title: Differential antagonism by conotoxin rho-TIA of contractions mediated by distinct alpha1-adrenoceptor subtypes in rat vas deferens, spleen and aorta.
abstract: The ability of the conotoxin rho-TIA, a 19-amino acid peptide isolated from the marine snail Conus tulipa, to antagonize contractions induced by noradrenaline through activation of alpha1A-adrenoceptors in rat vas deferens, alpha1B-adrenoceptors in rat spleen and alpha1D-adrenoceptors in rat aorta, and to inhibit the binding of [125I]HEAT (2-[[beta-(4-hydroxyphenyl)ethyl]aminomethyl]-1-tetralone) to membranes of human embryonic kidney (HEK) 293 cells expressing each of the recombinant rat alpha1-adrenoceptors was investigated. rho-TIA (100 nM to 1 microM) antagonized the contractions of vas deferens and aorta in response to noradrenaline without affecting maximal effects and with similar potencies (pA2 approximately 7.2, n=4). This suggests that rho-TIA is a competitive antagonist of alpha1A- and alpha1D-adrenoceptors with no selectivity between these subtypes. Incubation of rho-TIA (30 to 300 nM) with rat spleen caused a significant reduction of the maximal response to noradrenaline, suggesting that rho-TIA is a non-competitive antagonist at alpha1B-adrenoceptors. After receptor inactivation with phenoxybenzamine, the potency of rho-TIA in inhibiting contractions was examined with similar occupancies (approximately 25%) at each subtype. Its potency (pIC50) was 12 times higher in spleen (8.3+/-0.1, n=4) than in vas deferens (7.2+/-0.1, n=4) or aorta (7.2+/-0.1, n=4). In radioligand binding assays, rho-TIA decreased the number of binding sites (B(max)) in membranes from HEK293 cells expressing the rat alpha1B-adrenoceptors without affecting affinity (K(D)). In contrast, in HEK293 cells expressing rat alpha1A- or alpha1D-adrenoceptors, rho-TIA decreased the K(D) without affecting the B(max). It is concluded that rho-TIA will be useful for distinguishing the role of particular alpha1-adrenoceptor subtypes in native tissues.
authors.affiliation: Department of Pharmacology, Instituto de Biociências, UNESP, Botucatu, SP 18618-000, Brazil.
authors.forename: Vanessa
authors.initials: V
authors.lastname: Lima
authors.forename: André
authors.initials: A
authors.lastname: Mueller
authors.forename: Susana Y
authors.initials: SY
authors.lastname: Kamikihara
authors.forename: Vanessa
authors.initials: V
authors.lastname: Raymundi
authors.forename: Dianne
authors.initials: D
authors.lastname: Alewood
authors.forename: Richard J
authors.initials: RJ
authors.lastname: Lewis
authors.forename: Zhongjian
authors.initials: Z
authors.lastname: Chen
authors.forename: Kenneth P
authors.initials: KP
authors.lastname: Minneman
authors.forename: André S
authors.initials: AS
authors.lastname: Pupo
mesh_terms: D058646:Adrenergic alpha-1 Receptor Agonists; D058668:Adrenergic alpha-1 Receptor Antagonists; D000316:Adrenergic alpha-Agonists / Q000494:pharmacology; D000317:Adrenergic alpha-Antagonists / Q000494:pharmacology; D000818:Animals; D001013:Aorta, Thoracic / Q000187:drug effects / Q000502:physiology; D001667:Binding, Competitive / Q000187:drug effects; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D020916:Conotoxins / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D006801:Humans; D066298:In Vitro Techniques; D007457:Iodine Radioisotopes; D008297:Male; D009119:Muscle Contraction / Q000187:drug effects*; D009638:Norepinephrine / Q000494:pharmacology; D010879:Piperazines / Q000494:pharmacology; D011189:Potassium Chloride / Q000494:pharmacology; D011224:Prazosin / Q000494:pharmacology; D011869:Radioligand Assay; D051381:Rats; D017208:Rats, Wistar; D018340:Receptors, Adrenergic, alpha-1 / Q000378:metabolism*; D013154:Spleen / Q000187:drug effects / Q000502:physiology; D044024:Tetralones / Q000378:metabolism; D014649:Vas Deferens / Q000187:drug effects / Q000502:physiology
chemical_list: D058646:Adrenergic alpha-1 Receptor Agonists; D058668:Adrenergic alpha-1 Receptor Antagonists; D000316:Adrenergic alpha-Agonists; D000317:Adrenergic alpha-Antagonists; D020916:Conotoxins; D007457:Iodine Radioisotopes; D010879:Piperazines; D018340:Receptors, Adrenergic, alpha-1; D044024:Tetralones; C477745:conotoxin rho-TIA; C057446:5-methylurapidil; D011189:Potassium Chloride; C048008:2-(beta-(3-iodo-4-hydroxyphenyl)ethylaminomethyl)tetralone; C053428:BMY 7378; D009638:Norepinephrine; D011224:Prazosin