Corticotropin-releasing hormone receptor (CRHR)1 and CRHR2 are both trafficking and signaling receptors for urocortin.
Transport of urocortin, a potent satiety peptide, occurs at the blood-brain barrier of the mouse. Endocytosis of urocortin by the cerebral microvessel endothelial cells composing the blood-brain barrier is a rate-limiting step of this transport, but the cellular mechanisms involved have not been fully elucidated. The presence of both CRH receptors R1 and R2 in isolated cerebral microvessels shown in this study suggested that both subtypes might mediate urocortin transport. The roles of these two receptors in the endocytosis and signal transduction of urocortin were tested by overexpression studies in human embryonic kidney 293 cells. Both receptors led to a significant increase of binding and endocytosis of radiolabeled urocortin. CRHR1-mediated urocortin endocytosis was blocked by astressin (antagonist for both CRHRs), whereas CRHR2-mediated urocortin endocytosis was also blocked by antisauvagine 30 (selective CRHR2beta antagonist). Chlorpromazine, filipin, and nystatin had no effect on urocortin endocytosis, indicating the lack of significant involvement of clathrin or caveolae membrane microdomains. Both CRHR1 and CRHR2 were able to mediate the ligand-induced increase of cAMP production, suggesting that the overexpressed receptors were biologically active. Elevation of intracellular cAMP by forskolin or dibutyryl-cAMP, however, did not show acute modulation of the binding and endocytosis of urocortin. Despite the substantial intracellular degradation of endocytosed urocortin in cells overexpressing either CRHR1 or CRHR2, intact urocortin could be exocytosed during the 1-h study interval. We conclude that both CRHR1 and CRHR2 play a facilitatory role in the non-clathrin-, non-caveolae-mediated endocytosis and intracellular signal transduction of this potent peptide.
Tu
Hong
H
Pennington Biomedical Research Center, 6400 Perkins Road, Baton Rouge, Louisiana 70808, USA.
Kastin
Abba J
AJ
Pan
Weihong
W
D000818:Animals; D001921:Brain / Q000378:metabolism; D002478:Cells, Cultured; D003346:Corticotropin-Releasing Hormone / Q000378:metabolism*; D004705:Endocytosis; D042783:Endothelial Cells / Q000378:metabolism; D006801:Humans; D021962:Membrane Microdomains / Q000502:physiology; D051379:Mice; D008810:Mice, Inbred C57BL; D011499:Protein Processing, Post-Translational; D021381:Protein Transport; D018019:Receptors, Corticotropin-Releasing Hormone / Q000502:physiology*; D015398:Signal Transduction; D054832:Urocortins; D000099052:CRF Receptor, Type 1
C101531:CRF receptor type 2; D018019:Receptors, Corticotropin-Releasing Hormone; D054832:Urocortins; D000099052:CRF Receptor, Type 1; D003346:Corticotropin-Releasing Hormone