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 (1 A)-adrenoceptor is presented. The nucleotide sequence had an open reading f rame of 1401 bp that encoded a 466 aniino-acid protein with an estimated mo lecular mass of = 51.5 kDa. When the clone was expressed in Cos-1 cells, sp ecific high-affinity binding of [H-3]prazosin and [H-3]tamsulosin was obser ved. Chloroethylclonidine treatment of membranes slightly decreased the tot al binding with both radioligands. Binding competition experiments using [H -3]tamsulosin showed the following potency order: (a) for agonists: oxymeta zoline >> epinephrine > norepinephrine > methoxamine, and (b) for antagonis ts: prazosin greater than or equal to 5-methyl-urapidil = benoxathian > phe ntolamine >> BMY 7378 (8-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-8-aza spiro[4,5]decane-7,9-dione). Photoaffinity labeling using [I-125-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-increas ed production of [H-3]inositol phosphates that was blocked by prazosin. (C) 2001 Published by Elsevier Science B.V.