Insulin-Induced Alterations in Insulin Binding and Insulin Action in Primary Cultures of Rat Hepatocytes

Abstract

We have recently proposed that "down regulation" of the insulin receptor may be one of the many biological responses of a cell to insulin. In an attempt to further explore this hypothesis we have studied insulin action, binding, and degradation in freshly isolated hepatocytes from rats rendered insulin resistant by the administration of dexamethasone, 1.0 mg/kg every other day, for 1 and 4 wk, and in dexa-methasone-treated (0.1 ,uM for 24 h) primary cultures of hepatocytes from normal rats. Dexamethasone treatment for 1 and 4 wk resulted in significant hyperinsulinemia and euglycemia when compared with age-and weight-matched control animals. Freshly isolated hepatocytes from rats treated with dexamethasone for 1 wk bound less insulin than cells from control animals. This decrease in insulin binding was reflected in a decrease in the total number of cellular insulin receptors upon solubilization of the cells. Insulin action was evaluated by the ability of insulin to stimulate the uptake of a-aminoisobutyric acid. The basal rate of aminoisobutyrate uptake in freshly isolated hepatocytes was enhanced by 1 wk of dexamethasone treatment, and although there was an apparent shift to the right in the dose-response curve for insulin-stimulated aminoisobutyrate uptake, at no insulin concentration was there a significant difference in the uptake by hepatocytes from control and dexa-methasone-treated animals. This was true whether expressed as a percentage or absolute increment above basal. Insulin degradation was enhanced in hepato-cytes from animals treated with dexamethasone for 1 wk but could not account for the observed changes in insulin binding. Hepatocytes from animals treated with dexameth-asone for 4 wk were resistant to insulin with regard to aminoisobutyrate uptake, yet both insulin binding and insulin degradation returned to the levels observed in hepatocytes from control animals. Primary cultures of hepatocytes from normal rats exposed to dexamethasone, 0.1 ,uM, in vitro for 24 h were similar to hepatocytes from rats treated with dexamethasone for 4 wk in that they were insulin resistant with regard to aminoisobutyrate uptake and had normal to increased insulin binding. Insulin degradation was also similar. These cells were resistant to the ability of insulin, 0.1 ,uM, to down regulate its receptor whereas parallel cultures treated with insulin in the absence of dexamethasone had a 52% decrease in insulin binding. These data indicate that hepatocytes that are insulin responsive respond to in vivo hyperinsulinemia by a decrease in the number of insulin receptors and by increased insulin degradation. Hepatocytes rendered resistant to insulin both in vivo and in vitro are resistant to these effects of insulin. These studies emphasize the importance of postbinding events in the modulation of insulin binding, action, and degradation, and support the hypothesis that down regulation of the hepatocyte insulin receptor is one of the many biological actions of insulin. They also help explain how a cell can be insulin re...

Pathogenesis of Selective Insulin Resistance in Isolated Hepatocytes

“…Conversely, insulin-stimulated de novo lipogenesis was unchanged relative to vehicle treatment (Fig. 2D), consistent with previous data in rat hepatocytes (11). Moreover, CHI-treated cells showed a non-significant trend toward increased rates of DNL in the basal and insulin-stimulated states relative to control cells.…”

“…Although previous studies investigating expression of lipogenic genes in primary hepatocytes suggested this possibility, these studies did not measure DNL directly (10,32,33,57). On the other hand, older studies that also provide evidence that DNL continues unabated in the face of CHI did not correlate these data with lipogenic gene expression or compare them to GP (8,11,16).…”

“…The kinetics of CHI-induced InsR down-regulation in cultured hepatocytes is well described (8,11,14,16,24). Incubation of primary hepatocytes with 100 nM insulin (or lower concentrations) (24) recapitulates the effects of CHI, by decreasing InsR number by ϳ50% in 24 h for up to 72 h (16).…”

“…That is, the ED 50 of insulin signaling required to blunt glucose production (GP) is lower than that required to stimulate de novo lipogenesis (DNL), such that IR overcomes the former more quickly than it does the latter. The primary cause of this IR may be compensatory CHI itself, which is known to dampen the insulin response in target cells (7)(8)(9)(10)(11). CHI that arises as an early manifestation of insulin resistance may trigger partial down-regulation of InsR in liver (12,13), leading to a situation in which suppression of HGP is achieved at the price of stimulating DNL.…”

Long-term effect of insulin on glucose transport and insulin binding in cultured adipocytes from normal and obese humans with and without non-insulin-dependent diabetes.

“…Thus, it is likely that insulin sensitizes the insulin-resistant adipocytes. Whereas, it desensitizes the normal adipocytes and the evidence to support this contention is overwhelming in various animal cell models (17)(18)(19)(20)(21)(22)(23)(24). To test these hypotheses directly, we have developed a primary culture system for insulin-responsive human adipocytes from control and morbidly obese subjects with or without NIDDM.…”