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Fluctuations of calcium, phosphorus, sodium, potassium, and chlorine in beta cells were followed during rat islet perifusion with tolbutamide and related to insulin secretion. In 24 paired experiments two chambers containing 100 islets were perifused with buffered medium containing 4.2 mM glucose alone or with added tolbutamide (200 micrograms/ml). Effluent was collected frequently for insulin determinations. At eight different time intervals from 0 to 20 min islets were acutely fixed, prepared for scanning electron microscopy and beta cells in islet tissue were identified. Element content in 480 single cells was measured by energy dispersive x-ray analysis. Tolbutamide elicited typical monophasic insulin release that exceeded control islet secretory rates from 2 to 6 min with a peak value at 3 min. This pattern was preceded by monophasic calcium accumulation in beta cells that abruptly rose 150% above control cells at 1 min and declined to base line by 4 min. The rapid ascent of calcium was associated with significant depressions of sodium and potassium content without alterations of cell phosphorus. Chlorine fell at 2 min and then rose greater than 50% above control cells at 4 min. After 6 min insulin secretion and element content remained near control levels. We conclude that monophasic calcium accumulation in beta cells is the earliest, most predictive event of islet insulin secretion after a tolbutamide stimulus. Oscillations of beta cell sodium and potassium reciprocally relate to calcium, and an elevation of chlorine content is a relatively late phenomenon in the stimulus-secretion coupling process.

Tolbutamide perifusion of rat islets. Sequential changes in calcium, phosphorus, sodium, potassium, and chlorine in single beta cells.