Permeabilization reveals classical stimulus-secretion coupling in bovine parathyroid cells.
Unlike most secretory cells, high extracellular Ca2+ inhibits, rather than stimulates, PTH release from parathyroid cells. To gain more direct access to the potentially unique secretory apparatus of this cell type, we developed techniques for permeabilizing parathyroid cells using electroshock. Exposure of dispersed bovine parathyroid cells to five 2-kV discharges results in more than 90% uptake of trypan blue and rapid loss of intracellular 86Rb, documenting permeabilization of the plasma membrane. Unlike intact parathyroid cells, a high Ca2+ concentration increases PTH release 2.8-fold from permeabilized cells, with half-maximal and maximal stimulation at 10(-5) and 4 X 10(-4) M Ca2+, respectively, a pattern similar to classical stimulus-secretion coupling. Moreover, PTH release was low at Ca2+ concentrations equivalent to those in intact cells at low extracellular Ca2+ levels (approximately 2 X 10(-7) M). Cellular mechanisms other than changes in the cytosolic Ca2+ concentration per se, therefore, presumably mediate low Ca2+-stimulated PTH release in the intact cell. Diacylglycerol and protein kinase C may play a role in this process, since dioctanoylglycerol (10(-4) M) and 12-O-tetradecanoyl phorbol 13-acetate (10(-6) M) each enhance PTH release from permeabilized cells 2- to 4-fold at a Ca2+ concentration (approximately 2 X 10(-7) M) equivalent to that present in the intact cell at low extracellular Ca2+ concentrations.
Oetting
M
M
Endocrine-Hypertension Unit, Duke University Medical School, Durham, North Carolina 27710.
LeBoff
M S
MS
Levy
S
S
Swiston
L
L
Preston
J
J
Chen
C
C
Brown
E M
EM
D000818:Animals; D002118:Calcium / Q000494:pharmacology; D002417:Cattle; D002463:Cell Membrane Permeability; D000242:Cyclic AMP / Q000378:metabolism / Q000494:pharmacology; D004558:Electric Stimulation; D004789:Enzyme Activation; D019204:GTP-Binding Proteins / Q000502:physiology; D010280:Parathyroid Glands / Q000166:cytology / Q000378:metabolism*; D010281:Parathyroid Hormone / Q000378:metabolism; D011493:Protein Kinase C / Q000378:metabolism
D010281:Parathyroid Hormone; D000242:Cyclic AMP; D011493:Protein Kinase C; D019204:GTP-Binding Proteins; D002118:Calcium