Hypocretin stimulates [(35)S]GTP gamma S binding in Hcrtr 2-transfected cell lines and in brain homogenate.
In vitro functional analyses of hypocretin/orexin receptor systems were performed using [(125)I]hypocretin radioreceptor and hypocretin-stimulated [(35)S]GTP gamma S binding assay in cell lines expressing human or canine (wild-type and narcoleptic-mutation) hypocretin receptor 2 (Hcrtr 2). Hypocretin-2 stimulated [(35)S]GTP gamma S binding in human and canine Hcrtr 2 expressing cell lines, while cell lines expressing the mutated canine Hcrtr 2 did not exhibit specific binding for [(125)I]hypocretin or hypocretin-stimulated [(35)S]GTP gamma S. In rat brain homogenates, regional specific hypocretin-stimulated [(35)S]GTP gamma S binding was also observed. Hypocretin-stimulated [(35)S]GTP gamma S binding, may thus be a useful functional assay for hypocretin receptors in both cell lines and brain tissue homogenates.
Shiba
T
T
Center for Narcolepsy, Stanford University, 701B Welch Rd., Palo Alto, CA 94304, USA.
Ozu
M
M
Yoshida
Y
Y
Mignot
E
E
Nishino
S
S
D000818:Animals; D001921:Brain / Q000378:metabolism*; D016466:CHO Cells; D002460:Cell Line; D006224:Cricetinae; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism*; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D009479:Neuropeptides / Q000378:metabolism*; D064446:Orexin Receptors; D000068797:Orexins; D011485:Protein Binding; D051381:Rats; D043562:Receptors, G-Protein-Coupled; D018013:Receptors, Neuropeptide / Q000235:genetics / Q000378:metabolism*; D013462:Sulfur Radioisotopes; D014162:Transfection
C000596229:HCRT protein, human; C578625:Hcrtr2 protein, rat; D047908:Intracellular Signaling Peptides and Proteins; D009479:Neuropeptides; D064446:Orexin Receptors; D000068797:Orexins; D043562:Receptors, G-Protein-Coupled; D018013:Receptors, Neuropeptide; D013462:Sulfur Radioisotopes; D016244:Guanosine 5'-O-(3-Thiotriphosphate)