Modulation of stimulus-secretion coupling in porcine adrenal chromaffin cells by receptor-mediated increases in protein kinase C activity

Catecholamine (CAT) secretion by adrenal chromaffin cells is primarily triggered by nicotinic receptor-dependent increases in cytosolic Ca ²⁺. The principal aim of the present study was to determine whether pituitary adenylate cyclase activating peptide (PACAP), which is coreleased with acetylcholine from the splanchnic nerve, can modulate nicotinic receptor-dependent Ca ²⁺ signaling and catecholamine secretion in porcine adrenal medullary chromaffin (PAMC) cells. Activation of protein kinase C (PKC) with phorbol myristate acetate (PMA) dose- and time-dependently inhibited nicotine (NIC)-induced Ca ²⁺ transients. At 100 nM PMA, peak Ca ²⁺ levels were reduced by 27% ± 2% (P < 0.05) and 41% ± 3% (P < 0.05) after 10 and 20 min exposure, respectively. The inhibitory effects of PMA were significantly reduced by preincubation with the PKC inhibitor staurosporine. KCl-induced Ca ²⁺ transients were also reduced by 20 min PMA treatment (Δ -27% ± 4%; P < 0.05), suggesting that PKC affects voltage-gated Ca ²⁺ channel activity. Pretreatment with PACAP also resulted in both time- and concentration-dependent suppression of Ca ²⁺ transients. After 20 min exposure to 1 μM PACAP, NIC- and KCl-induced transients were reduced by 36% ± 5% (P < 0.05) and 51% ± 6% (P < 0.05), respectively. These effects could also be prevented by staurosporine pretreatment. NIC-induced CAT secretion was significantly reduced by pretreatment with both PMA (Δ -56% ± 2%; P < 0.05) and PACAP (Δ-53% ± 7%; P < 0.05). This suppressive effect on secretion could be prevented by pre-treatment with staurosporine. These data suggest that, in addition to having direct stimulatory effects on catecholamine synthesis and secretion, PACAP can also negatively modulate nicotinic receptor-dependent Ca ²⁺ signaling and secretion in PAMC cells. (C) 2000 Wiley-Liss, Inc.