Differential immunohistochemical staining of Na+/K+-ATPase have been carried out in different sections of Goldfish nephrons (Chasiotis et al. 2012). Differential Na+/K+-ATPase staining has also been carried out for different sections of nephrons of other fish species and basolateral localization of Na+/K+-ATPase has been reported for ion transports. In addition, specific patterns of distribution of Na+, K+-ATPase along the nephron have been reported for several other vertebrate groups (Piepenhagen et al. 1995; Kwon et al. 1998; Sturla et al. 2003). In Goldfish kidney, immunostaining response for Na+/K+-ATPase is primarily restricted to the basement membrane of renal epithelial cells in the proximal region of the nephron as well as the basolateral membrane of renal epithelial cells in the distal and collecting tubules (Chasiotis et al. 2012). In current study, expression of Na+/K+-ATPase was verified in all sections of nephron tubules.
Interestingly, in both killifish and rainbow trout, compatibility with freshwater or seawater environments does not cause immunohistochemical changes in the kidney at the microscopic level either for light or electron microscopy (Katoh et al. 2008). Two important NKCC isoforms, including the NKCC1 secretory form and the NKCC2 absorption form, can be found in vertebrates. In dogfish kidneys, however, the basolateral isoform (NKCC1) has been observed which is distributed in the proximal tubules while the NKCC2 isoform is observed apically from the proximal tubules to the collecting tubules (Biemesderfer et al. 1996). Secretory isoform (NKCC1) has also been found basolaterally in the large rectum of fish (Lytle et al. 1992). The anti-NKCC antibody used in the present study (T4) detects both secretory and absorption isoforms in a variety of animal tissues (Lytle et al. 1992). Therefore, in the present study, we cannot distinguish between these isoforms in different regions of the nephron. Katoh et al. Suggested that in the rainbow trout kidney only NKCC2 is present and located apically, while in the killifish kidney NKCC2 isoform is present apically and the NKCC1 isoform is present basolaterally (Katoh et al. 2008).
The killfish kidney is divided into four regions in terms of ion transport: the first part of the proximal tubule, the second part of the proximal tubule, the distal tubule and the collecting tubule, each of which differs in terms of NKCC and Na+/K+-ATPase function. NKCC is present basally in the first and second parts of the proximal tubule where they pump ions from interstitial tissue into epithelial cells, while in the distal and collecting tubules NKCC is present in the apical part of the cells and pumps ions from the lumen into the epithelial cells. Na+/K+-ATPase is also present basolaterally in the second part of the proximal tubule as well as distal tubules and collecting tubules (Katoh et al. 2008). In the same study by Katoh et al. rainbow trout kidneys were examined for localization and function of NKCC and Na+/K+-ATPase where Na+/K+ -ATPase was located in first and second part of proximal tubule, distal tubules and collecting tubules, although with higher intensities along the distal tubule compared to proximal tubule. NKCC was only reported apically in the distal and collecting tubules (Katoh et al. 2008). In the present study, detection of Na+/K+/2Cl expression was limited to collecting tubules and collecting ducts with higher intensities in the latter.
NHE3 is located in the apical membrane ionocytes in the gills of freshwater fish and seawater fish (Choe et al. 2005). Immunohistochemically localization of NHE3 responded positively in the brush border of intestinal epithelium (jejunum, ileum, and colon) and in the renal tubules (proximal tubule and thick ascending limb) (Mahnensmith and Aronson 1985).
In a study carried out by Kato et al. on the expression of the third isoform of Na+/H+ exchanger in Triakis scyllium, the kidneys were divided into bundle and sinus regions from immunohistochemical viewpoint. In fixed sections of the Triakis kidney, NHE3-specific signals were detected in the apical membrane of the renal tubules in both the bundle and sinus regions (Li et al. 2013). The sinus region consists of second (proximal tubules and intermediate tubules) and a fourth (end of distal tubules) rings. Proximal tubules could be distinguished with their large diameter, long epithelium and the presence of a brush border that can be stained with anti-Na/K -ATPase antibody in the basolateral part of these cells. Distal tubes were distinguished with their lower diameter and thinner epithelium as well as the absence of brush border which leads to high staining response to anti-Na/K -ATPase antibody. In the sinus region of the Triakis kidney, specific signals for NHE3 were detected in part of the proximal as well as end of distal tubules.
In other words, NHE3 were both positive and observed in both tubules. NHE3-negative proximal tubes were often observed in the sinusoidal region near the bundle region. In the bundle region, the five tubular sections are placed side by side to conduct the flow and are separated by a sheath from the adjacent bundle region (Li et al. 2013). NHE3-specific signals were found in the apical membrane of one of the five tubes.
NHE3-positive tubes were detected in the bundle area with relatively large diameters consisting of long epithelial cells with no brush border and reacted strongly to Na/K-ATPase localization (Li et al. 2013). These results clearly indicate that NHE3 is expressed in the first part of distal tubule (Li et al. 2013). In our current study, Na+/H+ Exchanger expression was detected only at the apex of epithelial cells of proximal tubules, collecting tubes and collecting ducts with high, moderate and low intensities, respectively, while distal tubules did not respond to Na+/H+ exchanger immunolocalization.