SHORT AND LONG FORM GAMMA(2) SUBUNITS OF THE GABA(A) BENZODIAZEPINE RECEPTORS/

Three novel antisera to the gamma(2) subunit of the gamma-aminobutyric acid(A) (GABA(A)) receptor/benzodiazepine receptor (GABA(A)R/BZDR) co mplex have been made. Anti-gamma(2S) and anti-gamma(2L) are specific a ntibodies to synthetic peptides that recognize the gamma(2S) (short) a nd gamma(2L)y (long) forms, respectively, of the gamma(2) subunit. An antibody (anti-gamma(2)IL2) to staphylococcal protein A fusion protein of the large intracellular loop (gamma(2)IL) located between the puta tive transmembrane segments M3 and M4 of gamma(2S) recognizes both gam ma(2S) and gamma(2L) subunits. The antibodies immunoprecipitated both the solubilized and affinity-purified GABA(A)R/BZDR from rat and bovin e brain. Immunoblots with membranes from rat brain cerebral cortex as well as with affinity-purified receptor from bovine cortex show that a nti-gamma(2S) and anti-gamma(2L) recognize peptides of 45,000 and 47,0 00 M(r), respectively. Immunoprecipitation experiments indicate that g amma(2S) is more prevalent in hippocampus, whereas gamma(2L) is more a bundant in cerebellum. Intermediate values for each form are found in the cerebral cortex. The results suggest that in the rat brain there i s a considerable amount of colocalization of gamma(2S) and gamma(2L) i n the same receptor complex. In the cerebral cortex, 15% of the BZDRs contain both gamma(2S) and gamma(2L) subunits and 41-48% of the gamma( 2L) subunit coexists with gamma(2S) in the same receptor complex. In c erebellum, in 27% of the clonazepam-insensitive and 39% of the clonaze pam-insensitive BZDRs the gamma(2S) and gamma(2L) coexist in the same receptor complex. The latter are presumably localized in granule cells and also contain alpha(6). In addition, almost all (93%) the clonazep am-insensitive BZDRs that contain gamma(2L) also contain a gamma(2S) s ubunit in the same receptor complex. The most likely interpretation of the results is that there is an important population of granule cell receptors that contain alpha(6) gamma(2S) and gamma(2L) coexisting in the same receptor complex. Nevertheless, 31% of the cerebellar recepto rs that contain alpha(6) subunit(s) have neither gamma(2S) nor gamma(2 L) subunits. There are also species differences with respect to the re lative abundance of gamma(2S) and gamma(2L) These results might be rel evant for understanding the molecular mechanisms underlying some of th e GABA(A)R/BZDR-mediated effects of ethanol intoxication involving cer ebellar granule cells.