The dynamics and interrelationships of glucagon and insulin secretion were studied in the isolated perfused rat pancreas by utilizing a series of compounds that stimulate the release of both hormone. Leucine, arginine, prostaglandins F2α and E2, bovine growth hormone, and isoproterenol were administered individually over 60-second intervals. The release of glucagon preceded that of insulin in response to all compounds tested. The rapidity of glucagon release varied in response to different secretagogues; the time course of insulin release was fairly constant. The timing and the magnitude of glucagon and insulin release did not correlate statistically. Conclusions: (1) pancreatic alpha cells respond more rapidly than beta cells to the same stimulus; (2) antecedent release of glucagon is not the principal mediator of insulin release in response to stimuli common to both hormones; and (3) endogenous glucagon may at best modify the release of insulin evoked by certain secretagogues.