Alpha 1D L-type Ca(2+)-channel currents: inhibition by a beta-adrenergic agonist and pituitary adenylate...

In this study the subunits of the dihydropyridine-sensitive L-type Ca2+ channels (L-channels) expressed in rat pinealocytes were characterized using reverse transcription (RT)-PCR analysis, and the modulation of these channels by adrenergic agonists and by pituitary adenylate cyclase-activating polypeptide (PACAP) was studied using the patch-clamp technique. RT-PCR analysis showed that rat pinealocytes expressed alpha 1D, alpha 2b, beta 2, and beta 4 Ca(2+)-channel subunit mRNAs. Other alpha 1 subunit transcripts were either not expressed or present at very low levels, indicating that the pinealocytes express predominantly alpha 1D L-channels. Electrophysiological studies confirmed that the pineal expressed a single population of L-channels. The L-channel currents were inhibited by two agonists that elevate cyclic AMP: the beta-adrenergic agonist isoproterenol and PACAP. Similar inhibition was observed with a cyclic AMP analogue, 8-bromo-cyclic AMP. The presence of a cyclic AMP antagonist, Rp-adenosine 3',5'-cyclic monophosphorothioate, blocked the inhibition by isoproterenol and PACAP. Norepinephrine, a mixed alpha- and beta-adrenergic agonist, also inhibited the L-channel currents, but the inhibition was smaller. The smaller inhibition by norepinephrine was secondary to the simultaneous activation of alpha- and beta-adrenergic receptors. These results indicate that (a) pinealocytes express predominantly alpha 1D L-channels, and (b) the beta-adrenergic agonist isoproterenol and PACAP inhibit the L-channel currents through elevation of cyclic AMP. However, an alpha-adrenergic-mediated mechanism also appears to be involved in the effect of norepinephrine on the L-channel currents.