Cellular/Molecular Contributions of the GABAA Receptor 6 Subunit to Phasic and Tonic Inhibition Revealed by a Naturally Occurring

GABAA receptors (GABARs) are heteromultimeric proteins composed of five subunits. The specific subunit composition determines critical properties of a GABAR such as pharmacological sensitivities and whether the receptor contributes to synaptic or extrasynaptic forms of inhibition. Classically, synaptic but not extrasynaptic GABARs are thought to respond to benzodiazepines, whereas the reverse has been suggested for ethanol. To examine the effects of subunit composition on GABAR function in situ, we took advantage of two naturally occurring alleles of the rat gene for GABAR subunit 6 (Gabra6 100R and Gabra6 100Q). Depending on their subunit partners, these two variants of 6 can lead to differential sensitivities to benzodiazepines and ethanol. An examination of synaptic and extrasyn-aptic GABA-mediated currents in cerebellar granule cells from Gabra6 100R/100R and Gabra6 100Q/100Q rats uncovered marked allele-dependent differences in benzodiazepine sensitivity. Unexpectedly, we found that the benzodiazepines flunitrazepam and diazepam enhanced extrasynaptic inhibitionmediated by subunit-containing GABARs in Gabra6 100Q/100Q rats. Complementary experiments on recombinant GABARs confirmed that, at subsaturating [GABA], flunitrazepam potentiates6/ subunit-containing GABARs. Based on data and a simple theoretical analysis, we estimate that the average extrasynaptic [GABA] is160 nM in perfused slices. These results (1) demonstrate contributions of 6 subunits to both synaptic and extrasynaptic GABA responses, (2) establish that subunit-containing GABARs are benzodiazepine sensitive at subsaturating [GABA] and, (3) provide an empirical estimate of extrasynaptic [GABA] in slices. Key words: granule cell; cerebellum; benzodiazepine; flunitrazepam; flumazenil; extrasynaptic inhibitio