Characterization of NK3 receptors in rabbit isolated iris sphincter muscle.
1. Tachykinin NK3 receptors were characterized in the rabbit isolated iris sphincter muscle by use of autoradiography and in vitro functional studies. 2. [125I]-[MePhe7]-neurokinin B (NKB) (1nM), a selective NK3 receptor agonist, specifically labelled a population of NK3 receptors that were uniformly distributed throughout the rabbit iris sphincter muscle. This labelling was inhibited by unlabelled [MePhe7]-NKB (1 microM) but not by the NK1 receptor antagonist CP 99994 (1 microM). 3. In the presence of CP 99994 (1 microM), the selective NK3 receptor agonists senktide (n = 14) and [Pro7]-NKB (n = 4), and the natural preferred ligand for the NK3 receptor, NKB (n = 8), were potent contractile agents in the rabbit iris sphincter muscle. They all produced monophasic concentration-effect curves with pD2 values of 9.53 +/- 0.08, 8.56 +/- 0.09 and 9.75 +/- 0.09, and nH values of 0.93 +/- 0.03, 1.53 +/- 0.17 and 0.76 +/- 0.06, respectively. [MePhe7]-NKB (n = 12) was also a potent agonist, but produced shallow concentration-effect curves which appeared biphasic (nH = 0.45 +/- 0.04). 4. Contractile responses to senktide were surmountably antagonized in a concentration-dependent manner by the selective non-peptide NK3 receptor antagonist, SR 142801 (3-30 nM; pA2 = 8.9; slope = 0.99) and the non-peptide NK2/NK3 receptor antagonist, SR 48968 (3-30 microM; pA2 = 6.1; slope = 1.5). These pA2 values were consistent with functional rabbit NK3 receptors more closely resembling guinea-pig and human NK3 receptors, than rat NK3 receptors. SR 142801 (10-100 nM) and SR 48968 (3 and 30 microM) inhibited responses to low (< or = 1 nM) but not higher (> 1 nM) concentrations of [MePhe7]-NKB, and concentration-effect curves to [MePhe7]-NKb became steeper and monophasic in the presence of either antagonist. 5. SR 142801 (3-30 nM) and SR 48968 (3-30 microM) also surmountably antagonized concentration-effect curves to [Pro7]-NKB and NKB, although results were more difficult to interpret, since the relationship between log concentration-ratios and the concentration of antagonist used did not adhere to the Schild equation. However, analysis of data with the lowest concentration of SR 142801 (3 nM) tested against NKB, and SR 48968 (3 microM) tested against [Pro7]-NKB and NKB, yielded apparent pA2 estimates of 9.3, 6.8 and 6.4, respectively, consistent with blockade of NK3 receptors. 6. SR 142801 (100 nM) had no effect on contractions induced by transmural nerve stimulation (2 Hz, 0.3 ms, 20 V for 30 s), whereas CP 99994 (1 microM) abolished these responses. 7. Phenoxybenzamine pretreatment (20 microM, 10 min) markedly reduced maximum responses to [MePhe7]-NKB (from 101 +/- 6.2% to 38 +/- 9.5% reference contraction, n = 4) and induced a marked (10 fold) rightward shift in the concentration-effect curve. The residual responses to [MePhe7]-NKB after phenoxybenzamine pretreatment were unaffected by 1 microM CP 99994 (maximum response = 41 +/- 9.4%, n = 4). 8. These results demonstrate autoradiographically and functionally, the presence of NK3 receptors in rabbit iris sphincter muscle that mediate contractile responses to NK3 receptor agonists, but not to sensory trigeminal nerve simulation. The present data with senktide and selective NK3 receptor antagonists suggest that functional rabbit NK3 receptors more closely resemble human and guinea-pig NK3 receptors than rat NK3 receptors. However, the pharmacological profiles of [MePhe7]-NKB, SR 142801 and SR 48968 suggest the presence of an 'atypical' NK3 receptor or a heterogeneous population of NK3 receptors in this tissue.
Medhurst
A D
AD
Department of Neurology Research, SmithKline Beecham Pharmaceuticals, Harlow, Essex.
Parsons
A A
AA
Roberts
J C
JC
Hay
D W
DW
D000317:Adrenergic alpha-Antagonists / Q000494:pharmacology; D000818:Animals; D001345:Autoradiography; D001549:Benzamides / Q000494:pharmacology; D004558:Electric Stimulation; D066298:In Vitro Techniques; D007498:Iris / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D009119:Muscle Contraction / Q000187:drug effects; D009130:Muscle, Smooth / Q000187:drug effects / Q000378:metabolism*; D015287:Neurokinin B / Q000494:pharmacology; D010446:Peptide Fragments / Q000494:pharmacology; D010643:Phenoxybenzamine / Q000494:pharmacology; D010880:Piperidines / Q000494:pharmacology; D011817:Rabbits; D018041:Receptors, Neurokinin-2 / Q000037:antagonists & inhibitors; D018042:Receptors, Neurokinin-3 / Q000819:agonists / Q000037:antagonists & inhibitors / Q000187:drug effects*; D013373:Substance P / Q000031:analogs & derivatives / Q000494:pharmacology
D000317:Adrenergic alpha-Antagonists; D001549:Benzamides; D010446:Peptide Fragments; D010880:Piperidines; D018041:Receptors, Neurokinin-2; D018042:Receptors, Neurokinin-3; D010643:Phenoxybenzamine; C054724:senktide; D013373:Substance P; C073839:SR 48968; D015287:Neurokinin B; C091554:SR 142801