alpha-adrenergic stimulation of cytosolic Ca2+ oscillations and exocytosis in identified rat corticotrophs.
1. The patch clamp technique was used in conjunction with a fluorescent Ca2+ indicator (indo-1, or indo-1FF) to measure simultaneously cytosolic Ca2+ concentration ([Ca2+]i), ionic current and changes in membrane capacitance in single rat corticotrophs identified with the reverse haemolytic plaque assay. 2. Application of the adrenocorticotropin (ACTH) secretagogue noradrenaline (NA; norepinephrine), triggered [Ca2+]i oscillation in corticotrophs via alpha-adrenergic receptors and the guanosine trisphosphate (GTP) binding protein-coupled phosphoinositide pathway. 3. Simultaneous measurement of [Ca2+]i and capacitance shows that exocytosis was triggered during the first cycle of NA-induced [Ca2+]i oscillation and the mean increase in cell membrane surface area was 1.4 +/- 0.3 % (n = 6). 4. When Ca2+ was directly released from the inositol 1,4, 5 trisphosphate (IP3)-sensitive store via flash photolysis of caged IP3, the mean increase in cell surface area was 1.5 +/- 0.5 % (n = 6). Thus, NA-stimulated ACTH secretion in rat corticotrophs is closely coupled to intracellular Ca2+ release. 5. Large and rapid elevation of [Ca2+]i (>15 microM) via flash photolysis of caged Ca2+ triggered two phases of exocytosis: a rapid exocytic burst that was complete in approximately 100 ms and a slow burst that continued for many seconds. 6. The rapid exocytic burst reflected the exhaustion of a pool of readily releasable granules and, on average, increased the cell surface by 2.8 +/- 0.1 % (n = 14). 7. We suggest that the relatively weak exocytic response in corticotrophs during intracellular Ca2+ release may be partially attributed to a smaller pool of readily releasable granules.
Department of Pharmacology, 9-70 Medical Science Building, University of Alberta, Edmonton, Alberta, Canada T6G 2H7. Amy.Tse@U.Alberta.ca
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D000316:Adrenergic alpha-Agonists / Q000494:pharmacology*; D000324:Adrenocorticotropic Hormone / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects / Q000502:physiology; D003600:Cytosol / Q000187:drug effects / Q000378:metabolism* / Q000528:radiation effects; D004558:Electric Stimulation; D004594:Electrophysiology; D005089:Exocytosis / Q000187:drug effects* / Q000528:radiation effects; D019204:GTP-Binding Proteins / Q000378:metabolism; D066298:In Vitro Techniques; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism; D008297:Male; D008564:Membrane Potentials / Q000502:physiology; D009638:Norepinephrine / Q000494:pharmacology; D018408:Patch-Clamp Techniques; D010782:Photolysis; D010902:Pituitary Gland / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley
D000316:Adrenergic alpha-Agonists; D015544:Inositol 1,4,5-Trisphosphate; D000324:Adrenocorticotropic Hormone; D019204:GTP-Binding Proteins; D002118:Calcium; D009638:Norepinephrine