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Posterior isolated gills of Neohelice (Chasmagnathus) granulatus were symmetrically perfused with hemolymph-like saline of varying [HCO3-] and pH. Elevating [HCO3-] in the saline from 2.5 to 12.5 mmol/l (pH 7.75 in both cases) induced a significant increase in the transepithelial potential difference (Vte), a measure of ion transport. The elevation in [HCO3-] also induced a switch from acid secretion (-43.7 ± 22.5 µequivµkg-1µh-1) in controls to base secretion (84.7 ± 14.4 µequivµkg-1µh-1). The HCO3--induced Vte increase was inhibited by basolateral acetazolamide (200 µmol/l), amiloride (1 mmol/l), and ouabain (5 mmol/l) but not by bafilomycin (100 nmol/l). The Vte response to HCO3- did not take place in Cl--free conditions; however, it was unaffected by apical SITS (2 mmol/l) or DIDS (1 mmol/l). A decrease in pH from 7.75 to 7.45 pH units in the perfusate also induced a significant increase in Vte, which was matched by a net increase in acid secretion of 67.8 ± 18.4 µequiv kg-1 h-1. This stimulation was sensitive to basolateral acetazolamide, bafilomycin, DIDS, and Na+-free conditions, but it still took place in Cl--free saline. Therefore, the cellular response to low pH is different from the HCO3--stimulated response. We also report V-H+-ATPase- and Na+-K+-ATPase-like immunoreactivity in gill sections for the first time in this crab. Our results suggest that carbonic anhydrase (CA), basolateral Na+/H+ exchangers and Na+-K+-ATPase and apical anion exchangers participate in the HCO3--stimulated response, while CA, apical V-H+-ATPase and basolateral HCO3--dependent cotransporters mediate the response to low pH. [ABSTRACT FROM AUTHOR]

We investigated the involvement of carbonic anhydrase (CA) in mediating V-H+-ATPase translocation into the basolateral membrane in gills of alkalotic Squalus acanthias. Immunolabeling revealed that CA is localized in the same cells as V-H+-ATPase. Blood plasma from dogfish injected with acetazolamide [30 mg/kg at time (t) = 0 and 6 h] and infused with NaHCO3 for 12 h (1,000 +µeq·kg-1·h-1) had significantly higher plasma HCO+ concentration than fish that were infused with NaHCO3 alone (28.72 ± 0.41 vs. 6.57 ± 2.47 mmol/l, n = 3), whereas blood pH was similar in both treatments (8.03 ± 0.11 vs. 8.04 ± 0.11 pH units at r = 12 h). CA inhibition impaired V-H+-ATPase translocation into the basolateral membrane, as estimated from immunolabeled gill sections and Western blotting on gill cell membranes (0.24 ± 0.08 vs. 1.00 ± 0.28 arbitrary units, n = 3; P < 0.05). We investigated V-H+-ATPase translocation during a postfeeding alkalosis ("alkaline tide"). Gill samples were taken 24-26 h after dogfish were fed to satiety in a natural-like feeding regime. Immunolabeled gill sections revealed that V-H+-ATPase translocated to the basolateral membrane in the postfed fish. Confirming this result, V-H+-ATPase abundance was twofold higher in gill cell membranes of the postfed fish than in fasted fish (n = 4-5; P < 0.05). These results indicate that 1) intracellular H+ or HCO+ produced by CA (and not blood pH or HCO3-) is likely the stimulus that triggers the V-H+-ATPase translocation into the basolateral membrane in alkalotic fish and 2) V-H+-ATPase translocation is important for enhanced HCO3- secretion during a naturally occurring postfeeding alkalosis. [ABSTRACT FROM AUTHOR]