Intracellular Ca2+ regulation by the leech giant glial cell.
1. We have measured the intracellular Ca2+ concentration, [Ca2+]i, and the intracellular Na+ concentration, [Na+]i, with the fluorescent dyes fura-2 (for Ca2+) and SBFI (for Na+) in situ in giant glial cells of the central nervous system of the leech Hirudo medicinalis. 2. The basal [Ca2+]i was 79 +/- 35 nM (n = 27) in cells voltage clamped at -70 to -80 mV, and 75 +/- 29 nM (mean +/- S.D., n = 82) in unclamped cells at a mean membrane potential of -67 +/- 6 mV. 3. Removal of external Na+ evoked a small reversible [Ca2+]i increase of 29 +/- 21 nM (n = 27) in cells voltage clamped at -70 to -80 mV, and of 35 +/- 18 nM (n = 37) in unclamped cells. This [Ca2+]i increase, and the time constant of the subsequent [Ca2+]i recovery after Na+ re-addition, did not change significantly with the holding potential between -110 and -60 mV. 4. The basal [Na+]i was 5.6 +/- 1.3 mM (n = 18). Increasing [Na+]i by inhibiting the Na+-K+ pump with 100 microM ouabain had no effect on the [Ca2+]i rise upon removal of external Na+. 5. The time course of recovery from a [Ca2+]i load mediated by voltage-dependent Ca2+ influx during depolarization in high K+ was unaffected by the removal of external Na+. 6. Cyclopiazonic acid (10 muM), an inhibitor of the endoplasmic reticulum Ca2+-ATPase, caused a transient increase in [Ca2+]i of 28 +/- 11 nM (n = 5), and significantly slowed the recovery from imposed [Ca2+]i loads. 7. Iontophoretic injection of orthovanadate, an inhibitor of P-type ATPases including the plasma membrane Ca2+-ATPase, caused a persistent increase in the basal [Ca2+]i of 163 +/- 101 nM (n = 5) in standard saline, and of 427 +/- 338 nM in Na+-free saline (n = 5). Vanadate injection significantly slowed the recovery from [Ca2+]i loads. Removal of external Na+ during vanadate injection induced an additional, reversible [Ca2+]i increase of 254 +/- 64 nM (n = 3). 8. The results suggest that the low basal [Ca2+]i in these glial cells is predominantly maintained by a Ca2+-ATPase in the plasma membrane. This ATPase is also the main Ca2+ extruder after an intracellular Ca2+ load, while intracellular stores appear to contribute little to this recovery. A Na+-Ca2+ exchanger seems to play a minor role in the maintenance of basal [Ca2+]i in these cells, but becomes prominent when the plasma membrane Ca2+-ATPase is blocked.
Nett
W
W
Abteilung fur Allgemeine Zoologie, FB Biologie, Universitat Kaiserslautern, Postfach 3049, D-67653 Kaiserslautern, Germany. wnett@rhrk.uni-kl.de
Deitmer
J W
JW
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors; D048429:Cell Size / Q000502:physiology; D004594:Electrophysiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration / Q000187:drug effects; D007211:Indoles / Q000494:pharmacology; D007425:Intracellular Membranes / Q000378:metabolism*; D007865:Leeches; D009457:Neuroglia / Q000166:cytology / Q000378:metabolism* / Q000502:physiology; D009994:Osmolar Concentration; D012964:Sodium / Q000378:metabolism / Q000494:pharmacology; D019284:Thapsigargin / Q000494:pharmacology; D014638:Vanadates / Q000494:pharmacology
D004791:Enzyme Inhibitors; D007211:Indoles; D014638:Vanadates; D019284:Thapsigargin; D012964:Sodium; D000252:Calcium-Transporting ATPases; D002118:Calcium; C000543:cyclopiazonic acid