Elsevier

Brain Research

Volume 922, Issue 2, 20 December 2001, Pages 173-179
Brain Research

Research report
Selective mGluR5 receptor antagonist or agonist provides neuroprotection in a rat model of focal cerebral ischemia

https://doi.org/10.1016/S0006-8993(01)03062-1Get rights and content

Abstract

Activation of group I metabotropic glutamate receptors (mGluR) has been implicated in the pathophysiology of acute central nervous system injury. However, the relative roles of the two group I subtypes, mGluR1 or mGluR5, in such injury has not been well examined. We compared the effects of treatment with the newly developed, selective mGluR5 antagonist 2-methyl-6-phenylethynylpyridine (MPEP) and the selective mGluR5 agonist (R,S)-2-chloro-5-hydroxyphenylglycine (CHPG) in a rat intraluminal filament model of temporary middle cerebral artery occlusion (MCAo). Rats were administered MPEP or CHPG i.c.v. beginning 15 or 135 min after induction of ischemia for 2 h. Infarct size was measured after either 22 or 70 h of reperfusion, and neurological function was quantified at 2, 24, 48 and 72 h. Treatment with MPEP or CHPG at 15 min reduced 24 h infarct volume by 61 and 44%, respectively. The neuroprotective effects were dose dependent. Delaying MPEP treatment until 135 min eliminated the neuroprotective effects. In other studies, using early MPEP treatment (15 min) at optimal doses, infarct volume was reduced by 44% at 72 h and this was correlated with significant neurological recovery. These data suggest that both MPEP and CHPG are neuroprotective when administered after focal cerebral ischemia. In separate, recent studies we found that although MPEP does act as an mGluR5 antagonist and blocks agonist induced phosphoinositide hydrolysis, it also serves as a non-competitive NMDA antagonist; in contrast, other results indicate that CHPG mediated neuroprotection may reflect anti-apoptotic activity. Therefore, both types of compounds may prove to have therapeutic potential for the treatment of stroke.

Introduction

Glutamate, a major excitatory neurotransmitter in the mammalian central nervous system, interacts with both ionotropic glutamate receptors and metabotropic glutamate receptors (mGluR). A role for ionotropic glutamate receptors in neuronal cell death following cerebral ischemia is well established [8], [12]. Because such receptors play such a critical role in fast synaptic transmission, blockade of these receptors may be associated with substantial side effects. Recent evidence suggests that mGluR, which are G-protein coupled receptors, may provide an effective alternative approach for reducing glutamate mediated cell death. Furthermore, the fact that mGluRs are primarily localized in the CNS may serve to limit certain peripheral side effects [9], [20]. It should also be noted that blockade of mGluRs seems to have only a modest impact on fast excitatory transmission [7].

There are eight mGluR subtypes, which have been divided into three major groups on the basis of sequence homology, signal transduction pathways, and pharmacological sensitivities [20], [21]. Group I mGluR includes mGluR1 and mGluR5; activation of these receptors causes stimulation of phospholipase C, resulting in phosphoinositide (PI) hydrolysis and intracellular calcium mobilization [20], [21]. The role of group I mGluR in neurodegeneration remains controversial. Whereas antagonists of these receptors are consistently neuroprotective, agonists have been found to either amplify or attenuate neuronal cell death [16]. Previous studies have suggested that the neurotoxic effects of putative group I ligands may be modulated primarily by the mGluR1 [6], [14], [15], but until recently, selective subtype specific antagonists have not been available to address this issue. The potent and selective mGluR1 antagonists (S)-4-carboxypheylglycine (AIDA) reduces traumatic neuronal injury in vivo and in vitro, and attenuates the delayed degeneration of vulnerable neurons in gerbils subjected to transient global ischemia [6], [19]. In addition, the selective mGluR1a antagonists (+)-2-methyl-4-carboxyphenylglycine (LY367385) and 7-hydroxyiminoclopropan[b]chromen-1a-carboxylic acid ethyl ester (CPCCOEt) have recently been described as being neuroprotective [2], [6]. From these studies it may be concluded that activation of mGluR1a receptors contributes to glutamate neurotoxicity and postischemic cell death, conditions that largely reflect necrotic cell death. In contrast to mGluR1, mGluR5 activation may serve to attenuate apoptotic cell death [3], [4]. This conclusion was partly based on studies using antisense oligonucleotides directed at mGluR5 in cerebellar granule cells cultures subjected to low levels of potassium [4]. In addition, activation of mGluR5 protects cultured granule cells against apoptotic death [1], [4].

Recent development of more selective mGluR1 and mGluR5 agonists/antagonists has provided tools to further address such hypotheses. These include the mGluR5 agonist (R,S)-2-chloro-5-hydroxyphenylglycine (CHPG) and antagonist 6-methyl-2-(phenylethynyl)-pyridine (MPEP) [5], [7]. CHPG has been shown to activate only mGluR5, but not mGluR1, in transfected cells; in vitro it has been shown to reduce neuronal apoptosis [5]. MPEP has been described as a selective non-competitive mGluR5 antagonist with no appreciable agonist or antagonist activity at recombinant mGlu1b, group II or III mGluRs [7]. Furthermore, MPEP does not act at the extracellular glutamate binding site of mGluR5 receptors common to all known competitive mGluR antagonists. Rather it interacts with transmembrane domains III and VII of mGluR5 receptors [18], which makes MPEP less sensitive to the ambient concentration of glutamate. In the present study, we compared the effects of MPEP or CHPG treatment in a rat intraluminal filament model of temporary middle cerebral artery occlusion (MCAo).

Section snippets

Surgical procedures

Male Sprague–Dawley rats (260–300 g; Charles River Lab., Raleigh, VA, USA) were used in this study. Anesthesia was induced by 5% halothane and maintained at 2% halothane delivered in oxygen. Body temperature was maintained normothermic (37±1°C) throughout all surgical procedures by means of a homeothermic heating system (Harvard Apparatus, South Natick, MA, USA). Food and water were provided ad libitum before and after surgery, and the animals were individually housed under a 12-h light–dark

2-h MCAo in vehicle-treated rats

MCAo with 22 h reperfusion resulted in significant core infarction within the temporal/parietal cortex and underlying striatum of the ipsilateral (injured) hemisphere. Ischemic damage generally extended from the most rostral forebrain sections to the final caudal sections and was greatest in the area around the bregma (Fig. 1). Total and core infarct volumes averaged 303±17 and 199±14 mm3, respectively. At 2 h after MCAo, neurological function (10.0±0.0) and EEG activity were markedly reduced.

Discussion

The intraluminal filament model of MCAo used in this study produces ischemia through temporary occlusion of the MCA with subsequent reperfusion at controlled time points. We have shown that both the selective mGluR5 agonist (CHPG) and antagonist (MPEP) have neuroprotective effects in this model. However, the neuroprotective mechanisms are likely to be different. Our in vivo observations are supported by in vitro results, which also showed that treatment with CHPG protects against apoptotic cell

Acknowledgments

This study was supported by grants from the National Institutes of Health (RO1NS37313) and the Department of Defense (DAMD-17-99-2-9007).

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