Expression of key ion transporters in the gill and esophageal- gastrointestinal tract of euryhaline mozambique tilapia oreochromis mossambicus acclimated to fresh water, seawater and hypersaline water

10.1371/journal.pone.0087591PLoS ONE91-POLN

Immunohistochemical localization of transporters in anterior and posterior intestine of tilapia acclimated to different salinities.

<p>Representative micrographs of immunolocalization of Nka (green) with Nkcc/Ncc (red) (a–d, f–h) from FW (a, f), SW (b, g) and HSW (c, h) acclimated tilapia. (d) A representative higher magnification micrograph of Nkcc/Ncc staining of the brush border of enterocytes with basolateral Nka staining from the anterior intestine of SW-acclimated fish. (e) Apical Cftr (red) double labeling with Nka (green) in the anterior intestine of a FW-acclimated fish. Panels (a–e) are sections of anterior intestine (AI) while panels (f–h) are sections of posterior intestine (PI). Sections are counter stained with the nuclear stain DAPI and overlaid with the DIC image for tissue orientation. Scale bar 100 µm (a–c, f–h); 25 µm (d,e).</p

Gene expression of <i>cystic fibrosis transmembrane conductance regulator</i> (<i>cftr</i>) and <i>sodium potassium ATPases</i> (<i>nka</i>).

<p>Expression levels of <i>cftr</i> (a), <i>nka</i>-α1 (b), <i>nka</i>-α3 (c) in gills and EGI tract of tilapia acclimated to freshwater (FW), seawater (SW) and hypersaline water (HSW). Each histogram bar represents the mean ± s.d. of the expression levels (log₁₀ copies of transcript per 50 ng cDNA). Expression levels labeled with different lowercase alphabets are significantly different (one-way ANOVA followed by Duncan's post-hoc test; <i>P</i><0.05) within the same tissue.</p

Immunohistochemical localization of transporters in gills of tilapia acclimated to different salinities.

<p>Representative micrographs of immunohistolocalization of Nka (green) with either Nkcc/Ncc (a–c; red) or Cftr (d–f; red) in the gills of tilapia acclimated to FW (a,d), SW (b,e) or HSW (c,f). Co-localization of red and green fluorochromes results in yellow-orange staining. Higher magnification (10×) of boxed areas in (a–f) correspond to panels (a′–f′). Sections are counter stained with the nuclear stain DAPI and overlaid with the DIC image for tissue orientation. Arrows indicate apical staining and arrowheads tubular system (basolateral) staining. Scale bar 100 µm (a–f), 10 µm (a′–c′).</p

Gene expression of <i>sodium (potassium) chloride</i> (<i>nkcc/ncc</i>) <i>cotransporters</i>.

<p>Expression levels of <i>nkcc1a</i> (a), <i>nkcc1b</i> (b), <i>nkcc2</i> (c), <i>ncc</i> (d), in the gills and EGI tract of tilapia acclimated to freshwater (FW), seawater (SW) and hypersaline water (HSW). Each histogram bar represents the mean ± s.d. of the expression levels (log₁₀ copies of transcript per 50 ng cDNA). Expression levels labeled with different lowercase alphabets are significantly different (one-way ANOVA followed by Duncan's post-hoc test; <i>P</i><0.05) within the same tissue.</p

Normalized luminosity indices of Nkcc/Ncc and Nka immunostaining in the gills, anterior intestine (AI) and posterior intestine (PI) of Mozambique tilapia.

*<p>Normalized Luminosity Index (mean±SEM) is expressed as a ratio of the luminosity of Alexa 488 (Nka) or Alexa 568 (Nkcc/Ncc) to DAPI (nuclei) staining within total field (0.307 mm²); the DAPI nuclear staining is used to correct for differences between fields.</p>**<p>The data were analyzed by one-way ANOVA followed by <i>post-hoc</i> Duncan's multiple range test (<i>P</i><0.05 is considered significant). Different alphabets indicate statistical significance from each other.</p

Gene expression profiles in gills and gastro-intestinal tract of tilapia acclimated different salinities.

<p>Heatmap representing quantity (log₁₀ copies per 50 ng cDNA) of <i>nkcc1a</i>, <i>nkcc1b</i>, <i>nkcc2</i>, <i>ncc</i>, <i>cftr</i>, <i>nka-α1</i>, and <i>nka-α3</i> in the gills and EGI tract segments, including the esophagus, stomach, anterior intestine (AI), middle intestine (MI), and posterior intestine (IG) of tilapia acclimated freshwater (FW), seawater (SW) and hypersaline water (HSW). Different alphabets indicates significant (<i>P</i><0.05) differential gene expression between the salinity conditions (refer to text for the scoring system) within a tissue.</p