Quantal secretion of catecholamines measured from individual bovine adrenal medullary cells permeabilized with digitonin.
Secretion of catecholamines from individual bovine adrenal medullary cells grown in primary culture has been investigated with a carbon-fiber microelectrode placed adjacent to the cells. Oxidation of catecholamines at the electrode surface results in changes in current, which give a real-time measure of catecholamine secretion. Chemical agents are introduced to the individual cells by pressure ejection from micropipettes. When incubated in Ca(2+)-containing buffers, secretion is not observed. However, permeabilization of the cell by exposure to 20 microM digitonin for approximately 15 s results in a Ca(2+)-dependent secretion, and the contents of individual vesicles are detected in the form of sharp spikes. The rate at which spikes occur is a function of the Ca2+ concentration in the external media and reaches a maximum at 19 microM Ca2+. The area of the spikes range from 0.1 to greater than 10 picocoulombs, but the majority are less than 2 picocoulombs, corresponding to less than 6 x 10(6) molecules detected per spike. Histograms of the spike areas are essentially independent of the Ca2+ concentration, indicating that the population of vesicles which undergo exocytosis is the same for all concentrations. Exocytotic secretion can be distinguished from nonexocytotic release by analysis of the shape of the spikes.
Jankowski
J A
JA
Department of Chemistry, University of North Carolina, Chapel Hill 27599-3290.
Schroeder
T J
TJ
Holz
R W
RW
Wightman
R M
RM
D000313:Adrenal Medulla / Q000166:cytology / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002395:Catecholamines / Q000378:metabolism*; D002413:Cations, Divalent; D002417:Cattle; D002478:Cells, Cultured; D002837:Chromaffin Granules / Q000378:metabolism; D004072:Digitonin / Q000494:pharmacology*; D004563:Electrochemistry; D005089:Exocytosis; D007150:Immunohistochemistry
D002395:Catecholamines; D002413:Cations, Divalent; D004072:Digitonin; D002118:Calcium