Intracellular voltage recordings in bovine non-pigmented ciliary epithelial cells in primary culture.
Bovine non-pigmented ciliary epithelial cells (NPE) have been isolated by a technique of selective adhesion to tissue culture plastic. NPE cells in primary culture proliferated and maintained epithelial-like morphology for about 4 weeks in tissue culture medium containing 10% fetal calf serum. If grown for longer than 4 weeks in serum-containing medium, cells changed their morphology and became elongated and spindle-shaped. Membrane potentials were measured using conventional microelectrodes. In NPE cells of epithelial-like shape, replacing extracellular Na+ induced a transient hyperpolarization of the membrane potential, while in elongated cells of spindle-shaped morphology an immediate depolarization was observed. We therefore only used epithelial-like NPE for further experiments. In these cells the mean membrane potential was -40.3 +/- 0.5 mV (n = 36). Relative K+ conductance was increased by extracellular alkalinization. Removing extracellular K+ led to a depolarization and readdition of K+ to K+ depleted cells resulted in a hyperpolarization. Both voltage responses were sensitive to ouabain, indicating that Na+/K+ ATPase is inhibited by K+ replacement, and that there is overshoot-activation of the pump when K+ is readded. Extracellular Cl- replacement led to a DIDS sensitive, transient depolarization, which is compatible with a stilbene-sensitive Cl(-)-conductance. Removing HCO3- led to a Na+ dependent and DIDS-sensitive depolarization. However, the electrical response on replacement of extracellular Na+ was not influenced by DIDS or the extracellular HCO3(-)-concentration.
Helbig
H
H
Institut für Klinische Physiologie, Klinikum Steglitz, Berlin, FRG.
Korbmacher
C
C
Wohlfarth
J
J
Coca-Prados
M
M
Wiederholt
M
M
D000109:Acetylcholine / Q000494:pharmacology; D000818:Animals; D001639:Bicarbonates / Q000494:pharmacology; D002417:Cattle; D002478:Cells, Cultured; D002924:Ciliary Body / Q000502:physiology*; D004847:Epithelial Cells; D006863:Hydrogen-Ion Concentration; D008564:Membrane Potentials; D011188:Potassium / Q000494:pharmacology; D012964:Sodium / Q000494:pharmacology
D001639:Bicarbonates; D012964:Sodium; D000109:Acetylcholine; D011188:Potassium