α(1D) L-type Ca²⁺-channel currents: Inhibition by a β-adrenergic agonist and pituitary adenylate cyclase-activating polypeptide (PACAP) in rat pinealocytes

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 (PA-CAP) was studied using the patch-clamp technique. RT-PCR analysis showed that rat pinealocytes expressed α(1D), α(2b), β₂, and β₄ Ca²⁺-channel subunit mRNAs. Other α₁ subunit transcripts were either not expressed or present at very low levels, indicating that the pinealocytes express predominantly α(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 β-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 α- and β-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 α- and β-adrenergic receptors. These results indicate that (a) pinealocytes express predominantly α(1D) L- channels, and (b) the β-adrenergic agonist isoproterenol and PACAP inhibit the L-channel currents through elevation of cyclic AMP. However, an α- adrenergic-mediated mechanism also appears to be involved in the effect of norepinephrine on the L-channel currents.