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The object of the present study was to determine the effect of thyroxine (T4) treatment of sheep on protein synthesis and associated energy costs in skeletal muscle and hepatocytes. Protein synthesis, and ouabain-sensitive and cycloheximide-sensitive respiration in isolated intercostal muscle and hepatocytes were determined in sheep after 5 weeks of daily injections of either saline or T4. Plasma T4 and total triiodothyronine (T3) concentrations were doubled and free T3 concentrations were quadrupled by T4 injections. The fractional rates of protein synthesis increased in isolated external intercostal muscle and hepatocytes from hyperthyroid sheep. Fractional rates of protein synthesis in isolated external intercostal muscle and hepatocytes were linearly correlated with plasma free T3 concentrations. Total oxygen consumption of muscle and hepatocytes was unaffected by T4 injections. Ouabain-sensitive respiration increased in hepatocytes and muscle of T4-treated animals. Cycloheximide-sensitive respiration was elevated in hepatocytes from hyperthyroid sheep. Cycloheximide-sensitive respiration in muscle was unaffected by T4 treatment. The present experiment demonstrates that T4 increases protein synthesis in ruminants. The energy expenditure in support of Na+, K(+)-ATPase and protein synthesis in skeletal muscle and hepatocytes may account for 34-60% of total cellular energy expenditure.