Regulation of the vasopressin V2 receptor by vasopressin in polarized renal collecting duct cells

J H Robben, N V A M Knoers, P M T Deen

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Abstract

Binding of arginine-vasopressin (AVP) to its V2 receptor (V2R) in the basolateral membrane of principal cells induces Aquaporin-2-mediated water reabsorption in the kidney. To study the regulation of the V2R by dDAVP in a proper model, a polarized renal cell line stably-expressing V2R-GFP was generated. Labeled AVP-binding studies revealed an equal basolateral vs. apical membrane distribution for V2R-GFP and endogenous V2R. In these cells, GFP-V2R was expressed in its mature form and localized for 75% in the basolateral membrane and for 25% to late endosomes/lysosomes. dDAVP caused a dose- and time-dependent internalization of V2R-GFP, which was completed within 1 h with 100 nM dDAVP, was prevented by coincubation with a V2R antagonist, and which reduced its half-life from 11.5 to 2.8 h. Semiquantification of the V2R-GFP colocalization with E-cadherin (basolateral membrane), early endosomal antigen-1 (EEA-1) and lysosome-associated membrane protein-2 (LAMP-2) in time revealed that most dDAVP-bound V2R was internalized via early endosomes to late endosomes/lysosomes, where it was degraded. The dDAVP-internalized V2R did not recycle to the basolateral membrane. In conclusion, we established the itinerary of the V2R in a polarized cell model that likely resembles the in vivo V2R localization and regulation by AVP to a great extent.

Original languageEnglish
Pages (from-to)5693-9
Number of pages7
JournalMolecular Biology of the Cell
Volume15
Issue number12
DOIs
Publication statusPublished - Dec-2004
Externally publishedYes

Keywords

  • Animals
  • Cell Line
  • Cell Polarity
  • Dogs
  • Glycosylation
  • Kidney Tubules, Collecting/cytology
  • Lysosomes/drug effects
  • Organelles/metabolism
  • Protein Processing, Post-Translational/drug effects
  • Receptors, Vasopressin/genetics
  • Time Factors
  • Vasopressins/pharmacology

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