David Gazzieri

Hydrogen sulfide causes vanilloid receptor 1-mediated neurogenic inflammation in the airways

1 Hydrogen sulfide (H2S) is described as a mediator of diverse biological effects, and is known to produce irritation and injury in the lung following inhalation. Recently, H2S has been found to cause contraction in the rat urinary bladder via a neurogenic mechanism. Here, we studied whether sodium hydrogen sulfide (NaHS), used as donor of H2S, produces responses mediated by sensory nerve activation in the guinea‐pig airways. 2 NaHS evoked an increase in neuropeptide release in the airways that was significantly attenuated by capsaicin desensitization and by the transient receptor potential vanilloid 1 (TRPV1) antagonist capsazepine. In addition, NaHS caused an atropine‐resistant contraction of isolated airways, which was completely prevented by capsaicin desensitization. Furthermore, NaHS‐induced contraction was reduced by TRPV1 antagonism (ruthenium red, capsazepine and SB366791), and was abolished by pretreatment with the combination of tachykinin NK1 (SR140333) and NK2 (SR48968) receptor antagonists. 3 In anesthetized guinea‐pigs, intratracheal instillation of NaHS increased the total lung resistance and airway plasma protein extravasation. These two effects were reduced by TRPV1 antagonism (capsazepine) and tachykinin receptors (SR140333 and SR48968) blockade. 4 Our results provide the first pharmacological evidence that H2S provokes tachykinin‐mediated neurogenic inflammatory responses in guinea‐pig airways, and that this effect is mediated by stimulation of TRPV1 receptors on sensory nerves endings. This novel mechanism may contribute to the irritative action of H2S in the respiratory system.

Neurogenic responses mediated by vanilloid receptor‐1 (TRPV1) are blocked by the high affinity antagonist, iodo‐resiniferatoxin

Stimulation of the vanilloid receptor‐1 (TRPV1) results in the activation of nociceptive and neurogenic inflammatory responses. Poor specificity and potency of TRPV1 antagonists has, however, limited the clarification of the physiological role of TRPV1. Recently, iodo‐resiniferatoxin (I‐RTX) has been reported to bind as a high affinity antagonist at the native and heterologously expressed rat TRPV1. Here we have studied the ability of I‐RTX to block a series of TRPV1 mediated nociceptive and neurogenic inflammatory responses in different species (including transfected human TRPV1). We have demonstrated that I‐RTX inhibited capsaicin‐induced mobilization of intracellular Ca2+ in rat trigeminal neurons (IC50 0.87 nM) and in HEK293 cells transfected with the human TRPV1 (IC50 0.071 nM). Furthermore, I‐RTX significantly inhibited both capsaicin‐induced CGRP release from slices of rat dorsal spinal cord (IC50 0.27 nM) and contraction of isolated guinea‐pig and rat urinary bladder (pKB of 10.68 and 9.63, respectively), whilst I‐RTX failed to alter the response to high KCl or SP. Finally, in vivo I‐RTX significantly inhibited acetic acid‐induced writhing in mice (ED50 0.42 μmol kg−1) and plasma extravasation in mouse urinary bladder (ED50 0.41 μmol kg−1). In in vitro and in vivo TRPV1 activated responses I‐RTX was ∼3 log units and ∼20 times more potent than capsazepine, respectively. This high affinity antagonist, I‐RTX, may be an important tool for future studies in pain and neurogenic inflammatory models.

Development of the First Ultra-Potent “Capsaicinoid” Agonist at Transient Receptor Potential Vanilloid Type 1 (TRPV1) Channels and Its Therapeutic Potential

Olvanil (N-9-Z-octadecenoyl-vanillamide) is an agonist of transient receptor potential vanilloid type 1 (TRPV1) channels that lack the pungency of capsaicin and was developed as an oral analgesic. Vanillamides are unmatched in terms of structural simplicity, straightforward synthesis, and safety compared with the more powerful TRPV1 agonists, like the structurally complex phorboid compound resiniferatoxin. We have modified the fatty acyl chain of olvanil to obtain ultra-potent analogs. The insertion of a hydroxyl group at C-12 yielded a compound named rinvanil, after ricinoleic acid, significantly less potent than olvanil (EC50 = 6 versus 0.7 nM), but more versatile in terms of structural modifications because of the presence of an additional functional group. Acetylation and phenylacetylation of rinvanil re-established and dramatically enhanced, respectively, its potency at hTRPV1. With a two-digit picomolar EC50 (90 pM), phenylacetylrinvanil (PhAR, IDN5890) is the most potent vanillamide ever described with potency comparable with that of resiniferatoxin (EC50, 11 pM). Benzoyl- and phenylpropionylrinvanil were as potent and less potent than PhAR, respectively, whereas configurational inversion to ent-PhAR and cyclopropanation (but not hydrogenation or epoxidation) of the double bond were tolerated. Finally, iodination of the aromatic hydroxyl caused a dramatic switch in functional activity, generating compounds that behaved as TRPV1 antagonists rather than agonists. Since the potency of PhAR was maintained in rat dorsal root ganglion neurons and, particularly, in the rat urinary bladder, this compound was investigated in an in vivo rat model of urinary incontinence and proved as effective as resiniferatoxin at reducing bladder detrusor overactivity.

Expression and functional pharmacology of the bradykinin B1 receptor in the normal and inflamed human gallbladder

Background and aims: It has recently been described that bradykinin B2 receptors are expressed in the human gallbladder and that their activation induces a powerful contraction, especially in acute cholecystitis tissues. Here the role of the B1 receptor in the contractility of control and inflamed human gallbladder was investigated. Methods: Strips of human gallbladder from either acute gallstone cholecystitis or elective gastro-entero-pancreatic surgery (control) were assessed in vitro and processed for reverse transcription-PCR analysis. Cumulative concentration–response curves with the selective B1 receptor agonist, Lys-Des-Arg9-bradykinin, cholecystokinin and carbachol were performed in control and cholecystitis specimens. Results: Lys-Des-Arg9-bradykinin concentration-dependently contracted strips of control gallbladders and its motor effect was higher in inflamed gallbladders. Lys-Des-Arg9-bradykinin-induced contraction was not altered by pretreatment with the selective bradykinin B2 receptor antagonist, HOE140 (1 μM), the NK1 (SR140333), NK2 (SR48968) and NK3 (SR142801) tachykinin receptor antagonists (all 1 μM), the muscarinic acetylcholine receptor antagonist, atropine (1 μM), and the cyclo-oxygenase inhibitor, indomethacin (5 μM). In contrast, the Lys-Des-Arg9-bradykinin-induced motor response was significantly reduced by the selective B1 receptor antagonist, R-715. Finally, quantitative real-time PCR analysis indicated that B1 receptor mRNA levels were significantly higher in cholecystitis smooth muscle specimens, when compared with that observed in control tissues. Conclusions: Bradykinin B1 receptor has an important role as a spasmogen of human gallbladder, and selective antagonists of the B1 receptor may represent a valid therapeutic option to control pain in patients with acute cholecystitis.