Molecular cloning and functional expression of the guinea pig alpha(1a)-adrenoceptor.
In the present paper, the cloning and expression of the guinea pig alpha(1A)-adrenoceptor is presented. The nucleotide sequence had an open reading frame of 1401 bp that encoded a 466 amino-acid protein with an estimated molecular mass of approximately 51.5 kDa. When the clone was expressed in Cos-1 cells, specific high-affinity binding of [(3)H]prazosin and [(3)H]tamsulosin was observed. Chloroethylclonidine treatment of membranes slightly decreased the total binding with both radioligands. Binding competition experiments using [(3)H]tamsulosin showed the following potency order: (a) for agonists: oxymetazoline >>epinephrine>norepinephrine>methoxamine, and (b) for antagonists: prazosin> or 5-methyl-urapidil=benoxathian>phentolamine>>BMY 7378 (8-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-8-azaspiro[4,5]decane-7,9-dione). Photoaffinity labeling using [(125)I-aryl]azido-prazosin revealed a major broad band with a molecular mass between 70 and 80 kDa. The receptor was functional, as evidenced by an epinephrine-increased production of [(3)H]inositol phosphates that was blocked by prazosin.
González-Espinosa
C
C
Departamento de Biología Celular, Instituto de Fisiología Celular, Universidad Nacional Autónoma de México, Apartado Postal 70-248, México, D.F. 04510, Mexico.
Romero-Avila
M T
MT
Mora-Rodríguez
D M
DM
González-Espinosa
D
D
García-Sáinz
J A
JA
D000322:Adrenergic Agonists / Q000494:pharmacology; D018674:Adrenergic Antagonists / Q000494:pharmacology; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001667:Binding, Competitive; D019556:COS Cells; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics; D004305:Dose-Response Relationship, Drug; D004837:Epinephrine / Q000494:pharmacology; D015870:Gene Expression; D006168:Guinea Pigs; D008729:Methoxamine / Q000494:pharmacology; D008969:Molecular Sequence Data; D009638:Norepinephrine / Q000494:pharmacology; D010075:Oxathiins / Q000494:pharmacology; D010109:Oxymetazoline / Q000494:pharmacology; D010646:Phentolamine / Q000494:pharmacology; D010879:Piperazines / Q000494:pharmacology; D011224:Prazosin / Q000378:metabolism / Q000494:pharmacology; D018340:Receptors, Adrenergic, alpha-1 / Q000235:genetics* / Q000378:metabolism; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013449:Sulfonamides / Q000378:metabolism; D000077409:Tamsulosin; D014316:Tritium
D000322:Adrenergic Agonists; D018674:Adrenergic Antagonists; D018076:DNA, Complementary; D010075:Oxathiins; D010879:Piperazines; D018340:Receptors, Adrenergic, alpha-1; D013449:Sulfonamides; D014316:Tritium; C057446:5-methylurapidil; D010109:Oxymetazoline; C050877:benoxathian; D000077409:Tamsulosin; D008729:Methoxamine; C053428:BMY 7378; D009638:Norepinephrine; D011224:Prazosin; D004837:Epinephrine; D010646:Phentolamine