The mechanisms involved in autocrine ATP release from cultured astrocytes isolated from the rat cortex were investigated using an online bioluminescence technique. Astrocytes released ATP in response to application of 10 microM uridine triphosphate, which was blocked by the non-specific purinergic receptor antagonist suramin. Intracellular pathways of the uridine triphosphate-stimulated ATP release were seen to involve inositol triphosphate and calcium with the assistance of the Golgi-complex and cytoskeleton as the release was inhibited by phospholipase C antagonist lithium, endoplasmic reticulum calcium-dependent ATPase inhibitor thapsigargin, F-actin interruptor cytochalasin D and Golgi-complex interruptor brefeldin A. The uridine triphosphate-stimulated ATP release was also potently blocked by exocytosis inhibitor botulinum toxin A and anion transporter blockers furosemide and glibenclamide. These results suggest that calcium-dependent exocytosis and transportation via anion transporters are the predominant secretion mechanisms for uridine triphosphate-stimulated ATP release from cortical astrocytes.