Capacitance increases of dissociated tilapia prolactin cells in response to hyposmotic and depolarizing stimuli

Abstract

Prolactin (PRL) is the major hormonal mediator of adaptation to hyposmotic conditions. In tilapia ( Oreochromis mossambicus), PRL cells are segregated to the rostral pars distalis of the anterior pituitary facilitating the nearly pure culture of dissociated PRL cells. Membrane capacitance ( C m) was recorded at 1 Hz or higher for tens of minutes as a surrogate monitor of PRL secretion by exocytosis from cells under perforated patch clamp. The study compares secretory responses to trains of depolarizing clamps (100 at 2.5 Hz, from −70 to +10 mV for 100 ms) to the physiological stimulus, exposure to hyposmotic medium, here a switch from 350 to 300 mOsm saline ([Ca 2+] 15 mM). Two-thirds of cells tested with each stimulus responded. In response to depolarizing clamps, C m increased linearly at an average rate of 7.2 fF/s. The increase was also linear in response to hyposmotic perfusion, but the average rate was 0.68 fF/s. Response to depolarization was reversibly blocked in Ca 2+-omitted saline, or in saline with 30 μM Cd 2+. It was unaffected by 0.1 μM tetrodotoxin. By contrast, responses were reduced but not absent during perfusion of hyposmotic saline with Ca 2+-omitted; 30 μM Cd 2+ appeared to enhance the hyposmotic response. BAPTA-AM eliminated responses to both stimuli, confirming that secretion was dependent on increases of intracellular [Ca 2+]. Together with previous observations from this laboratory of [Ca 2+] i with simultaneous collection and immunoassay of perfusate for PRL, we conclude that depolarization and hyposmotic stimuli initiate secretion by independent mechanisms.