EFFECT OF GLUCOSE AND INSULIN DEPRIVATION ON DIFFERENTIATION AND CARBOHYDRATE-METABOLISM OF RABBIT PROXIMAL TUBULAR CELLS IN PRIMARY CULTURE

Rabbit proximal tubule cells in primary culture revert from gluconeoge nesis to glycolysis. To determine whether glucose and insulin deprivat ion of the culture medium could prevent this metabolic conversion with out a loss of differentiation, rabbit proximal tubule cells were cultu red in hormonally defined medium free of glucose and insulin and compa red to rabbit proximal tubule cells cultured in medium supplemented wi th 17.5 mM glucose and 5 mug/ml insulin. In the two culture conditions , RPT cells grew at a similar rate and reached confluency within 4-5 d ays. Patterns of enzyme activity, including brush-border hydrolases, N -acetyl-beta-D-glucosaminidase and glutathione-S-transferases as a fun ction of culture time were comparable in the two media. During the gro wth phase in glucose-and insulin-free medium, cells showed higher sodi um-dependent glucose uptake. Scanning electron microscopy revealed a h igh density of microvilli at confluency regardless of the culture cond itions. In both the presence and absence of glucose and insulin, the a ctivities of gluconeogenic enzymes, phosphoenolpyruvate carboxykinase and fructose-1,6-bisphosphatase, as well as basal and pyruvate-stimula ted glucose production fell markedly as a function of time. By contras t, glucose and insulin deprivation greatly reduced both the lactate pr oduction rate and the activities of glycolytic enzymes, pyruvate kinas e, hexokinase and lactate dehydrogenase.