Insulin binding in differentiating rat preadipocytes in culture

Binding, degradation, and antilipolytic effect of insulin were studied during the differentiation of preadipocytes into unilocular adipocytes. The precursor cells were isolated from the stromal-vascular fraction of adult rat epididymal fat pads and were cultured according to methods previously described. Under appropriate conditions the cells attained full morphological maturation after 6 days. A gradual increase in insulin binding was found concomitant with the morphological development of the preadipocytes into adipocytes. This increase was due to an enhanced number of binding sites whether expressed per cell or per unit cell surface area. The presence of a high insulin concentration (1.67 micrograms/ml or 278 nM) in the culture medium did not prevent this effect. The receptor density, expressed per unit surface area, was higher in the newly developed univacuolar cells than in mature fat cells from the same rat. The increased receptor density was also reflected by a leftward shift in the dose-response curve for the antilipolytic effect of insulin. In parallel with the increased binding, insulin degradation also increased. The lipolytic response to catecholamine also showed a gradual increase with development. When expressed per unit surface area, newly formed cells exhibited a considerably greater response (approximately 3.4 times) than mature cells from the same animals. The maximal antilipolytic effect of insulin in new cells was of the same order as in old cells when the data were expressed per unit cell surface area. Thus, the data show that developing adipocyte precursors gain membrane properties similar to those of mature fat cells. This cell system may serve as a useful model for studying receptor formation and factors that regulate hormone responsiveness.