Mercury toxicity in the shark (Squalus acanthias) rectal gland: apical CFTR chloride channels are inhibited by mercuric chloride.
In the shark rectal gland, basolateral membrane proteins have been suggested as targets for mercury. To examine the membrane polarity of mercury toxicity, we performed experiments in three preparations: isolated perfused rectal glands, primary monolayer cultures of rectal gland epithelial cells, and Xenopus oocytes expressing the shark cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel. In perfused rectal glands we observed: (1) a dose-dependent inhibition by mercury of forskolin/3-isobutyl-1-methylxanthine (IBMX)-stimulated chloride secretion; (2) inhibition was maximal when mercury was added before stimulation with forskolin/IBMX; (3) dithiothrietol (DTT) and glutathione (GSH) completely prevented inhibition of chloride secretion. Short-circuit current (Isc) measurements in monolayers of rectal gland epithelial cells were performed to examine the membrane polarity of this effect. Mercuric chloride inhibited Isc more potently when applied to the solution bathing the apical vs. the basolateral membrane (23 +/- 5% and 68 +/- 5% inhibition at 1 and 10 microM HgCl2 in the apical solution vs. 2 +/- 0.9% and 14 +/- 5% in the basolateral solution). This inhibition was prevented by pre-treatment with apical DTT or GSH; however, only the permeant reducing agent DTT reversed mercury inhibition when added after exposure. When the shark rectal gland CFTR channel was expressed in Xenopus oocytes and chloride conductance was measured by two-electrode voltage clamping, we found that 1 microM HgCl2 inhibited forskolin/IBMX conductance by 69.2 +/- 2.0%. We conclude that in the shark rectal gland, mercury inhibits chloride secretion by interacting with the apical membrane and that CFTR is the likely site of this action.
Ratner
Martha A
MA
Nephrology Division, Department of Internal Medicine, Yale University School of Medicine, New Haven, Connecticut 06510, USA.
Decker
Sarah E
SE
Aller
Stephen G
SG
Weber
Gerhard
G
Forrest
John N
JN
D015056:1-Methyl-3-isobutylxanthine / Q000494:pharmacology; D000818:Animals; D005576:Colforsin / Q000494:pharmacology; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000037:antagonists & inhibitors* / Q000096:biosynthesis / Q000235:genetics; D004229:Dithiothreitol / Q000494:pharmacology; D004284:Dogfish / Q000502:physiology*; D004847:Epithelial Cells / Q000187:drug effects; D005978:Glutathione / Q000494:pharmacology; D066298:In Vitro Techniques; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D008627:Mercuric Chloride / Q000633:toxicity*; D018408:Patch-Clamp Techniques; D012491:Salt Gland / Q000166:cytology / Q000187:drug effects*; D014981:Xenopus / Q000235:genetics / Q000378:metabolism
D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D005576:Colforsin; D008627:Mercuric Chloride; D005978:Glutathione; D004229:Dithiothreitol; D015056:1-Methyl-3-isobutylxanthine