title: Mechanism underlying corticotropin-releasing hormone (CRH) triggered cytosolic Ca2+ rise in identified rat corticotrophs.
abstract: 1. The patch-clamp technique was used in conjunction with the fluorescent Ca2+ indicator indo-1 to measure simultaneously cytosolic Ca2+ concentration ([Ca2+]i) and membrane potential in single rat corticotrophs identified with the reverse haemolytic plaque assay. 2. Application of the adrenocorticotropin (ACTH) secretagogue, corticotropin-releasing hormone (CRH), triggered a sustained [Ca2+]i elevation and membrane depolarization. 3. The CRH action was mediated via the cAMP-dependent protein kinase cascade. Both the CRH-induced depolarization and [Ca2+]i elevation could be mimicked by extracellular application of the adenylate cyclase activator forskolin or the membrane-permeable cAMP analogue, 8-(4-chlorophenylthio)-adenosine-3',5'-cyclic monophosphate (8-CPT-cAMP). Intracellular adenosine cyclic 3',5'-(Rp)-phosphothioate (Rp-cAMPS), a protein kinase A inhibitor, abolished the CRH effects. 4. Voltage-clamp studies suggest that the CRH-triggered depolarization was due to the reduction of background K+ conductances. The CRH-sensitive current was Ca2+ independent and was insensitive to the K+ channel blockers tetraethylammonium (TEA) or 4-aminopyridine (4-AP), but could be partially inhibited by Ba2+. 5. The CRH-triggered steady-state depolarization stimulated extracellular Ca2+ entry via voltage-gated Ca2+ channels and raised [Ca2+]i. CRH failed to stimulate [Ca2+]i rise in cells that were voltage clamped at their resting potential. Removal of extracellular Ca2+ or inhibition of Ca2+ channels by Ni2+ abolished the [Ca2+]i rise. 6. Voltage-clamp studies of voltage-gated Ca2+ channels using Ba2+ as charge carrier show that approximately 90% of the channels were available for activation at the resting potential. CRH did not enhance the voltage-gated Ca2+ channels.
authors.affiliation: Department of Pharmacology, University of Alberta, Edmonton, Canada.
authors.forename: A K
authors.initials: AK
authors.lastname: Lee
authors.forename: A
authors.initials: A
authors.lastname: Tse
mesh_terms: D000818:Animals; D001464:Barium / Q000493:pharmacokinetics / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D005576:Colforsin / Q000494:pharmacology; D003346:Corticotropin-Releasing Hormone / Q000494:pharmacology*; D000242:Cyclic AMP / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004553:Electric Conductivity; D015640:Ion Channel Gating / Q000502:physiology; D008297:Male; D018408:Patch-Clamp Techniques; D010903:Pituitary Gland, Anterior / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D011188:Potassium / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D019789:Tetraethylammonium / Q000494:pharmacology; D013873:Thionucleotides / Q000494:pharmacology
chemical_list: D013873:Thionucleotides; D005576:Colforsin; C016957:adenosine-3',5'-cyclic phosphorothioate; D001464:Barium; C015929:8-((4-chlorophenyl)thio)cyclic-3',5'-AMP; D019789:Tetraethylammonium; D003346:Corticotropin-Releasing Hormone; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D011188:Potassium; D002118:Calcium