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Intracellular voltage recordings in bovine non-pigmented ciliary epithelial cells in primary culture

1989; Taylor & Francis; Volume: 8; Issue: 8 Linguagem: Inglês

10.3109/02713688909000869

1460-2202

Horst Helbig, Christoph Korbmacher, J Wohlfarth, Miguel Coca‐Prados, Michael Wiederholt,

Planarian Biology and Electrostimulation

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 stil-bene-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.