Inhibitory effects of ethanol on the calcium-dependent potentiation of vasoactive intestinal peptide-stimulated cAMP and cGMP accumulation in rat pinealocytes

Abstract

In rat pinealocytes, ethanol has been shown recently to inhibit the α 1-adrenergic potentiation of vasoactive intestinal peptide (VIP)-stimulated cyclic AMP (cAMP) and cyclic GMP (cGMP) responses, with the cGMP response being more sensitive to the inhibition. Two intracellular events known to be of importance to the potentiation mechanism are activation of protein kinase C and elevation of intracellular calcium ([Ca 2+] i). In this study, we examined the effects of ethanol on these two intracellular mechanisms with an activator of protein kinase C, and two agents that elevate [Ca 2+] i, depolarizing concentrations of K + and A23187. Using dispersed pinealocytes, ethanol (up to 175 mM) was found to have no effect on the 4β-phorbol 12-myristate 13-acetate (PMA, an activator of protein kinase C) potentiated VIP-stimulated cAMP response. As for the cGMP response, full potentiation requires both activation of protein kinase C and simultaneous elevation of [Ca 2+] i. This could be achieved by stimulating the VIP-treated cells with a combination of PMA and 15 mM K +. In the presence of ethanol, the amplification effect of PMA and K + on the VIP-stimulated cGMP response was inhibited with an ic 50 value of 125 mM. In contrast, similar concentrations of ethanol had no effect on the corresponding cAMP response. These findings suggest that the potentiation of cAMP response by protein kinase C is not affected by ethanol. When depolarizing concentrations of K + were used to potentiate the VIP-stimulated cAMP and cGMP accumulation, ethanol inhibited the K + potentiation of VIP-stimulated cAMP and cGMP responses with ic 50 values of 50 and 30 mM, respectively. The A23187 potentiation of VIP-stimulated cGMP response was also sensitive to the inhibitory effect of ethanol with an ic 50 value of 120 mM. In comparison, the corresponding ic 50 value for the cAMP response was > 175 mM. Based on our findings, we conclude that ethanol likely inhibits a Ca 2+-dependent event(s) that is critical to the α 1-adrenergic-mediated potentiation of VIP-stimulated cAMP and cGMP responses.