title: The V-ATPase is expressed in the choroid plexus and mediates cAMP-induced intracellular pH alterations.
abstract: The cerebrospinal fluid (CSF) pH influences brain interstitial pH and, therefore, brain function. We hypothesized that the choroid plexus epithelium (CPE) expresses the vacuolar H+-ATPase (V-ATPase) as an acid extrusion mechanism in the luminal membrane to counteract detrimental elevations in CSF pH. The expression of mRNA corresponding to several V-ATPase subunits was demonstrated by RT-PCR analysis of CPE cells (CPECs) isolated by fluorescence-activated cell sorting. Immunofluorescence and electron microscopy localized the V-ATPase primarily in intracellular vesicles with only a minor fraction in the luminal microvillus area. The vesicles did not translocate to the luminal membrane in two in vivo models of hypocapnia-induced alkalosis. The Na+-independent intracellular pH (pHi) recovery from acidification was studied in freshly isolated clusters of CPECs. At extracellular pH (pHo) 7.4, the cells failed to display significant concanamycin A-sensitive pHirecovery (i.e., V-ATPase activity). The recovery rate in the absence of Na+amounted to <10% of the pHirecovery rate observed in the presence of Na+Recovery of pHiwas faster at pHo7.8 and was abolished at pHo7.0. The concanamycin A-sensitive pHirecovery was stimulated by cAMP at pH 7.4 in vitro, but intraventricular infusion of the membrane-permeant cAMP analog 8-CPT-cAMP did not result in trafficking of the V-ATPase. In conclusion, we find evidence for the expression of a minor fraction of V-ATPase in the luminal membrane of CPECs. This fraction does not contribute to enhanced acid extrusion at high extracellular pH, but seems to be activated by cAMP in a trafficking-independent manner.
authors.affiliation: Department of Biomedicine, Health, Aarhus University, Aarhus, Denmark.
authors.forename: Henriette L
authors.initials: HL
authors.lastname: Christensen
authors.affiliation: Center for Systems Biology, Program in Membrane Biology/Nephrology Division, Massachusetts General Hospital and Harvard Medical School, Boston, Massachusetts.
authors.forename: Teodor G
authors.initials: TG
authors.lastname: Păunescu
authors.affiliation: Department of Biomedicine, Health, Aarhus University, Aarhus, Denmark.
authors.forename: Vladimir
authors.initials: V
authors.lastname: Matchkov
authors.affiliation: Department of Cellular and Molecular Medicine, Faculty of Health and Medical Sciences University of Copenhagen, Copenhagen, Denmark.
authors.forename: Dagne
authors.initials: D
authors.lastname: Barbuskaite
authors.affiliation: Center for Systems Biology, Program in Membrane Biology/Nephrology Division, Massachusetts General Hospital and Harvard Medical School, Boston, Massachusetts.
authors.forename: Dennis
authors.initials: D
authors.lastname: Brown
authors.affiliation: Department of Cellular and Molecular Medicine, Faculty of Health and Medical Sciences University of Copenhagen, Copenhagen, Denmark.
authors.forename: Helle H
authors.initials: HH
authors.lastname: Damkier
authors.affiliation: Department of Biomedicine, Health, Aarhus University, Aarhus, Denmark jp@ana.au.dk.
authors.forename: Jeppe
authors.initials: J
authors.lastname: Praetorius
mesh_terms: D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000008:administration & dosage / Q000031:analogs & derivatives / Q000378:metabolism; D000818:Animals; D001921:Brain / Q000502:physiology; D002462:Cell Membrane / Q000737:chemistry* / Q000378:metabolism; D002555:Cerebrospinal Fluid / Q000737:chemistry / Q000201:enzymology / Q000502:physiology; D002831:Choroid Plexus / Q000737:chemistry / Q000166:cytology / Q000378:metabolism* / Q000648:ultrastructure; D000242:Cyclic AMP / Q000031:analogs & derivatives / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000008:administration & dosage / Q000009:adverse effects; D005434:Flow Cytometry; D006863:Hydrogen-Ion Concentration / Q000187:drug effects*; D007424:Intracellular Fluid / Q000737:chemistry* / Q000201:enzymology / Q000378:metabolism; D018942:Macrolides / Q000008:administration & dosage / Q000009:adverse effects; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D012333:RNA, Messenger / Q000378:metabolism; D012964:Sodium / Q000378:metabolism; D013873:Thionucleotides / Q000378:metabolism; D025262:Vacuolar Proton-Translocating ATPases / Q000378:metabolism*
chemical_list: D004791:Enzyme Inhibitors; D018942:Macrolides; D012333:RNA, Messenger; D013873:Thionucleotides; D015124:8-Bromo Cyclic Adenosine Monophosphate; C015929:8-((4-chlorophenyl)thio)cyclic-3',5'-AMP; C036978:concanamycin A; D012964:Sodium; C044258:8-chloro-cyclic adenosine monophosphate; D000242:Cyclic AMP; D025262:Vacuolar Proton-Translocating ATPases
keywords: BCECF fluorometry; cerebrospinal fluid; fluorescence‐activated cell sorting; pH regulation