Regulation of ATP-citrate lyase in primary cultures of adult rat hepatocytes.

Hepatocytes were isolated from adult rats and cultured on a substratum of nylon mesh coated with rattail collagen (Sirica, A. E., Richards, W., Tsukada, Y., Sattler, C. A., and Pitot, H. C. (1979) Boc. N&Z. Acud. Sci. U. S. A. 76, 283-287). The addition of insulin (1 pM) to the culture medium increased ATP-citrate lyase activity 2.6-fold, whereas addition of glucagon (0.1 PM) decreased the activity by 68%. Dibutyryl-CAMP (0.1 IDM) mimicked the effect of glucagon, and glucagon supplementation of culture medium already containing dibutyryl-CAMP had no additive effect. The increase in enzyme activity caused by insulin is the result of new enzyme protein synthesis, as evidenced by the increased incorporation of radioactivity into ATP-citrate lyase immunoprecipitated from extracts of hepatocytes that had been maintained in media containing C3H]leutine. The half-life of ATP-citrate lyase in culture was determined to be 18 h and was unchanged by the presence of insulin or glucagon in the culture medium. When hepatocytes isolated from thyroidectomized rats were placed in culture, they exhibited decreased levels of ATP-citrate lyase activity, which could be restored by the addition of 3,3’,5-triiodo-L-thyronine (10 pM) to the culture medium. Insulin did not induce enzyme activity in the cells unless the hepatocytes were first treated with triiodothyronine 36 h prior to and during their exposure to insulin; this suggests that the prolonged deprivation of a specific hormone can affect the response of a cell to other hormones.