Characterization of NK 3 receptors in rabbit isolated iris sphincter muscle

Tachykinin NK 3 receptors were characterized in the rabbit isolated iris sphincter muscle by use of autoradiography and in vitro functional studies. [ 125 I]‐[MePhe 7 ]‐neurokinin B (NKB) (1 n m ), a selective NK 3 receptor agonist, specifically labelled a population of NK 3 receptors that were uniformly distributed throughout the rabbit iris sphincter muscle. This labelling was inhibited by unlabelled [MePhe 7 ]‐NKB (1 μ m ) but not by the NK 1 receptor antagonist CP 99994 (1 μ m ). In the presence of CP 99994 (1 μ m ), the selective NK 3 receptor agonists senktide ( n =14) and [Pro 7 ]‐NKB ( n =4), and the natural preferred ligand for the NK 3 receptor, NKB ( n =8), were potent contractile agents in the rabbit iris sphincter muscle. They all produced monophasic concentration‐effect curves with pD 2 values of 9.53±0.08, 8.56±0.09 and 9.75±0.09, and n H values of 0.93±0.03, 1.53±0.17 and 0.76±0.06, respectively. [MePhe 7 ]‐NKB ( n =12) was also a potent agonist, but produced shallow concentration‐effect curves which appeared biphasic (n H =0.45±0.04). Contractile responses to senktide were surmountably antagonized in a concentration‐dependent manner by the selective non‐peptide NK 3 receptor antagonist, SR 142801 (3–30 n m ; pA 2 = 8.9; slope = 0.99) and the non‐peptide NK 2 /NK 3 receptor antagonist, SR 48968 (3–30 μ m ; pA 2 =6.1; slope=1.5). These pA 2 values were consistent with functional rabbit NK 3 receptors more closely resembling guinea‐pig and human NK 3 receptors, than rat NK 3 receptors. SR 142801 (10–100 n m ) and SR 48968 (3 and 30 μ m ) inhibited responses to low (1 n m ) but not higher (>1 n m ) concentrations of [MePhe 7 ]‐NKB, and concentration‐effect curves to [MePhe 7 ]‐NKB became steeper and monophasic in the presence of either antagonist. SR 142801 (3–30 n m ) and SR 48968 (3–30 μ m ) also surmountably antagonized concentration‐effect curves to [Pro 7 ]‐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 n m ) tested against NKB, and SR 48968 (3 μ m ) tested against [Pro 7 ]‐NKB and NKB, yielded apparent pA 2 estimates of 9.3, 6.8 and 6.4, respectively, consistent with blockade of NK 3 receptors. SR 142801 (100 n m ) had no effect on contractions induced by transmural nerve stimulation (2 Hz, 0.3 ms, 20 V for 30 s), whereas CP 99994 (1 μ m ) abolished these responses. Phenoxybenzamine pretreatment (20 μ m , 10 min) markedly reduced maximum responses to [MePhe 7 ]‐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 [MePhe 7 ]‐NKB after phenoxybenzamine pretreatment were unaffected by 1 μ m CP 99994 (maximum response=41±9.4%, n =4). These results demonstrate autoradiographically and functionally, the presence of NK 3 receptors in rabbit iris sphincter muscle that mediate contractile responses to NK 3 receptor agonists, but not to sensory trigeminal nerve stimulation. The present data with senktide and selective NK 3 receptor antagonists suggest that functional rabbit NK 3 receptors more closely resemble human and guinea‐pig NK 3 receptors than rat NK 3 receptors. However, the pharmacological profiles of [MePhe 7 ]‐NKB, SR 142801 and SR 48968 suggest the presence of an ‘atypical’ NK 3 receptor or a heterogeneous population of NK 3 receptors in this tissue.British Journal of Pharmacology (1997) 120 , 93–101; doi: 10.1038/sj.bjp.0700867