K. Yamamoto et al., REGULATION OF NA,K-ADENOSINE TRIPHOSPHATASE GENE-EXPRESSION BY SODIUM-IONS IN CULTURED NEONATAL RAT CARDIOCYTES, The Journal of clinical investigation, 92(4), 1993, pp. 1889-1895
Na,K-ATPase (Na,K-pump) plays an important role in the regulation of i
ntracellular ion composition. The purpose of this study is to determin
e whether Na+ regulates the levels of mRNA coding for Na,K-ATPase alph
a and beta subunits in cultured neonatal rat cardiocytes. We measured
intracellular Na+ levels ([Na+]i) in cardiocytes using a Na+-sensitive
fluorescence dye (SBFI). 1 mM ouabain caused a significant increase i
n [Na+]i in cardiocytes; from 12.8+/-0.3 to 28.8+/-1.8 mM. Exposure of
cardiocytes to 1 mM ouabain resulted in a three- to fourfold increase
in alpha1, alpha2, and alpha3 mRNA accumulation, and an approximate t
wo-fold increase in beta1 mRNA accumulation. A maximum elevation was r
eached at 60 min in both cases. The ouabain-induced alpha1 mRNA accumu
lation was still observed in the Ca2+-free culture medium. Exposure of
cardiocytes to 10 muM monensin in the absence of extracellular Ca2+ a
lso resulted in a threefold increase in alpha1 mRNA accumulation. The
increased alpha1 mRNA expression by 1 mM ouabain was associated with a
fourfold increase in alpha1 subunit protein accumulation. Transfectio
n experiments with chimeric plasmids containing 5'-flanking sequences
of alpha1, alpha2, and alpha3 isoform genes and a luciferase reporter
gene revealed that 1 mM ouabain caused a twofold increase in luciferas
e activity in each alpha system. These results suggest that Na + direc
tly regulates Na,K-ATPase gene expression in cardiocytes. The transfec
tion study further supports the premise that Na+-responsive elements a
re located within the 5'-flanking sequences of each alpha isoform gene
.