Molecular and Functional Analyses of Transport Proteins for Organic Anions

The blood-cerebrospinal fluid (CSF)-barrier (CSFB) is formed by the Choroid Plexūs (CP). Together with the blood brain barrier it represents an important barrier between brain and circulating blood. Since it can only be passed by small, lipid-soluble and uncharged molecules it is of particular interest to study active transport processes for charged substances across this tissue, especially because many drugs are substrates for these transport proteins. Transporters in the CP are similar to those in the kidney. Therefore, I performed comparative experiments with model organic anions in isolated proximal kidney tubules of killfish and CP of rat and spiny dogfish shark. ·First, I studied transport of the fluorescent cAMP analog fluo-cAMP. Inhibition studies indicate transport mediated by the export proteins Mrp2 and Mrp4. In contrast to transport of the Mrp2-substrate fluorescein-methotrexate (FL-MTX), transport is not regulated by proteinkinase (PK) A, and PKC. For better characterization of kinetic parameters I studied transport of fluo-cAMP in membrane vesicles of MRP2 and MRP4 overexpressing cells. Both MRPs transported fluo-cAMP with similar affinites. Different from results obtained from experiments using renal proximale tubules, fluo-cAMP transport in CP seems not to be mediated by Mrp4, whereas a participation of Mrp1 is likely. Regulation of fluo-cAMP transport in rat CP is not mediated by PKA, PKC or mitogen activated PK (MAPK). ·Transport of Texas Red (TR) and FL-MTX was studied in CP of rat and spiny dogfish shark. Transport of TR across rat CP is a two-step mechanism, which is metabolism-driven, Na+-dependent, potential independent and also independent from Mrp1, Mrp2 and Mrp4. In dogfish shark CP TR transport is Na+-independent, with apical uptake apparently mediated by an Oat-protein, whereas basolateral efflux seems to be mediated by Oatp2 and Mrp1. ·Apical uptake of FL-MTX in rat CP is also Na+-independent with an potential independent basolateral efflux, which is presumably mediated via Mrp1 and Oatp2. Activation of PKA results in an activation of FL-MTX efflux. In dogfish shark FL-MTX is transported via the same transport proteins. But in contrast to rat CP, activation of PKA results in a reduced efflux of FL-MTX, whereas activation of PKC leads to a downregulation of FL-MTX transport.

Level thesis:degree_level

thesis.doctoral

Grantor dc:publisher

Universität Heidelberg

Year

2006

Author dc:creator

• Reichel, Valeska

Contributors dc:contributor

• Fricker, Gert

Repository record source_url

http://www.ub.uni-heidelberg.de/archiv/6122

OAI identifier oai:identifier

oai:archiv.ub.uni-heidelberg.de:6122