INSULIN REGULATES PROTEIN-METABOLISM IN MOUSE BLASTOCYSTS

Mouse blastocysts, in vitro, endocytosed 100 mug/ml I-125-labelled bovine serum albumin (BSA) at a rate equivalent to 192 +/- 27 mul/hr/mg embryonic protein over the first 20 min. Insulin stimulated this initial uptake by 30% (P < 0.05). After this time, accumulation of I-125-labelled BSA began to plateau as the endocytosed I-125-labelled BSA was catabolized and I-125 was released from the cells. Insulin caused an approximately 72% (P < 0.05) increase in the amount of uncatabolized I-125-labelled BSA remaining in insulin-treated blastocysts after 2 hr as compared to control blastocysts. Insulin partially inhibited catabolism of endocytosed I-125-labelled BSA during the first 2 hr following transfer to nonradioactive medium. After this time, degradation ceased in both control and insulin-treated blastocysts, leaving a small, uncatabolized protein pool remaining in the embryos; however, as a result of insulin's inhibitory effects on the initial catabolic rate, the uncatabolized protein pool was 30% (P < 0.05) larger in insulin-treated blastocysts after the 4 hr chase. Insulin inhibited endogenous protein degradation in blastocysts by 37% (P < 0.05). Combined with previous studies showing a 90% increase in endogenous protein synthesis in blastocysts following short-term stimulation with insulin (Harvey and Kaye, 1988), these results suggest that insulin acts to increase the endogenous protein reserves in the embryo. Dose-response studies indicated an EC50 of 0.5 pM for insulin's stimulation of I-125-labelled BSA accumulation, consistent with action via its own receptor. Insulin-like growth factor-1 (IGF-1) also stimulated protein accumulation at concentrations similar to those observed with insulin, suggesting that IGF-1 may act via its own receptor rather than the insulin receptor to exert its effects on endocytosis. (C) 1993 Wiley-Liss, Inc.