spike ventral tegmental area units were either silent (4/12) and unresponsive to dopamine and GABA or spontaneously active (range: 0.89–34.13 imp/s) and inhibited by GABA and, in some cases (2/8), by dopamine; all were phasically activated during movement and glutamate iontophoresis. It appears that ventral tegmental area neurons, including those with long-duration spikes, do not comprise a uniform population in awake, unrestrained rats. Type I, long-spike units match the characteristics of histochemically-identified dopamine neurons, and they appear to express dopamine autoreceptors, which may explain the relatively slow, stable rate of activity and the limited responsiveness to excitatory inputs. Although the nature of the other long-spike units in our sample is unclear, they may include dopamine neurons without autoreceptors as well as non-dopamine cells. The heterogeneity of ventral tegmental area neurons is an important consideration for further attempts to assess the role of the mesocorticolimbic dopamine system in motivated behaviour.

Dopamine enhances glutamate-induced excitation of rat striatal neurons by cooperative activation of D1 and D2 class receptors

Although dopamine (DA) usually inhibits firing of rat striatal neurons in vivo, iontophoresis of DA with low ejection currents can also potentiate glutamate-evoked activity. We used extracellular single cell recording to examine the DA receptor subtypes involved in such potentiation. At low iontophoretic currents (1–8 nA), both the DA D1 class receptor agonist SKF 38393 and the DA D2 class receptor agonist quinpirole mimicked the ability of DA to facilitate glutamate-induced activity. Acute depletion of DA abolished the excitatory modulation produced by either D1 or D2 agonists but not by DA. Co-administration of SKF 38393 and quinpirole restored the facilitation of glutamate effects in DA-depleted rats. Stimulation of both D1 and D2 class receptors appears to be required for DA to enhance glutamate-induced firing of striatal neurons.

Glutamate receptor regulation of rat nucleus accumbens neurons in vivo.

Extracellular single cell recording and microiontophoretic techniques were used to characterize the roles of ionotropic and metabotropic glutamate receptors (iGluRs and mGluRs) in glutamate-induced excitation of rat nucleus accumbens (NAc) neurons in vivo. Pulse-ejected glutamate (16-128 nA) induced a current-dependent increase in the firing of quiescent NAc neurons. A stronger excitatory response to alpha-amino-3-hydroxy-5-methyl-4-isoxazole-proprionic acid (AMPA) was observed at much lower ejection currents (0.1-6.4 nA). Compared to AMPA and glutamate, N-methyl-D-aspartate (NMDA) induced a much less potent excitation in a narrow current range (1-4 nA) and only when neurons were previously "primed" with other excitatory amino acids (EAAs). Higher ejection currents of all three EAA agonists drove NAc neurons into a state of apparent depolarization block. AMPA-evoked firing was selectively blocked by the AMPA receptor antagonist 6,7-dinitroquinoxaline-2,3-dione (DNQX) whereas NMDA-induced activity was selectively prevented by the NMDA receptor antagonist 2-amino-5-phosphonovalerate (D-AP5). DNQX, but not D-AP5, significantly attenuated glutamate-evoked activity. The mGluR receptor agonist (1S,3R)-1-aminocyclopentane-1,3-dicarboxylic acid (1S,3R-t-ACPD) failed to evoke activity of NAc neurons, but significantly reduced the excitatory effects of other EAAs. This modulatory effect of 1S,3R-t-ACPD was consistently blocked by the selective mGluR antagonist L(+)-2-amino-3-phopsonopropionic acid (L-AP3) whereas another mGluR antagonist (RS)-4-carboxy-3-hydroxy phenylglycine (4C3HPG) was inconsistent in this regard. These results indicate that the excitatory effects of glutamate on rat NAc neurons in vivo are primarily mediated by non-NMDA iGluRs and that mGluRs function to dampen excessive glutamate transmission through iGluRs.

Electrophysiological and Pharmacological Characteristics of Ionotropic Glutamate Receptors in Medial Vestibular Nucleus Neurons: A Whole Cell Patch Clamp Study in Acutely Dissociated Neurons

A patch clamp study was performed to determine which subtype of ionotropic glutamate receptors is involved in the glutamate-induced excitation of the medial vestibular nucleus (MVN) neurons. Whole cell recording was performed on MVN neurons that were acutely dissociated by enzymatic and mechanical treatments. Application of glutamate at a concentration of 100 fiM produced a current with a reversal potential of approximately 0 mV. The glutamate-induced current was completely blocked by 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX, 10 μM), a non-N-methyl-D-aspartate (NMDA)-receptor antagonist. Application of α-amino-3-hydroxy-5-methyl-isoxazole-4-propionic acid (AMPA) and kainic acid (KA), non-NMDA-receptor agonists, at concentrations of 30 and 100 μM produced a concentration-dependent depolarization concomitantly with an increase in firing rates during current clamp recording. During voltage clamp recording, glutamate, AMPA and KA elicited a concentration-dependent current with an equilibrium potential of approximately 0 mV. To clarify whether NMDA receptors are present in MVN neurons, the effects of glycine on the glutamate- and NMDA-induced current were examined. Two types of NMDA receptor-mediated current (types 1 and 2) were obtained in terms of the difference in sensitivity to both magnesium ion and MK-801, which act on the NMDA-receptor channel. In the type 1 neurons, the NMDA-induced current was not apparently blocked by magnesium ion or MK-801, although a larger current was obtained in the absence of magnesium ion. In the type 2 neurons, marked blockade of the NMDA-induced current was seen in the presence of magnesium ion and MK-801, as previously reported in other neurons of the central nervous system. These findings indicate the presence of both non-NMDA and NMDA receptors, which are involved in primary afferent transmission, in the MVN neuron, and two distinct types of NMDA receptors.