Effects of insulin and phospholipase a on glucose transport across the plasma membrane of free adipose cells

Abstract

Evidence is presented that d-glucose enters the basement membrane-free, isolated rat adipose cell by a facilitated transport mechanism which was stimulated to similar degrees by low concentrations of insulin and Naja naja snake venom. The available evidence permits the provisional conclusion that the snake venom factor responsible for the effects on glucose transport is phospholipase A (phosphatide acylhydrolase, EC 3.1.1.4). Evidence is also presented that insulin and phospholipase A stimulated the same rate-limiting step in the overall process of glucose entry into adipose cells. Relatively high concentrations of Naja venom phospholipase A drastically inhibited glucose utilization by free adipose cells, and cells so treated were insensitive to insulin. The effects on glucose utilization of both low and high concentrations of the phospholipase were modified by exogenous phosphatidyl serine and choline. Evidence was obtained for and against the suggestion that the effects of phospholipase A on facilitated transport of glucose may be due to hydrolysis of plasma membrane phospholipids. Phospholipase A stimulated facilitated transport; stimulations due to the enzyme were partially reversed by venom antiserum, and such cells remained sensitive to insulin ; stimulations due to insulin were partially reversed by anti-insulin serum, and such cells remained sensitive to phospholipase A ; and, semiquantitative Chromatographic analysis of adipose cell phospholipids revealed no obvious increase in the amount of lysophosphatidyl choline following incubation of cells with low concentrations of phospholipase A. On the other hand, low concentrations of exogenous lysophosphatidyl choline and ethanolamine stimulated, and higher concentrations inhibited, glucose utilization by adipose cells; calcium ions were required for the effects of phospholipase A on adipose cells ; and kinetic analysis of the stimulatory effects of phospholipase A on glucose transport suggested preliminary enzymatic catalytic activity. Insulin, itself, was devoid of phospholipase A-like activity with phospholipid substrates. However, the stimulation of glucose transport by one preparation of highly active, glucagon-free insulin exhibited kinetics resembling those seen with phospholipase A.