PHORBOL-MYRISTATE ACETATE-INDUCED HYPERTROPHY OF NEONATAL RAT CARDIACMYOCYTES IS ASSOCIATED WITH DECREASED SARCOPLASMIC-RETICULUM CA2-EXPRESSION AND CALCIUM REUPTAKE( ATPASE (SERCA2) GENE)

The decreased expression of the sarcoplasmic reticulum Ca2+-ATPase ass ociated with cardiac hypertrophy was investigated in cultured neonatal rat cardiac myocytes. Northern blot analysis indicated a significant 55-60% decrease in Ca2+-ATPase mRNA levels and after 12 and 24 h of tr eatment with the phorbol ester phorbol myristate acetate (PMA). Myocyt es treated with the phorbol ester for 80 h showed a significant 34% de crease (relative to Vehicle-treated control cells) in the levels of Ca 2+-ATPase protein, and a significant 38% increase in the levels of alp ha-sarcomeric actin, as assessed by Western blot analysis using specif ic antibodies. Immunocytochemistry of myocytes treated for 72 h with t he phorbol ester revealed a hypertrophied cell morphology, and showed a marked decrease in Ca2+-ATPase staining intensity. Contractile calci um transients were evaluated through the use of indo-1. It was found t hat the t(1/2) for the decline of calcium transient was significantly prolonged by PMA treatment (0.51 +/- 0.15) when compared to controls ( 0.38 +/- 0.17, P<0.001). Treatment of myocytes with endothelin-1 also led to a 35% decrease in sarcoplasmic reticulum Ca2+-ATPase mRNA level s. It is concluded that phorbol ester treatment of neonatal rat cardia c myocytes induces similar changes in Ca2+-ATPase gene expression as o bserved in vivo in the hypertrophied and failing heart. The observed p rolongation in t(1/2) for [Ca2+](i) decline might be due to the observ ed depressed levels for sarcoplasmic reticulum Ca2+-ATPase in PMA trea ted cells. (C) 1996 Academic Press Limited