Establishment of highly specific radioimmunoassays for neurokinin A and neurokinin B and determination of tissue distribution of these peptides in rat central nervous system

Highly specific radioimmunoassays (RIAs) for neurokinin A (NKA) and neurokinin B (NKB) were developed. Antisera were produced by the procedure which involved immunization with NKA or NKB, both conjugated with keyhole limpet hemocyanin, and treatments with a tolerogenic conjugate of kassinin and a copolymer of D-glutamic acid and D-lysine (D-GL) to inhibit the production of cross-reactive antibodies against common C-terminal region of tachykinins. Cross-reactivities of anti-NKA antiserum (R704), thus produced, with NKB, kassinin, eledoisin were 12.6%, 10.6% and 11.5%, respectively. This was in sharp contrast with those of antiserum obtained from the rabbit not treated with kassinin-D-GL, these values corresponding to 129.0%, 42.5% and 94.4%, respectively. The cross-reactivities of R704 with substance P and physalaemin were 0.3% and 1.5%, respectively. This antiserum also bound 35.6% of neuropeptide K which contains NKA at its C-terminal. More importantly, anti-NKB antiserum (R707) obtained by the above tolerizing regimen was highly specific for NKB and the cross-reactivities with NKA, neuropeptide K, kassinin and other tachykinins were all less than 0.001%. RIAs using these specific antisera allowed us to measure directly NKA and NKB in tissue extracts without their fractionation by chromatography prior to RIAs. Measurements of immunoreactive NKA and NKB in different rat brain regions and spinal cord revealed that they are present with various ratios (NKA/NKB: 1.1-9.9) depending on the region.     

A regulatory role for neurokinin B in placental physiology and pre-eclampsia

Tachykinin dogma has assumed, so far, that neurokinin B (NKB) is a neuropeptide that is not produced in any peripheral tissue even though its endogenous receptor, NK3, has been found in a number of locations throughout the human body. We have found an abundant source of peripheral NKB in the human and rat placenta. In this review we describe the discovery of NKB in the placenta and examine its possible role in placental physiology and pre-eclampsia (PE). Excessive secretion of placental NKB into the maternal circulation during the third trimester of pregnancy has been found in women suffering from PE. This may provide the key to the cause of the multiple and complex symptoms associated with this potentially life-threatening illness. We also reveal the structural organisation of the human NKB gene for the first time as well as discussing putative mechanisms for its control.     

Neurokinin A, neurokinin B and neuropeptide K in the rabbit iris: a study comparing different extraction methods

Neurogenic inflammation in the rabbit eye is thought to be partly mediated by tachykinins released from trigeminal sensory nerve fibres. In the present study we have investigated the occurrence of neurokinin A (NKA), neurokinin B (NKB), neuropeptide K (NPK) and related immunoreactive components in the rabbit iris-ciliary complex using neutral and different types of acidic media for extraction, reverse phase high performance liquid chromatography (RP-HPLC) and radioimmunoassay (RIA). The immunoreactive material detected with an antiserum reacting almost equally well with NKA, NKB and NPK consisted mainly of NKA, and small amounts of NPK but almost no NKB. Acidic media seemed to be more effective than neutral media for extraction of NKA and NPK. Acid extraction yielded also an NKA-immunoreactive component which eluted immediately before NKA while neutral extracts, on the other hand, contained a component which appeared behind NKA, i.e. in the position of NKA-(3-10) and NKA-(4-10). The present results indicate that NKA but not NKB may play a role in neurogenic inflammation in the rabbit eye.     

Substance P Immunoreactivity Exhibits Frequent Colocalization with Kisspeptin and Neurokinin B in the Human Infundibular Region

Neurons synthesizing neurokinin B (NKB) and kisspeptin (KP) in the hypothalamic arcuate nucleus represent important upstream regulators of pulsatile gonadotropin-releasing hormone (GnRH) neurosecretion. In search of neuropeptides co-expressed in analogous neurons of the human infundibular nucleus (Inf), we have carried out immunohistochemical studies of the tachykinin peptide Substance P (SP) in autopsy samples from men (21-78 years) and postmenopausal (53-83 years) women. Significantly higher numbers of SP-immunoreactive (IR) neurons and darker labeling were observed in the Inf of postmenopausal women than in age-matched men. Triple-immunofluorescent studies localized SP immunoreactivity to considerable subsets of KP-IR and NKB-IR axons and perikarya in the infundibular region. In postmenopausal women, 25.1% of NKB-IR and 30.6% of KP-IR perikarya contained SP and 16.5% of all immunolabeled cell bodies were triple-labeled. Triple-, double- and single-labeled SP-IR axons innervated densely the portal capillaries of the infundibular stalk. In quadruple-labeled sections, these axons formed occasional contacts with GnRH-IR axons. Presence of SP in NKB and KP neurons increases the functional complexity of the putative pulse generator network. First, it is possible that SP modulates the effects of KP and NKB in axo-somatic and axo-dendritic afferents to GnRH neurons. Intrinsic SP may also affect the activity and/or neuropeptide release of NKB and KP neurons via autocrine/paracrine actions. In the infundibular stalk, SP may influence the KP and NKB secretory output via additional autocrine/paracrine mechanisms or regulate GnRH neurosecretion directly. Finally, possible co-release of SP with KP and NKB into the portal circulation could underlie further actions on adenohypophysial gonadotrophs.     

Several mediators appear to interact in neurogenic inflammation

Plasma protein extravasation was studied in the rat abdominal skin. Substance P (SP), neurokinin A (NKA) and B (NKB) were found to induce extravasation with a threshold dose of about 1 pmol. Calcitonin gene-related peptide (CGRP) caused no or little extravasation alone but it potentiated the action of SP, NKA, NKB, and physalaemin. The potentiation of the SP-induced extravasation was unaffected by pretreatment with capsaicin, indomethacin or compound 48/80, it was reduced by neuropeptide Y or pretreatment with mepyramine plus cimetidine, and was abolished in streptozotocin diabetic rats. CGRP augmented extravasation induced by histamine, reduced the effect of ATP or adenosine and did not alter extravasation by serotonin, bradykinin or neurotensin. These results indicate that in addition to SP the novel mammalian tachykinins NKA and NKB may be considered as mediator candidates for neurogenic plasma extravasation. CGRP is a possible mediator of antidromic vasodilation. Furthermore, CGRP potentiates the extravasation caused by coexisting tachykinins and could thereby augment neurogenic inflammation. The diverse interactions of CGRP with other inflammatory mediators suggest multiple sites of action.     

神经激肽B及其生殖内分泌作用

神经激肽B(neurokinkin B,NKB)是速激肽家族的一员,主要通过其受体NK3R发挥作用。NKB及NK3R在神经系统分布广泛。NKB具有使空腔脏器平滑肌收缩、松弛血管、降低平均动脉压、减慢心率、兴奋离体大鼠脊髓神经元及收缩瞳孔括约肌缩小瞳孔等生物学作用。近年来NKB在生殖内分泌中的调控作用越来越受到关注,关于其调节下丘脑-垂体-性腺轴(HPGA)的作用的研究也越来越深入。本文综述了NKB及其受体的分布范围、生理功能、NKB在生殖内分泌调控过程中的作用,其具体的作用机制还有待于进一步研究。     

Degradative enzymes modulate airway responses to intravenous neurokinins A and B

We studied the effects of the neutral endopeptidase (NEP) inhibitor thiorphan (1.7 mg/kg iv) and the angiotensin-converting enzyme (ACE) inhibitor captopril (5.7 mg/kg iv) on airway responses to rapid intravenous infusions of neurokinin A (NKA) and neurokinin B (NKB) in anesthetized, mechanically ventilated guinea pigs. The dose of NKA required to decrease pulmonary conductance to 50% of its base-line value (ED50GL) was fivefold less (P less than 0.0001) in animals treated with thiorphan compared with controls. NKA1-8, a product resulting from cleavage of NKA by NEP, had no bronchoconstrictor activity. Similar results were obtained by using NKB as the bronchoconstricting agent. Captopril had no significant effect on airway responses to NKA or NKB. In contrast, both thiorphan and captopril decrease the ED50GL for substance P (SP). We also compared the relative bronchoconstrictor potency of NKA, NKB, and SP. In control animals, the rank order of ED50GL values was NKA much less than NKB = SP. NKA also caused a more prolonged bronchoconstriction than SP or NKB. Thiorphan had no effect on the rank order of bronchoconstrictor potency, but in animals treated with captopril, the rank order of ED50GL values was altered to NKA less than SP less than NKB. These results suggest that degradation of NKA and NKB by NEP but not by ACE is an important determinant of the bronchoconstriction induced by these peptides. The degradation by ACE of SP but not NKA or NKB influences the observed relative potency of the three tachykinins as bronchoactive agents.     

A Population of Kisspeptin/Neurokinin B Neurons in the Arcuate Nucleus May Be the Central Target of the Male Effect Phenomenon in Goats

Exposure of females to a male pheromone accelerates pulsatile gonadotropin-releasing hormone (GnRH) secretion in goats. Recent evidence has suggested that neurons in the arcuate nucleus (ARC) containing kisspeptin and neurokinin B (NKB) play a pivotal role in the control of GnRH secretion. Therefore, we hypothesized that these neurons may be the central target of the male pheromone. To test this hypothesis, we examined whether NKB signaling is involved in the pheromone action, and whether ARC kisspeptin/NKB neurons receive input from the medial nucleus of the amygdala (MeA)—the nucleus suggested to relay pheromone signals. Ovariectomized goats were implanted with a recording electrode aimed at a population of ARC kisspeptin/NKB neurons, and GnRH pulse generator activity, represented by characteristic increases in multiple-unit activity (MUA) volleys, was measured. Pheromone exposure induced an MUA volley and luteinizing hormone (LH) pulse in control animals, whereas the MUA and LH responses to the pheromone were completely suppressed by the treatment with an NKB receptor antagonist. These results indicate that NKB signaling is a prerequisite for pheromone action. In ovariectomized goats, an anterograde tracer was injected into the MeA, and possible connections between the MeA and ARC kisspeptin/NKB neurons were examined. Histochemical observations demonstrated that a subset of ARC kisspeptin/NKB neurons receive efferent projections from the MeA. These results suggest that the male pheromone signal is conveyed via the MeA to ARC kisspeptin neurons, wherein the signal stimulates GnRH pulse generator activity through an NKB signaling-mediated mechanism in goats.     

Neurokinin A and neurokinin B in the human retina

Very recently, the authors found levels of neurokinin (NK) A-like immunoreactivities in the human retina which were more than five times higher than those of substance P (SP). The present study aimed to find out how many of these immunoreactivities can be attributed to NKA and NKB and then the exact distribution pattern of both NKA and NKB was evaluated in the human retina and compared with that of SP. For this purpose, NKA-like immunoreactivities were characterized in the human retina by reversed phase HPLC followed by radioimmunoassay using the K12 antibody which recognizes both NKA and NKB. Furthermore, the retinae from both a 22- and 70-year-old donor were processed for double-immunofluorescence NKA/SP and NKB/SP. The results showed that NKA contributes to approximately two thirds and NKB to approximately one third of the immunoreactivities measured with the K12 antibody. NKA was found to be localized in sparse amacrine cells in the proximal inner nuclear layer, in displaced amacrine cells in the ganglion cell layer with processes ramifying in stratum 3 of the inner plexiform layer and also in sparse ganglion cells. By contrast, staining for NKB was only observed in ganglion cells and in the nerve fiber layer. Double-immunofluorescence revealed cellular colocalization of NKA with SP and also of NKB with SP. Thus, the levels of NKA and NKB are more than three and two times higher than those of SP, respectively. Whereas the distribution pattern of NKA is typical for neuropeptides, the localization of NKB exclusively in ganglion cells is atypical and unique.     

GPR54-Dependent Stimulation of Luteinizing Hormone Secretion by Neurokinin B in Prepubertal Rats

Kisspeptin, neurokinin B (NKB) and dynorphin A (Dyn) are coexpressed within KNDy neurons that project from the hypothalamic arcuate nucleus (ARC) to GnRH neurons and numerous other hypothalamic targets. Each of the KNDy neuropeptides has been implicated in regulating pulsatile GnRH/LH secretion. In isolation, kisspeptin is generally known to stimulate, and Dyn to inhibit LH secretion. However, the NKB analog, senktide, has variously been reported to inhibit, stimulate or have no effect on LH secretion. In prepubertal mice, rats and monkeys, senktide stimulates LH secretion. Furthermore, in the monkey this effect is dependent on kisspeptin signaling through its receptor, GPR54. The present study tested the hypotheses that the stimulatory effects of NKB on LH secretion in intact rats are mediated by kisspeptin/GPR54 signaling and are independent of a Dyn tone. To test this, ovarian-intact prepubertal rats were subjected to frequent automated blood sampling before and after intracerebroventricular injections of KNDy neuropeptide analogs. Senktide robustly induced single LH pulses, while neither the GPR54 antagonist, Kp-234, nor the Dyn agonist and antagonist ({"type":"entrez-nucleotide","attrs":{"text":"U50488","term_id":"1277101","term_text":"U50488"}}U50488 and nor-BNI, respectively) had an effect on basal LH levels. However, Kp-234 potently blocked the senktide-induced LH pulses. Modulation of the Dyn tone by {"type":"entrez-nucleotide","attrs":{"text":"U50488","term_id":"1277101","term_text":"U50488"}}U50488 or nor-BNI did not affect the senktide-induced LH pulses. These data demonstrate that the stimulatory effect of NKB on LH secretion in intact female rats is dependent upon kisspeptin/GPR54 signaling, but not on Dyn signaling.     

迷走神经切断和NK3受体拮抗剂干预在辣椒素诱导小鼠咳嗽中的作用

目的:观察右侧迷走神经切断及NK3受体拮抗剂对辣椒素诱导小鼠咳嗽的作用及其机制。方法:将48只小鼠随机分为4组,分别为:右侧迷走神经切断术组、右侧假手术组、SR 142801腹腔注射组和生理盐水对照组。辣椒素雾化制作小鼠咳嗽模型后,分别进行迷走神经切断术、假手术、SR142801腹腔注射及生理盐水腹腔注射,SR142801组及生理盐水对照组腹腔注射每日1次,连续7天。第8天计数各组所有小鼠咳嗽次数,检测所有小鼠肺组织中SP(substance P,P物质)、NKA(neurokinin A,神经肽A)、NKB(neurokinin B,神经肽B)表达水平。结果:右侧迷走神经切断组术后咳嗽次数[(6.92±1.78)次]较术前[(7.83±2.48)次]显著降低(P0.01),较假手术组[(7.58±2.43)次]降低(P0.05)。右侧迷走神经切断组术后SP、NKA、NKB水平较对照组显著降低(P0.05),SR 142810组干预后咳嗽次数[(8.67±1.37)次]较干预前[(8.33±2.15)次]无明显降低(P0.05)。SR 142801组腹腔注射后NKB[(8.32±0.86)pg/m L]较生理盐水对照组[(8.83±0.80)pg/m L]无明显降低(P0.05)。结论:迷走神经切断可以抑制辣椒素诱导的咳嗽,其机制主要与减少迷走神经相关神经肽SP、NKA、NKB的表达有关,而NK3受体拮抗剂SR142801对辣椒素诱导的咳嗽无明显抑制作用。     

Bimodal effects of chronically administered neurokinin B (NKB) on in vivo and in vitro cardiovascular responses in female rats

The in vivo cardiovascular effects of acutely administered neurokinin B (NKB) have been attributed both to direct effects on vascular tone and to indirect effects on central neuroendocrine control of the circulation. We proposed: 1) that a modest long-term increase in plasma NKB levels would decrease mean arterial pressure (MAP) due to attenuated peripheral vascular tone, and 2) that chronic high-dose NKB would increase MAP, due to increased sympathetic outflow which would override the peripheral vasodilation. We examined the in vivo and in vitro cardiovascular effects of chronic peripheral NKB. Low- (1.8 nmol/h) or high- (20 nmol/h) dose NKB was infused into conscious female rats bearing telemetric pressure transducers. MAP, heart rate (HR) and the pressor responses to I.V. phenylephrine (PE, 8 microg) and angiotensin II (Ang II, 150 ng) were measured. Concentration-response curves of small mesenteric arteries were constructed to PE using wire myography. Low-dose NKB reduced basal MAP (88+/-2 mm Hg to 83+/-2 mm Hg), did not affect resting HR, reduced the pressor responses to PE, and attenuated the maximal constriction of mesenteric arteries to PE and KCl. By contrast, high-dose NKB increased basal MAP (86+/-1 mm Hg to 89+/-1 mm Hg), increased HR (350+/-3 beats/min to 371+/-3 beats/min), increased the pressor responses to Ang II and, contrary to our hypothesis, increased the maximum contractile responses of mesenteric arteries to PE and KCl. The cardiovascular effects of NKB are thus dose-dependent: whereas chronic low-dose NKB directly modulates vascular tone to reduce blood pressure, chronic high-dose NKB induces an increase in blood pressure through both central (indirect) and peripheral (direct) pathways.     

Neurokinin B- and substance P-like immunoreactivity are co-localized in enteric nerves of rat ileum

The tachykinins (TKs) substance P (SP), neurokinin A (NKA), and neurokinin B (NKB) have conserved C-terminal sequences and mediate similar physiological responses by activating neurokinin receptors found on neural and smooth muscle cells. Many enteric nerves express preprotachykinin A (PPT A) mRNA and synthesize SP and NKA. However, it is unclear if NKB is synthesized in enteric neurons as many antibodies developed against NKB also recognize other TKs. Therefore, the cellular distribution of NKB-like-immunoreactivity (NKB-ir) in rat ileum was examined using selective antisera raised against either synthetic Cys10-NKB or peptide 2 (P2), a non-tachykinergic peptide sequence in NKB precursor protein. NKB-ir and P2-ir had a similar distribution in varicose nerve fibers in submucosal and myenteric ganglia and almost all ganglia contained immunoreactive nerves. Few submucosal or myenteric neuronal somata contained strong immunoreactivity. Preabsorption of NKB or P2 antisera with their respective cognate peptides, but not with other TK peptides, abolished specific immunostaining. Finally, co-localization of NKB-/P2-ir with SP-ir suggested that most NKB-/P2-ir nerve fibers contain SP-ir, but some SP-ir nerves do not contain detectable NKB-/P2-ir. These results indicate that PPT B products P2 and NKB are localized in a subpopulation of enteric nerves containing TKs encoded by PPT A. Stimulation of these nerves may release NKB to activate local neurokinin receptors.     

[3H]Neurokinin B and 125I-Bolton Hunter Eledoisin Label Identical Tachykinin Binding Sites in the Rat Brain

[3H]Neurokinin B ([3H]NKB) of high specific activity (75 Ci/mmol) was synthesized for study of its binding to crude synaptosomes from the rat cerebral cortex. The specific binding of [3H]NKB (75% of total binding) was temperature dependent, saturable, and reversible. Scatchard analyses and Hill plots showed the existence of a single population of noninteracting binding sites (KD = 4.3 nM; Bmax = 123 fmol/mg of protein). Competition studies indicated the following rank order of potencies among tachykinins: NKB greater than eledoisin (E) greater than kassinin greater than physalaemin greater than neurokinin A (NKA) greater than substance P (SP), a result suggesting that NKB might be the endogenous ligand for [3H]NKB binding sites. It is of interest that 127I-Bolton Hunter (BH) NKA (127I-BHNKA) was much more potent than NKA in inhibiting the specific binding of [3H]NKB, which raises certain questions concerning the use of 125I-BHNKA as a ligand for NKA binding sites in the brain. These results, as well as those obtained with different SP analogues, show a close similarity to those obtained previously with 125I-BHE binding to cortical synaptosomes. This suggested that the two ligands labeled identical binding sites. In addition, using either [3H]NKB or 125I-BHE as ligands, similar displacement curves were obtained with increasing concentrations of NKB and 127I-BHE. The similarity of the [3H]NKB and 125I-BHE binding sites was further confirmed by comparison of their localization on rat brain sections by autoradiography. The distribution of binding sites for [3H]NKB and 125I-BHE was identical throughout the brain, and the highest density of binding sites for the two ligands was found in layers IV and V of the cerebral cortex, the paraventricular nucleus of the hypothalamus (magnocellular part), and the ventral tegmental area.     

Spinal action of neurokinins in the rat: effects on mean arterial pressure, heart rate, and vascular permeability

In urethane-anaesthetized rats, the intrathecal administration of 6.5 nmol of substance P (SP), neurokinin A (NKA), or neurokinin B (NKB) at the T8-T10 level of the spinal cord enhances mean arterial pressure and heart rate. However, in the pentobarbital-anaesthetized rat, while NKB produces no effect on mean arterial pressure, NKA produces a biphasic change and SP, a depressor response. All three neurokinins elicit a tachycardia. The following rank order of potency SP greater than or equal to NKA greater than NKB is observed in relation to these cardiovascular responses when either one of the two anaesthetics is used. The low cardiovascular activity of NKB cannot be attributed to its hydrophobicity, as the water soluble analogue of NKB, [Arg0]NKB, elicits a response as weak as the native peptide. In pentobarbital-anaesthetized rats, the intrathecal administration of 6.5 nmol of SP, also enhances plasma protein extravasation in cutaneous tissues of the back, the hind paws, and the ears. In this response NKA and NKB are either inactive (skin of hind paws) or less potent than SP (ears and dorsal skin). These findings agree with the hypothesis that in the rat spinal cord, the neurokinin receptor producing changes in mean arterial pressure, heart rate, and vascular permeability is of the NK-1 subtype.     

Functional proteomics of neurokinin B in the placenta indicates a novel role in regulating cytotrophoblast antioxidant defences

Neurokinin B (NKB) has recently been demonstrated to be secreted from the placenta in abnormally high amounts in preeclampsia (PE) and to cause hypertension in rats, suggesting it may be a mediator of some pathophysiological features of PE. It is also known that NKB receptors exist in the placenta. To determine the effect of high levels of NKB on the placenta, we have performed proteomics on five separate preparations of cultured purified human term cytotrophoblast cells. The results showed a statistically significant decrease in 20 proteins, of which five were unknown proteins. Proteins important in antioxidant defenses that decreased were thioredoxin, cyclophilin A, cytokeratin 1, and peroxiredoxin 5. Two proteins that inhibit intravascular anticoagulation, cytokeratin 1 and annexin 11 were also decreased. Pathways involving pro-inflammatory cytokine activation of NF-kappa B are opposed by Raf kinase inhibitor protein, which was also decreased. Cofilin 1, a protein involved in defense against bacteria, was also decreased. Among other proteins that were suppressed by NKB were proteasome proteins, desmoplakin, and calgizzarin. Western blots confirmed the decrease in cytokeratin 1 and cyclophilin A protein after NKB exposure. In PE, there is reduced antioxidant activity and increased intravascular coagulation. The findings that high levels of NKB, similar to those observed in PE, can impair these two classes of activity support the hypothesis that high NKB levels may contribute to the pathogenesis of PE.     

Neurokinin B causes concentration-dependent relaxation of isolated human placental resistance vessels

Placental neurokinin B (NKB) was recently identified as the causative agent in preeclampsia, a condition characterized by increased maternal and feto-placental vascular resistance. We hypothesized that NKB should constrict placental resistance vessels. Placentas were obtained from normotensive pregnancies. Immediately after delivery, stem villous arteries (300 microm diameter, 1.2 mm long) were dissected from macroscopically normal tissue in cold HEPES-physiological salt solution (PSS), mounted on a wire myograph system, and bathed in HEPES-PSS at 37 degrees C. After determination of the passive-tension internal circumference characteristics, the arteries were set to 90% of the internal circumference they would have under a normal physiological transmural pressure. Cumulative concentration-response curves were constructed for NKB (1 x 10(-12) to 1 x 10(-5) mol/l). Since there was no constrictive response to NKB, cumulative constrictive concentration-response curves were constructed to the thromboxane A(2) mimetic U46619 (1 x 10(-9) to 1 x 10(-5) mol/l). The vessels were then pre-constricted to 80% of maximal response and exposed to cumulative concentrations of NKB (1 x 10(-12) to 1 x 10(-6) mol/l). NKB caused a concentration-dependent relaxation (Maximal response NKB, 51+/-5%, n=5; time control, 12+/-6%, n=4; P<0.05). Removal of the endothelium did not alter the vasodilatory response to NKB. We conclude that, contrary to our hypothesis, NKB causes an endothelium-independent relaxation of the placental resistance vessels. We propose that NKB plays a role in the maintenance of high placental blood flow in normal pregnancy.     

Tachykinin receptor subtype that mediates the increase in vascular permeability in guinea pig skin

To determine the tachykinin receptor subtype that mediates the increase in vascular permeability, we examined the rank order of potency of tachykinins for inducing plasma extravasation in guinea pig skin and the specificity of tachykinin-induced tachyphylaxis of the responses. Plasma extravasation of the skin induced by tachykinins was NK-1 (SP-P)-type response from the rank order of potency of mammalian and nonmammalian tachykinins. Tachyphylaxis of the vascular response was induced by intradermal preinjection of mammalian tachykinins and was tachykinin-specific. In substance P (SP) tachyphylaxis (where SP was preinjected), the response to SP, not to neurokinin A (NKA) or neurokinin B (NKB), was decreased. In NKA tachyphylaxis and NKB tachyphylaxis, the response to NKA, not to SP or NKB, and the response to NKB, not to SP or NKA, were decreased, respectively. Thus, we conclude that the apparent NK-1-type response is mediated through three mammalian tachykinin receptors, NK-1, NK-2, and NK-3, which are specifically stimulated by their preferred agonist, SP, NKA, and NKB, respectively.