CALCIUM AND GTP-GAMMA-S AS SINGLE EFFECTORS OF SECRETION FROM PERMEABILIZED RAT MAST-CELLS - REQUIREMENTS FOR ATP

Treatment with metabolic inhibitors and addition of exogenous MgATP ex erted different effects on secretion from streptolysin-O-permeabilized mast cells, responding to calcium and GTP-gamma-S as single effectors (i.e., independently of each other). These effects were also strongly dependent on the experimental conditions. Thus cells, triggered by Ca 2+ at the time of permeabilization, did not require MgATP, but after m etabolic inhibition rapidly became absolutely dependent on its provisi on, requiring high (> mM) concentrations. AMP-PNP was not effective. A fter longer treatment with metabolic inhibitors, the absolute dependen ce on MgATP was also exhibited by cells responding to dual effectors ( i.e., Ca2+ and GTP-gamma-S applied together). In contrast. calcium ind ependent secretion due to GTP-gamma-S was more resistant to metabolic inhibition, exhibiting no absolute requirements for MgATP. Once the re sponsiveness to GTP-gamma-S had been lost, it could not bc restored by addition of MgATP. MgATP, in fact, inhibited the response of permeabi lized cells to GTP-gamma-S. This effect could be mimicked by AMP-PNP. When permeabilized cells were washed before triggering, MgATP (0.1-1 m M concentration range) was no longer inhibitory but stimulatory. These differences between Ca2+- and GTP-gamma-S-induced responses indicate that ATP utilization is essential to the calcium, but not to the guani ne nucleotide, pathway to secretion. The rate of the response to calci um/MgATP was much slower in the absence than in the presence of GTP-ga mma-S. The onset of secretion occurred after an initial delay. This la g phase was abolished by addition of GTP-gamma-S, suggesting that a GT P-binding protein may control a reaction which constitutes a rate-limi ting step in the secretory process.