Further observations on the in vitro stimulation by insulin of fat synthesis by lactating mammary gland slices.

The incorporation of [carboxy-14C] acetate into fatty acids by rat-liver slices in vitro was studied by Bloch & Kramer (1948), who showed that the rate of incorporation was increased by pyruvate and still further enhanced if insulin was present as well. The insulin effect was, however, not seen if glucose replaced pyruvate, nor did it occur in acetate alone. Later, Bloch (1948) reported that these results could not be reproduced regularly with other strains of rats, and he concluded that further work was necessary before the general significance of this insulin effect, believed to be particularly related to the utilization of pyruvate for fatty acid synthesis, could be evaluated. Following upon. this work we were able, by using lactating mamary gland tissue of the rat, which is an eminently favourable medium for the study of in vitro fat synthesis (Folley & French, 1949a, 1950), to demonstrate a marked stimulating effect of insulin upon in vitro fat synthesis in this tissue in mixtures of acetate and glucose, as judged by respiratory data and acetate uptake (Balmain, French & Folley, 1950). A definite but less marked effect was observed in glucose alone. Our results thus differ in certain respects from those of Bloch & Kramer (1948)asdotherecent resultsofBrady& Gurin (1950), who have reported that insulin increases the incorporation of 14C into long-chain fatty acids by ratliver slices in presence of methylor carboxyllabelled acetate alone, but not in acetate plus pyruvate. In this paper we report further observations upon the stimulating action of insulin in vitro on fat synthesis by lactating mammary gland slices. The phenomenon has now been studied in mammary tissue from the mouse, rabbit and ewe, as well as more extensively in tissue from the rat, especially in relation to the functional state of the mammary gland. The important question of the mechanism of this remarkable in vitro insulin effect has also been considered in a preliminary way. The active mammary gland normallysecretes large quantities of neutral fat in the form of glycerides, and it would therefore appear that the problem of the net fat synthesis which mammary gland slices are clearly capable ofeffecting in vitro under suitable conditions (Folley & French, 1950) is essentially a problem of glyceride formation rather than merely of fatty acid synthesis. That being so, the question ofthe origin of the glycerol must be taken into consideration, and the possibility of the availability of glycerol being critical for the synthesis of neutral fat, and at one remove of fatty acids, in mammary tissue, naturally arises. In view ofthe possibility that the stimulating action of insulin on fat synthesis in this tissue, demonstrated by the present work, might, partly at any rate, be connected with the regulation of the supply of glycerol, we have studied the effects of the addition of glycerol on the respiratory metabolism and acetate uptake of lactating mammary tissue. Some ofthis work has already been communicated in preliminary form (Balnain & Folley, 1951).