| Popis: |
Regulation of systemic catecholamine levels is critical for maintenance of integral physiological functions such as heart rate, insulin secretion and regulation of blood flow within the body. The adrenal medulla, the innermost layer of the adrenal gland, contains neurosecretory chromaffin cells. Chromaffin cells receive synaptic excitation through the sympathetic splanchnic nerve to elicit catecholamine release into the circulation. Under basal sympathetic tone, splanchnic-released acetylcholine evokes chromaffin cells to fire action potentials, leading to synchronous phasic catecholamine release. Under elevated splanchnic firing, experienced under the sympatho-adrenal stress response, chromaffin cells undergo desensitization to cholinergic excitation. Yet, stress evokes a persistent and elevated adrenal catecholamine release. Sustained stress-evoked release has been shown to depend on splanchnic release of a peptide transmitter, Pituitary Adenylate Cyclase-Activating Peptide (PACAP), which acts through a mechanistically distinct pathway from cholinergic excitation. In this study, I investigate the cellular and tissue-level mechanisms through which PACAP evokes excitation and sustained secretion. I report that PACAP stimulates sustained catecholamine release through a PKC-dependent functional recruitment of nickel-sensitive T-type Cav3.2 channels. To investigate this stimulus path, I utilized perforated- patch clamp electrophysiology, immunohistochemistry, and carbon fiber amperometry. I show that despite abundant immunohistochemical staining for Cav3.2, there is little nickel-sensitive current present in the unstressed adrenal medulla. Subsequently, I show that PACAP-dependent PKC activation, functionally recruits latent Cav3.2 channels to a functional, current-producing form. The functional recruitment of Cav3.2 generates a tonic Ca2+ window current to support sustained secretion. Moreover, previous studies showed that chronic stress results in an electrical remodeling of the adrenal medulla by up-regulation of connexin protein expression. Other studies report that phospho-regulation of existing gap junction channels acts to increase junctional conductance. I tested the hypothesis that PACAP-mediated excitation up-regulates cell-cell electrical coupling to enhance chromaffin cell excitability. I utilized electrophysiological recordings to measure the effects of PACAP stimulation on cell coupling. I report that PACAP excitation increases electrical coupling and the spread of electrical excitation between adrenal chromaffin cells. These findings show that PACAP acts as a secretagogue and remodels electrical remodeling of the medulla to adapt to the organism’s needs during acute sympathetic stress. |
