Temperature and PMA affect different phases of exocytosis in bovine chromaffin cells

Amperometry was used to study secretory kinetics of single bovine chromaffin cells stimulated by transient depolarizations at different temperatures. The initial rate of release was moderately enhanced when the temperature was raised from 18 to 22 and 37 degrees C. Secretion increased drastically at a later period, 5-10 s after the initiation of stimulus. Interestingly, incubation of the cells with phorbol 12-myristate 13-acetate (PMA) clearly enhanced fast secretory components. In addition, the rate of secretion of the slower component recruited by prolonged depolarizations (t > 30 s) was unaffected at the range of temperatures normally used in secretory experiments (22-37 degrees C). A 'counting events' analysis of secretion, which avoids the influence of event charge changes, showed specific increases in a population of vesicles fusing between 7 and 12 s over the same range of temperatures, and a marked increase in vesicles fusing during the initial phase (1-5 s), of PMA-treated cell secretion. An analysis of temperature influence on transient components released by high sucrose, the secretion elicited by cell permeabilization with digitonin, and studies of the individual characteristics of amperometric events, allow us to conclude that an increase in the size of a secondary-released vesicle population is the main factor contributing to temperature-dependent enhancement of secretion, in clear contrast to the enhancement of fast releasable pools caused by phorbol esters.