The administration of a low-carbohydrate\high-saturated-fat (LC\HF) diet for 28 days or starvation for 48 h both increased pyruvate dehydrogenase kinase (PDHK) activity in extracts of rat hepatic mitochondria, by approx. 2.1-fold and 3.5-fold respectively. ELISAs of extracts of hepatic mitochondria, conducted over a range of pyruvate dehydrogenase (PDH) activities, revealed that mitochondrial immunoreactive PDHKII (the major PDHK isoform in rat liver) was significantly increased by approx. 1.4-fold after 28 days of LC\HF feeding and by approx. 2-fold after 48 h of starvation. The effect of LC\HF feeding to increase hepatic PDHK activity was retained through hepatocyte preparation, but was decreased on 21 h culture with insulin (100 µ-i.u.\ml). A sustained (24 h) 2-4-fold elevation in plasma insulin concentration in i o (achieved by insulin infusion via an osmotic pump) suppressed the effect of LC\HF feeding so that hepatic