mGluR7-like receptor and GABA_B receptor activation enhance neurotoxic effects of N-methyl-d-aspartate in cultured mouse striatal GABAergic neurones

Abstract

Presynaptic metabotropic glutamate receptors (mGluRs) of group III constitute possible targets for putative neuroprotective drugs acting against glutamate excitotoxic insults. Indeed, in glutamatergic cerebellar granule neurones in culture, high concentrations of l-2-amino-4-phosphonobutyrate (l-AP4, above 0.3 mM, thus activating mGluR7) inhibit NMDA-induced cell death. In contrast, in striatal cultures which are enriched in GABAergic neurones, we show that high concentrations of l-AP4 increased neuronal death in control as well as in NMDA-stimulated cultures. Moreover, similar results were obtained with the GABA_BR agonist, baclofen. Both the neuroprotective effects in cerebellar granule cells and the neurotoxic effects in striatal neurones were mediated via Gi-Go-coupled mGluRs, suggesting that these effects were probably mediated by mGluR7a or b and GABA_BR expressed in these neurones. In striatal neurones, we found that l-AP4 and baclofen inhibited both basal and NMDA-stimulated GABA release. These inhibitions of GABA release may be responsible for the increase in basal and NMDA-stimulated neuronal death. Indeed, blockade of GABA_A receptors with bicuculline increased neuronal death of control and NMDA-treated striatal cultures. Taken together, these results suggest that l-AP4 and baclofen, via mGluR7 and GABA_BR, reduced the neuroprotective effect of GABA present in striatal cultures acting via GABA_A receptors. Although caution must be taken when extrapolating from in vitro to in vivo situations, the present experiments and the recent observations that mGluR7 and GABA_BR are expressed in heterologous synapses, should be taken into consideration when evaluating the neuroprotective action of future mGluR7 specific agonists or GABA_BR specific antagonists.

Introduction

One of the classical roles of metabotropic receptors is the control of transmitter release at a number of central synapses (Conn and Pin, 1997). mGluRs generally decrease but in a few cases stimulate transmitter release (Herrero et al., 1992, Liou et al., 1996, Rodriguez-Moreno et al., 1998). The possible cellular mechanisms used by mGluRs to control transmitter release are multiple. They include inhibition of Ca²⁺ channels (Chavis et al., 1994, Takahashi et al., 1996), activation of presynaptic K⁺ channels (Saugstad et al., 1996), direct control of the synaptic release machinery (Gereau and Conn, 1995, Manzoni and Bockaert, 1997, Chavis et al., 1998) and alteration of Ca²⁺ release from presynaptic Ca²⁺ stores (Cochilla and Alford, 1998).

mGluR2-3 (group II) and mGluR4, 7 (group III) are mainly localised presynaptically. They reduce the strength of glutamatergic (Bradley et al., 1996, Shigemoto et al., 1996, Shigemoto et al., 1997, Conn and Pin, 1997) and possibly GABAergic synaptic transmission (Gereau and Conn, 1995, Conn and Pin, 1997, Kinoshita et al., 1998, Lujan et al., 1998), especially during high synaptic activity (Scanziani et al., 1997). Glutamate may spread from the release site and activate mGluR2-3 which are localised at the periphery of the synapse (Shigemoto et al., 1997). In addition, the role of mGluR7, another group III mGluR, is certainly original since it is localised in the presynaptic grid and is only activated by glutamate concentrations around 1 mM (Okamoto et al., 1994, Shigemoto et al., 1996). It is possible that mGluR7 receptors are activated only when very high, almost pathological glutamate release occurs for example during epileptic or ischemia episodes. mGluR7 activation will be the last opportunity for neurones to be protected from glutamate-induced excitotoxic death. Indeed, we have recently shown that mGluR7-like receptors are able to decrease NMDA receptor-induced glutamate release and cell death of cerebellar granule neurones (Lafon-Cazal et al., 1999).

Even more enigmatic is the role of mGluR7 which has been recently found in the grid of GABAergic terminals contacting either GABAergic or glutamatergic neurones (Lujan et al., 1998). Therefore, we were interested in finding out what would be the influence of such receptors on NMDA-induced cell death in GABAergic neurones.

Moreover, the results led us to consider the involvement of the recently cloned GABA_B receptor (Kaupmann et al., 1997). This metabotropic receptor was shown to present characteristics similar to those of mGluR7 concerning the presynaptic location on auto- as well as hetero-synapses and the role on the modulation of the neurotransmitter release (Bonanno et al., 1993, Waldmeier et al., 1994).

In this report, in a series of parallel experiments, we first compared the effects of l-AP4 (at a high concentration ≥0.3 mM) and baclofen, agonists of mGluR7 and GABA_B receptors respectively, on the NMDA-induced cell death in striatal and cerebellar granule neurones in culture. These two neuronal models are enriched in GABAergic and glutamatergic neurones, respectively. We then showed the expression of both metabotropic receptors, mGluR7 and GABA_BR in these two neuronal models, which were likely to be responsible for the present effects.