Adenosine 3',5'-cyclic monophosphate-mediated enhancement of calcium-evoked prolactin release from electrically permeabilised 7315c tumour cells.
1. The 7315c tumour cell was used as a model system for the investigation of adenosine 3',5'-cyclic monophosphate (cyclic AMP)-mediated enhancement of calcium-evoked prolactin release. 2. 7315c cells were permeabilised by subjecting the cells to intense electric fields. Studies investigating the penetration of the dye ethidium bromide indicated that the cells were completely permeabilised after 2 discharges of 2000 volts and that the pores remained open for at least 30 min before beginning to reseal. These permeabilisation parameters were consistent with those which gave maximal calcium-stimulated prolactin release. 3. In the absence of calcium and in the presence of EGTA (1 mM), permeabilised 7315c cells secreted prolactin at a rate of 0.23 ng min-1 per 10(6) cells. When EGTA was replaced by 1.5 mM calcium, permeabilised cells secreted prolactin at a rate of 2.20 +/- 0.30 ng min-1 per 10(6) cells in the first 5 min of exposure. Maximal calcium-dependent prolactin secretion from permeabilised cells occurred at 37 degrees C. 4. The amount of prolactin secreted, in a 5 min incubation at 37 degrees C, from permeabilised cells depended upon the free calcium concentration in the permeabilisation medium. Calcium stimulated prolactin release from permeabilised cells in the concentration range 0.1-10 microM (half maximal = 5.8 microM). When permeabilised cells were exposed to cyclic AMP (100 microM) for 5 min prior to and during a 5 min challenge with various concentrations of calcium, the amount of prolactin secreted at each effective concentration of calcium was increased. However, cyclic AMP did not alter the potency of calcium as a stimulant of prolactin secretion. 5. The results suggest that cyclic AMP potentiates calcium-evoked secretion from 7315c cells, not by increasing the entry of calcium into the cytosol, but at a step in the secretory process, distal to calcium entry, which modulates the ability of an increase in cytosolic calcium concentration to stimulate prolactin release.
Guild
S
S
Department of Pharmacology, University of Glasgow.
Frey
E A
EA
Pocotte
S L
SL
Kebabian
J W
JW
D000818:Animals; D002118:Calcium / Q000502:physiology*; D002463:Cell Membrane Permeability; D000242:Cyclic AMP / Q000502:physiology*; D004533:Egtazic Acid / Q000494:pharmacology; D004558:Electric Stimulation; D004996:Ethidium; D011388:Prolactin / Q000378:metabolism*; D011863:Radioimmunoassay; D051381:Rats; D014407:Tumor Cells, Cultured
D004533:Egtazic Acid; D011388:Prolactin; D000242:Cyclic AMP; D004996:Ethidium; D002118:Calcium