Ca(2+)-dependent stimulatory effect of pituitary adenylate cyclase- activating polypeptide on catecholamine secretion from cultured porcine adrenal medullary chromaffin cells

Abstract

Pituitary adenylate cyclase-activating polypeptide (PACAP) stimulates catecholamine secretion from cultured porcine adrenal medullary chromaffin cells in a dose-dependent manner with the half-maximal and maximal doses of 30 nM and 1 microM, respectively. Either removal of extracellular Ca2+ or addition of Gd3+, an inorganic Ca2+ channel blocker, very potently inhibits PACAP-induced catecholamine secretion. Both nicardipine (1 microM) and methoxyverapamil (1 microM), blockers of voltage-dependent Ca2+ channels, are also effective in inhibiting PACAP-induced catecholamine secretion. When the intracellular free Ca2+ concentration ([Ca2+]i) is measured in a fura 2-loaded single chromaffin cell, PACAP is found to cause a sustained increase in [Ca2+]i by mobilizing Ca2+ from both extra- and intracellular pools. It is also found that PACAP stimulates the production of inositol phosphates in a dose-dependent manner, which is not abolished by removal of extracellular Ca2+ unlike the case of nicotine. PACAP increases cAMP content in chromaffin cells in a dose-dependent manner. Removal of extracellular Ca2+ enhances PACAP-induced cAMP production but strongly inhibits PACAP-induced catecholamine secretion. Pretreatment of cells with adenosine-3':5'-monophosphothioate, cyclic, Rp-isomer, a cAMP antagonist, does not block PACAP-induced catecholamine secretion. The addition of forskolin or 3-isobutyl-1-methylxanthine does not enhance the PACAP-induced catecholamine secretion. These results indicate that PACAP activates voltage-dependent Ca2+ channels and phospholipase C as well as adenylate cyclase in cultured porcine adrenal medullary cells and strongly suggest that PACAP-induced catecholamine secretion is mainly mediated by activation of voltage-dependent Ca2+ channels.

Pituitary Adenylate Cyclase-Activating Polypeptide and Its Receptors: 20 Years after the Discovery

“…In adrenal chromaffin cells, PACAP exerts a stimulatory action on catecholamine secretion Isobe et al, 1993;Chowdhury et al, 1994;Guo and Wakade, 1994;Houchi et al, 1994;Perrin et al, 1995;Neri et al, 1996;Jorgensen et al, 2000;Fukushima et al, 2002;Douglas et al, 2008;Valiante et al, 2008). PACAP also stimulates the release of brain natriuretic peptide and enkephalins, two regulatory peptides that are cosequestered with catecholamines in chromaffin granules (Babinski et al, 1996;Hahm et al, 1998) as well as a 15-fold increase in VIP mRNA expression (Lee et al, 1999a).…”

“…PACAP-induced catecholamine secretion is significantly greater after the induction of hypoglycemia and PACAP(Ϫ/Ϫ) mouse exhibit a delayed normalization of plasma glucose levels in response to insulin injection , suggesting that PACAP may play a beneficial role in glucose counter-regulatory mechanisms in the adrenal medulla during hypoglycemia. The effect of PACAP on catecholamine secretion is associated with activation of AC and elevation of cytosolic calcium concentrations (Isobe et al, 1993;Houchi et al, 1994;Isobe et al, 1994;Perrin et al, 1995;Chamoux et al, 1998). Incubation of adrenomedullary cells in calcium-free medium or blockage of voltage-operated calcium channels suppresses the PACAP-evoked stimulation of catecholamine secretion (Isobe et al, 1993;Houchi et al, 1995; PITUITARY ADENYLATE CYCLASE-ACTIVATING POLYPEPTIDE Przywara et al, 1996;O'Farrell and Marley, 1997), indicating that the effect of PACAP on chromaffin cells is mediated through calcium influx.…”

“…The effect of PACAP on catecholamine secretion is associated with activation of AC and elevation of cytosolic calcium concentrations (Isobe et al, 1993;Houchi et al, 1994;Isobe et al, 1994;Perrin et al, 1995;Chamoux et al, 1998). Incubation of adrenomedullary cells in calcium-free medium or blockage of voltage-operated calcium channels suppresses the PACAP-evoked stimulation of catecholamine secretion (Isobe et al, 1993;Houchi et al, 1995; PITUITARY ADENYLATE CYCLASE-ACTIVATING POLYPEPTIDE Przywara et al, 1996;O'Farrell and Marley, 1997), indicating that the effect of PACAP on chromaffin cells is mediated through calcium influx. Concurrently, in bovine and human, PACAP increases calcium levels from ryanodine/caffeine-sensitive calcium stores (Houchi et al, 1995;Tanaka et al, 1996Tanaka et al, , 1998Shibuya et al, 1999;Payet et al, 2003).…”

Pituitary Adenylate Cyclase‐Activating Polypeptide Causes Ca2+ Release from Ryanodine/Caffeine Stores Through a Novel Pathway Independent of Both Inositol Trisphosphates and Cyclic AMP in Bovine Adrenal Medullary Cells

“…This view has been supported by the following findings: (a) the adrenal gland contains the second highest concentration of PA-CAP among peripheral organs (Arimura et al, 1991); (b) PACAP-like immunoreactivity is found in the nerve endings innervating the adrenal medulla, as well as in adrenal medullary cells (Tabarin et al, 1994;Frödin et al, 1995;Shiotani et al, 1995); (c) the adrenal medullary cells express PACAP type-I receptor (PACAP-preferring receptor) mRNA (Spengler et al, 1993); and (d) PACAP increases catecholamine release, intracellular cyclic AMP (cAMP), inositol trisphosphates (1P 3), and the intracellular Ca 2~concentra-tion ([Ca2~I~) (Watanabe et al, 1992;Isobe et al, 1993;Chowdhury et al, 1994;Guo and Wakade, 1994;Tanaka et al, 1996). PACAP type-I receptors are seven-helix receptors, and five splicing variants with insertions at the C-terminal end of the third intracellular ioop are reported (PACAP-R, PACAP-R-hip, PACAP-R-hop1, PACAP-R-hop2, PACAP-R-hip-hopl) (Spengler et al, 1993).…”

Stimulatory Effect of Pituitary Adenylate Cyclase-Activating Polypeptide on Catecholamine Synthesis in Cultured Bovine Adrenal Chromaffin Cells: Involvements of Tyrosine Hydroxylase Phosphorylation Caused by Ca2+ Influx and cAMP

“…In addition to brain, PACAP is wide ly distributed in other organs such as the testis, gastroin testinal tract and lung (3). The concentration of PACAP is also relatively high in the adrenal gland, in which the peptide stimulated the secretion of catecholamines mainly by the activation of voltage-dependent Ca 21 channels (4,5). Tyrosine hydroxylase, the rate-limiting enzyme in cat echolamine synthesis, is phosphorylated and activated by cAMP/forskolin (an activator of adenylate cyclase), phor bol esters (activators of protein kinase C) or the elevation of the intracellular concentration of Ca 21 ([Ca 21]i) caused by nicotinic receptor stimulation, high K+-in duced depolarization or Ca 21 ionophores (6 8).…”