Functional evidence for NO‐synthase activation by substance P through a mechanism not involving classical tachykinin receptors in guinea‐pig ileum in vitro

Abstract

1 This study tested the hypothesis that a nitric oxide synthase (NOS) was activated in guinea-pig ileum in vitro in response to substance P (SP), and attempted to characterize the tachykinin receptor involved in this activation by the use of selective receptor agonists and antagonists. 2 Strips of guinea-pig ileum (8 x 2 mm) were superfused (Krebs, 370C, 2 ml min-') with: (i) tachykinin receptor agonists: SP, GR 73,632 (NK1), GR 64,349 (NK2), senktide (NK3), and neuropeptide (NP)'y; (ii) tachykinin receptor antagonists: CP 99,994 (NKI), SR 48,968 (NK2), SR 142,801 (NK3); (iii) nerverelated agents: carbachol (CCh), atropine, tetrodotoxin (TTX), hexamethonium; (iv) NOS inhibitors: 4 Under baseline conditions, L-NAME (1 ,M), L-NMMA (1 Mm) and AG (1 Mm), failed to contract the muscle strip. In contrast, when superfused for 3 min, 10 min after SP (0.1 gM), they induced a transient contraction of the strip (e.g. for 1 gM L-NAME: 50 to 70 s duration; amplitude 73+ 12%, n = 24). 5 The NOS inhibitor-induced contractile response was not obtained after KCl (60 mM), GR 73,632, GR 64,349, senktide or CCh (all up to 1 gM). In contrast, this contractile response was obtained after NPy (1 gM). 48,968 and SR 142,801 (1 gM) respectively, starting 5 min before SP, did not modify the response to L-NAME, superfused 10 min after SP (0.1 gM). The contractile response to L-NAME (1 gM) was blocked by atropine (1 Mm), superfused either before or after SP. In contrast, it persisted after TTX or hexamethonium (1 gM) superfused in the same conditions. 7 The amplitude of NOS inhibitor-induced contraction (1 gM) was dependent on the concentration of priming SP (1 pM to 1 Mm). In contrast, the contractile response to NOS inhibitors (1 nm to 10 gM) of the ileum strip primed with SP (0.1 gM) was not concentration-related. 8 L-NAME-induced contraction was prevented by continuous superfusion of L-Arg (1 gM), but not DArg (1 gM). In addition, the NO donor, sodium nitroprusside (1 gM) and the NO scavenger, methaemoglobin (10 Mg ml-'), both prevented the contractile response to L-NAME. 9 In summary, SP and to a lesser extent NP'y, exert a permissive action allowing contractile stimulating effects of L-NAME, L-NMMA and AG, in guinea-pig ileum in vitro, by a mechanism which apparently does not involve tachykinin NKI, NK2 and NK3 receptors. This action is likely to result from the activation of a NO-synthase by SP in the vicinity of intestinal myocytes. Thus, L-NAME, L-NMMA or AG, by blocking this SP-induced NO production, unveiled a smooth muscle contraction which involves a cholinoceptor (atropine-sensitive) mechanism.

Substance P primes the formation of hydrogen peroxide and nitric oxide in human neutrophils

“…However, detailed reports on the priming role of SP for the neutrophil NADPH oxidase system are limited [8,15,16,23,24], and studies describing the effect of SP on NO production in human neutrophils have not been done. There is recent evidence that SP directly stimulates NO production in a variety of cell types under in vitro conditions [25,26]. In this report, we examined the modulatory role of SP on the NADPH oxidase and nitric oxide pathways in human neutrophils and found that SP is a significant priming agent for production of O 2 Ϫ and H 2 O 2 , in response to PMA, and NO in response to PMA and fMLP.…”

Protein kinase A facilitates relaxation of mouse ileum via phosphorylation of neuronal nitric oxide synthase

“…In our studies, we used substance P to maintain reliable contractions during the concentration–response curve. Substance P stimulates guinea pig ileum NO synthesis and up‐regulates cAMP in astrocytoma cells (Fowler & Brannstrom, ; Garcia‐Villar, Dupuis, Martinolle, Fioramonti, & Bueno, ). Thus, substance P may influence cAMP‐dependent nNOS S1412 phosphorylation.…”

Multidisciplinary approach to nitric oxide signaling: Focus on gastrointestinal and central nervous system

“…-Cyclic GMP-dependent opening of apamin-sensitive K + channels or other types of ion channels to produce hyperpolarization and relaxation; -Indirectly by inhibiting the release of the neurotransmitters acetylcholine and substance P 41 ;…”