Real-time measurements of acetylcholine-induced release of ATP from bovine medullary chromaffin cells

Abstract

The luminescent oxidation of luciferin has been used to monitor acetylcholine-induced ATP release from cultured bovine chromaffin cells. Acetylcholine (1-100 microM) evoked ATP release of up to 30% of the total cellular ATP. This secretion required external free calcium and could also be elicited by K+-induced membrane depolarization. The size of the cytosolic ATP compartment was estimated as 5% of the ATP in the cell by solubilising the cell membrane using digitonin (20 microM) or by application to the cells of brief pulses (2 microseconds) of high electric field (2000 V/cm). Blockers of the voltage-gated Ca2+ channel effectively blocked K+-induced ATP release, while the acetylcholine antagonists d-tubocurarine and beta-bungarotoxin inhibited the acetylcholine-induced release of ATP. These data support the concept that ATP is released together with the catecholamines by exocytosis of chromaffin granule contents.