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Evaluation of alpha1-adrenoceptors in the rabbit iris: pharmacological characterization and expression of mRNA.

ABSTRACT: Subtypes of alpha1-adrenoceptor in rabbit iris have been examined in functional, binding and molecular biological experiments. In functional studies, exogenous and endogenous noradrenaline produced contractions of the iris dilator muscle. The contractile responses to noradrenaline were competitively antagonized by a range of alpha1-adrenoceptor antagonists (pA2 values): prazosin (8.1), WB4101 (8.2), BMY7378 (5.9), YM617 (9.5), JTH-601 (8.8), HV723 (7.8) and KMD-3213 (9.8). The same order of inhibitory potency was seen in the adrenergic responses to electrical stimulation. This affinity profile corresponds well to that of the putative alpha1L-adrenoceptor, which has been proposed in lower urinary tract tissues. In binding studies on rabbit iris membrane however, prazosin, KMD-3213 and WB4101 displayed high affinity (pKd or pKi: 9.6, 10.3, 9.6, respectively), and BMY7378 displayed low affinity (pKi: 6.9). These results show that the binding sites typically correspond to alpha1A-adrenoceptor subtype in character, and we could not detect the significant amount of alpha1L-adrenoceptor subtype. The expression of the three distinct mRNAs that encode proteins of alpha1a-, alpha1b- and alpha1d-adrenoceptors was studied using reverse transcription-polymerase chain reaction (RT-PCR). RT-PCR demonstrated the strongest expression of the alpha1a-adrenoceptor, weak expression of the alpha1b-adrenoceptor and undetectable expression of the alpha1d-adrenoceptor. The present study suggests that alpha1A-adrenoceptor is a major subtype detectable in binding and RT-PCR studies in rabbit iris, but that the adrenergic contractions of iris dilator muscle are mediated via activation of alpha1-adrenoceptor subtype having low affinity for prazosin and WB4101.

SUBMITTER: Nakamura S

PROVIDER: S-EPMC1760646 | biostudies-other | 1999 Jul

REPOSITORIES: biostudies-other

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Project description:BackgroundThere are three turkey β-adrenoceptors: the original turkey β-adrenoceptor from erythrocytes (tβtrunc, for which the X-ray crystal structure has recently been determined), tβ3C and tβ4C-receptors. This study examined the similarities and differences between these avian receptors and mammalian receptors with regards to binding characteristics and functional high and low affinity agonist conformations.Methodology/principal findingsStable cell lines were constructed with each of the turkey β-adrenoceptors and 3H-CGP12177 whole cell binding, CRE-SPAP production and (3)H-cAMP accumulation assays performed. It was confirmed that the three turkey β-adrenoceptors are distinct from each other in terms of amino acid sequence and binding characteristics. The greatest similarity of any of the turkey β-adrenoceptors to human β-adrenoceptors is between the turkey β3C-receptor and the human β2-adrenoceptor. There are pharmacologically distinct differences between the binding of ligands for the tβtrunc and tβ4C and the human β-adrenoceptors (e.g. with CGP20712A and ICI118551). The tβtrunc and tβ4C-adrenoceptors appear to exist in at least two different agonist conformations in a similar manner to that seen at both the human and rat β1-adrenoceptor and human β3-adrenoceptors. The tβ3C-receptor, similar to the human β2-adrenoceptor, does not, at least so far, appear to exist in more than one agonist conformation.Conclusions/significanceThere are several similarities, but also several important differences, between the recently crystallised turkey β-adrenoceptor and the human β-adrenoceptors. These findings are important for those the field of drug discovery using the recently structural information from crystallised receptors to aid drug design. Furthermore, comparison of the amino-acid sequence for the turkey and human adrenoceptors may therefore shed more light on the residues involved in the existence of the secondary β-adrenoceptor conformation.

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Evaluation of alpha1-adrenoceptors in the rabbit iris: pharmacological characterization and expression of mRNA.

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