spacer gif spacer gif spacer gif spacer gif spacer gif
QUICK SEARCH:   [advanced]


spacer gif
     Home     Help     Feedback     Subscriptions     Archive     Search     Table of Contents    

This Article
Right arrow Summary Freely available
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Kern, G.
Right arrow Articles by Pelster, B.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Kern, G.
Right arrow Articles by Pelster, B.
Social Bookmarking
Add to CiteULike   Add to Complore   Add to Connotea   Add to Del.icio.us   Add to Digg   Add to Reddit   Add to Technorati   Add to Twitter  
What's this?

Mechanisms of acid secretion in pseudobranch cells of rainbow trout (Oncorhynchus mykiss)

G. Kern, S. T. Bösch, E. Unterhuber and B. Pelster*

Institut für Zoologie und Limnologie, Universität Innsbruck, Technikerstrasse 25, A-6020 Innsbruck, Austria



View larger version (122K):

[in a new window]
  		Fig. 1. A cluster of adhesive mitochondria-rich cells isolated from the pseudobranch. Scale bar, 50 µm.

 


View larger version (53K):

[in a new window]
  		Fig. 2. Immunohistochemical localization of cytokeratin (red signal) in freshly prepared pseudobranch cells. FITC-labeled streptavidin was used to stain mitochondria (green signal). Three groups of cells were identified. The most intense staining was observed in small and largely undifferentiated cells (A,B). (A) Confocal images with three optical sections of the cells. The top image shows a x,z section, bottom left a x,y section, and bottom right a y,z section. Within each image the locations of the two accompanying sections are indicated by white cross-sectional bars, i.e. in the x,y section (bottom left) the horizontal bar indicates the location of the x,z section (top image), and the vertical bar the location of the y,z section (bottom right). (B) A three-dimensional reconstruction. (C) Cells from a small population of intermediate cell type, with the typical mitochondrial arrangement of pseudobranch cells, but smaller and more spheroid. These cells also stained positive for cytokeratins (cell marked with an arrow in C), but the staining was not as intense as in the undifferentiated cells. Apparently fully differentiated mitochondria-rich cells showed a very weak or no positive reaction. (D) A fully differentiated cell with a faint cytokeratin staining was observed. The staining only became visible at very high amplification of the red channel. In C and D, confocal images are shown with three optical sections of the cells. The top image shows a x,z section, bottom left a x,y section, and bottom right a y,z section. The locations of the sections within the cell cluster are indicated by white cross-sectional bars (see explanation to A). Scale bars, 10 µm.

 


View larger version (78K):

[in a new window]
  		Fig. 3. Incubating fixed pseudobranch cells with FITC-labeled streptavidin resulted in a photostable staining of mitochondria. The confocal image shows three optical sections of a pseudobranch cell in the x,z (A), x,y (B) and y,z (C) planes. Within each image the locations of the two accompanying sections are indicated by white cross-sectional bars, i.e. in the x,y section (bottom left) the horizontal bar indicates the location of the x,z section (top image), and the vertical bar the location of the y,z section (bottom right). A similar picture was obtained by in vivo staining using the mitochondria-selective dyes Mitotracker green FM and Mitotracker orange. Scale bar, 10 µm.

 


View larger version (104K):

[in a new window]
  		Fig. 4. Immunocytochemical staining of the anion-exchanger 2 in pseudobranch cells. An antibody (rabbit anti rat AE2) directed against a specific 21-amino-acid peptide from rat showed cross-reactivity in pseudobranch tissue (A). Control sections with the primary antibody omitted showed no staining (B). Scale bars, 20 µm.

 


View larger version (32K):

[in a new window]
  		Fig. 5. Western-blot analysis of Na+/K+-ATPase (A), anion-exchanger 2 (B) and vacuolar-type ATPase (C) in pseudobranch tissue. The positions of molecular mass markers (kDa) are shown.

 


View larger version (84K):

[in a new window]
  		Fig. 6. Immunohistochemical localization of Na+/K+-ATPase in pseudobranch cells. The confocal image shows three optical sections of a pseudobranch cell in the (A) x,z, (B) the x,y and (C) the y,z planes. Within each image the location of the two accompanying sections is indicated by white cross-sectional bars, i.e. in the x,y section (bottom left) the horizontal bar indicates the location of the x,z section (top image), and the vertical bar the location of the y,z section (bottom right). Note that Na+/K+-ATPase immunoreactivity is located at all cell membranes. Scale bar, 10 µm.

 


View larger version (113K):

[in a new window]
  		Fig. 7. Immunohistochemical staining of subunit B of vacuolar-type ATPase using an antibody directed against a conserved region of the protein (A). A positive staining reaction was found in the membranes of most cells, especially in the tubular region. Control sections incubated without the primary antibody showed no staining at all (B). Scale bar, 10 µm.

 


View larger version (16K):

[in a new window]
  		Fig. 8. The effect of amiloride, an inhibitor of Na+/H+-exchange, on the rate of acidification of the external medium induced by acid secretion of isolated pseudobranch cells (A) and gill cells (B) as a function of time. Amiloride (50 µmoll-1) was added for the period indicated. Values are means ± S.E.M., N=12 (A); N=7 (B). Asterisks indicate significant differences from values before application of amiloride.

 


View larger version (18K):

[in a new window]
  		Fig. 9. The effect of bafilomycin, an inhibitor of V-ATPase, on the rate of acidification of the external medium induced by acid secretion of isolated pseudobranch cells (A) and gill cells (B) as a function of time. Bafilomycin (1 µmoll-1) was added for the period indicated. Values are means ± S.E.M., N=8 (A); N=9 (B). Asterisks indicate significant differences from the values before application of bafilomycin.

 


View larger version (18K):

[in a new window]
  		Fig. 10. The effect of DIDS, an inhibitor of anion exchange, on the rate of acidification of the external medium induced by acid secretion of isolated pseudobranch cells (A) and gill cells (B) as a function of time. DIDS (1 mmoll-1 in pseudobranch cells and 500 µmoll-1 in gill cells) was added for the period indicated. Values are means ± S.E.M., N=9 (A); N=11 (B). Asterisks indicate significant differences from values before application of DIDS.

 

Add to CiteULike CiteULike   Add to Complore Complore   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us   Add to Digg Digg   Add to Reddit Reddit   Add to Technorati Technorati   Add to Twitter Twitter    What's this?



© The Company of Biologists Ltd 2002