FISH GILL RESPIRATORY CELLS IN CULTURE - A NEW MODEL FOR CL--SECRETING EPITHELIA

Primary cultures of sea bass gill cells grown on permeable membranes f orm a confluent, polarized, functional tight epithelium as characteriz ed by electron microscopy and electrophysiological and ion transport s tudies. Cultured with normal fetal bovine serum (FBS) and mounted in a n Ussing chamber, the epithelium presents a small short-circuit curren t (I-sc: 1.4 +/- 0.3 mu A/ cm(2)), a transepithelial voltage (V-r) of 12.7 +/- 2.7 mV (serosal positive) and a high transepithelial resistan ce (R(t): 12302 +/- 2477 Omega x cm(2)). A higher degree of differenti ation and increased ion transport capacities are observed with cells c ultured with sea bass serum: numerous, organized microridges character istic of respiratory cells are present on the apical cell surface and there are increased I-sc (11.9 +/- 2.5 mu A/cm(2)) and V-t (25.9 +/- 1 .7 mV) and reduced R(t) (4271 +/- 568 Omega x cm(2)) as compared with FBS-treated cells. Apical amiloride addition (up to 100 mu M) had no e ffect on I-sc. The I-sc, correlated with an active Cl- secretion measu red as the difference between Cl-36(-) unidirectional fluxes, was part ly blocked by serosal ouabain, bumetanide, DIDS or apical DPC or NPPB and stimulated by serosal dB-cAMP. It is concluded that the chloride s ecretion is mediated by a Na+/K+/2Cl(-) cotransport and a Cl-/HCO3- ex changer both responsible for Cl- entry through the basolateral membran e and by apical cAMP-sensitive Cl- channels. This study gives evidence of a functional, highly differentiated epithelium in cultures compose d of fish gill respiratorylike cells, which could provide a useful pre paration for studies on ion transport and their regulation. Furthermor e, the chloride secretion through these cultures of respiratorylike ce lls makes it necessary to reconsider the previously accepted sea water model in which the chloride cells are given the unique role of ion tr ansport through fish gills.