Calcium-dependent effects of melatonin inhibition of glutamatergic response in rat striatum

The effects of melatonin, amlodipine, diltiazem (L-type Ca2+ channel blockers) and omega-conotoxin (N-type Ca2+ channel blocker) on the glutamate-dependent excitatory response of striatal neurones to sensory-motor cortex stimulation was studied in a total of 111 neurones. Iontophoresis of melatonin produced a significant attenuation of the excitatory response in 85.2% of the neurones with a latency period of 2 min. Iontophoresis of either L- or N-type Ca2+ channel blocker also produced a significant attenuation of the excitatory response in more than 50% of the recorded neurones without significant latency. The simultaneous iontophoresis of melatonin + amlodipine or melatonin + diltiazem did not increase the attenuation produced by melatonin alone. However, the attenuation of the excitatory response was significantly higher after ejecting melatonin + omega-conotoxin than after ejecting melatonin alone. The melatonin-Ca2+ relationship was further supported by iontophoresis of the Ca2+ ionophore A-23187, which suppressed the inhibitory effect of either melatonin or Ca2+ antagonists. In addition, in synaptosomes prepared from rat striatum, melatonin produced a decrease in the Ca2+ influx measured by Fura-2AM fluorescence. Binding experiments with [3H]MK-801 in membrane preparations from rat striatum showed that melatonin did not compete with the MK-801 binding sites themselves although, in the presence of Mg2+, melatonin increased the affinity of MK-801. The results suggest that decreased Ca2+ influx is involved in the inhibitory effects of melatonin on the glutamatergic activity of rat striatum.     

Effects of NMDA receptor antagonists on cocaine-conditioned motor activity in rats

NMDA receptor antagonists have been reported to affect learned behaviors conditioned with abused drugs, with the outcome dependent, in part, on the class of NMDA receptor antagonist used. The present study tested the ability of various site-selective NMDA receptor antagonists to modify cocaine-conditioned motor activity. Two procedures were used for independently assessing drug effects on spontaneous activity and expression of cocaine-conditioned behavior. In the conditioning experiments, rats were administered i.p. injections of cocaine (30 mg/kg) or saline paired with distinctive environments. Spontaneous horizontal activity was dose-dependently enhanced by dizocilpine (0.03-0.3 mg/kg) and memantine (1-30 mg/kg), but not by D-CPPene (3-(2-carboxypiperazin-4-yl)-1-propenyl-1-phosphonic acid; SDZ EAA 494; 1-10 mg/kg), ACEA-1021 (5-nitro-6,7-dichloro-1,4-dihydro-2, 3-quinoxalinedione; 3-56 mg/kg), or eliprodil (3-30 mg/kg). Higher doses of memantine, D-CPPene (1-10 mg/kg), eliprodil (3-30 mg/kg), or ACEA-1021 reduced vertical activity. Following five cocaine-environment pairings, rats displayed significant increases in motor activity when exposed to the cocaine-paired environment. The following antagonists were administered prior to the conditioning test: dizocilpine (MK-801; 0.03-0.1 mg/kg), memantine (1-10 mg/kg), D-CPPene (0.3-3 mg/kg), ACEA-1021 (3-10 mg/kg), and eliprodil (1-10 mg/kg). Of these, memantine, ACEA-1021 and, to the lesser degree, eliprodil attenuated expression of cocaine-conditioned motor activity at doses that did not significantly affect spontaneous motor activity. These results show that cocaine-conditioned behaviors can be selectively modulated by some, but not all, NMDA receptor antagonists.     

Morphine and amphetamine sensitization in rats demonstrated under moderate- and high-dose NMDA receptor blockade with MK-801 (dizocilpine)

Rationale: It has been inferred from indirect tests that MK-801, an NMDA receptor antagonist, blocks sensitization to amphetamine and to morphine. These inferences were made from studies where behavioral scores were not recorded after each drug treatment in the sensitization protocol. Objectives: We reinvestigated the role of NMDA receptors in sensitization to amphetamine or morphine more directly by taking locomotor and stereotypy scores after each of several treatments with MK-801 and amphetamine or morphine. Methods: Each male Long Evans rat was administered intraperitoneal injections of MK-801 (0.1 or 0.25 mg/kg) or saline followed 30 minutes later by amphetamine (0.75 mg/kg), morphine (1.25 mg/kg) or saline and placed immediately in a photocell chamber. Locomotion and stereotypy were measured simultaneously by photobeam breaks and direct observation, respectively. This procedure was repeated on days 1, 2, 3, 4, 5, 8, 11 and 27 for rats receiving amphetamine or saline as the second injection and on days 1–10, 13, 16 and 32 for rats receiving morphine or saline as their second injection (with no testing or treatment on intervening days). Results: The animals treated in the amphetamine condition and animals treated in the morphine condition all showed progressively greater locomotion and stereotypy over the first 5 (amphetamine) or 10 (morphine) test days; the sensitized response was seen regardless of whether the animals were pretreated with saline or with MK-801. Thus MK-801 failed to block the development of psychomotor sensitization seen with these treatment regimens. When, following initial sensitization, amphetamine or morphine was given in the absence of MK-801 (days 8 and 13 for amphetamine and morphine rats, respectively), there was no expression of the sensitized response; the sensitized response of animals previously treated in the MK-801 drug state was expressed only when the animal was tested in the MK-801 drug state. The sensitized response was still expressed, in animals tested in the appropriate drug condition, after a 2-week period in which no drugs were given, confirming that the changes underlying this form of sensitization were long-lasting and thus probably a consequence of some form of synaptic plasticity. Conclusions: Our data provide evidence that behavioral sensitization to amphetamine and to morphine can occur despite the presence of NMDA receptor blockade. These and previous findings suggest that the failure of expression of sensitization seen when MK-801 is withdrawn from a given psychomotor stimulant treatment regimen reflects, at least in part, the dependency of sensitization on the various conditions of training rather than dependency on some essential function of NMDA receptor activation. Received: 14 October 1999 / Accepted: 7 March 2000     

NMDA receptor antagonists can enhance or impair learning performance in animals

Summary The effects of NMDA receptor antagonism on learning and memory were investigated using competitive (DL-2-amino-7-phosphonoheptanoate, AP7) and non-competitive (MK 801) blockers in three different learning tasks. Administration (i.p.) of drugs prior to training resulted in impaired learning performance in the place-navigation and dark-avoidance paradigms, and improved performance in the step-down passive avoidance task; however, using this treatment protocol, the possibility of druginduced non-mnemonic effects modifying learning performance could not be excluded. Drug administration immediately post-trial had no effect in the place-navigation paradigm, and improved retention performance in the dark-avoidance and step-down avoidance tasks. The similar results obtained with both types of antagonist indicate that the observed effects are indeed due to NMDA receptor blockade, and hence that such blockade modifies learning in a task-dependent manner. Exclusion of non-mnemonic effects by using the post-trial treatment regime demonstrates that NMDA antagonists facilitate learning of passive avoidance tasks.     

颈动脉注射辣椒素激活脑干心血管相关核团的儿茶酚胺神经元

在外周压力感受器去神经支配的大鼠上，用Ｆos蛋白和酪氨酸羟化酶（ＴＨ）的双重免疫组化方法，研究辣椒素的效应是否通过激活脑干核团内儿茶酚胺能神经元而诱发。结果显示，颈动脉注射辣椒素诱发脑干中最后区（ＡＰ）、孤束核（ＮＴＳ）、巨细胞旁外侧核（ＰＧＬ）和蓝斑（ＬＣ）等多个部位出现大量ＦOS样免疫反应（ＦＬＩ）神经元和双标神经元，辣椒素受体阻断剂钌红（ＲＲ）或ＮＭＤＡ受体阻断剂ＭＫ－８０１可明显减弱此效应。以上结果表明，辣椒素的兴奋效应通过激活儿茶酚胺能神经元而诱发，辣椒素受体和／（或）谷氨酸介导这一效应。     

MK-801 can facilitate passive avoidance memory when retention is not present in control animals,and can fail to facilitate when it is present

The results confirm that NMDA receptor blockade can result in improved retention performance of mice in step-down passive avoidance. A series of behavioural variations and analyses revealed that memory in the task depended mainly on the appearance of the grid that had been associated with shock, rather than on the execution of an instrumental response or on the spatial locus of the punishment. When the grid was made invisible during retest, retention was never found. However, MK-801 didnot facilitate retention based on the appearance of the grid. In contrast, conditions were found in which, even though control animals showed no learning, MK 801 given after the learning trial facilitated retention. Thus, the dominant mode of learning and memory in the step-down task is insensitive to the drug, whereas the drug raises a weaker or alternative mode to above threshold levels.     

Ischemic brain injury in vitro: protective effects of NMDA receptor antagonists and calmidazolium

Excessive Ca2+ influx through NMDA receptor-coupled channels has been linked to neuronal cell death. Using an in vitro model of transient brain ischemia, we investigated possible protective effects of NMDA receptor antagonists ketamine or MK-801 and of calmidazolium, an inhibitor of intracellular Ca2(+)-activated proteins. Brain ischemia/recovery was simulated in isolated hippocampal slices and injury monitored by measurement of ATP levels. Omission of both glucose and oxygen (but not oxygen alone) for 20 min led to persistent ATP deficits after 4 h recovery. Addition of ketamine or MK-801 at 1 microM permitted ATP to recover within 1 h, as did addition of calmidazolium at 10 microM. Our findings are consistent with other reports that NMDA receptor antagonists can protect neuronal tissue from ischemic damage. The role of inappropriately activated Ca2(+)-mediated signaling processes in the mechanism(s) of such injury is suggested by the protection also seen with calmidazolium, an inhibitor of calmodulin and other structurally related proteins such as calpain(s) and protein kinase C. The inhibition of intracellular Ca2+ target proteins may be an alternative for protection of the brain against injury due to insults that activate NMDA receptors.     

Neuroprotection by glutamate receptor antagonists against seizure-induced excitotoxic cell death in the aging brain

We previously have identified phenotypic differences in susceptibility to hippocampal seizure-induced cell death among two inbred strains of mice. We have also reported that the age-related increased susceptibility to the neurotoxic effects of seizure-induced injury is regulated in a strain-dependent manner. In the present study, we wanted to begin to determine the pharmacological mechanism that contributes to variability in the response to the neurotoxic effects of kainate. Thus, we compared the effects of the NMDA receptor antagonist, MK-801 and of the AMPA receptor antagonist NBQX on hippocampal damage in the kainate model of seizure-induced excitotoxic cell death in young, middle-aged, and aged C57BL/6 and FVB/N mice, when given 90 min following kainate-induced status epilepticus. Following kainate injections, mice were scored for seizure activity and brains from mice in each age and antagonist group were processed for light microscopic histopathologic evaluation 7 days following kainate administration to evaluate the severity of seizure-induced injury. Administration of MK-801 significantly reduced the extent of hippocampal damage in young, mature and aged FVB/N mice, while application of NBQX was only effective at attenuating cell death in young and aged mice throughout all hippocampal subfields. Our results suggest that both NMDA and non-NMDA receptors are involved in kainate-induced cell death in the mouse and suggest that aging may differentially affect the ability of neuroprotectants to protect against hippocampal damage. Differences in the effectiveness of these two antagonists could result from differential regulation of glutamatergic neurotransmitter systems or ion channel specificity.     

Indacrinone (MK-196) - a specific inhibitor of the voltage-dependent CI- permeability in toad skin

The effect of indacrinone (MK-196) on Cl^- transport through toad (Bufo bufo) skin epithelium was studied by the voltage clamping technique. At the transepithelial potential, V = 50 mV (serosal bath grounded) the unidirectional fluxes, governed by a Cl^- self-exchange diffusion pathway, were not affected by 1 mM racemic MK-196 in the outer bath. Likewise at V = 0 mV, the unidirectional fluxes as well as the active (net) inward flux of Cl^- were unaffected by MK-196. Voltage clamping the epithelium in the physiological range of potentials activated a Cl^- specific passive conductance that saturated for V ±– 90 mV. The influx and efflux of Cl^- through this pathway were inhibited by MK-196, and the (passive) Cl^- current was inhibited in a dose-dependent way for [MK-196] ± 50 μM with about 70% inhibition for [MK-196] = I mM. The maximum Cl^- conductance was decreased without shifting the position along the V-axis of the inverted S-shaped conductance–voltage relationship. The time constants for the voltage-stimulated Cl^- conductance activation were not affected by MK-196 (50 μM ± [MK-196] ± 1 mM). The (+) and (–) isomers and racemic MK-196 affected the voltage-dependent Cl^- conductance in similar ways. It is concluded that MK-196 has the properties of a Cl^- channel blocker which is specific for the voltage-dependent Cl^- permeability of the epithelium. The time course for development of inhibition exhibited a fast (min) and a slow (h) component. The fast component may reflect a direct interaction of MK-196 with an extracellular site of the Cl^- channel, whereas the slow one may reflect impairment of a metabolic pathway regulating the Cl^- permeability, or an interaction of MK-196 with a cytoplasmic site of the anion permeation pathway.