Cytosolic Cl- ions in the regulation of secretory and endocytotic activity in melanotrophs from mouse pituitary tissue slices.
Cl- ions are known regulators of Ca2+ -dependent secretory activity in many endocrine cells. The suggested mechanisms of Cl- action involve the modulation of GTP-binding proteins, voltage-activated calcium channels or maturation of secretory vesicles. We examined the role of cytosolic Cl- ([Cl-]i) and Cl- currents in the regulation of secretory activity in mouse melanotrophs from fresh pituitary tissue slices by using the whole-cell patch-clamp. We confirmed that elevated [Cl-]i augments Ca2- -dependent exocytosis and showed that Cl- acts on secretory vesicle maturation. The latter process was abolished by a V-type H- -ATPase blocker (bafilomycin), intracellular 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid (DIDS), a Cl- channel blocker, and tolbutamide, a sulphonylurea implicated in secretory vesicle maturation. In a small subset of cells, block of plasmalemmal Cl- current by DIDS reversibly enhanced endocytosis. The direct activation of G-proteins by GTP-gamma-S, a non-hydrolysable GTP analogue, did not restore the impaired secretion observed in low [Cl-]i conditions. The amplitude of voltage-activated calcium currents was unaffected by the [Cl-]i. Furthermore, two Cl- -permeable channels, calcium-activated Cl- channels and GABAA receptors, appeared as major regulators of intracellular Cl- homeostasis. In conclusion, the predominant underlying mechanism of Cl- action is mediated by intracellular Cl- fluxes during vesicle maturation, rather than activation of G-proteins or modulation of voltage-activated Ca2+channels.
Turner
Jan-Eric
JE
European Neuroscience Institute-Göttingen, Waldweg 33, 37073 Göttingen, Germany.
Sedej
Simon
S
Rupnik
Marjan
M
D000465:Algorithms; D000818:Animals; D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling / Q000502:physiology*; D002462:Cell Membrane / Q000378:metabolism; D002712:Chlorides / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism*; D004594:Electrophysiology; D004705:Endocytosis / Q000502:physiology*; D005089:Exocytosis / Q000502:physiology; D066298:In Vitro Techniques; D008297:Male; D008543:Melanins / Q000378:metabolism*; D008564:Membrane Potentials / Q000502:physiology; D051379:Mice; D018408:Patch-Clamp Techniques; D010902:Pituitary Gland / Q000166:cytology / Q000378:metabolism / Q000502:physiology*; D011963:Receptors, GABA-A / Q000187:drug effects; D005680:gamma-Aminobutyric Acid / Q000502:physiology
D002712:Chlorides; D008543:Melanins; D011963:Receptors, GABA-A; D005680:gamma-Aminobutyric Acid; D002118:Calcium