Na⁺ and Ca²⁺ Homeostatic Mechanisms in Isolated Chloride Cells of the Teleost Oreochromis Mossambicus Analysed by Confocal Laser Scanning Microscopy

Chloride cells (CCs; recognised by the vital mitochondrial stain DASPEI) and pavement cells (PCs) isolated from tilapia opercular epithelium were adhered to Cell-Tak-coated glass coverslips and loaded with fluorescent probes for the measurement of intracellular concentrations of Na⁺ or Ca²⁺. Basal levels of cytosolic Na⁺ and Ca²⁺ ranged from 6.4 to 16.5 mmol l⁻¹ and from 76 to 110 nmol l⁻¹, respectively, and did not differ between CCs and PCs. In CCs, inhibition of Na⁺/K⁺-ATPase by ouabain or Cu²⁺ increased intracellular [Na⁺]. Replacing extracellular Na⁺ with N-methyl-D-glucamine⁺ led to a rise in cytosolic [Ca²⁺] that was dependent on the extracellular [Ca²⁺], indicating that a Na⁺/Ca²⁺ exchanger was operating in reverse mode (importing Ca²⁺). The forward mode of this exchanger could be demonstrated by inhibition with bepridil. The CC has various pathways for passive Na⁺ influx: a tetrodotoxin-sensitive pathway, an amiloride-sensitive pathway and other as yet unidentified pathways.