NMDA Receptor Channel Blocker Research Articles

Effects of extracellular matrix‐degrading proteases matrix metalloproteinases 3 and 9 on spatial learning and synaptic plasticity

Rats learning the Morris water maze exhibit hippocampal changes in synaptic morphology and physiology that manifest as altered synaptic efficacy. Learning requires structural changes in the synapse, and multiple cell adhesion molecules appear to participate. The activity of these cell adhesion molecules is, in large part, dependent on their interaction with the extracellular matrix (ECM). Given that matrix metalloproteinases (MMPs) are responsible for transient alterations in the ECM, we predicted that MMP function is critical for hippocampal-dependent learning. In support of this, it was observed that hippocampal MMP-3 and -9 increased transiently during water maze acquisition as assessed by western blotting and mRNA analysis. The ability of the NMDA receptor channel blocker MK801 to attenuate these changes indicated that the transient MMP changes were in large part dependent upon NMDA receptor activation. Furthermore, inhibition of MMP activity with MMP-3 and -9 antisense oligonucleotides and/or MMP inhibitor FN-439 altered long-term potentiation and prevented acquisition in the Morris water maze. The learning-dependent MMP alterations were shown to modify the stability of the actin-binding protein cortactin, which plays an essential role in regulating the dendritic cytoskeleton and synaptic efficiency. Together these results indicate that changes in MMP function are critical to synaptic plasticity and hippocampal-dependent learning.

Chronic hypoxia modulates NMDA-mediated regulation of the hypoxic ventilatory response in an amphibian, Bufo marinus

This study examined whether a hypoxia-tolerant amphibian, the Cane toad, undergoes mammalian-like ventilatory acclimatisation to hypoxia (VAH) and whether chronic hypoxia (CH) alters NMDA-mediated regulation of the acute hypoxic ventilatory response (HVR). Toads were exposed to 10 days of CH (10% O2) followed by acute hypoxic breathing trials or an intra-arterial injection of NaCN. Trials were conducted before and after i.p. treatment with an NMDA-receptor channel blocker (MK801). CH blunted the acute HVR but did not alter resting breathing. MK801 did not alter resting ventilation. In control animals, MK801 augmented breathing frequency (fR) during acute hypoxia by increasing the number of breaths per episode. This effect was attenuated following CH although MK801 did enhance the number of episodes per minute during acute hypoxia. MK801 enhanced the fR response to NaCN in both groups. The results indicate that CH did not produce mammalian-like VAH (i.e. increased resting ventilation and an augmented acute HVR) but did alter MK801-sensitive regulation of breathing pattern and the acute HVR.

Ethanol Exposure and Withdrawal Sensitizes the Rat Hippocampal CA1 Pyramidal Cell Region to ??-Amyloid (25-35)-Induced Cytotoxicity: NMDA Receptor Involvement

Millions of Americans suffer from Alzheimer's Disease (AD), which is characterized by significant neurological impairment and an accumulation in brain tissue of senile plaques consisting of beta amyloid (Abeta) peptide. The hippocampus, a region primarily responsible for learning and memory, appears to be particularly susceptible to AD-related injury and chronic alcohol abuse. Although certain risk factors for AD are known, it is unclear if alcohol abuse or dependence may contribute to neuropathology in AD. Recent research suggests that low-to-moderate consumption of alcohol may protect against development of AD, while alcohol dependence may increase risk of developing AD. Therefore, the current studies aimed to investigate the effects of exposure to 50 or 100 mM ethanol (EtOH) and withdrawal on hippocampal injury induced by Abeta peptide treatment. The present studies exposed organotypic hippocampal slice cultures to 50 or 100 mM ethanol (EtOH) for 10 days, after which the slices underwent ethanol withdrawal (EWD) in the presence of varying concentrations of Abeta 25-35 (0.1, 1, 10 microM), or 35-25 (200 microM), a negative control reverse sequence peptide. Cellular injury, as evidenced by uptake of propidium iodide (PI), was assessed for each subregion of the hippocampal complex (CA1, CA3, and dentate gyrus). Cellular injury in the CA1 pyramidal cell layer was significantly increased during withdrawal from exposure to 100 mM, but not 50 mM, EtOH. Exposure to Abeta in ethanol-naïve cultures did not produce significant cytotoxicity. However, exposure to Abeta during EWD from 100 mM produced marked increases in CA1 pyramidal cell region cytotoxicity, effects reversed by cotreatment with a nontoxic concentration of the NMDA receptor channel blocker MK-801 (20 microM). These data suggest that withdrawal from exposure to a high concentration of EtOH produces marked cellular injury in the hippocampus, particularly the CA1 subregion. Further, this EtOH exposure and withdrawal regimen sensitizes the hippocampus to the toxic effects of Abeta treatment in a manner reflecting over activity of NMDA receptor function.

Anti-allodynic interactions between NMDA receptor channel blockers and morphine or clonidine in neuropathic rats

Previous studies suggested that combining N-methyl-d-aspartate (NMDA) receptor antagonists with either μ-opioid agonist morphine or α2-adrenoreceptor agonist clonidine results in the significant synergistic enhancement of analgesic activity in the animal models of acute and neuropathic pain. When given alone, NMDA receptor antagonists, morphine and clonidine are capable of attenuating tactile allodynia associated with chronic nerve injury. The present study aimed to assess anti-allodynic effects of these compounds and to test additivity of these interactions using isobolographic analysis. Adult male Wistar rats with unilateral loose ligation of sciatic nerve developed significant tactile allodynia (between-paw difference of about 18–20 g). In separate groups of animals, dose-dependent anti-allodynic activity was confirmed for memantine (1.8–17.8 mg/kg), neramexane (1.8–17.8 mg/kg), morphine (1–10 mg/kg) and clonidine (0.01–0.1 mg/kg). In a subsequent series of experiments, memantine (or neramexane) and morphine (or clonidine) were co-administered at the fixed equi-effective dose ratios (six dose levels per drug combination). None of the tested combinations produced supra-additive, synergistic effects. In fact, memantine+clonidine, neramexane+clonidine and morphine+neramexane were producing simple additive effects, while morphine+memantine was characterized as the infra-additive combination. Thus, despite expectations based on previous studies, NMDA receptor channel blockers, memantine and neramexane, produce no synergistic interactions with either morphine or clonidine when administered acutely to rats with nerve injury-induced tactile allodynia.

Ketamine pre-treatment dissociates the effects of electroconvulsive stimulation on mossy fibre sprouting and cellular proliferation in the dentate gyrus

Electroconvulsive stimulation (ECS), the experimental analogue of electroconvulsive therapy (ECT), has been shown to produce both functional and structural effects in the hippocampal formation in infrahuman species. These changes may relate to the antidepressant and cognitive effects of ECT observed in patients treated for severe depressive disorders. Recent studies have described both enhanced neurogenesis in the dentate gyrus of the hippocampus and sprouting of mossy fibre projections from granule cells. The behavioural significance of these effects remains uncertain. In this study, we examined whether ketamine, a clinically available non-competitive NMDA receptor channel blocker, could block both of these “trophic” effects. Rats were given a course of eight spaced ECS or sham treatments under either halothane or ketamine anaesthesia. The thymidine analogue bromodeoxyuridine was administered to assess the degree of hippocampal cell proliferation and mossy fibre sprouting was quantified using the Timm staining method. Pre-treatment with ketamine dissociated these effects such that mossy fibre sprouting was attenuated significantly, while cell proliferation was unaffected. This dissociation may prove useful in determining the behavioural significance of these hippocampal changes, if any, for either the antidepressant or cognitive consequences of ECT.

Effect of agmatine on heteromeric N-methyl- d-aspartate receptor channels

Endogenous polyamines like spermine are known to have four distinct effects on recombinant N-methyl- d-aspartate (NMDA) receptor channels: voltage-dependent inhibition, glycine-dependent stimulation, glycine-independent stimulation and decreased affinity to the agonist ( l-glutamate). These effects are highly dependent on the constituting ɛ subunits (ɛ1–ɛ4) of the recombinant NMDA receptor channels. Agmatine reportedly inhibits native NMDA receptor channels in cultured hippocampal neurons. In the present investigation, the effects of agmatine on the ɛ/ζ heteromeric NMDA receptor channels expressed in Xenopus laevis oocytes were examined using the two-electrode voltage clamp method. Agmatine inhibited the four ɛ/ζ (ɛ1/ζ1, ɛ2/ζ1, ɛ3/ζ1 and ɛ4/ζ1) channels with similar sensitivity (an IC 50 value of about 300 μM at −70 mV). This effect was dependent on the membrane potential and was more pronounced at hyperpolarized membrane potentials (voltage-dependent inhibition). Agmatine did not exhibit other stimulatory (glycine-dependent and -independent effects) or inhibitory (decreased affinity to l-glutamate) effects. These properties are similar to the pharmacological profile of well-characterized NMDA receptor channel blockers like phencyclidine and ketamine. Thus, regarding the effects on the NMDA receptor channels, agmatine is not like other endogenous polyamines rather it acts as a channel blocker.

D‐Aspartate and NMDA, but not L‐aspartate, block AMPA receptors in rat hippocampal neurons

1 The amino acid, D-aspartate, exists in the mammalian brain and is an agonist at the N-methyl-D-aspartate (NMDA) subtype of ionotropic glutamate receptors. Here, for the first time, we studied the actions of D-aspartate on alpha-amino-3-hydroxyl-5-methyl-4-isoxazolepropionate receptors (AMPARs) in acutely isolated rat hippocampal neurons. 2 In the presence of the NMDA receptor channel blocker, MK801, D-aspartate inhibited kainate-induced AMPAR current in hippocampal neurons. The inhibitory action of D-aspartate on kainate-induced AMPAR current was concentration-dependent and was voltage-independent in the tested voltage range (-80 to +60 mV). 3 The estimated EC50 of the L-glutamate-induced AMPAR current was increased in the presence of D-aspartate, while the estimated maximum L-glutamate-induced AMPAR current was not changed. D-aspartate concentration-dependently shifted the dose-response curve of kainate to the right. Schild plot analysis indicated that D-aspartate acts competitively to block AMPARs. The K(b) for D-aspartate was estimated to be 0.93 mM. 4 D-aspartate also blocked L-glutamate-induced current in Xenopus laevis oocytes that expressed recombinant homomeric AMPARs. 5 NMDA possessed similar inhibitory action on AMPARs. However, L-aspartate had little inhibitory action on AMPARs. 6 D-Aspartate, but not L-aspartate, was found to reduce the amplitude of miniature excitatory postsynaptic current in cultured hippocampal neurons. 7 Our data are consistent with a model in which D-aspartate directly competes with kainate and L-glutamate in binding to the agonist binding site of AMPARs. The prevalence of D-aspartate in the brain suggests a possible role of D-aspartate in modulating AMPAR-mediated fast excitatory synaptic transmission.

Amantadine Inhibits NMDA Receptors by Accelerating Channel Closure during Channel Block

The channel of NMDA receptors is blocked by a wide variety of drugs. NMDA receptor channel blockers include drugs of abuse that induce psychotic behavior, such as phencyclidine, and drugs with wide therapeutic utility, such as amantadine and memantine. We describe here the molecular mechanism of amantadine inhibition. In contrast to most other described channel-blocking molecules, amantadine causes the channel gate of NMDA receptors to close more quickly. Our results confirm that amantadine binding inhibits current flow through NMDA receptor channels but show that its main inhibitory action at pharmaceutically relevant concentrations results from stabilization of closed states of the channel. The surprising variation in the clinical utility of NMDA channel blockers may in part derive from their diverse effects on channel gating.

The role of the NMDA receptor in alcohol relapse: a pharmacological mapping study using the alcohol deprivation effect

Modulators of glutamate receptors especially of the N-methyl- d-aspartate receptors (NMDARs) have recently been suggested as putative pharmacotherapeutic agents in the treatment of alcohol relapse. However, at present it is not clear, which binding and modulatory sites of the NMDAR are involved in relapse behavior. We, therefore, performed a pharmacological mapping study in long-term alcohol drinking rats using the alcohol deprivation effect (ADE) as a model for relapse behavior. In a comprehensive fashion, we studied dose–response curves, employing the following selective pharmacological agents: the NMDAR competitive antagonist CGP37849, the glycine binding site antagonist L-701.324, the NR2B subunit selective antagonist ifenprodil, which acts at the polyamine binding site, the NMDAR channel blocker neramexane, and ethanol, which acts as a functional antagonist at the NMDAR. Our data show that the animals' alcohol consumption inversely correlates with the dose of ethanol administered intraperitoneally. This indicates that under the present experimental conditions alcohol intake during an ADE is an entirely pharmacologically driven behavior that is not under the control of other factors such as taste or novelty of alcohol re-exposure. The effects of the administration of the aforementioned compounds were comparable to those of ethanol, suggesting a similar pharmacological impact on relapse behavior. Repeated administration of both competitive and uncompetitive NMDAR antagonist dose-dependently suppressed alcohol consumption during ADE. In addition, ifenprodil and L-701.324 dose-dependently reduced the expression of an ADE as well. In summary, the results suggest that an inhibition of NMDAR function in general, rather than a particular interference with a specific binding site of this receptor, is sufficient for the reduction of relapse behavior.

Effect of amantadine on motor memory consolidation in humans

Motor skills, once learned, need to be consolidated over time in order to become resistant to disruption or interference. In some instances, the consolidation phase can also include spontaneous gains in performance even in the absence of further rehearsal on a motor task. Clinical and behavioral evidence suggest that N-methyl-D-aspartate (NMDA)-receptor activity is required for motor learning acquisition and behavioral synaptic plasticity. However, the involvement of NMDA receptors in motor consolidation, leading to stabilization of the recently formed motor memory, has not yet been assessed in humans. To address this issue, we used post-training administration of amantadine, a low-affinity NMDA-receptor channel blocker. In a double-blind design, 200 mg of amantadine or a matching placebo was given orally to two different groups of 11 healthy young volunteers each. The subjects were tested twice 24 h apart, using a motor adaptation paradigm consisting of an eight-target-pointing task. Comparison of the mean performance levels on this task revealed that subjects in both groups improved their performance levels significantly on Day 2 compared to Day 1, regardless of the treatment administered. Our data indicate that amantadine failed to block motor learning consolidation in subjects that had already learned the motor adaptation task. Thus, although required in some stages (e.g. acquisition) of motor memory processes, the present results suggest that NMDA-receptor activation may not be essential for consolidation of motor adaptation in humans.

Drug discrimination analysis of NMDA receptor channel blockers as nicotinic receptor antagonists in rats

Antagonists acting at the N-methyl-D-aspartate (NMDA) subtype of glutamate receptors inhibit various phenomena associated with exposures to nicotine (e.g., tolerance, sensitization, dependence, and intravenous self-administration). These effects are often discussed in terms of nicotine-induced glutamate release with subsequent glutamate-dependent stimulation of dopamine metabolism and neuronal plasticity in brain areas critically involved in drug-addiction mechanisms. However, it is also well established that certain types of NMDA receptor antagonists (channel blockers) potently bind to nicotinic receptors and may act as nicotinic receptor antagonists. The present study aimed to evaluate the discriminative-stimulus effects of the NMDA receptor channel blockers (+)MK-801, dextromethorphan, and memantine in rats trained to discriminate nicotine from its vehicle. Adult male Wistar rats were trained to discriminate 0.6 mg/kg nicotine from saline under a two-lever, fixed-ratio 10 schedule of food reinforcement. During test sessions, injections of (+)MK-801 (0.03--0.3 mg/kg, i.p.), dextromethorphan (30 mg/kg, s.c.), or memantine (1--10 mg/kg, i.p.) were co-administered with s.c. nicotine (0.075--0.6 mg/kg; interaction tests) or saline (generalization tests). Additional interaction and generalization tests were conducted with the selective nicotinic receptor antagonists mecamylamine (0.1--3 mg/kg, s.c.) and MRZ 2/621 (0.3--10 mg/kg, i.p.), and the mGlu5 receptor antagonist MPEP (3--10 mg/kg, i.p.). In generalization tests, none of the compounds produced any appreciable levels of substitution for nicotine. The nicotine discriminative-stimulus control was dose dependently attenuated by mecamylamine (ED(50)=0.67 mg/kg) and MRZ 2/621 (ED(50)=9.7 mg/kg). Both agents produced a marked downward shift in the nicotine dose-response curve. Memantine and MPEP slightly attenuated nicotine discriminative-stimulus effects, while (+)MK-801 and dextromethorphan did not affect the nicotine-appropriate responding. NMDA receptor channel blockers, such as (+)MK-801, dextromethorphan, and memantine, have minimal interactions with the discriminative-stimulus effects of nicotine.

NR2 subunit‐dependence of NMDA receptor channel block by external Mg2+

The vital roles played by NMDA receptors in CNS physiology depend critically on powerful voltage-dependent channel block by external Mg(2+) (Mg(2+)(o)). NMDA receptor channel block by Mg(2+)(o) depends on receptor subunit composition: NR1/2A receptors (receptors composed of NR1 and NR2A subunits) and NR1/2B receptors are more strongly inhibited by Mg(2+)(o) than are NR1/2C or NR1/2D receptors. We investigated the effects of Mg(2+)(o) on single-channel and whole-cell currents recorded from recombinant NR1/2D and NR1/2A receptors expressed in HEK293 and 293T cells. The main conclusions are as follows: (1) Voltage-dependent inhibition by Mg(2+)(o) of whole-cell NR1/2D receptor responses was at least 4-fold weaker than inhibition of NR1/2A receptor responses at all voltages tested. (2) Channel block by Mg(2+)(o) reduced the duration of NR1/2D receptor single-channel openings; this reduction was used to estimate the apparent blocking rate of Mg(2+)(o) (k(+,app)). The k(+,app) for NR1/2D receptors was similar to but moderately slower than the k(+,app) obtained from cortical NMDA receptors composed of NR1, NR2A and NR2B subunits at all voltages tested. (3) Mg(2+)(o) blocking events induced an additional component in the closed-duration distribution; this component was used to estimate the apparent unblocking rate of Mg(2+)(o) (k(-,app)). The k(-,app) for NR1/2D receptors was much faster than the k(-,app) for cortical receptors at all voltages tested. The voltage-dependence of the k(-,app) of NR1/2D and cortical receptors differed in a manner that suggested that Mg(2+)(o) may permeate NR1/2D receptors more easily than cortical receptors. (4) Mg(2+)(o) inhibits NR1/2D receptors less effectively than cortical receptors chiefly because Mg(2+)(o) unbinds much more rapidly from NR1/2D receptors.

Cytotoxic Effects of Exposure to the Human Immunodeficiency Virus Type 1 Protein Tat in the Hippocampus Are Enhanced by Prior Ethanol Treatment

Long-term ethanol exposure leads to increases in the expression and/or sensitivity of NMDA-type glutamate receptors, effects that may contribute to the development of cytotoxicity in the brain. The human immunodeficiency virus 1 (HIV-1) transcription factor Tat is one of many viral proteins that may contribute to the development of HIV-associated dementia (HAD) by indirectly or directly promoting excess function of NMDA receptors. Thus, these studies examined the hypothesis that long-term ethanol pre-exposure would sensitize the hippocampus to Tat-induced cytotoxicity in an NMDA receptor-dependent manner. Organotypic slice cultures of rat hippocampus were exposed to a recombinant 86-amino acid form of Tat (Tat1-86) or a Tat deletion mutant devoid of amino acids 31 to 61 (TatDelta31-61; 0.1-100 nM) for 24 hr alone or during withdrawal from 10 days of ethanol exposure (50 mM in culture medium). Additional cultures were exposed to NMDA (5 microM) or the NMDA receptor channel blocker MK-801 (1 microM) during these treatments. Cellular injury in the CA1, CA3, and dentate gyrus regions of slice cultures was assessed by microscopy of propidium iodide fluorescence. Twenty-four hours of withdrawal from ethanol exposure did not produce overt cellular injury in any region of slice cultures. However, NMDA-induced toxicity was markedly increased in ethanol-pre-exposed cultures, an effect prevented by MK-801 (1 microM) coexposure. Treatment of cultures with Tat1-86 alone (> or = 0.1 nM) produced modest toxicity in each region of hippocampal cultures that was also blocked by MK-801 coexposure. In contrast, exposure to TatDelta31-61 did not alter propidium iodide fluorescence. Exposure of cultures to Tat1-86 (> or = 0.1 nM) during ethanol withdrawal resulted in a marked potentiation of Tat's toxic effects in each region of slice cultures, particularly the CA1 region. This potentiation of Tat neurotoxicity was significantly attenuated by coexposure of cultures to MK-801 (1 microM). These results indicate that long-term ethanol exposure sensitizes the hippocampus to the cytotoxic effects of Tat in an NMDA receptor-dependent manner. This may suggest that HIV-1-positive individuals who are alcohol dependent possess a heightened risk for the development of HAD. Furthermore, the NMDA receptor, particularly allosteric modulatory sites such as polyamine-sensitive sites, may be a therapeutic target to be investigated in the treatment of HAD.

Effects of nicotinic and NMDA receptor channel blockers on intravenous cocaine and nicotine self-administration in mice

Previous studies have indicated that blockade of N-methyl- d-aspartate (NMDA) subtype of glutamate receptors prevents acquisition of instrumental behaviors reinforced by food and drugs such as morphine and cocaine. The present study aimed to extend this evidence by testing whether NMDA receptor channel blocker, memantine, would exert similar effects on acquisition of cocaine and nicotine self-administration in mice. Inasmuch as memantine also acts as nicotinic receptor channel blocker, this study assessed the effects of mecamylamine and MRZ 2/621 that are more selective nicotinic blockers. Adult male Swiss mice were allowed to self-administer cocaine (0.8–2.4 μg/infusion) or nicotine (0.08–0.32 μg/infusion) during the 30-min test. Pretreatment with memantine (0.1–10 mg/kg) prevented acquisition of nicotine but not cocaine self-administration. Pretreatment with mecamylamine (0.3–3 mg/kg) and MRZ 2/621 (0.3–10 mg/kg) produced dose-dependent suppression of both cocaine and nicotine self-administration. Taken together with the previous reports, these results indicate that nicotinic receptor blockers antagonize acute reinforcing effects of cocaine while NMDA receptor blockade may have limited effectiveness.

Effects of low-affinity NMDA receptor channel blockers in two rat models of chronic pain

In contrast to conventional opioid analgesics, antagonists acting at the N-methyl- d-aspartate (NMDA) subtype of glutamate receptors are capable of suppressing pain-related phenomena in chronic pain models while having little or no effect on acute nociception. One of the few clinically used NMDA receptor antagonists, memantine, differs from prototypic antagonists with psychotomimetic activity such as phencyclidine and (+)MK-801, in showing lower receptor affinity, faster unblocking kinetics and stronger voltage-dependency. Recently, a series of novel amino-alkyl-cyclohexanes was reported to interact with NMDA receptors in a manner similar to that of memantine. The present study aimed to evaluate the effects of these compounds as well as (+)MK-801 and memantine in two rat models of chronic pain and the rotarod test. Unlike (+)MK-801 and memantine, most of the tested compounds were inactive against tactile allodynia induced by sciatic nerve ligation. On the other hand, all tested drugs were found to inhibit formalin-induced grooming behavior—a model of chronic pain induction. In agreement with previous reports on the effects of NMDA receptor antagonists in similar assays, the late phase seemed to be inhibited to a greater extent than the early phase. For all tested compounds, inhibition of formalin-induced behaviors occurred at dose levels that were also producing significant motor deficits (rotarod test). These results confirm low efficacy of acute administration of NMDA receptor antagonists in the models of established pain states. Thus, studies on the prevention and management of chronic pain should focus on preemptive or long-term administration of NMDA receptor antagonists.

Caffeine withdrawal syndrome in social interaction test in mice: Effects of the NMDA receptor channel blockers, memantine and neramexane

Antagonists acting at N-methyl-D-aspartate (NMDA) receptors have been demonstrated repeatedly to attenuate the expression of drug and alcohol withdrawal syndromes. The present study aimed to evaluate the effects of NMDA receptor blockade on the expression of behavioural signs of caffeine withdrawal syndrome, assessed using the social interaction paradigm. Adult male Swiss mice were treated with increasing doses of caffeine (40-100 mg/kg, i.p., twice daily) for 8 days. Twenty-four hours after the last injection of caffeine, there were significant increases in duration and frequency of defensive behaviours, as well as decreased locomotor activity. These changes faded within 72 hours. Pretreatment with a single dose of caffeine (1 mg/kg; 24 h after the end of repeated caffeine administration and 30 min prior to the test) completely reversed these withdrawal-related changes. Separate groups of mice were treated i.p. with different doses of memantine (1, 3 or 10 mg/kg) or neramexane (MRZ 2/579; 1, 3 or 10 mg/kg) 24 h after the last caffeine injection. Both compounds dose-dependently reduced the expression of defensive behaviours while increasing motor activity. These data suggest that NMDA receptor blockade may counteract the acute behavioural effects of caffeine withdrawal.

Effects of low-affinity NMDA receptor channel blockers in two rat models of chronic pain

In contrast to conventional opioid analgesics, antagonists acting at the N-methyl-d-aspartate (NMDA) subtype of glutamate receptors are capable of suppressing pain-related phenomena in chronic pain models while having little or no effect on acute nociception. One of the few clinically used NMDA receptor antagonists, memantine, differs from prototypic antagonists with psychotomimetic activity such as phencyclidine and (+)MK-801, in showing lower receptor affinity, faster unblocking kinetics and stronger voltage-dependency. Recently, a series of novel amino-alkyl-cyclohexanes was reported to interact with NMDA receptors in a manner similar to that of memantine. The present study aimed to evaluate the effects of these compounds as well as (+)MK-801 and memantine in two rat models of chronic pain and the rotarod test. Unlike (+)MK-801 and memantine, most of the tested compounds were inactive against tactile allodynia induced by sciatic nerve ligation. On the other hand, all tested drugs were found to inhibit formalin-induced grooming behavior—a model of chronic pain induction. In agreement with previous reports on the effects of NMDA receptor antagonists in similar assays, the late phase seemed to be inhibited to a greater extent than the early phase. For all tested compounds, inhibition of formalin-induced behaviors occurred at dose levels that were also producing significant motor deficits (rotarod test). These results confirm low efficacy of acute administration of NMDA receptor antagonists in the models of established pain states. Thus, studies on the prevention and management of chronic pain should focus on preemptive or long-term administration of NMDA receptor antagonists.

Positive N-methyl- d-aspartate receptor modulation by selective glycine transporter-1 inhibition in the rat dorsal spinal cord in vivo

In this study we have employed the selective glycine transporter-1 (GlyT-1) and GlyT-2 transporter inhibitors R-(−)- N-methyl- N-[3-[(4-trifluoromethyl)phenoxy]-3-phenyl-propyl]glycine (1:1) lithium salt (Org 24598) and 4-benzyloxy-3,5-dimethoxy- N-[1-(dimethylaminocyclopently)methyl]benzamide (Org 25543), respectively, and microdialysis perfusion to determine the effect of GlyT transporter inhibition on extracellular amino acid concentrations in the lumbar dorsal spinal cord of the halothane-anaesthetised rat. Reverse dialysis of Org 24598 (0.1–10 μM) induced a concentration-related increase in extracellular glycine accompanied by a progressive increase in citrulline, but not aspartate, glutamate or GABA, efflux. Org 25543 (10 μM) by the same route induced a similar increase in glycine levels without affecting the efflux of other amino acids quantified. To test the hypothesis that the increase in citrulline efflux resulted from activation of the N-methyl- d-aspartate receptor (NMDA-R)/nitric oxide synthase (NOS) signalling cascade, the sensitivity was determined of GlyT-1 inhibition-induced effects to NMDA-R antagonism or NOS inhibition. Co-administration by reverse dialysis of the selective NMDA-R channel blocker MK-801 (0.5 mM) or the selective antagonist of the strychnine-insensitive glycine site, 7-chlorokynurenic acid (1 mM), with Org 24598 (10 μM) did not affect the uptake inhibition-induced increase in glycine efflux, but did significantly attenuate the increase in extracellular citrulline. Similarly, co-administration with Org 24598 of the isoform non-selective and selective neuronal NOS inhibitors N ω-nitro- l-arginine methyl ester (1 mM) or 1-(2-trifluoromethylphenyl)imidazole (0.2 mM), respectively, prevented Org 24598-induced citrulline efflux with no effect on increased glycine efflux. These data provide evidence that the observed increased in extracellular citrulline is a consequence of positive modulation of NMDA-R, secondary to increased extracellular glycine and support a protective role for GlyT-1 against fluctuations in extracellular glycine uptake at glutamatergic synapses in the dorsal spinal cord. Such a mechanism could be important to NMDA-R-mediated synaptic plasticity in the spinal cord and be of relevance to the clinical usage of GlyT-1 inhibitors.

Pre- and postsynaptic contributions of voltage-dependent Ca2+ channels to nociceptive transmission in rat spinal lamina I neurons.

Activation of voltage-dependent Ca2+ channels (VDCCs) is critical for neurotransmitter release, neuronal excitability and postsynaptic Ca2+ signalling. Antagonists of VDCCs can be antinociceptive in different animal pain models. Neurons in lamina I of the spinal dorsal horn play a pivotal role in the processing of pain-related information, but the role of VDCCs to the activity-dependent Ca2+ increase in lamina I neurons and to the synaptic transmission between nociceptive afferents and second order neurons in lamina I is not known. This has now been investigated in a lumbar spinal cord slice preparation from young Sprague-Dawley rats. Microfluorometric Ca2+ measurements with fura-2 have been used to analyse the Ca2+ increase in lamina I neurons after depolarization of the cells, resulting in a distinct and transient increase of the cytosolic Ca2+ concentration. This Ca2+ peak was reduced by the T-type channel blocker, Ni2+, by the L-type channel blockers, nifedipine and verapamil, and by the N-type channel blocker, omega-conotoxin GVIA. The P/Q-type channel antagonist, omega-agatoxin TK, had no effect on postsynaptic [Ca2+]i. The NMDA receptor channel blocker D-AP5 reduced the Ca2+ peak, whereas the AMPA receptor channel blocker CNQX had no effect. Postsynaptic currents, monosynaptically evoked by electrical stimulation of the attached dorsal roots with C-fibre and Adelta-fibre intensity, respectively, were reduced by N-type channel blocker omega-conotoxin GVIA and to a much lesser extent, by P/Q-type channel antagonist omega-agatoxin TK, and the L-type channel blockers verapamil, respectively. No difference was found between unidentified neurons and neurons projecting to the periaqueductal grey matter. This is the first quantitative description of the relative contribution of voltage-dependent Ca2+ channels to the synaptic transmission in lamina I of the spinal dorsal horn, which is essential in the processing of pain-related information in the central nervous system.

Chronic exposure to NMDA receptor and sodium channel blockers during development in monkeys and rats: long-term effects on cognitive function.

The effects of chronic administration of MK-801 (NMDA-receptor antagonist) and remacemide (sodium channel blocker) on monkey learning of several brain function tasks was assessed in juveniles (nine months old). Low (LO) and high (HI) doses of both drugs were given orally each day for 18 months. There were no adverse effects of any treatment on tests of short-term memory or motivation. HI doses of both MK-801 and remacemide delayed acquisition of a visual discrimination task (the remacemide effect was much greater). HI doses of remacemide alone severely disrupted learning task acquisition and this effect lasted for several months after dosing. Thus, in monkeys, chronic blockade of NMDA receptors is relatively well tolerated, whereas blockade of sodium channels (perhaps in conjunction with NMDA receptor blockade) has long-term-perhaps permanent-consequences. To further explore the roles of NMDA receptors and sodium channels in these effects, MK-801, phenytoin (sodium channel blocker), or both were administered to rats and the acquisition of tasks similar to those used in the monkey study were assessed. Dosing began at weaning and continued for nine months. Throughout the study, HI MK-801 subjects exhibited impaired performance in all tasks. Some effects of MK-801 were blocked completely by phenytoin. In the rat, blockade of sodium channels was well tolerated but blockade of NMDA receptors had significant and long-term (permanent?) adverse consequences. These data contrast markedly with those obtained for the monkey and suggest, at least for some drug classes, that the rat might not be a good predictor of effects in primates.