Effect of Estradiol on Enzymes of Carbohydrate Metabolism in Rat Uterus

The effect of estradiol on the activities of enzymes of the Krebs cycle, enzymes of glycolysis and the dehydrogenases of the pentose cycle in the rat uterus was estimated. Castration had no effect on the enzymes examined except on the glucose-6phosphate dehydi'ogenase, which was decreased. Estradiol at a dose of 30 Mg/rat decreased the activity of aconitase with isocitrate as substrate and of isocitric dehydrogenase. The dehydrogenases of the pentose cycle were increased after estradiol administration. It is assumed that estradiol administration does not stimulate the Krebs cycle in the rat uterus and that the additional energy for the synthetic reactions that occur in the uterus after estradiol stimulation might be derived through the activity of an estrogendependent transhydrogenase. (Endocrinology 78: 409, 1966) V results have been obtained for the effect of estrogens on the respiratory rate of rat uterus. The increase, if any, as the result of the hormone, has been very small (1-3). Recently, the effect of estradiol on citrate accumulation in the rat uterus has been studied by means of fluoroacetate poisoning 3 hr before sacrifice (4). At 8 and 15 hr after estradiol administration, there was no increase in citrate accumulation. At 24 hr a small increase was noted, but 96 hr after the hormone a large decrease in citrate accumulation in the uterus occurred. In order to ascertain whether the decreased citrate accumulation was an effect of the increased permeability of the uterine cells rather than a decreased activity of the Krebs cycle, the activities of the enzymes of the cycle were determined at intervals after estradiol administration. A few enzymes of glycolysis and the dehydrogenases of the pentose cycle were also measured. Materials and Methods Female adult albino rats, 180-230 g body weight, purchased from the Charles River Laboratories, were ovariectomized and used in the experiments 10 days after ovariectomy. Estradiol-17/8 suspended in 0.2 ml of 1 % ethanol0.05M phosphate buffer-saline solution (pH 7.5) was injected subcutaneously in a dose of 30 Mg/rat. Received September 7, 1965. 1 Fellow of the Population Council. Present address: Department of Zoology, Hebrew University of Jerusalem, Jerusalem, Israel. All enzyme assays were performed on the supernatant fraction obtained by centrifuging homogenized uteri at 450 Xg for 10 min. Enzyme activities were determined by following the changes in optical densities of the reaction mixtures at 30 C in a Beckman DK-2A recording spectrophotometer. The substrate concentrations used were at least 10 times higher than those required to yield maximal activities. Initial enzyme activities were measured under conditions in which activities were proportional to enzyme concentrations. The dehydrogenases that act on glucose-6phosphate andphosphogluconate (5),lactate (6), isocitrate (7), malate (8), as well as the malic enzyme (9), were measured spectrophotometrically at 340 m/x by established procedures. Aconitase and fumarase (10) were measured by following the increase in optical density at 240 mjx resulting from the formation of cis-aconitate and fumarate, respectively. Succinic dehydrogenase was determined by the K3Fe(CN)6 reduction method (11). Phosphoglucomutase and glucokinase were measured by coupled assays with added glucose-6-phosphate dehydrogenase (12). Pyruvate kinase was measured by a coupled assay with lactic dehydrogenase (13); since the activity of endogenous lactic dehydrogenase was very great, it was unnecessary to add lactic dehydrogenase to the reaction mixture. Protein was determined according to the method of Lowry et al. (14). Results and Discussion There was no change in the activities of the enzymes of the Krebs cycle in the uterus after castration (Table 1). The high dose of estradiol depressed the activities of aconitase with isocitrate as substrate and of isocitric 409 410 NOTES AND COMMENTS Volume 78 TABLE 1. Effect of estradiol-17/3 on enzymes of carbohydrate metabolism in the 450 Xg supernatant of rat uterus homogenate (values are means + SE of 3-6 determinations, expressed as m/umoles produced/min/mg protein)