AJP - Endocrinology and Metabolism, Vol 254, Issue 3 E365-E371, Copyright © 1988 by American Physiological Society
Relationships between cell surface insulin binding and endocytosis in adipocytes
A. L. Jochen
Department of Medicine, Medical College of Wisconsin, Milwaukee 53226.
Chymotrypsin substrate analogues, such as N-acetyl-Tyr ethyl ester, have
recently been demonstrated to inhibit the endocytic uptake of insulin in
isolated rat adipocytes. In this study, the effects of N-acetyl-Tyr ethyl
ester on cell surface insulin binding and dissociation were examined.
Surface-bound 125I-insulin was distinguished from intracellular
125I-insulin by the sensitivity of the former to rapid dissociation with an
acidic buffer (pH 3.0). Plateau levels of surface-bound insulin at 37
degrees C were increased 70% by inhibiting the internalization pathway.
This increase was temperature and insulin concentration dependent. Thus
differences in surface binding were small at 12 degrees C and also at high
(100-200 ng/ml) insulin concentrations. Inhibition of internalization with
N-acetyl-Tyr ethyl ester markedly slowed the loss of surface-bound insulin
observed during dissociation studies. After 20-30 min of dissociation, the
remaining levels of surface-bound insulin were three- to fourfold higher in
treated adipocytes compared with control adipocytes. Added unlabeled
insulin retained its ability to accelerate the dissociation of insulin in
N-acetyl-Tyr ethyl ester-treated cells. These observations indicate that
the internalization pathway is a quantitatively important factor in
determining levels of surface binding at 37 degrees C and in determining
the rate of deactivation of insulin binding.
