α-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 mu M) via flash photolysis of caged Ca2+ triggered two phases of exocytosis: a rapid exocytic burst that was complete in similar to 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.