Abstract
Chloride cells occur in both primary and secondary epithelia of SW- and FW-adapted toadfish. These cells are characterized by numerous mitochondria, an extensive basolateral tubular system, optimal positioning between the external environment and vascular space and contain high levels of Na/K-ATPase.Upon isolation of cells from the gill epithelium chloride cells lose their polarity. The tubular system pinches off from the plasma membrane and no longer communicates with the external milieu. Separation of isolated gill cell suspensions on a discontinuous Ficoll gradient results in the formation of a discrete band of cells composed of 50-70% chloride cells. This represents a 7 fold enrichment in the percentage of chloride cells over isolated cell suspension. This enrichment is reflected in the increase in the specific activity of Na/K-ATPase in chloride cell fractions (3-5x higher than isolated cell levels).Ouabain binding to isolated gill cells can be described by a simple bimolecular reaction model. {('3)H}ouabain binding is saturable and demonstrates the presence of a single class of high affinity binding sites. The binding constant (K(,B)) for ouabain binding is 0.25 x 10('-6) M and at equilibrium the binding capacity is 13 pmol/mg cell protein corresponding to 2.5 x 10('5) Na('+) pump sites/cell. Binding is stimulated after 45 min incubation with isoproterenol (20% over control binding levels). Furosemide does not affect control binding or isoproterenol stimulation.Cortisol has been implicated as the seawater adaptation hormone in teleosts. The current mechanism for steroid hormone action requires the presence of specific, high affinity cytoplasmic receptor proteins. Chloride cells specifically bind {('3)H}dexamethasone (K(,d) = 10('-9) M) in both whole and broken cell assays. The occurrence of specific glucocorticoid receptors in chloride cells implies a direct effect of the hormone on Na/K-ATPase activity at the level of the genome.