title: Molecular analysis of beta-adrenergic receptor subtypes in rat collecting duct: effects on cell cAMP and Ca2+ levels.
abstract: Expression and functional properties of beta-adrenergic receptors (beta-ARs) were studied in rat collecting tubules isolated by microdissection. Reverse transcription-polymerase chain reaction experiments demonstrated that the beta 1- and beta 2-AR mRNAs, but not the beta 3-subtype, are expressed in the cortical collecting duct (CCD). Quantitation of mRNAs, carried out using mutant RNAs as internal standards, further showed that beta 1- and beta 2-ARs transcripts are present at comparable amounts in CCD (3,000-4,000 copies/mm of tubular length), but reach 6-8 times lower levels in the outer medullary collecting duct (OMCD: beta 1, 480 +/- 180; beta 2, 590 +/- 110 copies/mm of tubular length). Functional studies, carried out in CCD, corroborated the expression of these two receptor subtypes. The rank order of potency of beta-agonists for stimulating adenosine 3',5'-cyclic monophosphate (cAMP) accumulation was isoproterenol > norepinephrine = epinephrine, and similar efficiencies were found for a beta 1- and a beta 2-antagonist to inhibit isoproterenol-dependent cAMP formation. Fura 2 fluorescence measurements revealed that isoproterenol (10 microM) induces a biphasic rise of intracellular free Ca2+ concentration ([Ca2+]i), consisting of an initial fast increase (delta [Ca2+]i = 122 nM) followed by a plateau phase (delta [Ca2+]i = 58 nM). In the absence of basolateral Ca2+, the initial peak was still observed, suggesting intracellular Ca2+ release. Norepinephrine and epinephrine, as well as selective beta 1- and beta 2-agonists, also increased [Ca2+]i in CCD. Only slight [Ca2+]i variations were produced by isoproterenol in the OMCD (delta [Ca2+]i = 21 nM) and the cortical thick ascending limb (delta [Ca2+]i = 25 nM). These results show that both beta 1- and beta 2-ARs are expressed in the collecting tubule and that they predominate in the CCD. The two receptor subtypes contribute to cAMP accumulation induced by beta-agonists. They also trigger [Ca2+]i variations, indicating their possible coupling to several transduction pathways in the rat CCD.
authors.affiliation: Départment de Biologie Cellulaire et Moléculaire, Commissariat à l'Energie Atomique Saclay, Gif sur Yvette, France.
authors.forename: B
authors.initials: B
authors.lastname: Mandon
authors.forename: E
authors.initials: E
authors.lastname: Siga
authors.forename: A
authors.initials: A
authors.lastname: Champigneulle
authors.forename: M
authors.initials: M
authors.lastname: Imbert-Teboul
authors.forename: J M
authors.initials: JM
authors.lastname: Elalouf
mesh_terms: D000262:Adenylyl Cyclases / Q000378:metabolism; D000318:Adrenergic beta-Agonists / Q000494:pharmacology; D000319:Adrenergic beta-Antagonists / Q000494:pharmacology; D000818:Animals; D001483:Base Sequence; D002118:Calcium / Q000378:metabolism*; D000242:Cyclic AMP / Q000378:metabolism*; D017931:DNA Primers; D004837:Epinephrine / Q000494:pharmacology; D007093:Imidazoles / Q000494:pharmacology; D007545:Isoproterenol / Q000494:pharmacology; D007672:Kidney Cortex / Q000378:metabolism*; D007685:Kidney Tubules, Collecting / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009399:Nephrons / Q000378:metabolism*; D009638:Norepinephrine / Q000494:pharmacology; D010869:Pindolol / Q000494:pharmacology; D016133:Polymerase Chain Reaction / Q000379:methods; D011412:Propanolamines / Q000494:pharmacology; D012333:RNA, Messenger / Q000032:analysis / Q000096:biosynthesis; D051381:Rats; D017207:Rats, Sprague-Dawley; D018342:Receptors, Adrenergic, beta-1 / Q000096:biosynthesis*; D018343:Receptors, Adrenergic, beta-2 / Q000096:biosynthesis*
chemical_list: D000318:Adrenergic beta-Agonists; D000319:Adrenergic beta-Antagonists; D017931:DNA Primers; D007093:Imidazoles; D011412:Propanolamines; D012333:RNA, Messenger; D018342:Receptors, Adrenergic, beta-1; D018343:Receptors, Adrenergic, beta-2; C026777:ICI 118551; D010869:Pindolol; C049763:CGP 20712A; D000242:Cyclic AMP; D000262:Adenylyl Cyclases; D007545:Isoproterenol; D002118:Calcium; D009638:Norepinephrine; D004837:Epinephrine