Ca2+ clearance mechanisms in neurohypophysial terminals of the rat.
The importance of intracellular calcium ([Ca2+]i) in the release of vasopressin (AVP) and oxytocin from the central nervous system neurohypopyhysial nerve terminals has been well-documented. To date, there is no clear understanding of Ca2+ clearance mechanisms and their interplay with transmembrane Ca2+ entry, intracellular [Ca2+]i transients, cytoplasmic Ca2+ stores and hence the release of AVP at the level of a single nerve terminal. Here, we studied the mechanism of Ca2+ clearance in freshly isolated nerve terminals of the rat neurohypophysis using Fura-2 Ca2+ imaging and measured the release of AVP by radioimmuno assay. An increase in the K+ concentration in the perfusion solution from 5 to 50 mM caused a rapid increase in [Ca2+]i and AVP release. Returning K+ concentration to 5 mM led to rapid restoration of both responses to basal level. The K+-evoked [Ca2+]i and AVP increase was concentration-dependent, reliable, and remained of constant amplitude and time course upon successive applications. Extracellular Ca2+ removal completely abolished the K+-evoked responses. The recovery phase was not affected upon replacement of NaCl with sucrose or drugs known to act on intracellular Ca2+ stores such as thapsigargin, cyclopiazonic acid, caffeine or a combination of caffeine and ryanodine did not affect either resting or K+-evoked [Ca2+]i or AVP release. By contrast, the plasma membrane Ca2+ pump inhibitor, La3+, markedly slowed down the recovery phase. The mitochondrial respiration uncoupler, carbonyl cyanide 3-chlorophenylhydrazone (CCCP), slightly but significantly increased the basal [Ca2+]i, and also slowed down the recovery phase of both [Ca2+]i and release responses. In conclusion, we show in nerve terminals that (i) Ca2+ extrusion through the Ca2+ pump in the plasma membrane plays a major role in the Ca2+ clearance mechanisms of (ii) Ca2+ uptake by mitochondria also contributes to the Ca2+ clearance and (iii) neither Na+/Ca2+ exchangers nor Ca2+ stores are involved in the Ca2+ clearance or in the maintenance of basal [Ca2+]i or release of AVP.
Sasaki
Naoko
N
Department of Physiology, University of Occupational and Environmental Health School of Medicine, Kitakyushu 807-8555, Japan.
Dayanithi
Govindan
G
Shibuya
Izumi
I
D000818:Animals; D001127:Arginine Vasopressin / Q000378:metabolism; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects / Q000502:physiology*; D000252:Calcium-Transporting ATPases / Q000187:drug effects / Q000378:metabolism; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone / Q000031:analogs & derivatives* / Q000494:pharmacology; D002462:Cell Membrane / Q000378:metabolism; D019069:Cell Respiration / Q000187:drug effects / Q000502:physiology; D003593:Cytoplasm / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D016257:Fura-2; D007811:Lanthanum / Q000494:pharmacology; D008297:Male; D008657:Metabolic Clearance Rate / Q000502:physiology; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D010904:Pituitary Gland, Posterior / Q000187:drug effects / Q000378:metabolism*; D011188:Potassium / Q000378:metabolism / Q000494:pharmacology; D017729:Presynaptic Terminals / Q000187:drug effects / Q000378:metabolism*; D011863:Radioimmunoassay; D051381:Rats; D017208:Rats, Wistar; D013574:Synaptosomes
D001127:Arginine Vasopressin; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone; D007811:Lanthanum; D000252:Calcium-Transporting ATPases; D011188:Potassium; D002118:Calcium; D016257:Fura-2