Login to view PhD Thesis

Enter your username and password here in order to log in on the website:


Forgot your password?

Sodium-dependent acid/base transport in regulation of intracellular pH in rat choroid plexus

Elena V. Bouzinova

Abstract

Sodium-dependent acid/base transport in regulation of intracellular pH in rat choroid plexus

It has previously been suggested that Na+ and HCO3  transport across the choroid plexus epithelium is key to formation of cerebrospinal fluid. Two Na+-dependent HCO3transporters have previously been shown to localize to the epithelial cells of the rat choroid plexus (NBCn1 and NCBE), and the mRNA for a third protein from the same family of proteins has also been detected (NBCe2). In addition to playing a role for the formation of cerebrospinal fluid these transporters are believed to be involved in regulation of cerebrospinal fluid pH as well as in intracellular pH of the epithelial cells of the choroid plexus and thus play a vital role for normal brain function.

The functional characterization of bicarbonate-dependent and –independent mechanisms involved in the regulation of intracellular pH in choroid plexus epithelial cells was aim for this thesis.

Using immunohistochemistry it was possible to localize NBCe2 to the apical membrane of the choroid plexus epithelial cells. Using microfluometry it was possible to demonstrate that steady-state intracellular pH in choroid plexus epithelial cells was dependent on the presence of CO2/HCO3-, and that both DIDS-sensitive and DIDS-insensitive sodium-dependent bicarbonate transporting mechanisms were present in choroid plexus epithelial cells.

In the absence of bicarbonate I was still able to demonstrate a substantial transport of acid equivalents across the rat choroid plexus epithelial cell membrane. This transport was only slightly reduced with inhibitors of Na,H-exchange. To test whether this was due to steric hindrance for the transport inhibitors to reach the Na,H-exchanger I developed an assay for assessment of intracellular pH in isolated choroid plexus epithelial cells. In the isolated cells a larger part of the transport was inhibited by the Na,H-exchange inhibitors, although in the isolated cells a substantial transport of acid equivalents was still present. To test whether the bicarbonate independent transport was due to residual transport via the bicarbonate transporters (despite the nominal absence of bicarbonate) I attempted to down-regulated the bicarbonate transporters using siRNAs, but this procedure did not inhibit the transport in the absence of CO2/HCO3-.