PMID: 7545030May 1, 1995Paper

Delineation of the endocytic pathway of substance P and its seven-transmembrane domain NK1 receptor

Molecular Biology of the Cell
E F GradyN W Bunnett

Abstract

Many of the actions of the neuropeptide substance P (SP) that are mediated by the neurokinin 1 receptor (NK1-R) desensitize and resensitize, which may be associated with NK1-R endocytosis and recycling. We delineated this endocytic pathway in transfected cells by confocal microscopy using cyanine 3-SP and NK1-R antibodies. SP and the NK1-R were internalized into the same clathrin immunoreactive vesicles, and then sorted into different compartments. The NK1-R was colocalized with a marker of early endosomes, but not with markers of late endosomes or lysosomes. We quantified the NK1-R at the cell surface by incubating cells with an antibody to an extracellular epitope. After exposure to SP, there was a loss and subsequent recovery of surface NK1-R. The loss was prevented by hypertonic sucrose and potassium depletion, inhibitors of clathrin-mediated endocytosis. Recovery was independent of new protein synthesis because it was unaffected by cycloheximide. Recovery required endosomal acidification because it was prevented by an H(+)-ATPase inhibitor. The fate of internalized 125I-SP was examined by chromatography. SP was intact at the cell surface and in early endosomes, but slowly degraded in perinuclear vesicles. We conclude that ...Continue Reading

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Citations

Dec 1, 1996·Histochemistry and Cell Biology·B R SouthwellJ B Furness
Dec 1, 1996·Neuroscience·E F GradyN W Bunnett
Nov 11, 2003·Life Sciences·Jennifer L Seachrist, Stephen S G Ferguson
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