[Insulin receptors in isolated cells of murine lactating mammary gland].
Insulin receptors in isolated cells from lactating mouse mammary gland were investigated by 125I-labeled insulin. A modified chloramin T-method after Hunter and Greenwood (1962) was used for insulin labelling with 125I; 125I-insulin with a high specific activity (80--120 muCi per microgram), high immunoreactivity and preserved biological properties was obtained. Three methods of labeled insulin purification were also studied: absorption on the cellulose column, gel chromatography on different types of Sephadex (G-25, G-75, G-100) and electrophoresis on polyacrylamide gel. The best conditons for iodination and purification of 125I-insulin suitable for studying the receptor-insulin binding in the isolated cells from mouse mammary gland were selected. Insulin binding to its receptor was found to be a specific reversible process, having high affinity and a tendency to saturation. The receptor-insulin-binding equilibrium was reached in 30 min at 24 degrees C. Specific binding was observed at a very low concentration of 125I-insulin (0.4.10(-9)M), close to the physiological level. Saturation was observed at a concentration of over 1.5.10(-9)M. Native, non-labeled insulin, at a concentration of 1 ng per ml lowered the labeled insulin binding by 10--20 per cent, whereas 10 ng per ml led to a 60 per cent lowering. The affinity constant of the process was about 10(9).M-1. Each cell had about 3 000 to 4 000 insulin binding sites.
D000818:Animals; D005260:Female; D007328:Insulin / Q000302:isolation & purification / Q000378:metabolism*; D007457:Iodine Radioisotopes; D007553:Isotope Labeling; D007774:Lactation; D008321:Mammary Glands, Animal / Q000166:cytology*; D051379:Mice; D011247:Pregnancy; D011972:Receptor, Insulin / Q000378:metabolism*
D007328:Insulin; D007457:Iodine Radioisotopes; D011972:Receptor, Insulin
Insulinovye retseptory v izolirovannykh kletkakh laktiruiushcheĭ molochnoĭ zhelezy mysheĭ.