Contributions of the GABA(A) receptor alpha 6 subunit to phasic and tonic inhibition revealed by a naturally occurring polymorphism in the alpha 6 gene
GABA(A) 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 alpha 6 (Gabra6(100R) and Gabra6(100Q)). Depending on their subunit partners, these two variants of alpha 6 can lead to differential sensitivities to benzodiazepines and ethanol. An examination of synaptic and extrasynaptic 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 inhibition mediated by delta subunit-containing GABARs in Gabra6(100Q/100Q) rats. Complementary experiments on recombinant GABARs confirmed that, at subsaturating [ GABA], flunitrazepam potentiates alpha 6/delta subunit-containing GABARs. Based on data and a simple theoretical analysis, we estimate that the average extrasynaptic [GABA] is similar to 160 nM in perfused slices. These results ( 1) demonstrate contributions of alpha 6 subunits to both synaptic and extrasynaptic GABA responses, ( 2) establish that delta subunit-containing GABARs are benzodiazepine sensitive at subsaturating [GABA] and, ( 3) provide an empirical estimate of extrasynaptic [GABA] in slices.