ACTIVATION OF PROTEIN-KINASE-C INCREASES CA-2+ SENSITIVITY OF SECRETORY RESPONSE OF GH3 PITUITARY-CELLS

The effect of protein kinase C on the secretory response of GH-3 pituitary cells to Ca2+ was investigated. Activation of protein kinase C with 100 nM 12-0-tetradecanoylphorbol 13-acetate (TPA) for 40 min reduced the rise in intracellular free calcium concentration ([Ca2+]i) stimulated by depolarization with high K+ but did not affect the three-fold increase in prolactin secretion stimulated by 50 mM K+. Both [Ca2+]i and prolactin release were measured for control and TPA-treated cells over a range of [Ca2+]i values attained by adding the acetoxymethyl ester of 1,2-bis(2-aminophenoxy)-ethane -N,N,N',N'-tetraacetic acid (BAPTA/AM) to reduce [Ca2+]i or high K+ with or without BAY K 8644 to increase [Ca2+]i. Half-maximal prolactin secretion occurred at lower [Ca2+]i concentrations for cells treated with TPA (approximately 160 nM) than for control cells (approximately 270 nM), but the rate of secretion at high [Ca2+], was the same. GH3 cells also secreted more prolactin in response to thyrotropin-releasing hormone (TRH) after protein kinase C activation, although TRH evoked a smaller Ca2+ transient. Fluorescence ratio imaging revealed that GH3 cells undergo spontaneous [Ca2+]i oscillations (4-12/min) and that TPA nearly abolishes [Ca2+]i oscillations as well as inhibits the increase in [Ca2+]i stimulated by depolarization. These results demonstrate that activation of protein kinase C increases the Ca2+ sensitivity of the secretory response in GH3 cells, causing up to a twofold increase in the rate of secretion at typical intracellular Ca2+ concentrations.