Characterization of tachykinin receptors in the uterus of the oestrogen‐primed rat

The aim of our study was to characterize the tachykinin receptor population in the oestrogen‐primed rat uterus. For this purpose, we investigated the receptor type(s) responsible for tachykinin‐induced contraction of longitudinally‐arranged smooth muscle layer. The effects of substance P (SP), neurokinin A (NKA), neurokinin B (NKB) and several of their analogues with well‐defined selectivities for tachykinin NK 1 , NK 2 and NK 3 receptors were studied and their inhibition by the selective nonpeptide tachykinin receptor antagonists ( S )1‐(2‐[3‐(3,4‐dichlorophenyl)‐1‐(3‐isopropoxyphenylacetyl)piperidin‐3‐yl]ethyl)‐4‐phenyl‐1‐azoniabicyclo[2.2.2]octane chloride (SR 140333, NK 1 ‐selective), ( S )‐N‐methyl‐N[4‐(4‐acetylamino‐4‐phenylpiperidino)‐2‐(3,4‐dichlorophenyl)butyl]benzamide (SR 48968, NK 2 ‐selective) and ( R )‐(N)‐(1‐(3‐(1‐benzoyl‐3‐(3,4‐dichlorophenyl)piperidin‐3‐yl)propyl)‐ 4‐phenylpiperidin‐ 4‐yl)‐N‐ methylacetamide (SR 142801, NK 3 ‐selective) was evaluated. Additionally, expression of tachykinin receptor mRNA was examined by using the reverse transcription‐polymerase chain reaction (RT‐PCR). SP, NKA, [Nle 10 ]‐NKA(4‐10), the analogue with selectivity at the tachykinin NK 2 receptor type, and NKB elicited concentration‐dependent contractions of the rat uterus. The pD 2 values were 5.95±0.19; 6.73±0.21; 7.53±0.12 and 5.76±0.21, respectively. The selective agonist for the tachykinin NK 1 receptor [Sar 9 Met(O 2 ) 11 ]‐SP produced a small phasic response in the nanomolar concentration range. The selective tachykinin NK 3 receptor agonist [MePhe 7 ]‐NKB failed to induce any significant contraction. In the presence of the neutral endopeptidase inhibitor phosphoramidon (1 μ M ), the log concentration‐response curves to exogenous tachykinins and their analogues were shifted significantly leftwards. The pD 2 values were 6.12±0.10, 8.04±0.07, 7.89±0.03 and 6.59±0.07 for SP, NKA, [Nle 10 ]‐NKA(4‐10) and NKB, respectively. In the presence of phosphoramidon (1 μ M ), [Sar 9 Met(O 2 ) 11 ]‐SP (1 n M –0.3 μ M ) induced concentration‐dependent contractions of increasing amplitude when only one concentration of drug was applied to each uterine strip and the pD 2 value was 7.61±0.89. [MePhe 7 ]‐NKB induced small, inconsistent contractions and, therefore, a pD 2 value could not be calculated. In experiments performed in the presence of phosphoramidon (1 μ M ), SR 48968 (3 n M –0.1 μ M ) caused parallel and rightward shifts in the log concentration‐response curves of NKA. The calculated p K B value was 9.16±0.08 and the slope of the Schild regression was 1.28±0.24. SR 48968 (0.1 μ M ) also antagonized responses to SP with an apparent p K B value of 7.63±0.13. SR 48968 (0.1 μ M ) inhibited contractions elicited by NKB (1 n M –3 μ M ) and [Nle 10 ]‐NKA(4–10) (0.1 n M –3 μ M ) but had no effect on the response evoked by [Sar 9 Met(O 2 ) 11 ]‐SP (0.1 μ M ). SR 140333 (0.1 μ M ) inhibited responses to SP with an apparent p K B value of 7.19±0.22. This compound did not significantly affect responses to NKA, [Nle 10 ]‐NKA(4‐10) and NKB, but suppressed [Sar 9 Met(O 2 ) 11 ]‐SP (0.1 μ M )‐induced contraction. SR 142801 (0.1 μ M ) had no effect on responses to natural tachykinins or their analogues. Total RNA was extracted from some of the uteri used in functional studies. RT‐PCR assays revealed single bands corresponding to the expected product sizes encoding cDNA for tachykinin NK 1 (587 base pairs) and NK 2 receptors (491 base pairs) ( n =6 different animals). A very low abundance transcript corresponding to the 325 base pairs product expected for the tachykinin NK 3 receptor was detected. The present data show that functionally active tachykinin NK 1 and NK 2 receptors are expressed in the oestrogen‐primed rat uterus. The NK 2 receptor type seems to be the most important one involved in the contractile responses elicited by tachykinins. NK 3 receptors are present in trace amounts and seem not to be involved in tachykinin‐induced contractions.British Journal of Pharmacology (1998) 123 , 259–268; doi: 10.1038/sj.bjp.0701613