Glucose metabolism in the superovulated rat ovary in vitro. Effects of luteinizing hormone and the role of glucose metabolism in steroidogenesis

Abstract

1. Superovulated rat ovary slices from rats treated with 20μg. of luteininzing hormone/100g. body wt. 2hr. before death and from control animals have been incubated in vitro. Output of Δ4-3-oxo steroids (0·2μmole/g. wet wt./hr. in control tissue) was linear for 4hr., and was increased by approx. 70% in slices from luteinizing hormone-treated rats. Rate of oxygen consumption (90·0±4·6μmoles/g. wet wt./hr.) was linear for 3hr. and unaltered by luteinizing hormone treatment or addition of glucose (1mg./ml.) to the medium. 2. In slices from control animals, steady-state rate of glucose uptake was 78·0±2·9μg. atoms of carbon/g. wet wt./hr.; steady-state rates of lactate output, pyruvate output and incorporation of [U−14C]-glucose carbon atoms into carbon dioxide and total lipid extract were 60·7±0·9, 2·4±0·1, 18·0±1·1 and 0·7±0·1μg. atom of carbon/g. wet wt./hr. and accounted for 104·5±1·9% of the glucose uptake. In slices from luteinizing hormone-treated rats, glucose uptake and outputs of lactate, pyruvate and [14C]carbon dioxide were increased by approx. 25%, and 108·4±3·2% of the glucose uptake could be accounted for. 3. The total lipid extract was separated by thin-layer chromatography and saponification. Of the 14C incorporated into this fraction during incubation with [U−14C]glucose 97% was found in the fractions containing glyceride glycerol and less than 3% in the fractions containing sterols, steroids or fatty acids. Appreciable quantities of 14C were incorporated into these lipid fractions from [1−14C]acetate. 4. From a consideration of the tissue glycogen content, the specific activities of [14C]lactate and glucose 6-phosphate (C-1) derived from [1−14C]-, [6−14C]- and [U−14C]-glucose, and the ratio of [14C]carbon dioxide yields from [1−14C]glucose and [6−14C]glucose, it was concluded that there was no appreciable glycogenolysis or flow through the pentose phosphate cycle. 5. In ovary slices from both control and luteinizing hormone-treated animals, glucose in vitro raised the incorporation rate of 14C from [1−14C]acetate into sterols and steroids. Luteinizing hormone in vivo stimulated the incorporation rate in vitro but only in the presence of glucose. 6. In slices incubated in medium containing [3H]water, [14C]sorbitol and glucose (1mg./ml.), the total water space (865±7·1μl./g.) and the extracellular water space (581±22μl./g.) were unchanged by luteinizing hormone treatment in vivo but the glucose space was raised from 540±23·6μl./g. to 639±31·3μl./g. 7. Luteinizing hormone treatment was found to lower the tissue concentration of the hexose monophosphates and to increase the total activity of hexokinase, glucose 6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase and possibly of phosphofructokinase. 8. The kinetic properties of a partially purified preparation of phosphofructokinase were found to be qualitatively similar to those from other mammalian tissues. 9. The results are discussed with reference to both the role of glucose metabolism in steroidogenesis and the mechanism by which luteinizing hormone increases the rate of glucose uptake.