Tachykinin receptors on human monocytes: their involvement in rheumatoid arthritis

Three types of tachykinin receptors, namely NK1, NK2 and NK3, are known to preferentially interact with substance P (SP), neurokinin A (NKA) and neurokinin B (NKB), respectively. Experimental evidence indicates that SP and NKA modulate the activity of inflammatory and immune cells, including mononuclear ones. This study evaluated the effects of mammalian tachykinins and selective tachykinin agonists and antagonists on human monocytes isolated from healthy donors: SP, NKA and NKB all evoked a dose-dependent superoxide anion (O2-) production and the NK2 selective agonist [beta-Ala8]-NKA(4-10) induced a full response. The NK3 selective agonist senktide was inactive, while the NK1 selective agonists septide and [Sar9Met(O2)11]SP displayed some effects. These results indicate that NK2 and also some NK1 receptors are present in monocytes isolated from healthy donors. The role of tachykinin receptor activation in rheumatoid arthritis was also investigated, by measuring O2- production and TNF-alpha mRNA expression in monocytes isolated from rheumatoid patients. Tachykinins enhanced the expression of this cytokine in both control and rheumatoid monocytes and NK2 receptor stimulation was shown to trigger an enhanced respiratory burst in monocytes from rheumatoid patients. In conclusion, these results indicate that NK2 and NK1 receptors are present on human monocytes, the former being preferentially involved in rheumatoid arthritis.     

Effects of selective tachykinin-receptor antagonists on tachykinin-induced airway mucus secretion in the rat

Tachykinins like substance P (SP), neurokinin A (NKA), neurokinin B (NKB) differentially stimulate airway mucus secretion with the following rank order of potency in rat trachea: SP>NKA>NKB. These differential actions are most likely due to different affinities to the tachykinin receptors, termed neurokinin (NK)(1), NK(2)and NK(3). In this study we characterized the receptor subtype responsible for the differential secretagogue effects in rat trachea by means of selective receptor antagonists and receptor agonists.SR 140333 [NK(1)-antagonist] completely inhibited SP action (283,29+/-21, 12%-->84,53+/-4, 09%; P<0,01) and significantly reduced the effects of NKA (179,08+/-17,34%-->118,86+/-6,7%; P<0,01) and NKB (171,89+/-5, 75%-->109,5+/-4,11%; P<0,01). SR 48968 [NK(2)-antagonist] did not affect SP action, but reduced the effects of NKA and NKB. SR 142801 [NK(3)-antagonist] did not change any effect of SP, NKA or NKB. [Sar(9)]SP (NK(1)-agonist) caused strong dose-dependent secretagogue effects similar to SP, [betaAla(8)]NKA (NK(2)-agonist) showed only slight and [Pro(7)]NKB (NK(3)-agonist) no effects.The present data suggest that the secretagogue effects elicited by tachykinins in rat trachea are mediated via NK(1)receptors.     

Participation of GABAA receptors in the modulation of experimental anxiety by tachykinin agonists and antagonists in mice

Mice were acutely intraperitoneally treated with diazepam (DZP), pentylenetetrazol (PTZ) or NaCl 0.9% (control group), and 15 min later, the DZP-treated group received substance P (SP), neurokinin A (NKA; NK1 and NK2 natural preferential agonists), [Trp7 beta-Ala8] NKA(4-10) (Trp-7; NK3 antagonist) or vehicle intracerebroventricularly, whereas the PTZ-treated group was intracerebroventricularly administered with FK 888, SR 48968 (NK1 and NK2 antagonists, respectively) or senktide (SENK--[succinil-Asp6, MePhe8] substance P(6-11); NK3 agonist), or vehicle immediately before they were submitted to the elevated plus-maze (EPM) test. Another group of animals was repeatedly treated with increasing doses of DZP or NaCl 0.9% intraperitoneally for 28 days, and 3 days after the last injection (test day), animals received DZP, FK 888, SR 48968, SENK or vehicle intracerebroventricularly, or DZP (NaCl 0.9%) intraperitoneally before the EPM evaluation. The anxiolytic action of the acute treatment with DZP was inhibited by the central administration of NKA and Trp-7 but not by SP. NK1 and NK2 antagonists, but not NK3 agonist, blocked the anxiogenic action of PTZ, as evaluated in the plus-maze test. Flumazenil (FLM), a benzodiazepine antagonist, was not able to inhibit the anxiolytic profile of action induced by the NK2 antagonist. Central administration of FK 888 and SR 48968 promoted anxiolytic effects in both control and DZP-withdrawn animals, suggesting a clear relationship between the GABAergic and the tachykinergic systems, mostly involving NK1 and NK2 receptors, in the modulation of experimental anxiety in mice.     

Permissive role of neurokinin NK(3) receptors in NK(1) receptor-mediated activation of the locus coeruleus revealed by SR 142801.

The present experiments investigated the role of neurokinin-1 (NK(1)) and neurokinin-3 (NK(3)) receptors on the activity of the locus coeruleus (LC)-noradrenergic system by using a dual probe microdialysis technique in anesthetized guinea pigs. The local application in the LC of the selective NK(1) receptor agonists [SAR(9),Met(O(2))(11)]-SP (10 microM) and septide (1 microM) as well as the selective NK(3) receptor agonist senktide (1 microM), enhanced the extracellular norepinephrine (NE) levels in the prefrontal cortex. The enhancing effect of [SAR(9),Met(O(2))(11)]-SP was completely blocked by the peripheral administration of the selective non peptide NK(1) and NK(3) receptor antagonists, GR 205171 (1 mg/kg, i.p.) and SR 142801 (0.1 mg/kg, i.p.), respectively, whereas SR 142806 (0.1 mg/kg, i.p.) the inactive enantiomer of SR 142801 had no effect. Moreover, the [SAR(9),Met(O(2))(11)]-SP-induced increase in LC DOPAC concentrations, is only antagonized by GR 205171. In contrast, only SR 142801 (0.3 mg/kg, i.p.) could block stereoselectively the senktide-evoked increase in NE levels. Both [SAR(9),Met(O(2))(11)]-SP and senktide effects were blocked by local infusion into the LC of SR 142801 (10(-9) M). These results demonstrate that stimulation of NK(1) and NK(3) receptors located in the LC area modulates the activity of the LC-NE system, and that the excitatory effects of NK(1) receptor agonists require NKB/NK(3) receptor activation in the LC.     

Contribution of NK3 tachykinin receptors to propulsion in the rabbit isolated distal colon.

The role of NK3 receptors in rabbit colonic propulsion has been investigated in vitro with the selective agonist, senktide, and two selective antagonists, SR142801 and SB222200. Peristalsis was elicited by distending a rubber balloon with 0.3 and 1.0 mL of water leading to a velocity of 2.2 and 2.8 mm s-1, respectively. At concentrations of 1 nM, senktide inhibited propulsion evoked by both distensions (range 25-40%), whereas at 6 and 60 nmol L-1 facilitated 'submaximal' propulsion by 30%. In the presence of Nomega-nitro-L-arginine (L-NNA, 200 micromol L-1), which per se caused a slight prokinetic effect, 1 nmol L-1 senktide markedly accelerated propulsion (range 35-50%). Hexamethonium (200 micromol L-1) had minor effects on propulsion. In its presence, 60 nmol L-1 senktide significantly inhibited propulsion induced by both stimuli (range 20-50%). SR142801 (0.3, 3 nmol L-1) and SB222200 (30, 300 nmol L-1) facilitated 'submaximal' propulsion (range 20-40%). Conversely, higher antagonist concentrations (SR142801: 30, 300 nM; SB222200: 1, 10 micromol L-1) inhibited propulsion to both distensions by 20%. A combination of SR142801 (300 nmol L-1) plus hexamethonium (200 micromol L-1) induced an approximately four-fold greater inhibition of propulsion than that induced by SR142801 alone. In conclusion, in the rabbit-isolated distal colon, a subset of NK3 receptors located on descending pathways mediates an inhibitory effect on propulsion by activating a NO-dependent mechanism. Another subset of NK3 receptors, located on ascending pathways mediates a facilitative effect involving a synergistic interaction with cholinergic nicotinic receptors.     

Tachykinin activation of human monocytes from patients with interstitial lung disease, healthy smokers or healthy volunteers

Three types of tachykinin receptors, NK(1), NK(2)and NK(3), have been described to preferentially interact with substance P (SP), neurokinin A (NKA) and neurokinin B (NKB) respectively. Experimental evidence indicates that SP and NKA modulate the activity of inflammatory and immune cells, including mononuclear ones, and points to their involvement in lung pathophysiology. We previously reported that NK(1)and NK(2)receptors are present on monocytes (MO) isolated from healthy donors or rheumatoid patients - a greater sensitivity to NK(2)receptor stimulation was observed in the latter condition. This study evaluated the effects of SP and NKA, as well as NK(1)and NK(2)selective agonists and antagonists, on MO obtained from healthy volunteers, healthy smokers or patients with interstitial lung diseases (e.g. sarcoidosis and idiopathic pulmonary fibrosis). Superoxide anion (O(2)(-)) production was chosen as a parameter of cell activation. SP and NKA dose-dependently evoked O(2)(-)production from MO in all the conditions evaluated, their effects being competitively antagonized by selective antagonists (CP 96 345 and MEN 10 627, respectively). When selective NK(1)and NK(2)agonists were used, [Sar(9)Met(O(2))(11)]SP, a selective NK(1)agonist, induced a more than doubled O(2)production in MO obtained from patients with interstitial lung diseases as compared to healthy volunteers, whereas MO isolated from healthy volunteers were more sensitive to NK(2)receptor stimulation.     

Intestinal motor stimulation by the 5-HT4 receptor agonist ML10302: differential involvement of tachykininergic pathways in the canine small bowel and colon

5-Hydroxytryptamine (5-HT)4 receptor agonists stimulate gut motility through cholinergic pathways, although there are data suggesting that noncholinergic (tachykininergic) excitatory pathways may also be involved. Differences may exist between the small bowel and colon. Our aims were: (i) to compare the prokinetic effect exerted by the 5-HT4 receptor agonist ML10302 in the canine small bowel and colon in vivo; and (ii) to investigate the role of tachykininergic pathways in mediating this response. In fasting, conscious dogs, chronically fitted with electrodes and strain-gauge force transducers along the small bowel and colon, intravenous injection of ML10302 (35 microg kg-1) immediately stimulated spike activity and significantly increased propagated myoelectrical events at both intestinal levels. In the small bowel, the effects of ML10302 were unchanged by previous administration of the selective NK1 tachykinin receptor antagonist SR140333, the NK2 tachykinin receptor antagonist SR48968, or the NK3 tachykinin receptor antagonist SR142801. In the colon, all tachykinin receptor antagonists significantly inhibited stimulation of spike and mechanical activity by ML10302, without affecting ML10302-induced propagated myoelectrical events. Atropine (100 microg kg-1 i.v.) suppressed the stimulatory effect of ML10302 at both intestinal levels. In conclusion, the 5-HT4 receptor agonist ML10302 induced significant prokinesia both in the small bowel and colon through activation of cholinergic pathways. Tachykininergic pathways are not involved in the ML10302-induced prokinesia in the small bowel, but they play an important role in mediating the colonic motor response to ML10302.     

Evidence for Modulation of DoDamine-neuronal Function by Tachykinin NK3 Receptor Stimulation in Gerbil Mesencephalic Cell Cultures

Primary cultures of gerbil mesencephalon were used for studying the modulation exerted by tachykinin NK₃ receptor activation on the activity of dopamine (DA) neurons. Dopamine neurons were identified by their ability to take up [³H]DA in a nomifensine-dependent manner. Moreover, tyrosine hydroxylase immunohistochemistry revealed that these neurons accounted for 5–7% of the total cell population. The NK₃ receptor agonists, senktide (EC₅₀= 0.58 nM) and [MePhe⁷]neurokinin B (EC₅₀= 3 nM), increased spontaneous [³H]DA release in a concentration-dependent manner. In contrast, tested at a supramaximal concentration (10-⁷ M), neither septide nor substance P were found to affect [³H]DA release. The senktide-evoked [³H]DA release was not observed when extracellular Ca²⁺ was chelated, but was unaffected by nomifensine. This indicates that this increase in [³H]DA outflow resulted more from an exocytotic process than from reversal of carrier-mediated DA uptake. Moreover, the senktide effect was unaffected by the Na⁺ channel blocker tetrodotoxin, a result suggesting a direct action of senktide on DA neurons. The non-peptide NK₃ receptor antagonist, SR 142801, shifted or blocked (ICs0 = 0.89 nM) the senktide-evoked [³H]DA release, while its (-)-antipode, SR 142806, was 80-fold less potent, in agreement with binding data. Selective antagonists for NK, (SR 140333) or NK₂ (SR 48968) receptors failed to reduce the senktide effect. Light scanning microscopic analysis of mesencephalic cells loaded with the Ca²⁺ sensitive dye, fluo-3, showed that senktide induced a rise in cytosolic Ca²⁺ in 8-10% of the cell population. The senktide-induced elevation in intracellular Ca²⁺ was rapid in onset and transient (at lo⁴ M) or more sustained with no further increase in fluorescence intensity (at 10^-7 M). The proportion of senktide-responsive cells was not significantly modified when extracellular Ca²⁺ was chelated, but was reduced by 87% in the presence of SR 142801 and by 75% in cultures that were pre-treated with the DA neurotoxin l-methyl-4-phenylpyridinium. The present study shows that enhancement of spontaneous [³H]DA release and intracellular Ca²⁺ mobilization may be observed after NK₃ receptor stimulation and that both biochemical events are likely to occur in DA neurons.     

Comparison in different tissue preparations of the in vitro pharmacological profile of RP 67580, a new non-peptide substance P antagonist.

We describe the effects of RP 67580, a new non-peptide substance P (SP) antagonist, on tachykinin-induced contractions of guinea-pig ileum, trachea and urinary bladder, rabbit pulmonary artery and rat portal vein. All NK1 agonists tested (SP, Septide, SPOMe and [Pro9]SP) contracted guinea-pig ileum, trachea and urinary bladder (pD2 = 7.5 to 9.1), but they had no effect on rabbit pulmonary artery or rat portal vein (pD2 < 6). RP 67580 inhibited these effects: guinea-pig ileum, pA2 = 7.1 to 7.6; guinea-pig trachea and urinary bladder, pKB = 6.3 to 6.8. The difference in RP 67580 activity in these tissues might be due to the existence of subtypes of NK1 receptors. RP 67580 (1 microns) did not affect the contractions induced by the two NK2 agonists, NKA and [Lys5, MeLeu9, Nle10]NKA(4-10) (pA2 < 6), except in guinea-pig ileum (pA2 = 7.3-7.5) where these two NK2 agonists interact apparently with NK1 receptors. In the tissue preparations used, RP 67580 (1 micron) was without effect on contractions induced by the NK3 agonists: NKB and senktide. These results indicate the high selectivity for NK1 receptors of RP 67580. In all cases, similar results were obtained with another non-peptide SP antagonist, (+/-) CP-96,345. The present work provides further evidence that RP 67580 and (+/-) CP-96,345 exert in vitro a potent, selective and competitive antagonistic action on NK1 receptors and suggests the existence of at least two distinct NK1 receptor subtypes in some guinea-pig peripheral organs.     

Pharmacological characterization of mucin secretion from CHO-K1-hNK(1)R cells

Numerous respiratory diseases increase mucin secretion from human airways. Several investigators hypothesize that mucin secretion from airway epithelium is NK(1)-receptor mediated. We have developed a mucin secretion assay using CHO-K1 cells transfected with the human NK(1)receptor (CHO-K1-hNK(1)R) that respond to NK(1)-specific agonists. Cells were labeled with [(3)H]-glucosamine and stimulated with agonists including Ac-[Arg(6), Sar(9), Met(O(2))(11)] Substance P(6-11) (ASMSP; NK(1)-specific), [beta-Ala(8)]-Neurokinin A(4-10) (BANK; NK(2)-specific), or human neutrophil elastase (HNE). Basal mucin secretion from CHO-K1-hNK(1)R and non-transfected cells was similar. Stimulation of CHO-K1-hNK(1)R, but not CHO-K1, with ASMSP or BANK concentration-dependently increased mucin secretion (pD(2)value[Emax] = 8.9(1)+/-0.1(3)[175%] and 7.56+/-0.05[100%], respectively). SR140333 (NK(1)antagonist), but not SR48968 (NK(2)antagonist), decreased ASMSP- and BANK-induced mucin release from CHO-K1-hNK(1)R. In these cells, endothelin-1, angiotensin II, serotonin, phenylephrine, senktide, and methacholine showed negligible effects on mucin secretion. A similar lack of effect of these agonists was observed in non-transfected CHO-K1 cells. HNE increased mucin release four to five fold in both cell types. These studies demonstrate that stimulation of CHO- K1-hNK(1)R with ASMSP and BANK causes robust and NK(1)-selective mucin release.     

Contractile effect of tachykinins on Suncus murinus (house musk shrew) isolated ileum

Recent studies used Suncus murinus to investigate the anti-emetic potential of NK(1) tachykinin receptor antagonists. However, the pharmacology of tachykinin receptors in this species has not been fully characterized. In the present studies, therefore, we examined a range of tachykinin receptor agonists for a capacity to induce contractions of the isolated ileum. The tachykinin NK1 receptor preferring agonists substance P, septide and [Sar9Met(O2)11] substance P, and the tachykinin NK2 preferring agonists neurokinin A and GR 64349 (Lys-Asp-Ser-Phe-Val-Gly-R-gamma-lactam-Leu-Met-NH2) caused concentration dependent contractions with EC50 values in the nanomolar range. However, the tachykinin NK3 preferring agonists neurokinin B and senktide (1nM-1microM) induced only weak contractions. The action of senktide, but not [Sar9Met(O2)11] substance P, septide, or GR 64349, was antagonized significantly by atropine (P<0.05); tetrodotoxin and hexamethonium were inactive. The tachykinin NK1 receptor antagonist CP-99,994 ((+)-[(2S,3S)-3-(2-methoxy-benzyl-amino)-2-phenylpiperidine]) (10-100nM) inhibited substance P- and septide-induced contractions non-competitively. The pA2 value estimated for CP-99,994 against septide was 7.3+/-0.1. It also non-competitively antagonized the contractile responses induced by [Sar9Met(O2)11] substance P with a pA2 of 7.4+/-0.1. CP-99,994 also had a slight inhibitory action on neurokinin A-induced contractions, but did not modify the action of GR 64349. Conversely, the tachykinin NK2 receptor antagonist, saredutant, competitively antagonized GR 64349-induced contractions with a pA2 of 7.34+/-0.02. On the other hand, the presence of both CP-99,994 and saredutant competitively antagonized substance P-induced contraction. The present studies indicate that tachykininNK1 and NK2 receptors exist in the ileum of S. murinus and are involved in mediating contractions directly on smooth muscle, whereas tachykinin NK3 receptors may play a minor role involving a release of acetylcholine.     

Control by tachykinin NK(2) receptors of CRF(1) receptor-mediated activation of hippocampal acetylcholine release in the rat and guinea-pig

In vivo microdialysis was employed to explore the effects of different selective non-peptides NK(1),NK(2) and NK(3) receptor antagonists on the corticotropin releasing factor (CRF)-induced release of acetylcholine (ACh) in the hippocampus of rats and guinea-pigs. In both species, the intracerebroventricular (i.c.v.) administration of CRF produced a time- and dose-dependent increase in hippocampal ACh release that was totally suppressed by an intraperitoneally (i.p.) pretreatment with the selective non-peptide CRF(1) receptor antagonist antalarmin (30 mg/kg). Pretreatment with the selective NK(2) receptor antagonist SR48968 (1mg/kg, i.p.) significantly reduced the increase of ACh induced by CRF. In contrast, its low-affinity enantiomer SR48965 (1mg/kg, i.p.) or the NK(1) receptor antagonist, GR205171 (1mg/kg, i.p.) did not exert any antagonist effect. Moreover, administration of the selective NK(3) receptor antagonist SR142801 (1mg/kg, i.p.) did not significantly reduce the CRF-induced hippocampal ACh release in guinea-pigs (the only species studied). The selective activity of SR48968 versus GR205171 or SR142801 indicates that NK(2) receptors play a major role in the control of CRF-induced hippocampal ACh release. Moreover, in freely moving rats, two sessions of stroking of the neck and back of the rat for 30 min, at 90 min intervals, known to be a stressful stimulus, produced a marked and reproducible increase in hippocampal ACh release. This effect was prevented by the administration of the two selective non-peptide CRF1 and NK(2) receptor antagonists antalarmin (30 mg/kg, i.p.) and SR48968 (1mg/kg, i.p.), respectively. This suggests that stress-induced activation of the hippocampal ACh system may be under the control of both endogenously released CRF and NKA, and opens the possibility of the existence of a functional interplay between the pathways containing these peptides as we observed in our experiments on anaesthetized animals.     

Neuropeptide gamma, the most potent contractile tachykinin in human isolated bronchus, acts via a 'non-classical' NK2 receptor.

Cumulative contractile response curves to neurokinin A (NKA) and neuropeptide gamma (NP gamma) were obtained in human isolated bronchus, in the presence of phosphoramidon 10 microM. NP gamma was approximately 10-fold more potent than NKA (pD2 values 8.6 +/- 0.4 and 7.3 +/- 0.3 respectively, n = 6; P less than 0.01). The NK1-selective agonist [Sar9, Met(O2)11]-SP and the NK3 selective agonist senktide produced negligible contraction. Response curves to NP gamma and NKA were unaffected by the NK2 subtype-selective antagonist MDL 29913 at 2 microM, but NP gamma-induced contraction was markedly inhibited by 20 microM MDL 29,913. Thus NP gamma is the most potent tachykinin in human isolated bronchus and its effects are mediated at a receptor which is not of the 'classical' NK2 subtype found in hamster urinary bladder.     

Identification of mechanisms for duodenal contraction induced by tachykinins in the rat

Mechanisms for contractile effects of tachykinins on muscle strips of rat duodenum were studied using substance P (SP), neurokinin A (NKA), neurokinin B (NKB) and neuropeptide K (NPK), and the tachykinin analogues SP methyl ester (SPME), Nle¹⁰-NKA(4–10) (NleNKA) and senktide (SENK) selective for neurokinin (NK)-1, NK-2 and NK-3 receptors, respectively. NK receptors responsible for smooth muscle contraction were identified using selective ligands to protect NK receptors combined with inactivation of residual receptors with N-ethylmaleimide. Tachykinins (10^?9 to 10^?5 M) caused dose-related contractions of the muscle strips. The order of potency of native tachykinins was NKA > NKB > SP > NPK in circular, and NKB > NKA > NPK > SP in longitudinal muscle, whereas that of selective tachykinin analogues was SENK > NleNKA > SPME. NKA, NleNKA and SENK were equieffective as acetylcholine, whereas SP, SPME, NKB and NPK were less effective. Spantide decreased the sensitivity to all tachykinin analogues. Atropine reduced the sensitivity to SENK only, whereas hexamethonium reduced the sensitivity to SENK and SPME, but not to NleNKA. Selective receptor protection with SPME, NleNKA or SENK protected contractions induced by SPME, NleNKA and SENK, respectively. Responses to tachykinin analogues were reduced in Ca²⁺-free medium. Thus, NKA is suggested to be the dominating tachykinin to stimulate contraction of the rat duodenum via NK receptors coupled to Ca²⁺-dependent signal transduction pathways. Of the receptors available, the NK-1 subtype involves a nicotinic transmission step, and the NK-3 subtype also a muscarinic step, whereas the NK-2 receptor subtype is not dependent on cholinergic mechanisms.     

Structure-activity studies of neurokinin A

A structure-activity study on neurokinin A and its C-terminal fragment NKA (4-10) has been performed in order to find selective agonists for the NK-2 receptor and identify chemical modifications suitable for protecting the peptides from degradation, while maintaining activity. Five series of compounds have been prepared and tested: 1. the complete series of the L-Ala monosubstituted analogues of NKA; 2. a series of NKA fragments from the C- or N-terminal; 3. the complete series of NKA (4-10) analogues monosubstituted with beta-Ala; 4. a series of NKA (4-10) analogues with monosubstitutions in pos. 4, 8, 10 or multisubstitutions in two or more of the same positions; and 5. a series of 6 NKA (4-10) analogues monosubstituted with 1-amino,1-cyclohexane carboxylic acid residue. It has been found that the most selective agonists for the NK-2 receptor system are [beta Ala8]NKA (4-10) and [Nle10]NKA (4-10). Protection from aminopeptidase may be obtained by acetylation of the N-terminal amide of NKA (4-10), while partial protection from endopeptidases should be expected from the presence of beta-Ala in position 8. Conformational constraints induced with 1,amino,1-cyclohexane carboxylic acid residue gave weakly active compounds. Multiple substitutions reduce rather than potentiating the favorable effects of the corresponding monosubstituted compounds.     

Neurokinin receptor modulation of respiratory activity in the rabbit

The respiratory role of neurokinin (NK) receptors was investigated in α-chloralose-urethane-anaesthetized, vagotomized, paralysed and artificially ventilated rabbits by using bilateral microinjections (30–50 nL) of NK receptor agonists and antagonists. Microinjections were performed in a region located just caudal to the rostral expiratory neurons. This region displayed features similar to those of the pre-Bötzinger complex (pre-BötC) of adult cats and rats, and proved to produce excitatory respiratory effects in response to microinjections of d,l -homocysteic acid. We used as agonists (0.1, 0.5 and 5 m m ) substance P (SP), the NK1 receptor agonists [Sar⁹, Met(O₂)¹¹]-SP and GR 73632, the NK2 receptor agonist NKA, the NK3 receptor agonist senktide, and as antagonists (5 m m ) the NK1 receptor antagonist CP-99,994 and the NK2 receptor antagonist MEN 10376. SP always increased respiratory frequency, but NK1 receptor agonists did not change respiratory variables. NKA and senktide at 5 m m increased respiratory frequency. CP-99,994 caused increases in respiratory frequency and did not antagonize the effects of SP. MEN 10376 prevented the respiratory responses induced by NKA and reduced those provoked by SP. SP or the NK1 receptor agonists (5 m m ) injected (1 μL) into the IV ventricle caused marked excitatory effects on respiration. The results suggest that NK2 and NK3, but not NK1, receptors are involved in the excitatory modulation of inspiratory activity within the investigated region and are consistent with the notion that the pre-BötC neurons are important components of the inspiratory rhythm-generating mechanisms.     

NK(3) receptors in the feline nucleus tractus solitarius are not involved with the muscle pressor response

Isometric muscle contractions cause an increase in mean arterial pressure and heart rate. Previously, we showed that substance P (SP) is released from sites in the feline medial nucleus tractus solitarius (mNTS) in response to isometric muscle contractions, and that it most likely interacted with NK(1) tachykinin receptors at these sites. This study was undertaken to determine whether other tachykinin receptors in this area of the brainstem are involved with the muscle pressor response. Receptor autoradiography, using [(125)I]Bolton-Hunter SP and [(125)I] [MePhe(7)] neurokinin B to label NK(1) and NK(3) receptors, respectively, indicated that NK(3) tachykinin receptors are as abundant as NK(1) and NK(3) receptors, respectively, indicated that NK(3) tachykinin receptors are as abundant as NK(1) receptors in this region of the feline brainstem Injections of the specific NK(3) receptor antagonist, SR 142801 (0.1 to 10 microM) into the mNTS did not modify the pressor response or the heart rate response to isometric muscle contractions. Injection of SR142801 into the NTS prior to the injection of the NK(1) antagonist, GR82334 did not affect the action of GR82334 to attenuate the muscle pressor reflex. We conclude that NK(3) receptors in the NTS are not involved with the regulation of cardiovascular function during activation of the muscle pressor response.     

Role of kinin B1 and B2 receptors and mast cells in post intestinal infection-induced hypersensitivity to distension

Distension of the rat intestine causes a capsaicin-sensitive, pressure-dependent depressor response which is indicative of nociception. A hypersensitivity of jejunal distension which possibly involves tachykinin NK2 receptors and is restricted to areas with mast cell hyperplagia is observed in rats infected 30 days previously with Nippostrongylus brasiliensis. This study aimed to further investigate the role of mast cells, tachykinins and kinins in this intestinal hypersensitivity. The activity of a mast cell stabilizer (doxantrazole), kinin antagonists (des-Arg 10-[Leu9]-kallidin, B1, HOE 140, B2) and tachykinin antagonists (CP 99, 994, NK1, SR 142801, NK3) were tested against the distension-induced depressor responses in control and post-infected rats. The 30-day post-infection-induced hypersensitivity was significantly reduced by the mast cell stabilizer doxantrazole. The hypersensitivity had resolved in 90-day post-infected rats when mast cells levels had normalized. Des-Arg 10-[Leu9]-kallidin and HOE 140 did not inhibit the depressor responses in controls but produced a significant inhibition in 30-day post-infected rats. CP 99,994 inhibited the depressor responses in post-infected rats with an equal potency to that in control rats. SR 142801 was inactive in both groups. In conclusion, mast cells and kinin-mediated nociception appear to be involved in post-infection intestinal hypersensitivity whereas tachykinin NK1 and NK3 receptors do not.     

NK(2)-receptor mediated contraction in monkey, guinea-pig and human airway smooth muscle

Neurokinins (NK) are implicated in airway pathology. Selective NK(2)-receptor antagonists may prove therapeutic in airway disease. We studied Neurokinin A (NKA) responses of isolated, cryopreserved cynomolgus monkey, fresh guinea pig, and fresh and cryopreserved human airways. NKA contracted monkey trachea (pD(2)= 7.9), guinea-pig bronchus (pD(2)= 8.8) and human bronchus (pD(2)= 7.1). Potency rank order (pK(b)) of NK(2)-antagonists, SR 48968 and GR 159897, and a dual NK(1)/NK(2)-antagonist, MDL 103392, against NKA responses in monkey trachea, guinea pig and human bronchus, respectively, were SR 48968 (9.29 +/- 0.11, 9.15 +/- 0.10 and 9.51 +/- 0.17) > GR 159897 (8.45 +/- 0.26, 8.19 +/- 0.13 and 8.57 +/- 0. 22) > MDL 103392 (6.55 +/- 0.13, 6.97 +/- 0.14 and 7.16 +/- 0.13). CP 99994 (1 microM), a NK(1)-receptor antagonist, was inactive against NKA responses in all three species. The NK(3)-antagonist SR 142801 (1 microM) was inactive against NKA in monkey trachea and guinea-pig bronchus, but demonstrated weak antagonist activity (pK(b)= 6.97 +/- 0.03) in human bronchus. These findings demonstrate that NK(2)-receptors mediate tracheal smooth muscle contraction to NKA in cynomolgus monkey and that the pharmacological responsiveness of airway NK(2)-receptors in the three species studied is similar. Furthermore, our results suggest that cryopreservation may extend the viability of human and non-human primate airway tissue for studies of neurokinin receptor pharmacology. Studies are needed to further determine the similarity in neurokinin pharmacology between fresh and cryopreserved airway tissue.     

Behavioural effects of selective tachykinin agonists in midbrain dopamine regions.

The effects of selective NK-1, NK-2 and NK-3 tachykinin agonists in midbrain dopamine cell containing regions were investigated in the rat. The NK-3 agonist senktide induced locomotion, rearing and sniffing following infusion into the substantia nigra pars compacta, and to a lesser extent in the ventral tegmental area. These behavioural responses were not seen following infusion of the selective NK-1 agonist [Sar9,Met (O2)11]SP or the NK-2 agonist [N1e10]NKA4-10. In contrast, grooming was induced only by the NK-1 agonist administered into the substantia nigra. Yawning, chewing mouth movements and wet dog shakes were all seen following infusion of senktide into the ventral tegmental area. These findings suggest that (i) dopamine-mediated behavioural responses seen following tachykinin administration into the midbrain are dependent upon stimulation of NK-3 tachykinin receptors, (ii) tachykinin-induced grooming is mediated by stimulation of NK-1 receptors and (iii) some of the previously described 5-HT mediated behaviours seen following administration of NK-3 tachykinin agonists are probably generated by stimulation of 5-HT cell bodies in the ventral tegmental area.