Transient receptor potential vanilloid 1, calcitonin gene-related peptide, and substance P mediate nociception in acute pancreatitis

The mechanism of pancreatitis-induced pain is unknown. In other tissues, inflammation activates transient receptor potential vanilloid 1 (TRPV1) on sensory nerves to liberate CGRP and substance P (SP) in peripheral tissues and the dorsal horn to cause neurogenic inflammation and pain, respectively. We evaluated the contribution of TRPV1, CGRP, and SP to pancreatic pain in rats. TRPV1, CGRP, and SP were coexpressed in nerve fibers of the pancreas. Injection of the TRPV1 agonist capsaicin into the pancreatic duct induced endocytosis of the neurokinin 1 receptor in spinal neurons in the dorsal horn (T10), indicative of SP release upon stimulation of pancreatic sensory nerves. Induction of necrotizing pancreatitis by treatment with L-arginine caused a 12-fold increase in the number of spinal neurons expressing the proto-oncogene c-fos in laminae I and II of L1, suggesting activation of nociceptive pathways. L-arginine also caused a threefold increase in spontaneous abdominal contractions detected by electromyography, suggestive of referred pain. Systemic administration of the TRPV1 antagonist capsazepine inhibited c-fos expression by 2.5-fold and abdominal contractions by 4-fold. Intrathecal, but not systemic, administration of antagonists of CGRP (CGRP(8-37)) and SP (SR140333) receptors attenuated c-fos expression in spinal neurons by twofold. Thus necrotizing pancreatitis activates TRPV1 on pancreatic sensory nerves to release SP and CGRP in the dorsal horn, resulting in nociception. Antagonism of TRPV1, SP, and CGRP receptors may suppress pancreatitis pain.     

TRPV1-mediated protection against endotoxin-induced hypotension and mortality in rats

This study was designed to test the hypothesis that the transient receptor potential vanilloid type 1 (TRPV1) channel, expressed primarily in sensory nerves, and substance P (SP), released by sensory nerves, play a protective role against lipopolysaccharide (LPS)-induced hypotension. LPS (10 mg/kg iv) elicited tachycardia and hypotension in anesthetized male Wistar rats, which peaked at 10 min and gradually recovered 1 h after the injection. Blockade of TRPV1 with its selective antagonist capsazepine (CAPZ, 3 mg/kg iv) impaired recovery given that the fall in mean arterial pressure (MAP) was greater 1 h after CAPZ plus LPS injections compared with LPS injection alone (45 +/- 5 vs. 25 +/- 4 mmHg, P < 0.05). Blockade of the neurokinin 1 (NK1) receptor with its selective antagonists RP-67580 (5 mg/kg iv) or L-733,060 (4 mg/kg iv) prevented recovery, considering that falls in MAP were not different 1 h after injections of NK1 antagonists plus LPS from their peak decreases (66 +/- 9 vs. 74 +/- 5 mmHg or 60 +/- 7 vs. 69 +/- 3 mmHg, respectively, P > 0.05). LPS increased plasma SP, norepinephrine (NE), and epinephrine (Epi) levels compared with vehicles, and the increases in plasma SP, NE, and Epi were significantly inhibited by CAPZ or RP-67580. The survival rate at 24 or 48 h after LPS injection (20 mg/kg ip) was lower in conscious rats pretreated with CAPZ or RP-67580 compared with rats treated with LPS alone (P < 0.05). Thus our results show that the TRPV1, possibly via triggering release of SP which activates the NK1 and stimulates the sympathetic axis, plays a protective role against endotoxin-induced hypotension and mortality, suggesting that TRPV1 receptors are essential in protecting vital organ perfusion and survival during the endotoxic condition.     

Regulation of epithelial transport in the jejunal mucosa of the guinea pig by neurokinins

This study characterizes the actions of the neurokinins and calcitonin-gene related peptide (CGRP) on electrolyte transport across the mucosa of the guinea pig jejunum in vitro in a modified Ussing chamber. By following changes in short circuit current (Isc) induced by substance P (SP) and neurokinins A & B (NKA & NKB) in the presence and absence of tetrodotoxin (TTX) and atropine, it was established that two distinct neurokinin receptors are involved in the regulation of electrolyte transport. NKA preferentially activates a neuronal receptor since the actions of this neurokinin were inhibited by both TTX and atropine. SP, whose actions were reduced to a lesser extent by TTX and atropine, is considered to activate preferentially a receptor on the epithelial cells. The third neurokinin, NKB, appears to act non-selectively on both the neuronal and epithelial receptors. CGRP, which per se did not affect Isc, markedly potentiated the increases in Isc induced by SP and NKB, and thus acts synergistically with the epithelial neurokinin receptor. These results suggest that two distinct neurokinin receptors (the NK-1 and the NK-2) regulate epithelial transport in the jejunal mucosa of the guinea pig, and furthermore indicate that at least one of the peptides found in enteric nerves (i.e. CGRP) modulates the actions of neurokinins on epithelial cells.     

Transient receptor potential ankyrin 1 channel localized to non-neuronal airway cells promotes non-neurogenic inflammation

Background

The transient receptor potential ankyrin 1 (TRPA1) channel, localized to airway sensory nerves, has been proposed to mediate airway inflammation evoked by allergen and cigarette smoke (CS) in rodents, via a neurogenic mechanism. However the limited clinical evidence for the role of neurogenic inflammation in asthma or chronic obstructive pulmonary disease raises an alternative possibility that airway inflammation is promoted by non-neuronal TRPA1.

Methodology/Principal Findings

By using Real-Time PCR and calcium imaging, we found that cultured human airway cells, including fibroblasts, epithelial and smooth muscle cells express functional TRPA1 channels. By using immunohistochemistry, TRPA1 staining was observed in airway epithelial and smooth muscle cells in sections taken from human airways and lung, and from airways and lung of wild-type, but not TRPA1-deficient mice. In cultured human airway epithelial and smooth muscle cells and fibroblasts, acrolein and CS extract evoked IL-8 release, a response selectively reduced by TRPA1 antagonists. Capsaicin, agonist of the transient receptor potential vanilloid 1 (TRPV1), a channel co-expressed with TRPA1 by airway sensory nerves, and acrolein or CS (TRPA1 agonists), or the neuropeptide substance P (SP), which is released from sensory nerve terminals by capsaicin, acrolein or CS), produced neurogenic inflammation in mouse airways. However, only acrolein and CS, but not capsaicin or SP, released the keratinocyte chemoattractant (CXCL-1/KC, IL-8 analogue) in bronchoalveolar lavage (BAL) fluid of wild-type mice. This effect of TRPA1 agonists was attenuated by TRPA1 antagonism or in TRPA1-deficient mice, but not by pharmacological ablation of sensory nerves.

Conclusions

Our results demonstrate that, although either TRPV1 or TRPA1 activation causes airway neurogenic inflammation, solely TRPA1 activation orchestrates an additional inflammatory response which is not neurogenic. This finding suggests that non-neuronal TRPA1 in the airways is functional and potentially capable of contributing to inflammatory airway diseases.     

ATP: a mediator for HCl-induced TRPV1 activation in esophageal mucosa

In esophageal mucosa, HCl causes TRPV1-mediated release of calcitonin gene-related peptide (CGRP) and substance P (SP) from submucosal neurons and of platelet-activating factor (PAF) from epithelial cells. CGRP and SP release was unaffected by PAF antagonists but reduced by the purinergic antagonist suramin. ATP caused CGRP and SP release from esophageal mucosa, confirming a role of ATP in the release. The human esophageal epithelial cell line HET-1A was used to identify epithelial cells as the site of ATP release. HCl caused ATP release from HET-1A, which was reduced by the TRPV1 antagonist 5-iodoresiniferatoxin. Real-time PCR demonstrated the presence of mRNA for several P2X and P2Y purinergic receptors in epithelial cells. HCl also increased activity of lyso-PAF acetyl-CoA transferase (lyso-PAF AT), the enzyme responsible for production of PAF. The increase was blocked by suramin. ATP caused a similar increase, confirming ATP as a mediator for the TRPV1-induced increase in enzyme activity. Repeated exposure of HET-1A cells to HCl over 2 days caused upregulation of mRNA and protein expression for lyso-PAF AT. Suramin blocked this response. Repeated exposure to ATP caused a similar mRNA increase, confirming ATP as a mediator for upregulation of the enzyme. Thus, HCl-induced activation of TRPV1 causes ATP release from esophageal epithelial cells that causes release of CGRP and SP from esophageal submucosal neurons and activation of lyso-PAF AT, the enzyme responsible for the production of PAF in epithelial cells. Repeated application of HCl or of ATP causes upregulation of lyso-PAF AT in epithelial cells.     

Fibroblast-like synovial cells from normal and inflamed knee joints differently affect the expression of pain-related receptors in sensory neurones: a co-culture study

Innervation of the joint with thinly myelinated and unmyelinated sensory nerve fibres is crucial for the occurrence of joint pain. During inflammation in the joint, sensory fibres show changes in the expression of receptors that are important for the activation and sensitization of the neurones and the generation of joint pain. We recently reported that both neurokinin 1 receptors and bradykinin 2 receptors are upregulated in dorsal root ganglion (DRG) neurones (the cell bodies of sensory fibres) in the course of acute and chronic antigen-induced arthritis in the rat. In this study, we begin to address mechanisms of the interaction between fibroblast-like synovial (FLS) cells and sensory neurones by establishing a co-culture system of FLS cells and DRG neurones. The proportion of DRG neurones expressing neurokinin 1 receptor-like immunoreactivity was not altered in the co-culture with FLS cells from normal joints but was significantly upregulated using FLS cells from knee joints of rats with antigen-induced arthritis. The proportion of DRG neurones expressing bradykinin 2 receptors was slightly upregulated in the presence of FLS cells from normal joints but upregulation was more pronounced in DRG neurones co-cultured with FLS cells from acutely inflamed joints. In addition, the expression of the transient receptor potential V1 (TRPV1) receptor, which is involved in inflammation-evoked thermal hyperalgesia, was mainly upregulated by co-culturing DRG neurones with FLS cells from chronically inflamed joints. Upregulation of neurokinin 1 receptors but not of bradykinin 2 and TRPV1 receptors was also observed when only the supernatant of FLS cells from acutely inflamed joint was added to DRG neurones. Addition of indomethacin to co-cultures inhibited the effect of FLS cells from acutely inflamed joints on neurokinin 1 receptor expression, suggesting an important role for prostaglandins. Collectively, these data show that FLS cells are able to induce an upregulation of pain-related receptors in sensory neurones and, thus, they could contribute to the generation of joint pain. Importantly, the influence of FLS cells on DRG neurones is dependent on their state of activity, and soluble factors as well as direct cellular contacts are crucial for their interaction with neurones.     

Cardioprotection by ischemic postconditioning is lost in isolated perfused heart from diabetic rats: Involvement of transient receptor potential vanilloid 1, calcitonin gene-related peptide and substance P

We previously found that the expression of transient receptor potential vanilloid 1 (TRPV1) and contents of calcitonin gene-related peptide (CGRP) and substance P (SP), two main neuropeptides released from TRPV1, were decreased in diabetic hearts. This study aimed to test whether decreased TRPV1, CGRP and SP levels were responsible for the loss of cardioprotection by ischemic postconditioning (IPostC) in isolated perfused heart from streptozotocin-induced diabetic rats. IPostC effectively protected non-diabetic hearts against ischemia/reperfusion injury by improving cardiac function and lowering creatine kinase (CK) and cardiac troponin I (cTnI) release, which could be abolished by inhibiting TRPV1, CGRP receptor or SP receptor. However, IPostC had no effect on cardiac function and the release of CK and cTnI in diabetic hearts regardless of whether TRPV1, CGRP receptor or SP receptor were inhibited. CGRP or SP-induced postconditioning significantly prevented both non-diabetic and diabetic hearts from ischemia/reperfusion injury by improving cardiac function and lowering CK and cTnI release. Additionally, IPostC markedly increased CGRP and SP release in non-diabetic hearts, which could be reversed with TRPV1 inhibition, but not CGRP receptor or SP receptor inhibition. However, IPostC failed to affect CGRP and SP release in diabetic hearts in the presence or absence of TRPV1, CGRP receptor or SP receptor inhibition. These results indicate that the loss of cardioprotection by IPostC during diabetes is partly associated with a failure to increase CGRP and SP release, likely due to decreased TRPV1 expression and CGRP and SP contents in diabetic hearts.     

Transient receptor potential vanilloid 1-mediated expression and secretion of endothelial cell-derived calcitonin gene-related peptide

Calcitonin gene-related peptide (CGRP), the principal transmitter in sensory nerves, could also be expressed in vascular endothelium. Transient receptor potential vanilloid 1(TRPV1), which modulates the synthesis and release of CGRP in sensory nerves, is also present in endothelial cells. The present study tested whether TRPV1 modulates the release and synthesis of CGRP in endothelial cells, and evaluated the protective effect of endothelial cell-derived CGRP. Human umbilical vein endothelial cells (HUVECs) were treated with capsaicin or hyperthermia. The level of CGRP mRNA was detected by RT-PCR, and protein level was measured by radioimmunoassay. Endothelial cell injury was induced by lysophosphatidylcholine, and evaluated by cell viability and lactate dehydrogenase activity. HUVECs expressed CGRP, both alpha- and beta-subtype. Capsaicin increased the level of CGRP in the culture medium, and up-regulated the expression of CGRP in endothelial cells. Hyperthermia also increased the level of CGRP mRNA. These effects were abolished by capsazepine, a competitive antagonist of TRPV1. Capsaicin significantly attenuated the endothelial cell damage induced by LPC, which was prevented and aggravated by capsazepine or CGRP(8-37,) antagonist of CGRP receptor. These results indicate that TRPV1 also regulates the expression and secretion of endothelial cell-derived CGRP, which affords protective effects on endothelial cells.     

Acceleration of ulcer healing by cholecystokinin (CCK): role of CCK-A receptors, somatostatin, nitric oxide and sensory nerves.

CCK exhibits a potent cytoprotective activity against acute gastric lesions, but its role in ulcer healing has been little examined. In this study we determined whether exogenous CCK or endogenously released CCK by camostate, an inhibitor of luminal proteases, or by the diversion of pancreatico-biliary secretion from the duodenum, could affect ulcer healing. In addition, the effects of antagonism of CCK-A receptors (by loxiglumide, LOX) or CCK-B receptors (by L-365,260), an inhibition of NO-synthase by N(G)-nitro-L-arginine (L-NNA), or sensory denervation by large neurotoxic dose of capsaicin on CCK-induced ulcer healing were examined. Gastric ulcers were produced by serosal application of acetic acid and animals were sacrificed 9 days after ulcer induction. The area of ulcers and blood flow at the ulcer area were determined. Plasma levels of gastrin and CCK and luminal somatostatin were measured by RIA and mucosal biopsy samples were taken for histological evaluation and measurement of DNA synthesis. CCK given s.c. reduced dose dependently the ulcer area; the threshold dose of CCK being 1 nmol/kg and the dose inhibiting this area by 50% being 5 nmol/kg. This healing effect of CCK was accompanied by a significant increase in the GBF at ulcer margin and the rise in luminal NO production, plasma gastrin level and DNA synthesis. Concurrent treatment with LOX, completely abolished the CCK-8-induced acceleration of the ulcer healing and the rise in the GBF at the ulcer margin, whereas L-365,260 remained without any influence. Treatment with camostate or diversion of pancreatic juice that raised plasma CCK level to that observed with administration of CCK-8, also accelerated ulcer healing and this effect was also attenuated by LOX but not by L-365,260. Inhibition of NO-synthase by L-NNA significantly delayed ulcer healing and reversed the CCK-8 induced acceleration of ulcer healing, hyperemia at the ulcer margin and luminal NO release, and these effects were restored by the addition to L-NNA of L-arginine but not D-arginine. Capsaicin denervation attenuated CCK-induced ulcer healing, and the accompanying rise in the GBF at the ulcer margin and decreased plasma gastrin and luminal release of somatostatin when compared to those in rats with intact sensory nerves. Detectable signals for CCK-A and B receptor mRNAs as well as for cNOS mRNA expression were recorded by RT-PCR in the vehicle control gastric mucosa. The expression of CCK-A receptor mRNA and cNOS mRNA was significantly increased in rats treated with CCK-8 and camostate, whereas CCK-B receptor mRNA remained unaffected. We conclude that CCK accelerates ulcer healing by the mechanism involving upregulation of specific CCK-A receptors, enhancement of somatostatin release, stimulation of sensory nerves and hyperemia in the ulcer area, possibly mediated by NO.     

Priming effects of mammalian tachykinins on human neutrophils.

The undecapeptide substance P (SP) is known to activate different cell types involved in inflammatory and immune processes. By evaluating primed stimulation of human neutrophils, we now demonstrate that SP (10 nM-0.1 mM) dose-dependently enhances superoxide anion production from cells stimulated by the phospholipid mediator Platelet Activating Factor (PAF). We also provide evidence that neurokinin A (NKA), which is released, as well as SP, from C fibers of sensory nerves, potentiates PAF-evoked superoxide anion generation, while neurokinin B (NKB) is ineffective.     

Local disruption of the celiac ganglion inhibits substance P release and ameliorates caerulein-induced pancreatitis in rats

Primary sensory neurons of the C and Adelta subtypes express the vanilloid capsaicin receptor TRPV1 and contain proinflammatory peptides such as substance P (SP) that mediate neurogenic inflammation. Pancreatic injury stimulates these neurons causing the release of SP in the pancreas resulting in pancreatic edema and neutrophil infiltration that contributes to pancreatitis. Axons of primary sensory neurons innervating the pancreas course through the celiac ganglion. We hypothesized that disruption of the celiac ganglion by surgical excision or inhibition of C and Adelta fibers through blockade of TRPV1 would reduce the severity of experimental pancreatitis by inhibiting neurogenic inflammation. Resiniferatoxin (RTX) is a specific TRPV1 agonist that, in high doses, selectively destroys C and Adelta fibers. Sprague-Dawley rats underwent surgical ganglionectomy or application of 10 microg RTX (vs. vehicle alone) to the celiac ganglion. One week later, pancreatitis was induced by six hourly intraperitoneal injections of caerulein (50 microg/kg). The severity of pancreatitis was assessed by serum amylase, pancreatic edema, and pancreatic myeloperoxidase (MPO) activity. SP receptor (neurokinin-1 receptor, NK-1R) internalization in acinar cells, used as an index of endogenous SP release, was assessed by immunocytochemical quantification of NK-1R endocytosis. Caerulein administration caused significant increases in pancreatic edema, serum amylase, MPO activity, and NK-1R internalization. RTX treatment and ganglionectomy significantly reduced pancreatic edema by 46% (P < 0.001) and NK-1R internalization by 80% and 51% (P < 0.001 and P < 0.05, respectively). RTX administration also significantly reduced MPO activity by 47% (P < 0.05). Neither treatment affected serum amylase, consistent with a direct effect of caerulein. These results demonstrate that disruption of or local application of RTX to the celiac ganglion inhibits SP release in the pancreas and reduces the severity of acute secretagogue-induced pancreatitis. It is possible that selectively disrupting TRPV1-bearing neurons could be used to reduce pancreatitis severity.     

瞬时受体电位香草酸亚型1激活释放的P物质在小鼠急性心肌梗死后凋亡中的保护作用

瞬时受体电位香草酸亚型1 (transient receptor potential vanilloid 1, TRPV1)在心肌缺血激活后可传导心绞痛信号和释放P物质(substance P, SP).SP是速激肽家族成员之一,主要通过结合并激活神经激肽1 (neurokinin 1,NK1)受体发挥作用. TRPV1和SP在缺血性心脏病中对心功能的恢复和重塑有一定保护作用,但对心肌梗死后凋亡的作用及具体机制尚不明确.本研究用TRPV1基因敲除(TRPV1-/- )小鼠和野生型(wide type, WT)小鼠建立心肌梗死模型,并外源性给予SP和NK1受体拮抗剂RP67580,用TTC染色法观察梗死的面积,TUNEL法检测心肌细胞凋亡指数,Western印迹方法检测caspase-3、Bcl-2、Bax、p53的蛋白表达.结果发现,心肌梗死24 h后,TRPV1-/-小鼠比WT小鼠梗死面积更大,凋亡指数和caspase-3活性更高,Bcl-2/Bax和p53蛋白表达更低. SP预处理可以明显缩小TRPV1-/-小鼠梗死面积,降低凋亡指数、caspase-3活性和升高Bcl-2/Bax比值,而在WT小鼠中改善不明显.外源性给予RP67580,阻断SP与NK1受体结合后,与相应对照组相比,WT小鼠梗死面积和凋亡指数更大,caspase-3蛋白表达更高,Bcl-2/Bax比值更低;TRPV1-/-小鼠与相应对照组比较,凋亡指数和caspase-3表达升高,Bcl-2/Bax比值降低.研究结果表明,SP可能介导了TRPV1在急性心肌梗死后凋亡中的保护作用.     

Role of transient receptor potential vanilloid 1 receptors in endotoxin-induced airway inflammation in the mouse

Airways are densely innervated by capsaicin-sensitive sensory neurons expressing transient receptor potential vanilloid 1 (TRPV1) receptors/ion channels, which play an important regulatory role in inflammatory processes via the release of sensory neuropeptides. The aim of the present study was to investigate the role of TRPV1 receptors in endotoxin-induced airway inflammation and consequent bronchial hyperreactivity with functional, morphological, and biochemical techniques using receptor gene-deficient mice. Inflammation was evoked by intranasal administration of Escherichia coli lipopolysaccharide (60 microl, 167 microg/ml) in TRPV1 knockout (TRPV1(-/-)) mice and their wild-type counterparts (TRPV1(+/+)) 24 h before measurement. Airway reactivity was assessed by unrestrained whole body plethysmography, and its quantitative indicator, enhanced pause (Penh), was calculated after inhalation of the bronchoconstrictor carbachol. Histological examination and spectrophotometric myeloperoxidase measurement was performed from the lung. Somatostatin concentration was measured in the lung and plasma with radioimmunoassay. Bronchial hyperreactivity, histological lesions (perivascular/peribronchial edema, neutrophil/macrophage infiltration, goblet cell hyperplasia), and myeloperoxidase activity were significantly greater in TRPV(-/-) mice. Inflammation markedly elevated lung and plasma somatostatin concentrations in TRPV1(+/+) but not TRPV1(-/-) animals. In TRPV1(-/-) mice, exogenous administration of somatostatin-14 (4 x 100 microg/kg ip) diminished inflammation and hyperreactivity. Furthermore, in wild-type mice, antagonizing somatostatin receptors by cyclo-somatostatin (4 x 250 microg/kg ip) increased these parameters. This study provides the first evidence for a novel counterregulatory mechanism during endotoxin-induced airway inflammation, which is mediated by somatostatin released from sensory nerve terminals in response to activation of TRPV1 receptors of the lung. It reaches the systemic circulation and inhibits inflammation and consequent bronchial hyperreactivity.     

Tachykinins stimulate a subset of mouse taste cells

The tachykinins substance P (SP) and neurokinin A (NKA) are present in nociceptive sensory fibers expressing transient receptor potential cation channel, subfamily V, member 1 (TRPV1). These fibers are found extensively in and around the taste buds of several species. Tachykinins are released from nociceptive fibers by irritants such as capsaicin, the active compound found in chili peppers commonly associated with the sensation of spiciness. Using real-time Ca(2+)-imaging on isolated taste cells, it was observed that SP induces Ca(2+) -responses in a subset of taste cells at concentrations in the low nanomolar range. These responses were reversibly inhibited by blocking the SP receptor NK-1R. NKA also induced Ca(2+)-responses in a subset of taste cells, but only at concentrations in the high nanomolar range. These responses were only partially inhibited by blocking the NKA receptor NK-2R, and were also inhibited by blocking NK-1R indicating that NKA is only active in taste cells at concentrations that activate both receptors. In addition, it was determined that tachykinin signaling in taste cells requires Ca(2+)-release from endoplasmic reticulum stores. RT-PCR analysis further confirmed that mouse taste buds express NK-1R and NK-2R. Using Ca(2+)-imaging and single cell RT-PCR, it was determined that the majority of tachykinin-responsive taste cells were Type I (Glial-like) and umami-responsive Type II (Receptor) cells. Importantly, stimulating NK-1R had an additive effect on Ca(2+) responses evoked by umami stimuli in Type II (Receptor) cells. This data indicates that tachykinin release from nociceptive sensory fibers in and around taste buds may enhance umami and other taste modalities, providing a possible mechanism for the increased palatability of spicy foods.     

The role of reactive oxygen species and capsaicin-sensitive sensory nerves in the pathomechanisms of gastric ulcers induced by stress.

Gastric microcirculation plays an important role in the maintenance of the gastric mucosal barrier and mucosal integrity. Sensory nerves are involved in the regulation of mucosal blood circulation and mucosal defense. Therefore, the ablation of these nerves by neurotoxic doses of capsaicin provides the possibility of determination of their role in gastric mucosal integrity. Stress ulceration represents a serious gastric lesions. Results of our previous experiments have indicated that water immersion and restraint stress (WRS) led to increased oxidative metabolism. Ablation of sensory nerves by high doses of capsaicin retards healing of gastric ulcers, but the role of reactive oxygen species (ROS) in the healing process has been little studied. Therefore, the aim of our present investigations was to determine the participation of ROS in sensory nerve activity during WRS. Experiments were carried out on 90 male Wistar rats and the area of gastric lesions was measured by planimetry. Colorimetric assays were used to determine gastric mucosal levels of malondialdehyde (MDA) and 4-hydroxynonenal (4-HNE), as well as superoxide dismutase (SOD) activity. We demonstrated that inactivation of sensory nerves resulted in magnification of gastric mucosal damage induced by the WRS. In this process, oxidative stress, as reflected by an increase of MDA and 4-HNE tissue concentrations (an index of lipid peroxidation), as well as decrease of SOD activity, could play an important role. Aspirin, applied in a low dose, exerts a protective activity, possibly due to its metabolites, which possess the anti-oxidant and ROS scavanging properties. Pentoxyfilline-induced gastroprotection and hyperemia depends upon attenuation of the oxidative stress. This protection and hyperemia were, at least in part, attenuated by ASA.     

Mechanistic studies of acid-evoked coughing in anesthetized guinea pigs

Experiments carried out in conscious guinea pigs suggest that citric acid-evoked coughing is partly mediated by transient receptor potential vanilloid type 1 (TRPV1) receptor-dependent activation of tachykinin-containing, capsaicin-sensitive C fibers. In vitro electrophysiological analyses indicate, however, that acid also activates capsaicin-sensitive and -insensitive vagal afferent nerves by a TRPV1-independent mechanism, and studies in anesthetized guinea pigs show that coughing evoked by acid is mediated by activation of capsaicin-insensitive vagal afferent nerves. In the present study, we have characterized the mechanisms of citric acid-evoked coughing in anesthetized guinea pigs. Drugs were administered directly to the Krebs buffer perfusing the extrathoracic trachea. Citric acid was applied topically to the tracheal mucosa, directly into the tracheal perfusate in increasing concentrations and at 1-min intervals. Citric acid dose dependently evoked coughing in anesthetized guinea pigs. This was mimicked by hydrochloric acid but not by sodium citrate. The coughing evoked by acid was nearly or completely abolished by TTX or by cutting the recurrent laryngeal nerves. Perfusing the trachea with a low Cl- buffer potentiated the acid-induced cough reflex. In contrast, prior capsaicin desensitization, 10 microM capsazepine, Ca2+-free perfusate, 0.1 microM iberiotoxin, 1 microM atropine, 10 microM isoproterenol, 10 microM albuterol, 3 microM indomethacin, 0.1 microM HOE-140, a combination of neurokinin1 (NK1; CP-99994), NK2 (SR-48968), and NK3 (SB-223412) receptor antagonists (0.1 microM each), a combination of histamine H1 (3 microM pyrilamine) and cysLT1 (1 microM ICI-198615) receptor antagonists, superior laryngeal nerve transection, or epithelium removal did not inhibit citric acid-evoked coughing. These and other data indicate that citric acid-evoked coughing in anesthetized guinea pigs is mediated by direct activation of capsaicin-insensitive vagal afferent nerves, perhaps through sequential activation of acid-sensing ion channels and chloride channels.     

Distribution of the vanilloid (capsaicin) receptor type 1 in the human stomach

Vanilloid receptor type 1 (TRPV1) is expressed in a capsaicin-sensitive and peptide-containing sub-population of primary sensory nerves that in the rat stomach seems involved in regulation of chlorhydropeptic secretion and gastroprotection. Our aim was to identify which cell types express TRPV1 in the human stomach in order to gain a better insight in the role of this receptor in the regulation of HCl secretion. Immunohistochemistry, by using three different commercially available anti-capsaicin antibodies, in situ hybridisation and Western blot analysis were performed on fragments surgically obtained from the gastric body on the large curvature. TRPV1 labelling was found in the parietal cells at the level of intra-cytoplasmatic granules matching mitochondrial features and distribution. Immunolabelled neurons and nerve fibres were also seen, the latter numerous in the submucosa and mucosa and often ending close to the parietal cells. TRPV1 presence was confirmed by Western blot analysis and in situ hybridisation. TRPV1 presence in nerve structures and parietal cells suggests the possibility of a combined effect of both neuronal and epithelial TRPV1 on chlorhydropeptic secretion. The presumed TRPV1 mitochondrial location inside parietal cells is in favour of the existence of a local pathway of auto-regulation of HCl secretion. Therefore, TRPV1 might modulate chlorhydropeptic secretion in the human stomach through more complex pathways than previously thought.     

Coexisting peptides in capsaicin-sensitive sensory neurons: release and actions in the respiratory tract of the guinea-pig

Release of substance P (SP) and neurokinin A (NKA), was demonstrated in the isolated perfused guinea-pig lung. Significant release was obtained by perfusion with capsaicin, high potassium, histamine, bradykinin dimethylphenylpiperazinium, and by electrical vagal nerve stimulation. Capsaicin-induced peptide release was not blocked by 1 microM clonidine. SP and NKA contracted respiratory smooth muscle, NKA being 42 times more potent. Both tachykinins were equipotent in relaxing pulmonary artery. It is concluded that multiple tachykinin can be released from capsaicin-sensitive sensory nerves in the respiratory tract, exerting multiple effects on the target tissues.