VPAC receptor modulation of neuroexcitability in intracardiac neurons: dependence on intracellular calcium mobilization and synergistic enhancement by PAC1 receptor activation

Pituitary adenylate cyclase-activating polypeptide (PACAP) and vasoactive intestinal polypeptide (VIP) have been found within mammalian intracardiac ganglia, but the cellular effects of these neuropeptides remain poorly understood. Fluorometric calcium imaging and whole cell patch clamp recordings were used to examine the effects of PACAP and VIP on [Ca2+]i and neuroexcitability, respectively, in intracardiac neurons of neonatal rats. PACAP and VIP evoked rapid increases in [Ca2+]i that exhibited both transient and sustained components. Pharmacological experiments using PAC1 and VPAC receptor-selective antagonists demonstrated that the elevations in [Ca2+]i result from the activation of VPAC receptors. The transient increases in [Ca2+]i were shown to be the product of Ca2+ mobilization from caffeine/ryanodine-sensitive intracellular stores and were not due to inositol 1,4,5-trisphosphate-mediated calcium release. In contrast, the sustained [Ca2+]i elevations were dependent on extracellular Ca2+ and were blocked by the transient receptor channel antagonist, 2-aminoethoxydiphenyl borate, which suggests that they are due to Ca2+ entry via store-operated channels. In addition to elevating [Ca2+]i, both PACAP and VIP depolarized intracardiac neurons, and PACAP was further shown to augment action potential firing in these cells. Depolarization of intracardiac neurons by the neuropeptides was dependent on activation of VPAC receptors and the concomitant increases in [Ca2+]i. Although activation of PAC1 receptors alone had no direct effects on neuroexcitability, PAC1 receptor stimulation potentiated the VPAC receptor-induced depolarizations. Furthermore, enhanced action potential firing was only observed upon concurrent stimulation of PAC1 and VPAC receptors, which indicates that these receptors act synergistically to enhance neuroexcitability in intracardiac neurons.