Localization of mRNA for the proton pump (H⁺-ATPase) and exchanger in the rainbow trout gill

Abstract

In situ hybridization was performed on sections of rainbow trout (Oncorhynchus mykiss) gill tissue using oligonucleotide probes complementary to the mRNA of the 3 1-kilodalton subunit of the bovine renal V-type H+-ATPase or rat kidney Band 3 anion exchanger (C1-IHCO, exchanger). This was conducted in conjunction with measurements of whole-body net acid fluxes and blood acid-base status during imposed conditions of respiratory acidosis (external hypercapnia) or metabolic alkalosis (NaHCO, infusion). A positive hybridization signal for the H+-ATPase mRNA was localized predominantly in lamellar epithelial cells and was less apparent in cells associated with the filament or interlamellar regioqs. The Hf -ATPase hybridization signal was enhanced during hypercapnic acidosis concurrently with a marked increase in whole-body net acid excretion. A positive hybridization signal for the C1-IHCOj exchanger mRNA was observed in epithelial cells on both the filament and lamella. During metabolic alkalosis induced by intra-arterial infusion of NaHCO,, there was a marked increase in the C1-/HCO; exchanger mRNA hybridization signal in cells on both the filament and lamella that occurred concurrently with a decrease in net acid excretion. The results of this study support the existence of a V-type Hf -ATPase and a Cl-/HCO; exchanger in rainbow trout gill epithelial cells and demonstrate that alterations in gene expression for the pump-exchanger may be a significant mechanism underlying the altered rates of net acid equivalent excretion during acid-base disturbances.

The role of branchial carbonic anhydrase in acid-base regulation in rainbow trout (Oncorhynchus mykiss)

“…Morphological adjustment of the branchial epithelium, specifically a reduction in the exposed surface area of MR cells, the presumed site of branchial Cl -/HCO 3 -exchangers (see below), constitutes one mechanism through which this reduction in Cl -/HCO 3 -exchange is achieved (Goss and Perry, 1993; reviewed by Goss et al, 1992Goss et al, , 1995Perry, 1997;Perry et al, 2003). The minor enhancement of proton extrusion during hypercarbia is supported by increased branchial H + -ATPase activity (Lin and Randall, 1993;Sullivan et al, 1995;Galvez et al, 2002) that reflects, at least to some degree, greater gene expression of the vacuolar H + -ATPase (Sullivan et al, 1996;Perry et al, 2000a,b). The results of the present study indicate that the abundance of the trout gill cytoplasmic CA isoform (tCAc) is also increased, albeit modestly, during hypercarbia (Fig.·4), at least in part owing to enhanced gene transcription (Fig.·3).…”

The involvement of SLC26 anion transporters in chloride uptake in zebrafish (Danio rerio) larvae

“…Prior to the demonstration of pendrin (SLC26A4)-like immunoreactivity on the elasmobranch gill (Piermarini et al, 2001), it was generally considered that branchial Cl -/HCO 3 -exchange was accomplished by one or more members of the SLC4 gene family. This conclusion was reinforced by the results of studies utilizing immunocytochemistry (Wilson et al, 2000), in situ hybridization (Sullivan et al, 1996) and data obtained from pharmacological - (Epstein et al, 1973;Perry et al, 1981a;Chang and Hwang, 2004;Preest et al, 2005). With the benefit of hindsight, however, the results of these former studies are subject to reinterpretation.…”

Ion uptake and acid secretion in zebrafish (Danio rerio)

“…On the other hand, it was proposed that the apical Cl -/HCO 3 -exchanger is the major player in the mechanism of Cl -uptake linked with basal secretion in fish gills, and anion exchanger (AE) 1 was one of the candidates proposed (Evans et al, 2005;Hwang and Lee, 2007;Evans, 2008). However, ISH and ICC data for the existence of AE1 in teleost gills (Sullivan et al, 1996;Wilson et al, 2000) were controversial when considering the specificity of the antibody or probe used and the absence of evidence for AE1 on the apical membrane of other acid/base-secreting epithelia Tresguerres et al, 2006). In our preliminary experiments on zebrafish, mRNA of an AE1 paralogue was localized in a specific group of skin/gill cells (Fig.…”