CA2-WATER-ADAPTED EURYHALINE TELEOST, FUNDULUS-HETEROCLITUS( TRANSPORT BY OPERCULAR EPITHELIUM OF THE FRESH)

We examined transepithelial transport of Ca2+ across the isolated oper cular epithelium of the euryhaline killifish adapted to fresh water. T he opercular epithelium, mounted in vitro with saline on the serosal s ide and fresh water (0.1 mmol . l(-1) Ca2+) bathing the mucosal side, actively transported Ca2+ in the uptake direction; net flux averaged 2 0-30 nmol . cm(-2). h(-1). The rate of Ca2+ uptake varied linearly wit h the density of mitochondria-rich cells in the preparations. Ca2+ upt ake was saturable, apparent K-1/2 of 0.348 mmol . l(-1), indicative of a multistep transcellular pathway. Ca2+ uptake was inhibited partiall y by apically added 0.1 mmol . l(-1) La3+ and 1.0 mmol . l(-1) Mg2+. A ddition of dibutyryl-cyclic adenosine monophosphate (0.5 mmol . l(-1)) + 0.1 mmol . l(-1) 3-isobutyl-1-methylxanthine inhibited Ca2+ uptake by 54%, but epinephrine, clonidine and isoproterenol were without effe ct. Agents that increase intracellular Ca2+, thapsigargin (1.0 mu mol . l(-1), serosal side), ionomycin (1.0 mu mol . l(-1), serosal side) a nd the calmodulin blocker trifluoperazine (50 mu mol . l(-1), mucosal side) all partially inhibited Ca2+ uptake. In contrast, apically added ionomycin increased mucosal to serosal unidirectional Ca2+ flux, indi cating Ca2+ entry across the apical membrane is rate limiting in the t ransport. Verapamil (10-100 mu mol . l(-1), mucosal side), a Ca2+ chan nel blocker, had no effect. Results are consistent with a model of Ca2 + uptake by mitochondria rich cells that involves passive Ca2+ entry a cross the apical membrane via verapamil-insensitive Ca2+ channels, int racellular complexing of Ca2+ by calmodulin and basolateral exit via a n active transport process. Increases in intracellular Ca2+ invoke a d ownregulation of transcellular Ca2+ transport, implicating Ca2+ as a h omeostatic mediator of its own transport.