Functional characterisation of tachykinin receptors mediating ion transport in porcine jejunum

Abstract In the present study, tachykinin receptors (designated NK 1 , NK 2 and NK 3 ) involved in regulation of ion transport in porcine jejunum were characterised. Stripped tissue preparations were mounted in Ussing chambers and short-circuited. Substance P produced a concentration dependent increase in short-circuit current, the relationship showing a double sigmoidal form. The non-peptide NK 1 receptor antagonist, CP 99,994 ((2 S ,3 S )-3-(2-methoxybenzyl)amino-2-phenylpiperidine), caused a dextral shift of the first sigmoidal response, indicating the involvement of an NK 1 receptor. This was further supported by a concentration-dependent response of the NK 1 receptor agonist [Sar 9 Met(O 2 ) 11 ]substance P with an EC 50 value of 235.0±53.9 nM. Increasing concentrations of CP 99,994 (0.1, 0.3 and 1 μM) produced a parallel dextral shift of the [Sar 9 Met(O 2 ) 11 ]substance P curve with a slope of the Schild regression significantly different from unity (1.59). The neurokinin A concentration–response curve, with an EC 50 value of 68.87±16.23 nM, was not significantly changed by the non-peptide NK 2 receptor antagonist SR 48,968 (( S )- N -methyl- N -(4-(4-acetylamino-4-phenylpiperidino)-2-(3,4-dichlorophenyl)butyl)bezamide). In additional studies, the peptide NK 2 receptor antagonists, GR 94,800 (PhCO–Ala–Ala–DTrp–Phe–DPro–Pro–NleNH 2 ) and PD 147,714 ((2,3-diOMeZ)-( S )Trp( S )αMePheGlyNH 2 ), did not change the response to neurokinin A. However, CP 99,994 totally inhibited neurokinin A responses at 0.5 μM and above. The NK 2 receptor agonist, [β-Ala 8 ]neurokinin A-(4–10), caused only an increase in short-circuit current in μM concentrations, whereas the NK 3 receptor agonist, senktide, did not elicit a response. These results indicate, that substance P and neurokinin A mediate ion transport in porcine jejunum through NK 1 receptors. However, tachykinins seem to activate another receptor. Two active conformers of the NK 1 receptor might be present.