Norepinephrine (NE) activates processes of hypertrophy and proliferation in cardiac myocytes and fibroblasts, respectively, in conjunction with expression of immediate early genes (IEG). Since adenosine 5'-triphosphate (ATP) is co-released with NE from sympathetic nerve endings in the heart, we studied the effects of extracellular ATP on IEG expression in cardiac cells and investigated the intracellular mechanisms of IEG induction by ATP. In response to micromolar quantities of extracellular ATP, levels of IEG mRNA increased at 15 min, peaked 30 min (5-8 fold), and declined to baseline by 1 hr in both cell types. ATP increased intracellular Ca2+ concentration (Cai) in cardiac myocytes (MYO) and fibroblasts (CAFB) loaded with Indo-1-am, whereas NE did not. The potency order of ATP analogues for increasing Cai and c-fos mRNA levels was: ATPrS is greater than ATP is greater than ADPbetaS = ADP is greater than 2-met-ATP is greater than AMP-PNP. Adenosine had no effect on either c-fos or Cai levels. In MYO, the ATP-induced increase in Cai and c-fos were inhibited by pretreatment with intracellular Ca2+ chelator, BAPTA-AM. However, pretreatment with BAPTA-AM did not inhibit the NE-induced increase in IEG in MYO. NE increased the rate of protein synthesis (incorporation of 14C- phenylalanine into TCA-precipitable protein) 2-3-fold, whereas ATP did not. In CAFB, overnight pretreatment with TPA or pretreatment with staurosporine for 30 min decreased the ATP-induced level of c-fos. Western blot analysis showed that ATP induced tyrosine phosphorylation of a 40-45-kDa protein. These data suggest that the ATP-induced increase in IEG occurs via activation of P2-purinergic receptors in both cell types, and that in CAFB multiple second messenger systems are involved. In MYO, IEG-induction by ATP, but not NE, is calcium dependent, suggesting that ATP and NE activate IEG expression by different intracellular signalling mechanisms. Furthermore, these results suggest that induction of c-fos is not sufficient to activate hypertrophy in MYO.
