MITOCHONDRIAL GENE-EXPRESSION IS IMPAIRED BY ETHANOL EXPOSURE IN CULTURED CHICK CARDIAC MYOCYTES

Objective: A depression in cytochrome c oxidase (COX) activity occurs following chronic embryonic ethanol exposure in vivo. The aim of this study was to examine the effect of chronic ethanol exposure on COX act ivity in isolated cardiac cells maintained in vitro. Additionally, the mechanism by which ethanol produces an impairment in COX activity was evaluated by examining mitochondrial gene expression. Methods: Sponta neously beating cardiac myocyte cultures were established from 10-day embryonic chick hearts. Various concentrations of ethanol (0-250 mM) w ere introduced at the time of plating and cells were harvested over 7 days. COX activity was determined in myocyte homogenates. The levels o f nuclear-encoded (COXIV) and mitochondrial-encoded (COXII) subunit pr oteins were measured by Western blotting. Relative levels of mitochond rial DNA and the mitochondrially-encoded COXIII mRNA were determined b y Southern and Northern blotting. Results: A consistent decrease in CO X activity in ethanol-exposed cardiac myocytes of approximately 30% wa s observed with an ethanol concentration of 25 mM. Increasing the etha nol concentration to 250 mM produced only a minor enhancement of this effect, while severely decreasing cellular viability, The content of t he mitochondrially-encoded COXII subunit was reduced by ethanol exposu re, while that of the nuclear-encoded COXIV subunit was unchanged. The content of the mitochondrially-encoded COXIII mRNA was unchanged by e thanol exposure. However, prolonged ethanol exposure produced an incre ase in mitochondrial DNA levels in cardiac myocytes. Conclusions: Etha nol exposure of cardiac myocytes produces deficits in COX activity in the absence of systemic variables, indicating that ethanol has a direc t effect on cardiac mitochondria. The ethanol-induced decrease in COX activity is correlated with a specific decrease in at least one mitoch ondrially-encoded gene product, COXII. No changes were observed in the level of the nuclear-encoded COXIV subunit, indicating that expressio n of this nuclear-encoded gene is not impaired by ethanol exposure. (C ) 1998 Elsevier Science B.V.