title: Cooling evokes redistribution of alpha2C-adrenoceptors from Golgi to plasma membrane in transfected human embryonic kidney 293 cells.
abstract: Cold-induced vasoconstriction in cutaneous blood vessels is mediated by increased constrictor activity of vascular alpha2-adrenoceptors (alpha2-ARs). In mouse cutaneous arteries, alpha2-AR constriction at 37 degrees C is mediated by alpha2A-ARs, whereas after cold exposure (28 degrees C), alpha2C-ARs are no longer silent and mediate the remarkable cold-induced augmentation of alpha2-AR responsiveness. The goals of the present study were to develop a cell model of cutaneous thermoregulation and to determine the mechanisms underlying the thermosensitivity of alpha2C-ARs. Human embryonic kidney 293 cells were transiently transfected with the mouse alpha2A- or alpha2C-AR. In cells expressing alpha2A-ARs, UK-14,304 (5-bromo-N-(4,5-dihydro-1H-imidazol-2-yl)-6-quinoxalinamine), an alpha2-AR agonist, inhibited (10 pM) and stimulated (1-10 nM) the accumulation of cAMP evoked by forskolin. Similar responses were obtained at 37 degrees C and 28 degrees C. In contrast, in cells expressing alpha2C-ARs, UK-14,304 did not affect forskolin-stimulated cAMP accumulation at 37 degrees C but did cause a concentration-dependent inhibitory effect at 28 degrees C. Subcellular fractionation revealed that at 37 degrees C alpha2C-ARs were localized predominantly to Golgi compartments, whereas alpha2A-ARs localized predominantly to the plasma membrane. After cooling (28 degrees C), alpha2C-ARs relocated from Golgi compartments to the plasma membrane, whereas the alpha2A-AR remained at the plasma membrane. Immunofluorescence microscopy confirmed that, at 37 degrees C, alpha2A-ARs were localized to the cell surface, whereas alpha2C-ARs colocalized with a trans-Golgi marker. Cooling did not affect localization of alpha2A-ARs, but shifted alpha2C-ARs to the cell surface. Moderate cooling, therefore, caused a selective redistribution of alpha2C-ARs from the Golgi compartments to the cell surface, allowing the rescue of the alpha2C-adrenergic functional response. This mechanism may explain the role of alpha2-ARs in thermoregulation of the cutaneous circulation.
authors.affiliation: Dorothy M. Davis Heart and Lung Research Institute, The Ohio State University, Columbus, Ohio 43210, USA.
authors.forename: S C
authors.initials: SC
authors.lastname: Jeyaraj
authors.forename: M A
authors.initials: MA
authors.lastname: Chotani
authors.forename: S
authors.initials: S
authors.lastname: Mitra
authors.forename: H E
authors.initials: HE
authors.lastname: Gregg
authors.forename: N A
authors.initials: NA
authors.lastname: Flavahan
authors.forename: K J
authors.initials: KJ
authors.lastname: Morrison
mesh_terms: D002462:Cell Membrane / Q000378:metabolism*; D002478:Cells, Cultured; D005455:Fluorescent Antibody Technique; D006056:Golgi Apparatus / Q000378:metabolism*; D006801:Humans; D018341:Receptors, Adrenergic, alpha-2 / Q000378:metabolism*; D015398:Signal Transduction; D013347:Subcellular Fractions; D013696:Temperature; D014162:Transfection
chemical_list: C547774:ADRA2C protein, human; C547775:Adra2c protein, mouse; D018341:Receptors, Adrenergic, alpha-2