Heterogeneity of the Ca2+ sensitivity of secretion in a pituitary gonadotrope cell line and its modulation by protein kinase C and Ca2+.
Modulation of the Ca2+ sensitivity and cooperativity of secretion is an important means of regulating neurotransmission and hormone secretion. Employing high-time resolution measurement of membrane capacitance (Cm) stimulated by step-like or ramp [Ca2+]i elevation, we have identified the co-existence of both a high and low Ca2+-sensitive exocytosis in an immortal pituitary gonadotrope cell line, LbetaT2. Ramp [Ca2+]i generated by slow uncaging elicited a biphasic C(m) response. The first phase of response, which represents a highly Ca2+-sensitive pool (HCSP) of vesicles, began to secrete at low [Ca2+]i concentration (<1 microM) with low Ca2+ cooperativity. In contrast, the second phase, which represents a lowly Ca2+-sensitive pool (LCSP) of vesicles, only exocytozed at higher [Ca2+]i (>5 microM) and displayed a steep Ca2+ cooperativity. The co-existence of vesicle populations with different Ca2+ sensitivities was further confirmed by flash photolysis stimuli. The size of the HCSP was approximately 30 fF under resting conditions, but was dramatically increased (approximately threefold) by application of phorbol-12-myristate-13-acetate (PMA, an activator of protein kinase C). Forskolin (an activator of protein kinase A), however, exerted no significant effect on the size of both HCSP and LCSP. GnRH (gonadotropin releasing hormone) augmented the size of both pools to a larger extent (5- and 1.7-fold increase for HCSP and LCSP, respectively). The heterogeneity of Ca2+ sensitivity from different pools of vesicles and its differential modulation by intracellular signals suggests that LbetaT2 cells are an ideal model to further unravel the mechanism underlying the modulation of Ca2+-sensing machineries for exocytosis.
Joint Laboratory of Institute of Biophysics and Huazhong University of Science and Technology, Huazhong University of Science and Technology, Wuhan, PR China.
Hui-Sheng
HS
Liu
Zhi-Tao
ZT
Hu
Ke-Ming
KM
Zhou
Ya-Ming
YM
Jiu
Hua
H
Yang
Zheng-Xing
ZX
Wu
Tao
T
Xu
D000818:Animals; D002118:Calcium / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D002413:Cations, Divalent / Q000737:chemistry; D002460:Cell Line; D005089:Exocytosis / Q000187:drug effects; D007987:Gonadotropin-Releasing Hormone / Q000494:pharmacology; D007700:Kinetics; D051379:Mice; D010902:Pituitary Gland / Q000187:drug effects / Q000201:enzymology* / Q000378:metabolism*; D011493:Protein Kinase C / Q000378:metabolism*
D002413:Cations, Divalent; D007987:Gonadotropin-Releasing Hormone; D011493:Protein Kinase C; D002118:Calcium