Agonist-stimulated Cyclic ADP Ribose

• Published in: The Journal of biological chemistry 1995-10, Vol.270 (43), p.25488-25494
• Main Authors: Kuemmerle, John F., Makhlouf, Gabriel M.
• Format: Article
• Language: English
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.270.43.25488

We have previously shown that agonist-induced Ca2+ mobilization in intestinal longitudinal muscle is mediated by ryanodine-sensitive, inositol 1,4,5-trisphosphate-insensitive sarcoplasmic Ca2+ channels. Ca2+ release via these channels is triggered by agonist-stimulated Ca2+ influx and results in Ca2+-induced Ca2+ release. The present study examined whether cyclic ADP-ribose (cADPR) is synthesized in response to stimulation of longitudinal muscle by agonists and modulates the activity of Ca2+ release channels. Cyclic ADPR bound with high affinity to dispersed longitudinal muscle cells (IC50 1.9 nM) and induced Ca2+ release (EC50 3.8 nM), increase in [Ca2+]i(EC50 2.0 nM), and contraction (EC50 1.1 nM); cADPR had no effect on circular muscle cells. The effects of cADPR were blocked by ruthenium red, dantrolene, and the specific antagonist, 8-amino-cADPR, and were augmented by caffeine but not affected by heparin. The binding of cADPR and its ability to stimulate Ca2+ release were dependent on the concentration of Ca2+. Cyclic ADPR was capable of stimulating Ca2+ release at subthreshold Ca2+ concentrations (25-100 nM) and of enhancing Ca2+-induced Ca2+ release. Longitudinal muscle extracts incubated with β-NAD+ produced a time-dependent increase in Ca2+-mobilizing activity identified as authentic cADPR by blockade of Ca2+ release with 8-amino-cADPR and ruthenium red. Ca2+ mobilizing activity was increased by cholecystokinin octapeptide (CCK-8) in a concentration-dependent fashion. The increase induced by CCK-8 was suppressed by the CCK-A antagonist, L364,718, nifedipine, and guanyl-5′-yl thiophosphate. The study shows that ADP-ribosyl cyclase can be stimulated by agonists and that cADPR can act as an endogenous modulator of Ca2+-induced Ca2+ release.