GABA(A) receptor subunit expression and assembly in cultured rat cerebellar granule neurons

The assembly of multisubunit GABA(A) receptors in specific neuronal populations is a complex process which is poorly understood. To begin to examine receptor assembly, α1, β2/3, and γ2 subunit polypeptide expression and association, as well as receptor binding, were examined in cultured rat cerebellar granule neurons. Western blots revealed two α1-immunoreactive proteins. A 39 kDa species was maximal at 2 days in culture and subsequently declined. In contrast, a 51 kDa polypeptide, the anticipated size of the mature α1 subunit, was first detected at 4 days and increased throughout the culture period. Additional studies demonstrated that the β2/3 and γ2 subunits were detectable at 2 days and attained maximal levels by 6 days. The level of [³H]Ro15-1788 binding, a measure of assembled receptors, rose in parallel with the increases in the 51 kDa α1, β2/3, and γ2 subunits. Moreover, the 51 kDa α1, β2/3, and γ2 subunits were associated in receptor complexes. However, immunohistochemical studies demonstrated the presence of substantial intracellular subunit staining. This finding suggests that only some of the subunits expressed in granule neurons contribute to functional GABA, receptors on the cell surface.