Tr. Jiang et al., ENDOTHELIN-DEPENDENT ACTIONS IN CULTURED AT-1 CARDIAC MYOCYTES - THE ROLE OF THE EPSILON-ISOFORM OF PROTEIN-KINASE-C, Circulation research, 78(4), 1996, pp. 724-736
The consequences of endothelin receptor activation were examined in at
rial tumor myocytes derived from transgenic mice (AT-1 cells). Endothe
lin-1 (endothelin) stimulates phosphoinositide hydrolysis in a dose-de
pendent manner. Endothelin also induces the rapid and transient transl
ocation of protein kinase C (PKC)-epsilon immunoreactivity from the so
luble to the particulate cell fraction. The subcellular distributions
of PKC alpha and PKC zeta (also expressed by AT-1 cells) are not influ
enced by endothelin. Using quantitative fluorescence microscopy with f
ura 2, we examined the effects of endothelin on intracellular calcium.
In electrically driven myocytes, endothelin induces a rapid and trans
ient increase in the amplitude of the calcium transient. This is block
ed by both phorbol Ir-myristate 13-acetate (PMA) pretreatment to downr
egulate PKC and the PKC inhibitor chelerythrine, arguing that PKC epsi
lon plays a critical role in endothelin receptor-dependent increases i
n intracellular calcium. Endothelin also stimulates mitogen-activated
protein kinase (MAPK). MAPK activation is markedly attenuated by pretr
eatment with PMA or pertussis toxin (PTX, to inactivate susceptible G
protein alpha subunits); it is completely prevented by combined pretre
atment with PMA and PTX. In contrast, it is not attenuated by chelatio
n of intracellular calcium with BAPTA. These findings indicate that th
e pathway for endothelin receptor stimulation of MAPK involves PKC eps
ilon and PTX-sensitive G protein(s). Thus, these studies identify a fu
nctional role for PKC epsilon as a mediator of endothelin receptor-dep
endent increases in cytosolic calcium and MAPK activity ill AT-I cells
. Accordingly, the AT-1 cell system should provide a uniquely useful m
odel to identify the intracellular targets for PKC epsilon and investi
gate their function in the regulation of intracellular calcium homeost
asis and the induction of the growth response in cardiac myocytes.