Stimulation of Endocannabinoid Formation in Brain Slice Cultures through Activation of Group I Metabotropic Glutamate Receptors

Kwang-Mook Jung,Janet Kim,Ken Mackie,Matthew Wallace,Darren Fegley,Regina Mangieri,Daniele Piomelli,Christopher Stapleton

doi:10.1124/mol.105.013961

Abstract

Activation of group I metabotropic glutamate (mGlu) receptors drives the endocannabinoid system to cause both short- and long-term changes of synaptic strength in the striatum, hippocampus, and other brain areas. Although there is strong electrophysiological evidence for a role of endocannabinoid release in mGlu receptor-dependent plasticity, the identity of the endocannabinoid transmitter mediating this phenomenon remains undefined. In this study, we show that activation of group I mGlu receptors triggers the biosynthesis of the endocannabinoid 2-arachidonoylglycerol (2-AG), but not anandamide, in primary cultures of corticostriatal and hippocampal slices prepared from early postnatal rat brain. Pharmacological studies suggest that 2-AG biosynthesis is initiated by activation of mGlu5 receptors, is catalyzed by phospholipase C (PLC) and 1,2-diacylglycerol lipase (DGL) activities, and is dependent on intracellular Ca2+ ions. Realtime polymerase chain reaction and immunostaining analyses indicate that DGL-beta is the predominant DGL isoform expressed in corticostriatal and hippocampal slices and that this enzyme is highly expressed in striatal neurons, where it is colocalized with PLC-beta1. The results suggest that 2-AG is a primary endocannabinoid mediator of mGlu receptor-dependent neuronal plasticity.

Highlights

MGlu receptor-dependent forms of synaptic modulation mediated by endocannabinoids have been implicated in prolonged changes in synaptic strength, ABBREVIATIONS: DSI, depolarization-induced suppression of inhibition; mGlu, metabotropic glutamate; 2-AG, 2-arachidonoylglycerol; PLC, phospholipase C; 1,2-DAG, 1,2-diacylglycerol; DGL, diacylglycerol lipase; DHPG, (S)-3,5-dihydroxyphenylglycine; LY367385 (S)-(ϩ)-␣-amino-amethylbenzeneacetic acid; BAPTA-AM, 1,2-bis(2-aminophenoxy)ethane-N,N,NЈ,NЈ-tetraacetic acid, acetoxymethyl ester; RHC80267, 1,6-bis(cyclohexyloximino carbonylamino)-hexane; U73122, 1- (6-[17␤-3-methoxyestra-1,3,5-(10)triene-17-yl]amino/hexyl) 1H-pyrroledione; HPLC, highperformance liquid chromatography; MS, mass spectrometry; 2-OG, 2-oleoylglycerol; PCR, polymerase chain reaction; MPEP, 2-methyl-6phenylethynyl-pyridine; 1,2-DAG, 1,2-diacylglycerol
We found that activation of mGlu5 receptors, which are expressed at high levels in both striatum and hippocampus (Testa et al, 1994), rapidly stimulates the biosynthesis of 2-AG, but not anandamide, suggesting that 2-AG plays a key role in mGlu5 receptor-initiated signaling events
The main finding of the present study is that activation of glutamate mGlu5 receptors stimulates 2-AG formation in rat corticostriatal and hippocampal slice cultures. mGlu5 receptors belong to the group I mGlu receptor subfamily, which includes the mGlu1 and mGlu5 subtypes (Conn and Pin, 1997). mGlu5 receptors are highly expressed in the striatum, nucleus accumbens, and hippocampus and more moderately in the neocortex (Testa et al, 1994)

Summary

Introduction

MGlu receptor-dependent forms of synaptic modulation mediated by endocannabinoids have been implicated in prolonged changes in synaptic strength, ABBREVIATIONS: DSI, depolarization-induced suppression of inhibition; mGlu, metabotropic glutamate; 2-AG, 2-arachidonoylglycerol; PLC, phospholipase C; 1,2-DAG, 1,2-diacylglycerol; DGL, diacylglycerol lipase; DHPG, (S)-3,5-dihydroxyphenylglycine; LY367385 (S)-(ϩ)-␣-amino-amethylbenzeneacetic acid; BAPTA-AM, 1,2-bis(2-aminophenoxy)ethane-N,N,NЈ,NЈ-tetraacetic acid, acetoxymethyl ester; RHC80267, 1,6-bis(cyclohexyloximino carbonylamino)-hexane; U73122, 1- (6-[17␤-3-methoxyestra-1,3,5-(10)triene-17-yl]amino/hexyl) 1H-pyrroledione; HPLC, highperformance liquid chromatography; MS, mass spectrometry; 2-OG, 2-oleoylglycerol; PCR, polymerase chain reaction; MPEP, 2-methyl-6phenylethynyl-pyridine; 1,2-DAG, 1,2-diacylglycerol Such as those occurring during the induction of long-term depression at corticostriatal synapses (Robbe et al, 2002; Gerdeman et al, 2003). 2-AG may be produced by PLC-␤–mediated cleavage of membrane phosphoinositides, which yields 1,2-diacylglycerol (1,2DAG), followed by diacylglycerol lipase (DGL)-catalyzed conversion of 1,2-DAG to 2-AG (Stella et al, 1997; Bisogno et al, 2003; Hashimotodani et al, 2005)

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