SYNTHESIS, INSERTION INTO THE PLASMA-MEMBRANE, AND TURNOVER OF ALPHA-BUNGAROTOXIN RECEPTORS IN CHICK SYMPATHETIC NEURONS

.alpha.-Bungarotoxin was used to identify an integral membrane protein in the plasma membrane of chick sympathetic neurons [from 11 to 14 day old embryos]. The synthesis, insertion into the plasma membrane, and turnover of the .alpha.-bungarotoxin receptor were studied using isotopically labeled amino acids (2H, 13C, 15N) to directly label receptor molecules. Neurons incubated in medium containing dense amino acids continued to insert unlabeled receptors from a pool of previously synthesized molecules for 2 h. Density-labeled receptors began to appear in the plasma membrane after this 2 h period. Synthesis of receptors, but not insertion into the surface, was blocked by cycloheximide (100 .mu.g/ml). Neither colchicine (0.05 .mu.g/ml) nor actinomycin D (5 .mu.g/ml) has any effect on .alpha.-bungarotoxin receptors synthesis or insertion. Autoradiographic studies revealed that receptors occur on growth cones, axons and cell bodies of single neurons and explanted ganglia. The rate of insertion of newly synthesized receptors into the plasma membrane of axons extending from explanted sympathetic ganglia was approximately the same as that into the cell body portion of the ganglion. Cytochalasin B (2 .mu.g/ml) rapidly disrupted growth cones but had no effect on receptor insertion. The growth cone is probably not the sole or even the primary site for insertion of this membrane protein. The kinetics of turnover of the .alpha.-bungarotoxin receptor were a 1st order exponential with t1/2 = 11 h. Neurons that had their surface receptors labeled with 125I-.alpha.-bungarotoxin produced [125I]iodotyrosine. This process was inhibited by low temperature (23.degree. C) and also by a metabolic inhibitor. This is interpreted as evidence that receptors turn over by a mechanism in which they are internalized and then proteolytically degraded.