Role of Ins(1,4,5)P3, cADP-ribose and nicotinic acid-adenine dinucleotide phosphate in Ca2+ signalling in mouse submandibular acinar cells

cADP-ribose (cADPr) and nicotinic acid-adenine dinucleotide phosphate (NAADP) are two putative second messengers; they were first shown to stimulate Ca2+ mobilization in sea urchin eggs. We have used the patch-clamp whole-cell technique to determine the role of cADPr and NAADP in relation to that of Ins(1,4,5)P-3 in mouse submandibular acinar cells by measuring agonist-evoked and second-messenger-evoked changes in Ca2+-dependent K- and Cl- currents. Both Ins(1,4,5)P-3 and cADPr were capable of reproducing the full range of responses normally seen in response to stimulation with acetylcholine (ACh). Low concentrations of agonist (10-20 nM ACh) or second messenger [1-10 muM Ins(1,4,5)P-3 or cADPr] elicited a sporadic transient activation of the Ca2+-dependent currents: mid-range concentrations [50-500 nM ACh, 50 muM Ins(1,4,5)P-3, or 50-100 muM cADPr] elicited high-frequency (approx. 2 Hz) trains of current spikes; and high concentrations [more than 500 nM ACh, more than 50 muM Ins(1,4,5)P-3 or more than 100 muM cADPr] gave rise to a sustained current response. The response to ACh was inhibited by antagonists of both the Ins(1,4,5)P-3 receptor [Ins(1,4,5)P3R] and the ryanodine receptor (RyR) but could be completely blocked only by an Ins(1,4,5)P3R antagonist (heparin). NAADP (50 nM to 100 muM) did not itself activate the Ca2+-dependent ion currents, nor did it inhibit the activation of these currents by ACh. These results show that, in these cells, both Ins(1,4,5)P3R and RyR are involved in the propagation of the Ca2+ signal stimulated by ACh and that cADPr can function as an endogenous regulator of RyR. Furthermore, although NAADP might have a role in hormone-stimulated secretion in pancreatic acinar cells, it does not contribute to ACh-evoked secretion in submandibular acinar cells.