Insulin-Dependent Apparent Translocation of Glucose Transport Activity: Studies by the Reconstitution Method

Abstract

Nearly a decade ago, we found that when rat epididymal adipocytes were exposed to [¹²⁵I]iodoinsulin for 10 min at 37°C, ¹²⁵I activity would associate with two subcellular structures that could be fractionated by sucrose density-gradient centrifugation into the plasma membrane fraction (peak 1) and a low-density microsomal fraction (peak 2).⁽¹⁾ The latter peak (peak 2 of ¹²⁵I activity) was later identified as the peak of the internalized hormone.^(2,3) During this study, however, we noticed several similarities between the characteristics of the peak 2 formation, i,e., the endocytotic internalization of the hormone-receptor complex, and those of the hormonal action on glucose transport. Thus, both reactions are very slow at a low temperature, such as 15°C,⁽²⁾ both reactions require adenosine triphosphate (ATP) not only for their development^(2,3) but also for their reversal,^(4,5) and both reactions are completed in approximately 5–10 min at 37°C when the hormone concentration is 1 nM.^(2,6) We therefore postulated as a working hypothesis that the complex of glucose and its transport carrier might be co-internalized along with the insulin-receptor complex. In theory, one can test this working hypothesis by measuring the distribution of glucose transport activity in subcellular fractions obtained from the basal and plus-insulin states of adipocytes using the aforementioned sucrose density-gradient centrifugation.