NKA-induced Contraction Research Articles

Efficient Ca 2+ mobilization induced by neurokinin A in rat vas deferens

Although both neurokinin A (NKA) and norepinephrine (NE) induced similar maximal contractions in the epididymal and the prostatic site of vas deferens, NKA affected sensitivity more potently than did NE in both sites. The NKA-induced contractions were more strongly inhibited by nicardipine, a dihydropyridine Ca 2+ entry blocker, or by elimination of extracellular Ca 2+ (Ca 2+ o) in both sites. However, ryanodine, which interferes with the release of intracellular C 2+ (Ca 2+ i), abolished the contractions caused by NKA in the prostatic site whereas it had no effect in the epididymal site. These results suggest that NKA-induced contraction utilizes both Ca 2+ i and Ca 2+ o in the prostatic site but mobilizes only Ca 2+ o in the epididymal site. Ca 2+ i concentration ([Ca 2+] i)was measured directly with a Ca 2+ -sensitive fluorescent dye, fura-2. In the epididymal site NKA induced contractions with smaller increase in [Ca 2+] i compared to that necessary for NE-induccd contractions. These results suggest that NKA utilizes Ca 2+ more efficiently than does NE and plays a role as a neuromodulator in rat vas deferens.

Leu 9ψ(CH 2NH)Leu 10]-Neurokinin A(4–10) (MDL 28, 564) distinguishes tissue tachykinin peptide NK 2 receptors

The neurokinin A analogue, MDL 28, 564 (AspSerPheValGlyLeuCH 2NHLeuNH 2), inhibited 125I-NKA binding to hamster urinary bladder NK 2 receptors with a K I of 130 nM. For rat submaxillary gland NK 1 receptors and cerebral cortical NK 3 receptors, the K I's for MDL 28,564 were>250 μM and >500 μM, respectively. MDL 28,564 did not relax dog carotid artery (NK 1 tissue) or contract rat portal vein (NK 3 tissue). In guinea-pig trachea tissues, MDL 28,564 stimulated phosphatidylinositol turnover and induced contraction with maximum effects similar to those of neurokinin A. In hamster urinary bladder tissue, MDL 28,564 stimulated phosphatidylinositol turnover with maximum effect only 10% of that of neurokinin A, did not produce sustained contraction itself and antagonized NKA-induced contraction. MDL 28,564 also produced full contraction in rabbit pulmonary artery (NK 2 tissue) but was inactive in rat vas deferens (NK 2 tissue). These data with MDL 28,564 are consistent with the NK 2 receptors in guinea-pig trachea and rabbit pulmonary artery being different from those in hamster urinary bladder and rat vas deferens.

Effect of thiorphan on response of the guinea-pig gallbladder to tachykinins

Tachykinins produced a concentration-related contraction of the isolated guinea-pig gallbladder, with a rank order of potency neurokinin A (NKA) > Arg-neurokinin B = neurokinin B (NKB) > substance P (SP). Only the effect of SP was potentiated by thiorphan (0.1–10 μM). A significant enhancement of the response to SP was also produced by captopril (1 μM). [Nle 10]NKA-(4–10) and [β-Ala 8]NKA-(4–10), selective NK-2 receptor agonists, were active, whereas [Pro 9]SP sulfone (selective NK-1 agonist) was almost ineffective. [MePhe 7]NKB (selective NK-3 agonist) had some activity but only at high concentrations. Septide was almost ineffective and DiMeC 7 had an action comparable to that of [MePhe 7]NKB. None of the effects induced by these synthetic tachykinin analogs were significantly potentiated by thiorphan. Capsaicin (10 μM) produced a contraction which was unaffected by thiorphan. Both capsaicin and NKA-induced contractions were antagonized by Spantide at concentrations (5–10 μM) which had no effect against the atropine-sensitive contractions produced by electrical field stimulation. Capsaicin (1 μM) produced a consistent release of SP-like immunoreactivity (SP-LI) and a second application of the drug had no further effect, indicating complete desensitization. SP-LI release by capsaicin was almost doubled in the presence of thiorphan. These findings indicate that NK-2 and possibly some NK-3 receptors mediate the contractile response of the guinea-pig gallbladder to tachykinins. Both exogenous and endogenous (released by capsaicin) SP were degraded to a significant extent in this organ via a thiorphan-sensitive mechanism, the identity of which remains to be established.

Royal academy of medicine in Ireland section of biological sciences

THE EFFECT OF SOME NEUROKININ A (NKA) ANALOGUES ON NKA-INDUCED CONTRACTION OF GUINEAPIG TRACHEAL SMOOTH MUSCLE IN VITRO J. M. Allen*, A. A. Abu Shanab, D. J. S. Guthrie, G. B. Irvine, B. Walker. *Biomedical Sciences Research Centre, University of Ulster, Coleraine, and Division of Biochemistry, The Queen's University of Belfast, Belfast. The peptide neurokinin A is found in the lung and is known to cause bronchoconstriction both in vivo and in vitro (rheodorssonNerheim et al, Acta. Physiol. Scand. 1985: 124, 129, Joos et al, Thorax. 1987: 243,901), but an understanding of its role in control of airway function in health and disease has been difficult because of the lack of specific antagonists. It was the purpose of this study to synthetise analogues of NKA and to test them for pharmacological antagonism of NKA-induced contraction of guinea-pig tracheal smooth muscle. We have previously reported that NKA analogue in which glycine 8 was replaced by aminoisobutyric acid (Aib) is an antagonist of NKA, but not Substance P, activity on tracheal smooth muscle from guinea-pig (Abu Shanab et al, Biochem. Soc. Trans. 1990: 18, 286). We now report the effect on biological activity of deletions at the N-terminus. NKA was purchased from Biosyn Ltd. (Belfast, N. Ireland) and the peptides [Ala s, Aib s, Leu 1~ NKA (4-10), [Ala ~, Aib 8, Leu 1~ NKA (2-10) and [Ala s, Aib s, Leu 1~ NkA, hereafter referred to as peptides 1, 2 and 3 respectively, were synthetised by solid phase methods. Isometric contraction of isolated guinea-pig tracheal rings in response to exogenous NKA (10 -1~ to lO-6M) was recorded as described previously (Abu Shanab et al, Biochem. Soc. Trans. 1990: 18, 286) and the effect of the peptide analogues on the NKA doseresponse relationship examined. In the presence of peptide 3 (10 -8 to 10-6M) NKA evoked contractions were reduced at all doses producing acharacteristic shift in the NKA dose-response curve suggestive of classical non-competitive antagonism, although contractions to substance P (SP; 10 -8 to 10-6M) were unaltered by analogue. In contrast, peptide 2 (104 to 1 ff6M) caused a dose-dependent shift to the right of the NKA doseresponse curve with no depression of the maximum NKA response. Peptide 2 showed no pharmaclogical antagonism to SP. A schild regression plot for peptide 2 antagonism of NKA evoked contractions yielded a straight line of slope -0.4 with an interscept (PA2) of 7.7 on the abscissa. Unlike peptides 2 and 3, peptide 1 (10-6M) itself caused contraction of the tracheal preparation. In conclusion, peptide analogues that selectively antagonise the activity of NKA on tracheal smooth muscle have been produced by replacement of Gly s in a NKA ,,~icgue that also had the substitutions Ala s and Leu 1~ Removal G~ the N-terminal residue did not abolish this selectivity although the nature of the antagonism was altered. Removal of a further two amino acid residues from the Nterminus resulted in the generation of agonist activity.