IN-VITRO CHARACTERIZATION OF TACHYKININ NK2-RECEPTORS MODULATING MOTOR-RESPONSES OF HUMAN COLONIC MUSCLE STRIPS

1 Human in vitro preparations of transverse or distal colonic circular smooth muscle were potently and dose-dependently contracted by neurok inin A (EC50, 4.9 nM). the tachykinin NK2-receptor selective agonist [ beta-Ala(8)]neurokinin A (4-10) ([beta-Ala(8)]NKA (4-10)) (EC50, 5.0 n M), neurokinin B (EC50, 5.3 nM) and substance P (EC50, 160 nM), but no t by the tachykinin NK1-receptor selective agonist [Sar(9)Met(O-2)(11) ] substance P, or the NK3-receptor selective agonists, senktide and [M ePhe(7)] neurokinin B. No regional differences between transverse and distal colon were observed in response to [beta-Ala(8)]NKA (4-10). 2 A tropine (1 mu M) and tetrodotoxin (1 mu M) did not significantly inhib it responses to [beta-Ala(8)]NKA (4-10), neurokinin A, substance P or neurokinin B.3 The newly developed non-peptide antagonists for tachyki nin NK2-receptors SR 48968, SR 144190 and its N-demethyl (SR 144743) a nd N,N-demethyl (SR 144782) metabolites, were used to challenge agonis t responses, as appropriate. SR 144190 and the metabolites all potentl y and competitively antagonized the response to [beta-Ala(8)]NKA (4-10 ), with similar potency (Schild plot pA(2) values 9.4, 9.4 and 9.3, sl ope = 1). SR 48968 antagonism was not competitive: the Schild plot slo pe was biphasic with a high (X intercept similar to 9.3) and a low (X intercept 8.4, slope 1.6) affinity site. Co-incubation of SR 48968 (10 , 100 nM) and SR 144782 (10 nM) produced additive effects; in this exp erimental condition, SR 48968 apparent affinity (pK(B)) was 8.2. In ad dition, SR 144782 (0.1 mu M) antagonized responses to neurokinin A, su bstance P and neurokinin B, with pK(B) consistent with its affinity fo r tachykinin NK2-receptors. The potent and selective NK1 and NK3-recep tor antagonists, SR 140333 and SR 142801 (both 0.1 mu M), failed to in hibit contractions induced by SP or NKB. 4 In conclusion, the in vitro mechanical responses of circular smooth muscle preparations from huma n colon are strongly consistent with the presence of non-neuronal tach ykinin NK2-receptors, but not tachykinin NK1- or NK3-receptors. Our fi ndings with SR 48968 suggest the existence of two tachykinin NK2-recep tor subtypes, that it seems to distinguish, unlike SR 144190 and its m etabolites. However, the precise nature of SR 48968 allotopic antagoni sm remains to be elucidated, since allosteric effects at the tachykini n NK2-receptor might well account for the complexity of the observed i nteraction.