The enzyme, dopanime Beta-hydroxylase (DBH), normally found only on the inner surface of the secretory granules of chromaffin cells, has been shown by immunofluorescence to be present on the surface membrane following stimulation of exocytosis by Ba2+. These techniques will be used to determine the time course of the appearance of DBH during stimulation and of its internalization afterwards, as well as how quickly after internalization it can be stimulated to reappear on the surface. Other experiments with video enchanced light microscopy will examine a the low density vesicles which appear on the surface of the cells in the presence of Ba2+, as well as the large vacuoles which take up fluorescent dextran from the solution. The release of chromaffin material by the adrenal medullary cell is considered a model for the release of transmitter by nerve cells, and these experiments should increase our understanding of this process. Although glial cells lack voltage dependent Na+ channels, they take up labelled saxitoxin with a high affinity, and the addition of scorpion toxin and veratridine gives a large increase in Na+ uptake. Since veratradine and scorpion toxin can open and keep open the voltage-dependent Na channel, the results suggest that glial cells may have a Na+ channel that lacks a voltage-dependent gate. Electrophysiological techniques will be used to determine if a non-voltage-dependent gate. Electrophysiological techniques will be used to determine if a non-voltage-dependent Na+ channel is present and, if so, to investigate its ionic selectivity. The characterization of a channel with these properties is of interest to studies of the molecular structure of the Na+ channel. The actions of several agonists and antagonists at the crayfish neuromuscular junction will be investigated. Dose response relationships will be measured for glutamate, quisqualate, aspartate and kainate in the presence of Con A, which blocks desensitization. Other studies will use noise analysis and patch clamp techniques to learn more about the mechanisms of the action of the agonists. These substances have profound effects on many synapses in higher animals, and the results of these experiments should assist in understanding these actions.
