Conventional in vitro studies of pepsinogen secretion have measured secretion into the bulk medium and have demonstrated the critical role of Ca²⁺ in the process. The present study was undertaken to obtain further details of the process of secretion and its relation to Ca²⁺ changes over very short time periods. The relation between Ca²⁺ mobilization and exocytosis in an isolated individual peptic cell of the bullfrog was investigated by a method to measure both intracellular Ca²⁺ ([Ca²⁺]_i), using a fluorescent Ca²⁺ indicator, fura 2, and exocytosis from single cells using a video microscope analyzing system. Bombesin (3.2 × 10⁷ M) and bethanechol (3.2 × 10⁴ M) caused a rapid increase in [Ca²⁺]_i (initial peak) and a corresponding high frequency of initial exocytosis. After the initial peak, [Ca²⁺]_i was maintained at a somewhat elevated level over the baseline (sustained phase), with a corresponding low frequency of exocytosis. Both the sustained phase of elevated [Ca²⁺]_i and the related exocytosis were eliminated by the depletion of extracellular Ca²⁺. Low concentrations of bombesin (3.2 × 10¹⁰ M) and bethanechol (3.2 × 10⁷ M) caused sustained low-amplitude Ca²⁺ oscillations with correspondingly low frequencies but also caused sustained exocytosis. These data show that 1) cellular response differs between high and low concentrations of stimulus, 2) there is a close relation between [Ca²⁺]_i and exocytosis, 3) exocytosis follows elevation of [Ca²⁺]_i by 14-45 s (n = 6), and 4) there is a significant positive correlation between the peak [Ca²⁺]_i and the number of exocytoses.