ATP can stimulate exocytosis in rat brown adipocytes without apparent increases in cytosolic Ca2+ or G protein activation

Extracellular ATP activates large increases in cell surface area and membra ne turnover in rat brown adipocytes (Pappone, P. A., and Lee, S. C, 1996. J . Gen. Physiol. 108:393-404). We used whole-cell patch clamp membrane capac itance measurements of membrane surface area concurrently with fura-2 ratio imaging of intracellular calcium to test whether these purinergic membrane responses are triggered by cytosolic calcium increases or G protein activa tion. Increasing cytosolic calcium with adrenergic stimulation, calcium ion ophore, or calcium-containing pipette solutions did not cause exocytosis. E xtracellular ATP increased membrane capacitance in the absence of extracell ular calcium with internal calcium strongly buffered to near resting levels . Purinergic stimulation still activated exocytosis and endocytosis in the complete absence of intracellular and extracellular free calcium, but endoc ytosis predominated. Modulators of G protein function neither triggered nor inhibited the initial ATP-elicited capacitance changes, but GTP gamma S or cytosolic nucleotide depletion did reduce the cells' capacity to mount mul tiple purinergic responses. These results suggest that calcium modulates pu rinergically-stimulated membrane trafficking in brown adipocytes, but that ATP responses are initiated by some other signal that remains to be identif ied.