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Marshall WS, Howard JA, Cozzi RR, Lynch EM: NaCl and fluid secretion by the intestine of the teleost Fundulus heteroclitus: involvement of CFTR. J Exp Biol. 2002 Mar;205(Pt 6):745-58.
Sections of posterior intestine of the euryhaline killifish Fundulus heteroclitus adapted to sea water were stimulated by the calcium ionophore ionomycin (1 micromol l (-1)) in combination with agents to elevate intracellular cyclic AMP levels, 0.5 mmol l (-1) dibutyryl-cyclic AMP (db-cAMP) with 0.1 mmol l (-1) 3-isobutyl-1-methylxanthine (IBMX). Intestinal bag preparations from recently fed animals (but not from overnight unfed animals) changed from fluid absorption (+18.9+/-8.30 microl cm (-2) h (-1), N=8) in the untreated control period to net fluid secretion after stimulation (-7.43+/-1.30 microl cm (-2) h (-1), N=8, P <0.01; means +/- S.E.M.), indicative of the capacity of teleost intestine to undergo secretion. Posterior intestinal pieces mounted in vitro in Ussing-style membrane chambers showed net Cl (-) uptake (+2.245+/-0.633 microequiv cm (-2) h (-1), N=7) that turned to net secretion following stimulation by ionomycin + db-cAMP + IBMX (-3.809+/-1.22 microequiv cm (-2) h (-1), N=7, P <0.01). Mucosal application of the anion channel blocker 1 mmol l (-1) diphenylamine-2-carboxylate (DPC) after ionomycin + db-cAMP + IBMX treatment significantly reduced serosal-to-mucosal unidirectional Cl (-) flux (P <0.001), net Cl (-) flux (P <0.05), short-circuit current (I (sc), P <0.001) and tissue conductance (G (t), P <0.001), while 0.1 mmol l (-1) 4,4'-diisothiocyano-2,2'-stilbene-disulphonic acid (DIDS, a blocker of anion exchange) was without effect. Stimulation by db-cAMP + IBMX (no ionomycin) significantly increased unidirectional fluxes, I (sc) and G (t) but did not produce net Cl (-) secretion. Ionomycin alone produced a transient increase in I (sc) but had no effect on G (t) and caused no significant changes in unidirectional or net Cl (-) fluxes. Addition of db-cAMP + IBMX after ionomycin treatment produced net secretion of Cl (-) and large increases in unidirectional fluxes and G (t). Cystic fibrosis transmembrane conductance regulator (CFTR) was immunocytochemically localized with a monoclonal mouse antibody to the carboxy terminus and found to be present in the cytoplasm and basolateral membranes of all enterocytes and in the brush-border membrane of some cells, whereas NKCC immunofluorescence, demonstrating the presence of the Na (+)/K (+)/2Cl (-) cotransporter, was present in the cytoplasm and brush-border membrane. We conclude that the teleost intestine is capable of salt and fluid secretion only if intracellular Ca (2+) and cyclic AMP pathways are stimulated together and that this secretion appears to involve activation of CFTR ion channels in the apical membrane of a subpopulation of enterocytes. |
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