GABAA α6-Containing Receptors Are Selectively Compromised in Cerebellar Granule Cells of the Ataxic Mouse, Stargazer

Helen L Payne,William M Connelly,Jane H Ives,Reinhard Lehner,Birgit Furtmuller,Werner Sieghart,Priyanka Tiwari,John M Lucocq,George Lees,Christopher L Thompson

doi:10.1074/jbc.m700111200

Abstract

Stargazer mice fail to express the gamma2 isoform of transmembrane alpha-amino-3-hydroxyl-5-methyl-4-isoxazolepropionate (AMPA) receptor regulatory proteins that has been shown to be absolutely required for the trafficking and synaptic targeting of excitatory AMPA receptors in adult murine cerebellar granule cells. Here we show that 30 +/- 6% fewer inhibitory gamma-aminobutyric acid, type A (GABA(A)), receptors were expressed in adult stargazer cerebellum compared with controls because of a specific loss of GABA(A) receptor expression in the cerebellar granule cell layer. Radioligand binding assays allied to in situ immunogold-EM analysis and furosemide-sensitive tonic current estimates revealed that expression of the extrasynaptic (alpha6betaxdelta) alpha6-containing GABA(A) receptor were markedly and selectively reduced in stargazer. These observations were compatible with a marked reduction in expression of GABA(A) receptor alpha6, delta (mature cerebellar granule cell-specific proteins), and beta3 subunit expression in stargazer. The subunit composition of the residual alpha6-containing GABA(A) receptors was unaffected by the stargazer mutation. However, we did find evidence of an approximately 4-fold up-regulation of alpha1betadelta receptors that may compensate for the loss of alpha6-containing GABA(A) receptors. PCR analysis identified a dramatic reduction in the steady-state level of alpha6 mRNA, compatible with alpha6 being the primary target of the stargazer mutation-mediated GABA(A) receptor abnormalities. We propose that some aspects of assembly, trafficking, targeting, and/or expression of extrasynaptic alpha6-containing GABA(A) receptors in cerebellar granule cells are selectively regulated by AMPA receptor-mediated signaling.

Highlights

The stargazer mutant mouse arose by virtue of a spontaneous viral transposon insertion into the stargazin gene [1]
TARP␥2 is reported to be heavily expressed in the cerebellum [2, 9] where it is largely restricted to the cerebellar granule cells (CGCs), neurons that normally exclusively express the TARP␥2 isoform of TARPs
We have revealed that it is the ␣6 subunit-containing GABAR subtypes that are selectively affected by the mutation, and these include the synaptic ␣6 subunit-containing GABAR subtypes (␣6␤x␥2 and ␣1␣6␤x␥2) and the extrasynaptic, tonic inhibition-conferring ␣6␤x␦ receptors [10, 24, 25], the latter being responsible for eliciting Ͼ97% of GABAR-mediated inhibition in CGCs and pivotal to information transfer in the cerebellum [26]

Summary

Introduction

The stargazer (stg) mutant mouse arose by virtue of a spontaneous viral transposon insertion into the stargazin gene [1]. TARP␥2 is reported to be heavily expressed in the cerebellum [2, 9] where it is largely restricted to the cerebellar granule cells (CGCs), neurons that normally exclusively express the TARP␥2 isoform of TARPs. mossy fiber-CGC synapses in stg are bereft of AMPARs and are subsequently electrically silent [7] leading to a CGC-specific deficit in brain-derived neurotrophic factor (BDNF) expression and signaling [4]. The abundance and distribution of the GABAR ␥2 subunit and the number of receptors that it contributes to are largely unaffected, as is the developmental maturation profile of the BZ-S subtype of cerebellar GABARs. our data imply that AMPA receptor activity has selective effects on GABAR subtypes expressed in cerebellar granule cells that may underpin homeostatic GABAergic responses to neuronal excitability

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