The contributions of plasma membrane Na+-Ca2+-exchange and the Ca2+-ATPase to the regulation of cytosolic calcium ([Ca2+]i) in a clonal pituitary cell line (AtT-20) of mouse corticotropes

• Published in: Life sciences (1973) Vol. 70; no. 6; pp. 681 - 698
• Main Author: Fiekers, J F
• Format: Journal Article
• Language: English
• Published: Netherlands 28.12.2001
• Subjects: Animals; Calcium-Transporting ATPases - antagonists & inhibitors; Calcium-Transporting ATPases - metabolism; Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone - pharmacology; Cell Line; Cell Membrane - enzymology; Clone Cells; Enzyme Inhibitors - pharmacology; Fura-2 - pharmacology; Hydroquinones - pharmacology; Image Processing, Computer-Assisted; Ionomycin - pharmacology; Lithium - pharmacology; Meglumine - pharmacology; Meglumine Antimoniate; Mice; Organometallic Compounds - pharmacology; Organophosphates - pharmacology; Pituitary Gland - cytology; Pituitary Gland - drug effects; Pituitary Gland - enzymology; Potassium Chloride - pharmacology; Signal Transduction; Sodium-Calcium Exchanger - antagonists & inhibitors; Sodium-Calcium Exchanger - metabolism; Tetraethylammonium - pharmacology; Thapsigargin - pharmacology
• ISSN: 0024-3205
• DOI: 10.1016/S0024-3205(01)01443-6

Single cell calcium microfluorimetry was used to examine the regulation of [Ca2+]i homeostasis in a clonal cell line of corticotropes (AtT-20 cells). Single cells, loaded with fura-2/AM, were exposed briefly to elevated potassium chloride (KCI, 40 mM, 5 sec). The time constant of decay of the [Ca2+]i signal was used as an index of [Ca2+]i extrusion and/or sequestration. Substitution of extracellular sodium with lithium, N-methyl-D-glucamine (NMDG), or Tris, increased resting levels of [Ca2+]i and significantly increased the time constant of [Ca2+]i decay by 40% compared to control indicating the participation of Na+-Ca2+-exchange. Prior exposure of single cells to thapsigargin (1 microM) or BuBHQ (10 microM). inhibitors of the SERCA Ca2+-ATPases, and/or the mitochondrial uncoupler FCCP (1 microM) did not significantly change the time constant of [Ca2+]i decay following KCl. Lanthanum ions (La3+), applied during the decay of the KCI-induced increase in [Ca2+]i, significantly increased the time constant of the return of [Ca2+]i to resting levels by 70% compared to control. Brief exposure of cells to sodium orthovanadate, an inhibitor of ATP-dependent pump activity, slowed and longer exposures prevented, the return of [Ca2+]i to resting levels. We conclude that neither intracellular SERCA pumps nor mitochondrial uptake contribute significantly to [Ca2+]i sequestration following a [Ca2+]i load and that the plasma membrane Ca2+-ATPase contributes to a greater extent than the Na+-Ca2+-exchanger to the return of [Ca2+]i to resting levels following a [Ca2+]i load under these experimental conditions.