Inflammatory oedema induced by Lachesis muta muta (Surucucu) venom and LmTX-I in the rat paw and dorsal skin

The ability of crude venom and a basic phospholipase A₂ (LmTX-I) from Lachesis muta muta venom to increase the microvascular permeability in rat paw and skin was investigated. Crude venom or LmTX-I were injected subplantarly or intradermally and rat paw oedema and dorsal skin plasma extravasation were measured. Histamine release from rat peritoneal mast cell was also assessed. Crude venom or LmTX-I induced dose-dependent rat paw oedema and dorsal skin plasma extravasation. Venom-induced plasma extravasation was inhibited by the histamine H₁ antagonist mepyramine (6 mg/kg), histamine/5-hydroxytriptamine antagonist cyproheptadine (2 mg/kg), cyclooxygenase inhibitor indomethacin (5 mg/kg), nitric oxide synthesis inhibitor l-NAME (100 nmol/site), tachykinin NK₁ antagonist SR140333 (1 nmol/site) and bradykinin B₂ receptor antagonist Icatibant (0.6 mg/kg). Platelet-activating factor (PAF) antagonist PCA4248 (5 mg/kg) had no effect. LmTX-I-induced skin extravasation was inhibited by cyproheptadine, mepyramine, indomethacin and PCA4248, while l-NAME and SR140333 had no effect. Additionally, both Lachesis muta muta venom and LmTX-I concentration-dependently induced histamine release from rat mast cells. In conclusion, Lachesis muta muta venom and LmTX-I increase microvascular permeability by mechanisms involving in vivo mast cell activation and arachidonic acid metabolites. Additionally, crude venom-induced responses also involve substance P, nitric oxide and bradykinin release, whether LmTX-I-induced responses involve PAF.     

The effect of a tachykinin NK1 receptor antagonist, SR140333, on oedema formation induced in rat skin by venom from the Phoneutria nigriventer spider.

1. The possibility that tachykinin NK1 receptors are involved in the plasma extravasation evoked by intradermal (i.d.) injection of Phoneutria nigriventer venom (PNV) in rat dorsal skin in vivo has been investigated. 2. Local oedema formation induced by the i.d. injection of test agents was measured by the extravascular accumulation of intravenously (i.v.) injected 125I-labelled human serum albumin over a 30 min period. 3. The tachykinin NK1 agonist, GR73632 (30 pmol per site), induced local oedema formation which was potentiated by co-injection with the neuropeptide vasodilator, calcitonin gene-related peptide (CGRP, 10 pmol per site). The non-peptide tachykinin NK1 receptor antagonist, SR140333 (0.03-1 nmol per site co-injected, i.d.) significantly inhibited (0.3 nmol per site, P < 0.05; 1 nmol per site, P < 0.001) local oedema formation induced by GR73632 with CGRP but not that induced by histamine (10 nmol per site) with CGRP. 4. PNV (0.03-0.3 microgram per site) injected i.d. induced dose-dependent local oedema formation. SR140333 (1 nmol per site, co-injected i.d.) inhibited oedema formation; with complete inhibition observed at doses of 0.03 microgram (P < 0.05) and 0.1 microgram (P < 0.001); and partial inhibition (50%) observed with the highest dose of PNV, 0.3 microgram (P < 0.05). 5. Local oedema formation induced by PNV was not affected by systemic pretreatment with the bradykinin B2 receptor antagonist, Hoe 140 (80 nmol kg-1, i.v.), which was used at a dose which significantly inhibited oedema formation by bradykinin (1 nmol per site). 6. Local oedema formation induced by PNV was significantly inhibited (P < 0.01) by co-injection of the histamine H1 receptor antagonist, mepyramine (2.5 nmol per site), together with the 5-hydroxytryptamine (5-HT) antagonist, methysergide (2.8 nmol per site). 7. In the presence of all three antagonists (mepyramine 2.5 nmol per site; methysergide, 2.8 nmol per site and SR140333 1 nmol per site), the plasma extravasation induced by PNV was further significantly inhibited (P < 0.001, when compared with PNV injected i.d. alone; P < 0.05 when compared with PNV co-injected with mepyramine and methysergide and P < 0.01, when compared with PNV co-injected with SR140333). 8. These results suggest that oedema formation evoked by i.d. PNV in rat skin may be partially mediated via a mechanism involving tachykinin NK1 receptors and that this effect is independent of histamine and 5-HT.     

The non-peptide NK1 receptor antagonist SR140333 produces long-lasting inhibition of neurogenic inflammation,but does not influence acute chemo- or thermonociception in rats

In anaesthetized rats, the neurokinin (NK)₁ receptor antagonist SR140333 (10–1000 g/kg) stereoselectively inhibited mustard oil-induced plasma protein extravasation in the dorsal skin of the hind paw. After s.c. administration of SR140333, inhibition of plasma protein extravasation was maximal 3 h after injection. A dose of 0.1 mg/kg i.v. or 1.0 mg/kg s.c. produced long-lasting inhibition which was still significant 24 h after treatment.Since systemic administration of SR140333 has been shown to inhibit nociceptive responses in anaesthetized rats, we wanted to evaluate a possible effect of SR140333 on chemo- and thermonociception in conscious rats. SR140333 (100 g/kg s.c) did not reduce the behavioral response of rats to the irritant effect of capsaicin in the wiping test, nor did it affect the thermal nociceptive threshold in the plantar test. Furthermore, the decrease in thermal nociceptive threshold which was produced by intraplanter injection of PGE₂, and which has been shown to be entirely dependent on capsaicin-sensitive afferents, was not affected by treatment with this NK₁ receptor antagonist.These results show that systemic administration of SR140333, at doses which cause inhibition of neurogenic inflammation, has no detectable effect on acute chemo- or thermonociception in conscious rats.     

Distinct tachykinin NK(1) receptor function in primate nucleus tractus solitarius neurons is dysregulated after second-hand tobacco smoke exposure

BACKGROUND AND PURPOSE

Second-hand tobacco smoke (SHS) exposure in children increases the risk of asthma and sudden infant death syndrome. Epidemiological and experimental data have suggested SHS can alter neuroplasticity in the CNS, associated with substance P. We hypothesized that exposure to SHS in young primates changed the effect of substance P on the plasticity of neurons in the nucleus tractus solitarius (NTS), where airway sensory information is first processed in the CNS.

EXPERIMENTAL APPROACH

Thirteen-month-old rhesus monkeys were exposed to filtered air (FA, n = 5) or SHS (n = 5) for >6 months from 50 days of their fetal age. Whole-cell patch-clamp recordings were performed on NTS neurons in brainstem slices from these animals to record the intrinsic cell excitability in the absence or presence of the NK₁ receptor antagonist, SR140333 (3 µM).

KEY RESULTS

Neurons were electrophysiologically classified based on their spiking onset from a hyperpolarized membrane potential into two phenotypes: rapid-onset spiking (RS) and delayed-onset spiking (DS) types. In RS neurons, SR140333 reduced the spiking response, similarly in both FA- and SHS-exposed animals. In DS neurons, SR140333 almost abolished the spiking response in FA-exposed animals, but had no effect in SHS-exposed animals.

CONCLUSIONS AND IMPLICATIONS

The contribution of NK₁ receptors to cell excitability depended on firing phenotype of primate NTS neurons and was disrupted by SHS exposure, specifically in DS neurons. Our findings reveal a novel NK₁ receptor function in the primate brainstem and support the hypothesis that chronic exposure to SHS in children causes tachykinin-related neuroplastic changes in the CNS.     

Are biological actions of neurokinin A in the adult brain mediated by a cross-talk between the NK1 and NK2 receptors?

Mice lacking the NK₁ receptor (NK₁R?/? mice) and selective, high-affinity, non-peptide, NK₁, NK₂ and NK₃ receptor antagonists were used to identify the tachykinin receptor subtype(s) mediating the central responses induced by neurokinin A (NKA). The peptides, substance P (SP), NKA and senktide and the antagonists were injected intracerebroventricularly (ICV) through an implanted cannula. NKA (50 pmol) was as potent as SP (50 pmol) in inducing grooming behaviour (face washing and hind limb grooming) in wild-type mice, but both peptides failed to induce behavioural responses in NK₁R?/? mice. In wild-type mice, the NK₁ receptor antagonist, RP 67580 (2 nmol), effectively inhibited grooming behaviour elicited by SP, but was inactive against grooming induced by NKA, which in turn was abolished after pre-treatment with the selective NK₂ receptor agonist, SR 48968 (2 nmol). Unlike NKA, the selective NK₂ receptor agonists, (β Ala⁸) NKA 4–10 and (NLeu¹⁰) NKA 4–10, injected ICV at doses of 50 or 100 pmol did not elicit any behavioural response in wild-type mice. The NK₃ receptor antagonist, SR 142801, inhibited behaviours induced by the NK₃ receptor agonist, senktide, but did not alter behavioural responses to either SP or NKA in wild-type mice. The present findings demonstrate that central biological actions of SP and senktide are mediated by activation of NK₁ and NK₃ receptors, respectively. Our results also indicate that NK₁ receptors are essential for generating central actions induced by NKA, which are most probably mediated by a cross-talk between the NK₁ and NK₂ receptors.     

Enhanced nerve‐stimulated muscarinic and neurokinin contractions of ileum from streptozotocin guinea‐pigs

相似文献   

Prejunctional Control of pH 6-Induced Bronchoconstriction by NK1, NK2, μ-Opioid, α2-Adrenoceptor and Glucocorticoid Receptors in Guinea-pig Isolated Perfused Lung

This study investigated the release of calcitonin-gene related peptide-like (CGRP) immunoreactivity and bronchoconstriction induced by pH 6 buffer in guinea-pig isolated perfused lung. Both pH 6-induced CGRP-like immunoreactivity and bronchoconstriction were completely abolished after systemic pretreatment with capsaicin. Pretreatment with the NK₂ receptor antagonist SR 48968 (5 times 10^?7M) completely inhibited bronchoconstriction and significantly reduced the immunoreactivity induced by the pH 6 buffer. The NK₁ antagonist SR 140333 (5 times 10^?7M) and, to a lesser extent the NK₁ antagonist CP 96345, morphine (5 times 10^?6M), the α₂-adrenoceptor agonist UK 14304 (10^?7M) and betamethasone (10^?6M) significantly reduced both pH 6-induced bronchial response and CGRP-like immunoreactivity overflow. The effects of morphine and UK14304 were partially reversed by naloxone (5 times 10^?5M) and idazoxan (5 times 10^?5M). Therefore, NK₁, NK₂, μ-opioid, α₂-adrenoceptor and glucocorticoid receptors seemed to have a prejunctional action on pH 6 buffer-induced CGRP-like immunoreactivity and bronchoconstriction.     

Inflammatory oedema induced by phospholipases A2 isolated from Crotalus durissus sp. in the rat dorsal skin: a role for mast cells and sensory C-fibers.

The ability of the phospholipases A(2) (PLA(2)s) from Crotalus durissus cascavella, Crotalus durissus collilineatus and Crotalus durissus terrificus venoms and crotapotin to increase the vascular permeability in the rat skin as well as the contribution of both mast cells and sensory C-fibers have been investigated in this study. Vascular permeability was measured as the plasma extravascular accumulation at skin sites of intravenously injected 125I-human serum albumin. Intradermal injection of crotalic PLA(2)s (0.05-0.5 microg/site) in the rat skin resulted in dose-dependent increase in plasma extravascular whereas crotapotin (1 microg/site) failed to affect this response. Co-injection of crotapotin (1 microg/site) did not modify the increased vascular permeability induced by the PLA(2)s (0.05-0.5 microg/site). Previous treatment (30 min) of the animals with cyproheptadine (2 mg/kg, i.p.) markedly reduced PLA(2) (0.5 microg/site)-induced oedema. In rats treated neonatally with capsaicin to deplete neuropeptides, the plasma extravasation induced by all PLA(2)s (0.5 microg/site) was also significantly reduced. Similarly, the tachykinin NK(1) receptor antagonist SR140333 (1nmol/site) significantly reduced the PLA(2)-induced oedema. In addition, the combination of SR140333 with cyproheptadine further reduced the increased plasma extravasation by PLA(2) from C. d. cascavella venom, but not by PLA(2) from C. d. terrificus and C. d. collilineatus venoms. Our results suggest that increase in skin vascular permeability by crotalic PLA(2)s is mediated by activation of sensory C-fibers culminating in the release of substance P, as well as by activation of mast cells which in turn release amines such as histamine and serotonin.     

Pharmacological characterization of Synoeca cyanea venom: An aggressive social wasp widely distributed in the Neotropical region

The venom of social wasps has been poorly studied so far, despite the high number of accidents in humans and assessment of the use of these wasps as a biological control of pests. The study of the pharmacological effects of the venom is of great importance since the poisoning is dangerous causing serious systemic effects, including death in the case of multiple attacks. In this study, the pharmacological activities of venom from the social wasp Synoeca cyanea were evaluated by the following assays: LD50 in mice, the behavioural effects and the hemorrhagic activity induced by the venom in mice, the oedematogenic activity in rat, the haemolysis in human blood, the stimulating effect on guinea-pig smooth muscle, and the antimicrobial activity. The aim was to determine the toxic effects of venom and to perform a comparative study with earlier work conducted with venom from other wasp species. Results showed that S. cyanea venom produced a potent dose-dependent oedema, as well as antibacterial and haemolytic activities, suggesting the presence of histamine, serotonin, kinins and other molecules related to increased vascular permeability and cytolytic activity in this venom. Despite previous studies with wasp venoms, S. cyanea venom presented a slight hemorrhagic effect. Data obtained in the smooth muscle assay also suggest the presence of BK or analogues in S. cyanea whole venom. The knowledge of symptoms and effects produced by S. cyanea venom is critical for health organizations, in order to improve clinical treatment in accidents caused by wasp stings.     

A role for the sensory neuropeptide calcitonin gene-related peptide in endothelial cell proliferation in vivo

BACKGROUND AND PURPOSE

We have tested the hypothesis that calcitonin gene-related peptide (CGRP) is a mediator of capsaicin-induced angiogenesis in vivo.

EXPERIMENTAL APPROACH

In a series of experiments, the knee joints of rats were injected with CGRP, capsaicin or vehicle control. Groups of animals (n = 6) were treated with the CGRP receptor antagonist BIBN4096BS and/or the NK₁ receptor antagonist SR140333. Endothelium, proliferating endothelial cell nuclei and macrophages were identified 24 h later in the synovium by immunohistochemistry and quantified by image analysis. mRNA for the receptors for CGRP and adrenomedullin were sought in normal and inflamed rat and human synovia using RT-PCR.

KEY RESULTS

Intra-articular CGRP injection increased the endothelial cell proliferation index, whereas macrophage infiltration and knee joint diameters were similar to saline-injected controls. CGRP-induced endothelial cell proliferation was dose-dependently inhibited by BIBN4096BS. mRNA for adrenomedullin and the CGRP receptor subunits were detected in normal and inflamed human and rat synovia. In capsaicin-induced synovitis, the increased endothelial cell proliferation index was partially blocked by administration of NK₁ or CGRP antagonists individually and was reduced to the level of saline controls by coadministration of both receptor antagonists.

CONCLUSIONS AND IMPLICATIONS

These data support the hypothesis that CGRP stimulates angiogenesis in vivo directly by activating CGRP receptors. Capsaicin-induced endothelial cell proliferation was completely blocked by coadministration of CGRP and NK₁ receptor antagonists, indicating that both CGRP and substance P may contribute to angiogenesis in this model of synovitis.     

Defaecation, intestinal fluid accumulation and motility in rodents: implications of cannabinoid CB1 receptors

We have studied the effect of SR141716A (0.1–5 mg/kg, i.p.), a cannabinoid CB₁ receptor antagonist, and WIN (0.1–5 mg/kg, i.p.), a cannabinoid receptor agonist, on acute defaecation and gastrointestinal transit in mice and on intraluminal fluid accumulation in the rat small intestine. SR141716A increased while WIN 55,212-2 decreased defaecation, gastrointestinal transit and fluid accumulation. A per se non-effective dose of SR141716A (0.3 mg/kg) counteracted the inhibitory effect of WIN 55,212-2 (1 mg/kg) on gastrointestinal functions studied. The effect of SR141716 on both intestinal fluid accumulation in rats and gastrointestinal transit in mice was inhibited by atropine (1 mg/kg, i.p.), but not by hexamethonium (1 mg/kg, s.c.), SR140333 (20 μg/kg, i.p.) or SR48968 (20 μg/kg, i.p.), antagonists of NK₁ and NK₂ receptors, respectively. These results suggest that intestinal fluid accumulation and motility are inhibited by endogenous cannabinoid(s) acting at the cannabinoid CB₁ receptors. This effect may be mediated by mechanisms involving muscarinic cholinoceptors. Received: 13 July 1998 / Accepted: 25 September 1998     

Capsaicin‐ and Mustard Oil‐Induced Extracellular Signal‐Regulated Protein Kinase Phosphorylation in Sensory Neurons in vivo: Effects of Neurokinins 1 and 2 Receptor Antagonists and of a Nitric Oxide Synthase Inhibitor

Abstract: Stimulation of primary sensory neurons with capsaicin or mustard oil leads to phosphorylation of extracellular signal‐regulated protein kinase 1/2 (p‐ERK1/2) via activation of transient receptor potential V1 (TRPV1) or TRPA1, respectively. p‐ERK1/2 was determined by Western immunoblotting in the dorsal root ganglia and in the sciatic nerve of rats following either systemic or perineural capsaicin treatment, or mustard oil application to the hind paw skin. To investigate the possible involvement of neurokinin 1 (NK₁) and NK₂ receptors as well as of nitric oxide, the selective antagonists, SR140333 for NK₁ and SR48968 for NK₂, and the nitric oxide synthase inhibitor, N^G‐nitro‐L‐arginine methyl ester (L‐NAME), were employed. The increase of p‐ERK1/2 after systemic capsaicin treatment was markedly attenuated by SR140333, while only the increase in the dorsal root ganglia was impaired by SR48968; in contrast, inhibition of nitric oxide synthase had no effect. Perineural capsaicin induced an increase in p‐ERK1/2 in the ipsilateral sciatic nerve and in the dorsal root ganglia. This effect was not influenced by SR140333 or L‐NAME. We found for the first time that mustard oil application to the hind paw skin caused an increase in p‐ERK1/2 in the sciatic nerve and in the dorsal root ganglia and only the phosphorylation in the latter was attenuated by SR140333 while L‐NAME showed no effect. From the present results, it may be assumed that capsaicin‐ or mustard oil‐induced p‐ERK1/2 in sensory neurons is not solely directly linked to TRPV1 or TRPA1 channels, but under certain conditions NK₁‐ and NK₂‐mediated mechanisms are involved.     

Involvement of sensory nerves and TRPV1 receptors in the rat airway inflammatory response to two environment pollutants: diesel exhaust particles (DEP) and 1,2-naphthoquinone (1,2-NQ)

The environmental chemical 1,2-naphthoquinone (1,2-NQ) is implicated in the exacerbation of airways diseases induced by exposure to diesel exhaust particles (DEP), which involves a neurogenic-mediated mechanism. Plasma extravasation in trachea, main bronchus and lung was measured as the local ¹²⁵I-bovine albumin accumulation. RT-PCR quantification of TRPV1 and tachykinin (NK₁ and NK₂) receptor gene expression were investigated in main bronchus. Intratracheal injection of DEP (1 and 5 mg/kg) or 1,2-NQ (35 and 100 nmol/kg) caused oedema in trachea and bronchus. 1,2-NQ markedly increased the DEP-induced responses in the rat airways in an additive rather than synergistic manner. This effect that was significantly reduced by L-732,138, an NK₁ receptor antagonist, and in a lesser extent by SR48968, an NK₂ antagonist. Neonatal capsaicin treatment also markedly reduced DEP and 1,2-NQ-induced oedema. Exposure to pollutants increased the TRPV1, NK₁ and NK₂ receptors gene expression in bronchus, an effect was partially suppressed by capsaicin treatment. In conclusion, our results are consistent with the hypothesis that DEP-induced airways oedema is highly influenced by increased ambient levels of 1,2-NQ and takes place by neurogenic mechanisms involving up-regulation of TRPV1 and tachykinin receptors.     

Erratum to: Involvement of sensory nerves and TRPV1 receptors in the rat airway inflammatory response to two environment pollutants: diesel exhaust particles (DEP) and 1,2-naphthoquinone (1,2-NQ)

The environmental chemical 1,2-naphthoquinone (1,2-NQ) is implicated in the exacerbation of airways diseases induced by exposure to diesel exhaust particles (DEP), which involves a neurogenic-mediated mechanism. Plasma extravasation in trachea, main bronchus and lung was measured as the local ¹²⁵I-bovine albumin accumulation. RT-PCR quantification of TRPV1 and tachykinin (NK₁ and NK₂) receptor gene expression were investigated in main bronchus. Intratracheal injection of DEP (1 and 5 mg/kg) or 1,2-NQ (35 and 100 nmol/kg) caused oedema in trachea and bronchus. 1,2-NQ markedly increased the DEP-induced responses in the rat airways in an additive rather than synergistic manner. This effect that was significantly reduced by L-732,138, an NK₁ receptor antagonist, and in a lesser extent by SR48968, an NK₂ antagonist. Neonatal capsaicin treatment also markedly reduced DEP and 1,2-NQ-induced oedema. Exposure to pollutants increased the TRPV1, NK₁ and NK₂ receptors gene expression in bronchus, an effect was partially suppressed by capsaicin treatment. In conclusion, our results are consistent with the hypothesis that DEP-induced airways oedema is highly influenced by increased ambient levels of 1,2-NQ and takes place by neurogenic mechanisms involving up-regulation of TRPV1 and tachykinin receptors.     

Differential role of tachykinin NK3 receptors on cholinergic excitatory neurotransmission in the mouse stomach and small intestine

Background and purpose:

Tachykinin NK₃ receptors are widely expressed in the mouse gastrointestinal tract but their functional role in enteric neuromuscular transmission remains unstudied in this species. We investigated the involvement of NK₃ receptors in cholinergic neurotransmission in the mouse stomach and small intestine.

Experimental approach:

Muscle strips of the mouse gastric fundus and ileum were mounted in organ baths for tension recordings. Effects of NK₃ agonists and antagonists were studied on contractions to EFS of enteric nerves and to carbachol.

Key results:

EFS induced frequency-dependent tetrodotoxin-sensitive contractions, which were abolished by atropine. The cholinergic contractions to EFS in the stomach were enhanced by the NK₃ antagonist SR142801, but not affected by the NK₃ agonist senktide or neurokinin B. The cholinergic contractions to EFS in the small intestine were not affected by SR142801, but dose-dependently inhibited by senktide and neurokinin B. This inhibitory effect was prevented by SR142801 but not by hexamethonium. SR142801, senktide or neurokinin B did not induce any response per se in the stomach and small intestine and did not affect contractions to carbachol.

Conclusions and implications:

NK₃ receptors modulate cholinergic neurotransmission differently in the mouse stomach and small intestine. Blockade of NK₃ receptors enhanced cholinergic transmission in the stomach but not in the intestine. Activation of NK₃ receptors inhibited cholinergic transmission in the small intestine but not in the stomach. This indicates a physiological role for NK₃ receptors in mouse stomach contractility and a pathophysiological role in mouse intestinal contractility.     

Involvement of vanilloid receptors and purinoceptors in the Phoneutria nigriventer spider venom-induced plasma extravasation in rat skin

Phoneutria nigriventer venom causes stimulation of capsaicin-sensitive primary afferent neurons in the rat dorsal skin, leading to neurogenic plasma protein extravasation due to the release of tachykinin NK(1) receptor agonist. In this study we further investigated the mechanisms involved in the venom-induced activation of capsaicin-sensitive primary afferent neurons. The plasma extravasation in response to venom intradermally injected was measured in Wistar rats as the local accumulation of i.v. injected 125I-labelled human serum albumin into skin sites. The tachykinin NK(1) receptor agonist, D-Ala-[L-Pro(9),Me-Leu(8)]substance P-(7-11) (GR73632; 10-100 pmol/site), induced a significant plasma leakage that was abolished by the selective tachykinin NK(1) receptor antagonist, (S)-1-[2-[3-(3,4-dichlorphenyl)-1 (3-isopropoxyphenylacetyl) piperidin-3-yl] ethyl]-4-phenyl-1 azaniabicyclo [2.2.2]octane chloride (SR140333; 1 nmol/site), whereas the leakage after venom (1-10 microgram/site) was significantly inhibited (but not abolished) by SR140333. The calcitonin gene-related peptide (CGRP) receptor antagonist, CGRP-(8-37), failed to further reduce the residual plasma extravasation induced by venom plus SR140333. The mu-opioid receptor agonist, [D-Ala(2), Me-Phe(4),Gly-ol(5)]enkephalin (DAMGO), and the local anaesthetic, lignocaine, had no effect on the venom-induced plasma extravasation. Similarly, the L-, N- and P/Q-type voltage-sensitive Ca(2+) channel blockers (verapamil, omega-conotoxin MVIIA and MVIIC, respectively) as well as the Na(+) channel blockers, tetrodotoxin and carbamazepine, had no effect on the venom-induced effect. Neither the systemic treatment nor the local injection of ruthenium red prevented the venom-induced plasma extravasation. However, the vanilloid receptor antagonist, N-[2-(4-chlorophenyl) ethyl]-1,3,4, 5-tetrahydro-7,8-dihydroxy-2H-2-benzazepine-2-carbothioamide (capsazepine; 120 micromol/kg, i.v.), reduced by 48% (P<0.05) the venom (10 microgram/site)-induced plasma extravasation. A significant inhibitory effect was also observed with the P(2) purinoceptor agonists, adenosine 5'-triphosphate (ATP; 10 and 30 nmol/site) and adenosine 5'-diphosphate (ADP; 10 nmol/site). The involvement of histamine and/or 5-hydroxytryptamine (5-HT) in the venom-induced plasma extravasation was ruled out since neither histamine and 5-HT receptor antagonists nor depletion of mast cells by compound 48/80 affected the venom response. This was further supported by the failure of venom to degranulate in vitro peritoneal mast cells. In conclusion, only vanilloid receptors and P(2) prejunctional purinoceptors had an inhibitory effect on the neurogenic plasma extravasation evoked by P. nigriventer venom in rat dorsal skin.     

Pharmacological characterization of mouse hind paw oedema induced by Bothrops insularis (jararaca ilhoa) snake venom.

Bothrops snake venoms produce marked local effects, including oedema, haemorrhage and necrosis. The ability of Bothrops insularis venom to induce oedema in mice was investigated. Venom was injected into hind paws and the change in volume over time was measured by plethysmometry. B. insularis venom (0.01-2.5 microg/paw) induced paw oedema which, at high doses (>/=0.5 microg/paw), was accompanied by haemorrhage. The peak oedematogenic response occurred 3 h after venom injection with all doses and decreased gradually thereafter, but was still elevated with high doses after 24 h. Pretreating the mice with cyproheptadine (histamine H(1) and serotonin 5-HT(2) receptor antagonist), mepyramine (histamine H(1) receptor antagonist), L-NAME (inhibitor of nitric oxide synthase), indomethacin and rofecoxib (inhibitors of cyclooxygenases), and dexamethasone (indirect inhibitor of PLA(2)) significantly attenuated venom-induced oedema, whereas methysergide, a serotonin 5-HT(1)/5-HT(2) receptor antagonist, had no effect. The administration of antivenom 30 min before or immediately after venom injection also significantly inhibited venom-induced oedema. These results show that B. insularis venom causes oedema in the mouse hind paw and that this response is mediated by histamine, nitric oxide, and arachidonic acid metabolites formed by cyclooxygenases 1 and 2. The neutralization by commercial antivenom indicates that the venom components responsible for oedema are recognized by the antivenom and share immunological identity with their counterparts in the venoms of mainland Bothrops species.     

Differential roles of neurokinin 1 and neurokinin 2 receptors in the development and maintenance of heat hyperalgesia induced by acute inflammation

1. Following induction of acute inflammation by intraarticular injection of kaolin and carrageenan into the knee joint in rats, there was a significant decrease in the withdrawal latency to radiant heat applied to the paw (i.e. heat hyperalgesia), an increased joint circumference and increased joint temperature.
2. A neurokinin₁ (NK₁) receptor antagonist (CP-99,994, 10 mM) had no effect on the paw withdrawal latency when it was administered spinally through a microdialysis fibre before the induction of inflammation. Pretreatment with a NK₂ receptor antagonist (SR48968, 1 mM) administered spinally through the microdialysis fibre prevented the heat hyperalgesia from developing in the early stages of the inflammation.
3. Post-treatment through the microdialysis fibre with the NK₁ receptor antagonist (0.0110 mM) was effective in reversing the heat hyperalgesia. In contrast, post-treatment spinally with the NK₂ receptor antagonist (0.011 mM) had no effect on the heat hyperalgesia. The inactive stereoisomers of the NK₁ receptor antagonist, CP100,263, or the NK₂ receptor antagonist, SR48965, administered at the same doses, had no effect on the joint inflammation or the heat hyperalgesia.
4. Pretreatment systemically with the NK₁ receptor antagonist (30 mg kg⁻¹) had no effect on the heat hyperalgesia or pain-related behaviour ratings where 0 is none and 5 is non weight bearing and complete avoidance of limb contact. Pretreatment with a NK₂ receptor antagonist (10 mg kg⁻¹) systemically prevented the heat hyperalgesia and pain-related behaviour ratings from developing in the early stages of the inflammation. The inactive stereoisomers of NK₁ receptor antagonist, CP100,263, or the NK₂ receptor antagonist, SR48965, administered at the same doses, had no effect on the joint inflammation or the heat hyperalgesia.
5. Post-treatment systemically with either the NK₁ (0.130 mg kg⁻¹) or the NK₂ (0.110 mg kg⁻¹) receptor antagonist resulted in a dose-dependent reversal of the heat hyperalgesia. Pain-related behaviour ratings were reduced by post-treatment only with the NK₁ receptor antagonist. The inactive stereoisomers of the NK₁ receptor antagonist, CP100,263, or the NK₂ receptor antagonist, SR48965, administered at the same doses, had no effect on the behavioural responses.
6. Direct pretreatment of the knee joint with either the NK₁ (30 mg) or the NK₂ (10 mg) receptor antagonist prevented the heat hyperalgesia from developing without affecting joint swelling. The inactive stereoisomers of the NK₁ receptor antagonist, CP100,263, or the NK₂ receptor antagonist, SR48965, administered at the same doses, had no effect on the joint inflammation or the heat hyperalgesia.
7. There appears to be a differential role for the spinal tachykinin receptors in the development and maintenance of the heat hyperalgesia associated with acute joint inflammation. The NK₂ receptors appear to be activated early in the development of the heat hyperalgesia and NK₁ receptors are involved in the maintenance of the heat hyperalgesia.
8. Peripherally, both NK₁ and NK₂ receptors are involved in the development of heat hyperalgesia and pain-related behaviour ratings induced by acute inflammation.

     

Characterization of sensory neurotransmission and its inhibition via α2B-adrenoceptors and via non-α2-receptors in rabbit iris

Summary To find out whether, and which type of, adrenoceptors mediate prejunctional inhibition of sensory neurotransmitter release from trigeminal fibres, the modulation of twitch response to electrical field stimulation on rabbit isolated iris was investigated. Evoked iris sphincter contractions consisted of a minor fast cholinergic and a large slow component. The latter was unaffected by atropine and guanethidine, hence nonadrenergic noncholinergic in nature (NANC), but nearly completely abolished by capsaicin pretreatment and by the neurokinin receptor antagonist spantide. The response was probably not mediated by NK₂ receptors as SR 48,968, an NK₂ selective nonpeptide antagonist, failed to reduce the response to the release of the endogenous neurokinin(s) (and exogenous substance P), but in part due to NK₁ receptor activation as shown by a reduction of response by CP 96,345, an NK₁ selective non-peptide antagonist, and in part perhaps mediated by NK₃ receptors. A small neurokinin receptor antagonist- and capsaicin-insensitive NANC contraction is probably not mediated by CGRP receptors.The ₂-adrenoceptor agonist oxymetazoline inhibited the evoked NANC response (22 nmol/1, IC₂₀; about 40%, maximum inhibition) without affecting the cholinergic response (up to 1 mol/1) or the postjunctional iris sensitivity to exogenous substance P. The inhibition was antagonized by rauwolscine (apparent -log K_B 8.04) and by the relatively _2B-adrenoceptor selective antagonist ARC-239 (-log K_B 8.51).The ₂- and imidazoline receptor agonist aganodine inhibited the evoked NANC response (0.25 mol/l, IC₂₀; about 30%, maximum inhibition) without affecting the postjunctional substance P responses. Rauwolscine 0.3 mol/l failed to antagonize this effect.It is concluded that the release of sensory neurotransmitter(s) from trigeminal fibres in the rabbit eye may be inhibited by _2B-adrenoceptors and by a non-₂-receptor, perhaps an imidazoline receptor.This study was supported by the Deutsche Forschungsgemeinschaft (Fu 163/3)Correspondence to H. Fuder at the above address     

Effect of SR 142801 on nitric oxide-dependent and independent responses to tachykinin NK3 receptor agonists in isolated guinea-pig colon

We have determined the ability of the novel nonpeptide tachykinin (TK) NK₃ receptor antagonist, SR 142801, [(S) -(N)-(1-(3-(1-benzoyl-3-(3,4-dichlorophenyl) piperidin-3-yl) propyl)-4-phenylpiperidin-4-yl)-N-methylacetamide] in inhibiting the nitric oxide (NO)-independent prejunctional inhibition of cholinergic twitches and the NO-dependent relaxation produced by the NK₃ receptor selective agonist, senktide, in the circular muscle of the guinea-pig proximal colon. Under moderate load (10 mN) and isometric recording of mechanical activity, single pulse electrical field stimulation (EFS) produced atropine- and tetrodotoxin-sensitive twitch contractions of mucosa-free circular muscle strips from the guinea-pig proximal colon. In the presence of NK₁ and NK₂ receptor antagonists (SR 140333 0.01 M and GR 94800 0.1 M, respectively) the NK₃ receptor selective agonist, senktide (EC₅₀ 33 pM) and the NK₃ receptor preferring natural TK, neurokinin B (NKB, EC₅₀ 13 pM) produced a concentration-dependent slowly developing inhibition of cholinergic twitches. Senktide (1 nM) did not affect the contractile response to acetylcholine (1 gM) indicating that depression of evoked twitches occurs prejunctionally. The inhibitory effect of senktide was 'unaffected when evoked in the presence of the cyclooxygenase inhibitor (S)-ketoprofen (10 M), guanethidine (10 M), naloxone (0.3 M), the GABA_B receptor antagonist 2-hydroxysaclofen (10 M) or the combined application of the adenosine A₁ and A₂ receptor antagonists, 8-cyclopentyl-1,3-dipropylxanthine (10 M) and 3,7-dimethyl-l-propargylxanthine (30 M) respectively.In the presence of NK₁ and NK₂ receptor antagonists, the NO-synthase inhibitor l-nitroarginine (L-NOARG 30-100 M) did not affect twitch inhibition induced by senktide (EC₅₀ 33 pM). The response to NKB (EC₅₀ 95 pM) was slightly reduced by L-NOARG, yet the bulk of the inhibitory effect of both agonists on cholinergic twitches was substantially independent of NO generation. SR 142801 (0.1–0.3 M) produced a moderate rightward shift of the concentration-response curve to senktide without depression of the Emax to the agonist, yielding an apparent pK_B value of 7.65.Under low resting tone (3 mN) and isotonic recording of mechanical activity, mucosa-free circular muscle strips from the guinea-pig proximal colon gained a high intrinsic tone suitable for testing the response to relaxant agents. In the presence of atropine (1 M), guanethidine (3 M), SR 140333 (0.01 [M) and GR 94800 (0.1 LM), senktide (EC₅₀ 50 pM) produced a concentration-dependent relaxation of the strips, which was blocked by L-NOARG. SR 142801 (0.01–0.1 M) produced a large rightward shift of the L-NOARG-sensitive concentration-response curve to senktide yielding an apparent pKB value of 8.62. Under isometric recording condition, SR 142801 (0.1 M) did not affect twitch inhibition produced by 3 nM clonidine. Under isotonic recording condition, SR 142801 did not affect the L-NOARG-sensitive relaxation produced by EFS.The present results indicate that NK₃ receptor stimulation produces a NO-dependent relaxation of the guinea-pig colon and a substantially NO-independent prejunctional inhibition of cholinergic twitches. The variable affinities of SR 142801 in antagonizing various senktide-induced neuromodulatory effects in the guinea-pig intestine suggest a possible intraspecies heterogeneity of NK₃ receptors in the enteric nervous system.