Estradiol stimulation of glucose transport in rat uterus
AuthorMeier, Daniel Alan
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A study of the mechanism of the estradiol-mediated increase in glucose transport in the uteri of ovariectomized rats was undertaken. An essential first step in this study was the characterization of the glucose transport process using plasma membrane vesicles. Uterine plasma membrane preparations were obtained by centrifugation on discontinuous sucrose gradients. The specific activity of the plasma membrane marker 5'-nucleotidase was increased 10-fold while the specific activity of an endoplasmic reticulum marker glucose-6-phosphatase was increased 3-fold. D-Glucose transport into plasma membrane vesicles was inhibited by sulfhydryl reagents, phloretin, and cytochalasin B. Uptake was prevented by high osmotic pressures. The Km of glucose transport was 12.2 +_ 1.1 mM. Transport was unaffected by sodium and was energy independent. 2-Deoxyglucose transport was determined in uteri of rats grouped by stages of the reproductive cycle, i.e., diestrus 1, diestrus 2, proestrus and estrus. The rate of 2-deoxyglucose transport was highest in proestrus and lowest in diestrus 1. The increase in glucose transport in ovariectomized rats was half-maximal at approximately 5 ng estradiol/rat and reached the maximal 2 to 3-fold response after 2 hours whether measured in whole tissue or in uterine plasma membrane vesicles. Estrone and estriol treatment resulted in a similiar 2-fold increase in glucose transport while progesterone and dihydrotestosterone had no effect. Injection of protein synthesis inhibitors cycloheximide and emetine resulted in an increase in the basal glucose transport rate while having no effect on the estradiolstimulated increase in glucose transport. Treatment with the transcriptional inhibitor actinomycin D resulted in a slight increase in glucose transport with no effect on the estradiol-stimulated increase in glucose transport. Antiestrogen treatment (nafoxidine and tamoxifen) resulted in a 85-90 % decrease in the total estradiol binding sites in the cytosol but did not effect the estradiol stimulated increase in 2-deoxyglucose transport in uterine tissue. Insulin injection (0.2 mg/rat) resulted in a 40 % increase in 2-deoxyglucose transport in 24 h-starved rats in contrast to the 200 % increase seen with estradiol treatment. Insulin and estradiol treatment together were not additive in regard to the increase in 2-deoxyglucose transport in tissue. Estradiol treatment did not change binding of [1251 ] insulin to uterine plasma membranes. Estradiol treatment resulted in a 3-fold increase in the Vmax with no apparent change in the Km for 2-deoxyglucose transport. Also, estradiol treatment did not result in an increase in the amount of glucose transporters in uterine plasma membranes as measured by antibodies raised against the glucose transporter protein from human erythrocytes. In summary, estradiol stimulates the rate of glucose transport in rat uterus by increasing the rate by which the transporter protein moves glucose across the plasma membrane .