Cyclic AMP-mediated regulation of GABA(A) receptor subunit expression in mature rat cerebellar granule cells: evidence for transcriptional and translational control.

Abstract

Exposure of rat cerebellar granule cells cultured to maturity in vitro to forskolin, N6,2'-O-dibutyryladenosine 3',5'-cyclic monophosphate (Bt2cAMP), and 3-isobutyl-1-methylxanthine (IBMX) down-regulated GABA(A) receptor alpha6 and beta3 subunits but up-regulated alpha1 and beta2 subunits with respect to vehicle-treated controls. Dideoxyforskolin had no effect on subunit expression. Protein kinase A inhibitors, H-89 and Rp-adenosine 3',5'cyclic monophosphothioate, prevented these effects on alpha1 but not alpha6 subunit expression. Flunitrazepam-sensitive [3H]Ro 15-4513 binding sites were increased by 144 +/- 20% following forskolin treatment. [3H]Ro 15-4513 photoaffinity labelling showed that the GABA(A) receptor alpha1 subunit was the principal locus of the increased flunitrazepam-sensitive [3H]Ro 15-4513 binding. Forskolin decreased flunitrazepam-insensitive [3H]Ro 15-4513 binding sites by 25 +/- 8% and resulted in a 20% decrease in the irreversible incorporation of radioactivity in the alpha6 subunit. Steady-state levels of GABA(A) receptor subunit mRNAs were determined by semiquantitative RT-PCR in forskolin-treated cultures. Forskolin, Bt2cAMP, and IBMX down-regulated GABA(A) receptor alpha6 subunit mRNA expression; alpha1 and beta3 mRNA levels were unaffected, whereas beta2 subunit mRNA was up-regulated. Dideoxyforskolin had no significant effect on alpha1, alpha6, beta2, and beta3 mRNA levels. Thus, in mature cerebellar granule cells, GABA(A) receptor expression can be regulated by intracellular cyclic AMP levels. This occurs at the level of gene transcription and/or translation by mechanisms that are only partially governed by protein kinase A.