Morphology and secretory activity of digitonin- and α-toxin-permeabilized chromaffin cells

Abstract

An ultrastructural examination of cultured bovine chromaffin cells permeabilized with Staphylococcus aureus α-toxin or digitonin revealed differences in the preservation of cell morphology. The toxin-treated cells closely resembled control cultured cells whereas digitonin-treated cells showed gradations in cytoplasmic densities suggesting extraction, some swelling of the endoplasmic reticulum and, occasionally, discontinuities in the plasma membrane and free granules in the extracellular medium. In both cell models, there was a swelling of the mitochondria. Horseradish peroxidase labelling of permeabilized cells marked the cytoplasm of digitonin-treated cells but only the surface of toxin-treated cells, demonstrating that larger lesions were caused by digitonin. In stimulated cells, the decrease in volumetric density of chromaffin granules correlated well with catecholamine release. The sites of secretory activity could be demonstrated in toxin-treated cells using horseradish peroxidase as a surface marker. Although both cell systems secrete catecholamines in response to calcium stimulation, their calcium requirements and the kinetics of release were different. In α-toxin-treated cells, 100μ free calcium induced maximal catecholamine release. In digitonin-treated cells, 20 μM evoked maximal release but secretion was blocked at 100μM. Catecholamine release terminated in digitonin-treated cells within 10 min but continued in at-toxin-treated cells for at least 60 min. In addition, the maximal release observed in toxin-treated cells (50%) was always greater than that observed in digitonin-permeabilized cells (20%). The results suggest that both exocytosis and granule translocation are operational in α-toxin-treated cells, but that the translocation step or the docking of granules at the plasma membrane may be impaired in digitonin-treated cells.