INVOLVEMENT OF CYTOPLASMIC CALCIUM AND PROTEIN-KINASES IN THE REGULATION OF ATRIAL-NATRIURETIC-FACTOR SECRETION BY CONTRACTION RATE AND ENDOTHELIN

To characterize the effects of the cellular events associated with con traction on atrial natriuretic factor (ANF) secretion, primary neonata l rat atrial myocytes were electrically paced to contract while being monitored for ANF release, cytoplasmic calcium, phosphoinositide hydro lysis, and protein kinase C activation. Similar measurements were also carried out in the presence of endothelin-1 (ET) for comparison of co ntraction-related and hormone-stimulated ANF secretion. Pacing (6-8 Hz ) immediately increased ANF secretion by 3-5-fold and the time-average d cytoplasmic calcium concentration (as monitored with indo-1 fluoresc ence) varied with pace frequency in a similar manner, suggesting that cytoplasmic calcium may play a key role in pace-induced ANF secretion. Furthermore, nifedipine and ryanodine, which inhibited the contractil e calcium transients, inhibited pace-induced ANF release, whereas Bay K 8644 increased both the calcium transients and ANF secretion. Pace-i nduced ANF release was also completely inhibited by KN-62, a specific inhibitor of Ca2+/calmodulin-dependent protein kinase II (CaMK) but wa s not inhibited by chelerythrine, a protein kinase C-selective inhibit or. Pace-induced ANF release averaged 40% of that elicited by ET which is known to require both PKC and CaMK for maximal effects on ANF secr etion. The effects of pacing and ET on ANF secretion were approximatel y additive. In contrast to pacing, ET strongly stimulated phosphoinosi tide hydrolysis, activated PKC, and did not increase cytoplasmic calci um. Thus, regulation of ANF secretion by contraction rate depends prim arily on the contractile calcium transients and CaMK and is independen t of PKC.