The role of substance P and bradykinin in the cough reflex and bronchoconstriction in guinea-pigs

Abstract

In this study we investigated the ability of aerosolized substance P to induce either cough or bronchoconstriction in guinea-pigs. We have also examined whether pre-treatment, by the inhaled route, of animals with a combination of the neutral endopeptidase inhibitor, phosphoramidon (10⁻³ M), and the diaminopeptidase IV inhibitor, diprotin A (10⁻³ M), enhances the airway response to substance P. Moreover, we also assessed whether aerosol pre-treatment of guinea-pigs with either substance P or bradykinin, at 10⁻⁴ M, affects the citric acid-induced cough and/or bronchoconstriction. Challenge of guinea-pigs with substance P only at 10⁻³ M resulted in significant bronchconstriction but only a weak and variable cough response (1.1±0.6; P>0.05). Pre-treatment of guinea-pigs with both phosphoramidon and diprotin A resulted in a small non-significant increase in the cough response (2.8±0.9 vs. 1.1±0.6; P>0.05) but significantly enhanced substance P-induced bronchoconstriction (P<0.05). Moreover, exposure of guinea-pigs to substance P (10⁻⁴ M) prior to citric acid challenge (0.6M) resulted in a significant (P<0.05) enhancement of the citric acid-induced bronchoconstriction but not the citric acid-induced cough (11.7±1.8 vs. 12.8±1.5; P>0.05). In contrast, exposure of guinea-pigs to bradykinin (10⁻⁴ M) prior to the citric acid challenge resulted in a significant enhancement of the cough response (9.2±1.9 vs. 25.8±2.5; P<0.05) but not the bronchoconstriction (P>0.05). These data do not support a major peripheral role for substance P in the cough reflex, although bradykinin is able to sensitize the cough reflex. Furthermore, these data suggest that bronchoconstriction, induced by citric acid, is not responsible for the cough associated with this irritant.

Introduction

Cough is generally considered as reflex defensive mechanism which is initiated through activation of sensory nerves in the wall of the respiratory tract, from the larynx down to the bronchi, in response to mechanical, chemical or inflammatory mediator stimulation (Adcock, 2003). Despite cough being a very common problem, the mechanisms contributing to this symptom remain poorly understood. However, there is evidence to support the involvement of airway vagal afferents such as sensory C-fibres and rapidly adapting receptors in the cough reflex, as well as other symptoms of respiratory diseases such as bronchospasm (Adcock, 2003).

In addition to their afferent function, C-fibres have also been shown to have an efferent function. These non-myelinated C-fibres contain the tachykinins substance P, neurokinin A and neurokinin B which, upon release, act on NK₁, NK₂, NK₃ receptors respectively to mediate several functions (Regoli et al., 1994). Whilst inhalation of citric acid stimulates both C-fibres and rapidly adapting receptors (RARs), capsaicin appears to stimulate only C-fibres and both these agents have been shown to induce cough, in several species including man, and also bronchoconstriction (Lalloo et al., 1995, Yasumitsu et al., 1996, Kondo et al., 1998, Undem et al., 2002, El-Hashim et al., 2004). Additionally, substance P has also been reported to stimulate RARs (Prabhakar et al., 1987, Matsumoto et al., 1994, Bonham et al., 1996) which are also reported to be involved in cough (Canning, 2002).

The fact that neurokinins have been reported to induce cough and that citric acid and capsaicin cause airway neuropeptide release, induce cough and bronchonstriction (the latter two being blocked by neurokinin receptor antagonists) has led to the belief that the peripherally released neuropeptides may be directly responsible for mediating both cough and bronchoconstriction induced by these irritants and that the bronchoconstriction may also, indirectly, enhance the cough response to these irritants (Kohrogi et al., 1988, Takahama et al., 1993, Ujiie et al., 1993, Yasumitsu et al., 1996).

A role for substance P in the peripheral initiation of the cough reflex is also suggested by several other lines of evidence. Tussive stimuli induce substance P release (Martins et al., 1991a, Martins et al., 1991b) and exposure of guinea-pigs to substance P, in concentrations as low as 10⁻¹⁷ M, has been reported to induce a cough response (Kohrogi et al., 1988, Takahama et al., 1993, Takahama et al., 1995). Furthermore, the neutral endopeptidase inhibitor, phosphoramidon, has been reported to enhance both substance P and histamine-induced cough (Sekizawa et al., 1995, Takahama et al., 1995). Finally, it has been shown that the neurokinin NK₁ receptor antagonist and the dual NK₁ and NK₂ peptide receptor antagonists, N²-[(4R)-4-hydroxy-1-(1-methyl-1 H-indol-3-yl)carbonyl-l-prolyl]-N-methyl-N-phenylmethyl-3-(2-naphthyl)-l-alaninamide) (FK888) and (N-[N²-[2,3-didehydro-N-methyl-N-[N-[3-(2-penthylphenyl)-propionyl]-l-threonyl]tyrosyl]-l-leucynyl]-d-phenylalanyl]-l-allo-threonyl]-l-asparaginyl]-l-serine-v-lactone) (FK224) respectively, are both effective anti-tussive agents in different types of guinea-pig models of cough (Ujiie et al., 1993, Sekizawa et al., 1995, Xiang et al., 1998), when administered by the inhaled route. Taken together, these studies would suggest a role for substance P in cough and would also imply that the mechanism of action of this peptide may be through a local airways effect.

Not all studies however, have been able to confirm that exposure to substance P, by the inhaled route, induces a cough response. For example, exposure of guinea-pigs to substance P at 10⁻⁴ M (Fox et al., 1996a, Fox et al., 1996b) or pigs, at 10⁻⁵ M, (Moreaux et al., 2000) does not result in a cough response. Similarly, no cough response has been reported when normal humans inhale substance P (Joos et al., 1987). Thus the peripheral role of substance P in eliciting the cough reflex is far from clear.

The release of some inflammatory mediators in the airways, such as substance P and bradykinin, may not only induce cough or bronchoconstriction but may also sensitize these airway responses to other stimuli (Fox et al., 1996a, Fox et al., 1996b, Moreaux et al., 2000, Boichot et al., 1996). However, whether exposure of animals to inflammatory mediators results in the simultaneous enhancement of both cough and bronchoconstriction is not known.

The aims of this study were to investigate the ability of substance P to induce cough and/or bronchonconstriction in guinea-pigs and whether inhibition of the enzymes neutral endopeptidase and diaminopeptidase IV, by phosphoramidon and diprotin A respectively, can enhance the effect of substance P-induced cough and/or bronchconstriction. Futhermore, we also investigated whether bradykinin and substance P can cause simultaneous enhancement of citric acid-induced cough or bronchoconstriction.