Direct neurite-mast cell communication in vitro occurs via the neuropeptide substance P.
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Communication between nerves and mast cells is a prototypic demonstration of neuroimmune interaction. However, whether mast cell activation occurs as a direct response to neuronal activation or requires an intermediary cell is unclear. Addressing this issue, we used an in vitro coculture approach comprising cultured murine superior cervical ganglia and rat leukemia basophilic cells (RBLs; possesses properties of mucosal-type mast cells). Following loading with the calcium fluorophore, Fluo-3, neurite-RBL units (separated by <50 nm) were examined by confocal laser scanning microscopy. Addition of bradykinin, or scorpion venom, dose-dependently elicited neurite activation (i.e., Ca2+ mobilization) and, after a lag period, RBL Ca2+ mobilization. Neither bradykinin nor scorpion venom had any direct effect on the RBLs in the absence of neurites. Addition of a neutralizing substance P Ab or a neurokinin (NK)-1 receptor antagonist, but not an NK-2 receptor antagonist, dose-dependently prevented the RBL activation that resulted as a consequence of neural activation by either bradykinin or scorpion venom. These data illustrate that nerve-mast cell cross-talk can occur in the absence of an intermediary transducing cell and that the neuropeptide substance P, operating via NK-1 receptors, is an important mediator of this communication. Our findings have implications for the neuroimmune signaling cascades that are likely to occur during airways inflammation, intestinal hypersensitivity, and other conditions in which mast cells feature. (+info)
Repeated challenge with dinitrobenzene sulphonic acid in dinitrofluorobenzene-sensitized mice results in vascular hyperpermeability in the trachea: a role for tachykinins.
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1. This study investigates the role of tachykinins in a repeated challenge with dinitrobenzene sulphonic acid (DNS) on the tracheal vascular permeability in dinitrofluorobenzene (DNFB)-sensitized mice. 2. DNFB-contact sensitization was followed by an intranasal (i.n.) challenge with DNS. A second challenge with DNS was administered 24 h after the first challenge. To assess changes in tracheal vascular permeability, Evans blue dye accumulation in tracheal tissue was measured. 3. A repeated challenge with DNS in DNFB-sensitized mice led to a 2.8 fold increase in tracheal vascular permeability when compared to DNFB-sensitized and vehicle-challenged mice or a 2.5 fold increase when compared to DNFB-sensitized single DNS-challenged mice (P<0.001, ANOVA). 4. RP67580 (10-9 mol mouse-1 i.v.) reduced the increased tracheal vascular permeability induced by a second exposure to DNS in DNFB-sensitized mice completely when injected 15 min before the second challenge (P<0.001, ANOVA). 5. The increased tracheal vascular permeability response induced by the second exposure to DNS could be mimicked with i.n. application of capsaicin (10-10 mol mouse-1) or substance P (SP) (10-12 mol mouse-1) to DNFB-sensitized and single DNS-challenged mice. 6. These results suggest that both tachykinin NK1 receptors and sensory nerves are involved in the development of vascular hyperpermeability changes found in the trachea of DNFB-sensitized mice after a repeated DNS-challenge. (+info)
Nociceptin-induced scratching, biting and licking in mice: involvement of spinal NK1 receptors.
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1. Intrathecal (i.t.) injection of nociceptin at small doses (fmol order) elicited a behavioural response consisting of scratching, biting and licking in conscious mice. Here we have examined the involvement of substance P-containing neurons by using i.t. injection of tachykinin neurokinin (NK)1 receptor antagonists and substance P (SP) antiserum. 2. Nociceptin-induced behavioural response was evoked significantly 5 - 10 min after i.t. injection and reached a maximum at 10 - 15 min. Dose-dependency of the induced response showed a bell-shaped pattern from 0.375 - 30.0 fmol, and the maximum effect was observed at 3.0 fmol. 3. The behavioural response elicited by nociceptin (3.0 fmol) was dose-dependently inhibited by intraperitoneal (i.p.) administration of morphine. 4. The NK1 receptor antagonists, CP-96,345, CP-99,994 and sendide, inhibited nociceptin-induced behavioural response in a dose-dependent manner. A significant antagonistic effect of [D-Phe7, D-His9]SP (6 - 11), a selective antagonist for SP receptors, was observed against nociceptin-induced response. The NK2 receptor antagonist, MEN-10376, had no effect on the response elicited by nociceptin. 5. Pretreatment with SP antiserum resulted in a significant reduction of the response to nociceptin. No significant reduction of nociceptin-induced response was detected in mice pretreated with NKA antiserum. 6. The N-methyl-D-aspartate (NMDA) receptor antagonists, dizocilpine (MK-801) and D(-)-2-amino-5-phosphonovaleric acid (APV) (D-APV), and L-NG-nitro arginine methyl ester (L-NAME), a nitric oxide (NO) synthase inhibitor, failed to inhibit nociceptin-induced behavioural response. 7. off present results suggest that SP-containing neurons in the mouse spinal cord may be involved in elicitation of scratching, biting and licking behaviour following i.t. injection of nociceptin. (+info)
Targeted ablation of secretin-producing cells in transgenic mice reveals a common differentiation pathway with multiple enteroendocrine cell lineages in the small intestine.
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The four cell types of gut epithelium, enteroendocrine cells, enterocytes, Paneth cells and goblet cells, arise from a common totipotent stem cell located in the mid portion of the intestinal gland. The secretin-producing (S) cell is one of at least ten cell types belonging to the diffuse neuroendocrine system of the gut. We have examined the developmental relationship between secretin cells and other enteroendocrine cell types by conditional ablation of secretin cells in transgenic mice expressing herpes simplex virus 1 thymidine kinase (HSVTK). Ganciclovir-treated mice showed markedly increased numbers of apoptotic cells at the crypt-villus junction. Unexpectedly, ganciclovir treatment induced nearly complete ablation of enteroendocrine cells expressing cholecystokinin and peptide YY/glucagon (L cells) as well as secretin cells, suggesting a close developmental relationship between these three cell types. In addition, ganciclovir reduced the number of enteroendocrine cells producing gastric inhibitory polypeptide, substance-P, somatostatin and serotonin. During recovery from ganciclovir treatment, the enteroendocrine cells repopulated the intestine in normal numbers, suggesting that a common early endocrine progenitor was spared. Expression of BETA2, a basic helix-loop-helix protein essential for differentiation of secretin and cholecystokinin cells was examined in the proximal small intestine. BETA2 expression was seen in all enteroendocrine cells and not seen in nonendocrine cells. These results suggest that most small intestinal endocrine cells are developmentally related and that a close developmental relationship exists between secretin-producing S cells and cholecystokinin-producing and L type enteroendocrine cells. In addition, our work shows the existence of a multipotent endocrine-committed cell type and locates this hybrid multipotent cell type to a region of the intestine populated by relatively immature cells. (+info)
Substance P and bradykinin activate different types of KCa currents to hyperpolarize cultured porcine coronary artery endothelial cells.
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1. Substance P and bradykinin, endothelium-dependent vasodilators of pig coronary artery, trigger in endothelial cells a rise in cytosolic Ca2+ concentration ([Ca2+]i) and membrane hyperpolarization. The aim of the present study was to determine the type of Ca2+-dependent K+ (KCa) currents underlying the endothelial cell hyperpolarization. 2. The substance P-induced increase in [Ca2+]i was 30 % smaller than that induced by bradykinin, although the two peptides triggered a membrane hyperpolarization of the same amplitude. The two agonists evoked a large outward K+ current of the same conductance at maximal stimulation. Agonists applied together produced the same maximal current amplitude as either one applied alone. 3. Iberiotoxin (50 nM) reduced by about 40 % the K+ current activated by bradykinin without modifying the substance P response. Conversely, apamin (1 microM) inhibited the substance P-induced K+ current by about 65 %, without affecting the bradykinin response. Similar results were obtained on peptide-induced membrane hyperpolarization. 4. Bradykinin-induced, but not substance P-induced, endothelium-dependent relaxation resistant to NG-nitro-L-arginine and indomethacin was partly inhibited by 3 microM 17-octadecynoic acid (17-ODYA), an inhibitor of cytochrome P450 epoxygenase. Similarly, the bradykinin-induced K+ current was reduced by 17-ODYA. 5. Our results show that responses to substance P and bradykinin result in a hyperpolarization due to activation of different KCa currents. A current consistent with the activation of large conductance (BKCa) channels was activated only by bradykinin, whereas a current consistent with the activation of small conductance (SKCa) channels was stimulated only by substance P. The observation that a similar electrical response is produced by different pools of channels implies distinct intracellular pathways leading to KCa current activation. (+info)
Substance P relaxes rat bronchial smooth muscle via epithelial prostanoid synthesis.
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BACKGROUND: Substance P is present in bronchial nerve fibres. The physiological actions of substance P are mediated via tachykinin NK(1) receptors. Immunochemical studies have demonstrated tachykinin NK(1) receptors in the rat airway epithelium. OBJECTIVE: To elucidate how epithelial tachykinin NK(1) receptors affect smooth muscle response to substance P. METHODS: Contractile response of isolated rat bronchial trunk with or without epithelium was recorded. RESULTS: In intact segments precontracted by 5-hydroxytryptamine, relaxation was induced by substance P and the nitric oxide donor, sodium nitroprusside. Removal of the epithelium abolished relaxation induced by substance P but did not affect relaxation induced by sodium nitroprusside. The cyclo-oxygenase inhibitor, indomethacin, but not the nitric oxide synthase inhibitor, L-N(G)-monomethylarginine, reduced the relaxation in response to substance P. CONCLUSIONS: Epithelial tachykinin NK(1) receptors mediate substance-P-induced relaxation of rat bronchial smooth muscle via release of prostanoids but not nitric oxide. (+info)
Interaction between substance P and gastrin-releasing peptide on thyrotropin secretion by rat pituitary in vitro.
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The effect of substance P (SP) on thyrotropin (TSH) secretion is controversial. In this study we evaluated the effect of SP on TSH secretion by hemipituitaries of 3-month-old Wistar rats in vitro and its interaction with gastrin-releasing peptide (GRP) at equimolar concentrations (1 microM and 10 microM). TSH release was measured under basal conditions and 30 min after incubation in the absence or presence of SP, GRP or both peptides. Pituitary TSH content was also measured in the pituitary homogenate after incubation. SP at both concentrations caused a significant (P<0.05) increase in TSH secretion compared with all other groups, which was approximately 60% (1 microM) and 85% (10 microM) higher than that of the control group (23.3 +/- 3.0 ng/ml). GRP at the lower concentration did not produce a statistically significant change in TSH secretion, whereas at the concentration of 10 microM it produced a 50% reduction in TSH. GRP co-incubated with substance P completely blocked the stimulatory effect of SP at both concentrations. Pituitary TSH content decreased in the SP-treated group compared to controls (0.75 +/- 0.03 microg/hemipituitary) at the same proportion as the increase in TSH secretion, and this effect was also blocked when GRP and SP were co-incubated. In conclusion, in an in vitro system, SP increased TSH secretion acting directly at the pituitary level and this effect was blocked by GRP, suggesting that GRP is more potent than SP on TSH secretion, and that this inhibitory effect could be the predominant effect in vivo. (+info)
Electron-immunocytochemical studies of perivascular nerves of mesenteric and renal arteries of golden hamsters during and after arousal from hibernation.
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Electron immunocytochemistry was used to examine perivascular nerves of hamster mesenteric and renal arteries during hibernation and 2 h after arousal from hibernation. Vessels from cold-exposed but nonhibernating, and normothermic control hamsters were also examined. During hibernation the percentage of axon profiles in mesenteric and renal arteries that were immunopositive for markers of sympathetic nerves, tyrosine hydroxylase (TH) and neuropeptide Y (NPY), were increased 2-3 fold compared with normothermic and cold control animals. This increase was reduced markedly only 2 h after arousal from hibernation. The small percentage of nitric oxide synthase-1-positive axon profiles found in mesenteric (but not renal) arteries was also increased during hibernation and returned towards control values after arousal. In contrast, the percentage of perivascular axons immunostaining for vasoactive intestinal polypeptide (VIP), a marker for parasympathetic nerves, was reduced in mesenteric arteries during hibernation. There was no labelling of perivascular nerves for substance P in either mesenteric or renal arteries. It is suggested that the increase in percentage of TH- and NPY-immunostained perivascular nerves may account for the increased vasoconstriction associated with high vascular resistance that is known to occur during hibernation. The reduction in the percentage of axons positive for VIP in hibernating animals would contribute to this mechanism since this neuropeptide is a vasodilator. (+info)