OBJECTIVE—Glucagon-like peptide-1 (GLP-1) increases intracellular Ca²⁺ concentrations ([Ca²⁺]_i), resulting in insulin secretion from pancreatic β-cells. The molecular mechanism(s) of the GLP-1–mediated regulation of [Ca²⁺]_i was investigated.

RESEARCH DESIGN AND METHODS—GLP-1–induced changes in [Ca²⁺]_i were measured in β-cells isolated from Cd38^+/+ and Cd38^−/− mice. Calcium-mobilizing second messengers were identified by measuring levels of nicotinic acid adenine dinucleotide phosphate (NAADP) and cyclic ADP-ribose (ADPR), using a cyclic enzymatic assay. To locate NAADP- and cyclic ADPR–producing enzyme(s), cellular organelles were separated using the sucrose gradient method.

RESULTS—A GLP-1–induced [Ca²⁺]_i increase showed a cooperative Ca²⁺ signal, i.e., an initial [Ca²⁺]_i rise mediated by the action of NAADP that was produced in acidic organelles and a subsequent long-lasting increase of [Ca²⁺]_i by the action of cyclic ADPR that was produced in plasma membranes and secretory granules. GLP-1 sequentially stimulated production of NAADP and cyclic ADPR in the organelles through protein kinase A and cAMP-regulated guanine nucleotide exchange factor II. Furthermore, the results showed that NAADP production from acidic organelles governed overall Ca²⁺ signals, including insulin secretion by GLP-1, and that in addition to CD38, enzymes capable of synthesizing NAADP and/or cyclic ADPR were present in β-cells. These observations were supported by the study with Cd38^−/− β-cells, demonstrating production of NAADP, cyclic ADPR, and Ca²⁺ signal with normal insulin secretion stimulated by GLP-1.

CONCLUSIONS—Our findings demonstrate that the GLP-1–mediated Ca²⁺ signal for insulin secretion in pancreatic β-cells is a cooperative action of NAADP and cyclic ADPR spatiotemporally formed by multiple enzymes.