The purpose of this study was to elucidate some of the mechanisms of control of the glucose transport step in adipose tissue. Glucose transport was studied by monitoring the conversion of [1-14C]glucose to 14CO2 in a system where glucose transport was made rate limiting by increasing the flux through the pentose phosphate pathway with phenazine methosulphate, an agent which results in rapid rates of reoxidation of NADPH. The maximum velocity for the apparent rate of glucose transport was increased significantly by insulin. There was no change in the glucose concentration required for half-maximal rates of 14CO2 production. Glucose transport was also monitored by directly measuring the rate of glucose uptake. Glucose uptake was increased by phenazine methosulphate. The intracellular glucose-6-phosphate concentration was decreased by phenazine methosulphate. These two agents, insulin and phenazine methosulphate, seemed to act by independent mechanisms as their optimal effects on glucose uptake were additive. The apparent rate of glucose transport was decreased by ATP which resulted in a decrease in maximal velocity but did not affect the affinity for glucose. This effect of ATP was seen in the presence of absence of insulin.

An indirect method for obtaining a reliable measure of the rate of glucose transport into adipocytes is described. Evidence is presented that altered levels of 3':5'-cyclic AMP can influence the transport of glucose into adipocytes. When cyclic AMP levels were lowered with antilipolytic agents (insulin, nicotinic acid, or clofibrate), rates of glucose transport were increased. In contrast, when adipose tissue levels of cyclic AMP were elevated by lipolytic hormones or theophylline, glucose transport was inhibited. Insulin no longer increased glucose transport when cyclic AMP levels were elevated by lipolytic agents. Agents that can raise cyclic AMP but inhibit lipolysis (procaine, amitryptyline, and phenylethylbiguanide) reduced the rate of glucose transport. Other data are presented that are consistent with the conclusion that cyclic AMP inhibits glucose transport into adipocytes.The relatively small intracellular water volume in adipose tissue makes it technically difficult to measure rates of glucose transport into adipocytes (1). This rate is usually determined by measuring the volume of distribution of added glucose or derivatives in cells rapidly separated from the incubation medium or by measuring countertransport from prelabeled adipocytes (1-9). However, these methods suffer from the disadvantage that incubation is required at reduced temperature or for very short time intervals. The magnitude of the response of the glucose transport system in adipocytes to insulin as observable by previously used methods is small compared to the observed increases in rates of glucose metabolism. Consequently, a considerable portion of information relating to glucose transport in adipose tissue has been inferred from indirect techniques in which glucose utilization has been measured (10-12). Even though these indirect techniques do not clearly dissociate effects on glucose transport from effects on subsequent glucose metabolism, it is generally accepted that insulin stimulates glucose transport in adipose tissue (for review, see ref. 13).Addition of insulin to adipose tissue results in a fall in levels of 3':5'-cyclic AMP (cAMP) (for review, see refs. 13 and 14). Evidence is lacking, however, that the insulin-induced decrease in cAMP is directly associated with increased glucose transport. The apparent absence of an influence of cAMP on glucose transport is incompatible with the possibility of a unitary concept that would link the major metabolic effects of insulin to its actions on intracellular cAMP levels (15).In this report, we first describe a method to determine the rate of glucose transport in adipocytes. This technique offers several advantages over those previously described in that incubation is permitted at normal temperatures for longer periods. Using this method, we then report observations demonstrating an inverse correlation between adipocyte cAMP levels and the rate of glucose transport. On the basis of these data, we discuss the hypothesis that the major actions of insulin in adipose tissue may ...