In vitro release of substance P from spinal cord slices by capsaicin congeners

The in vitro release of immunoreactive substance P (I-SP) by capsaicin and three congeners was studied on slices of rat spinal cord upper dorsal horn. Capsaicin and its congeners were all able to stimulate I-SP release, indicating that they act on chemosensitive primary afferents terminating in this region. A positive correlation was found between the I-SP releasing and pain-producing potencies of these compounds. This is in agreement with the concept that primary afferents containing substance P (SP) are involved in the transmission of nociceptive information.     

NMDA receptor activation modulates evoked release of substance P from rat spinal cord

The possible modulation exerted by glutamate on substance P (SP) release from the rat spinal cord has been investigated. The N-methyl-D-aspartate (NMDA) receptor agonist, NMDA (1 μM), increased SP basal outflow by 46.5±10.9% (n=3, P<0.01) without changing the evoked release of the peptide. Conversely, NMDA antagonists but not 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) inhibited both electrically-evoked and capsaicin-induced release of SP. In particular, D-2-amino-5-phosphonopentanoate (D-AP5; 50 μM) inhibited electrically-evoked and capsaicin-induced release of SP by 93±2.4% and 93.2±3.8% (n=12, P<0.01), respectively. Functional pharmacological evidence is provided for glutamate exerting a positive feedback on SP release evoked by C fibre stimulation via NMDA receptor activation.     

Adenosine analogs do not inhibit the potassium-stimulated release of substance P from rat spinal cord slices

Summary Adenosine agonists produce antinociception when injected directly onto the spinal cord of rats and mice. One mechanism to account for this effect could be inhibition of neurotransmitter release from nociceptive sensory neurons. Consequently, we studied whether these agents could inhibit the potassium stimulated release of one such transmitter, substance P, from rat spinal cord slices. A 2 cm section of lumbar spinal cord was dissected from male Sprague-Dawley rats, chopped into 0.5 × 0.5 mm sections and perfused at 37°C with a modified Krebs bicarbonate buffer containing either 3.5 mM, 30 mM, or 50 mM KCl in the presence and absence of various adenosine analogs. Perfusates, collected every 2 min, were assayed for substance P by radioimmunoassay. Exposure of tissue to 50 mM KC1 produced an approximate three-fold increase in the release of substance P over basal release. This increase in release was calcium dependent. Perfusion of spinal cord tissues with either adenosine (10^–3 M), N⁶-cyclohexyladenosine (10^–5 M or 5 × 10^–5 M), 5-N-ethylcarboxamide adenosine (10^–5 M) or L-N⁶-phenylisopropyladenosine (10^–5 M) did not significantly alter basal or potassium-stimulated release of SP when compared to controls. In contrast to the adenosine agonists, exposure of the spinal cord tissue to 10^–5 M morphine significantly reduced the potassium-stimulated release of substance P. Pretreatment of the slices with 10^–5 M theophylline or 8-phenyltheophylline did not significantly attenuate the inhibition of substance P release produced by morphine. Theophylline alone (10^–5 M) had no significant effect on either basal or potassium-stimulated release of SP. These studies demonstrate that adenosine does not inhibit the release of SP from spinal cord slices and does not appear to mediate the morphine-induced inhibition of SP release. The results suggest that the mechanism of the antinociceptive effects of adenosine at the level of the spinal cord is not via inhibition of substance P release. Send offprint requests to M. R. Vasko at the above address     

Effect of the tachykinin NK1 receptor antagonists, RP 67580 and SR 140333, on electrically-evoked substance P release from rat spinal cord.

1. The effects of the non-peptide tachykinin NK1 receptor antagonists, RP 67580, SR 140333, CP-96,345 and CP-99,994 have been investigated on electrically-evoked release of substance P-like immunoreactivity (SP-LI) from rat spinal cord slices. 2. RP 67580 (10 nM) and SR 140333 (1 nM), perfused 5 min prior to and during 8 min stimulation of the dorsal roots (20 V, 0.5 ms, 1 Hz), significantly enhanced SP-LI release by 213 +/- 43 (n = 8) and 203 +/- 31 (n = 5) % of control evoked release (187 +/- 16% of basal outflow, n = 22) respectively. Neither compound modified basal outflow of SP-LI (15.3 +/- 2.5 fmol/8 ml, n = 10). 3. RP 67580 (10 nM) did not modify electrically-evoked release of calcitonin gene-related peptide-LI from rat spinal cord slices. 4. CP-96,345 (10 nM) and CP-99,994 (1 and 10 nM) did not alter electrically-evoked SP-LI release; however, they both inhibited release at 1 microM. Inhibition was also induced by 1 microM RP 67580 but not 1 microM SR 140333. 5. The effect of the NK1 receptor agonists, [Sar9Met (O2)11]SP and [Sar9]SP, could not be tested on SP-LI release due to interference with the substance P radioimmunoassay (RIA). The other NK1 receptor agonists used, GR 73632, [Pro9]SP and septide, which did not interfere with the RIA, increased SP-LI basal outflow by 1807 +/- 713% (n = 3), 1259 +/- 160% (n = 3) and 620 +/- 69% (n = 3) at 10 nM, 1 nM and 1 microM, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)     

The effects of substance P and related peptides on alpha-amylase release from rat parotid gland slices.

1 The effects of substance P and related peptides on amylase release from rat parotid gland slices have been investigated. 2 Supramaximal concentrations (1 microM) of substance P caused enhancement of amylase release over the basal level within 1 min; this lasted for at least 40 min at 30 degrees C. 3 Substance P-stimulated amylase release was partially dependent on extracellular calcium and could be inhibited by 50% upon removal of extracellular calcium. 4 Substance P stimulated amylase release in a dose-dependent manner with an ED50 of 18 nM. 5 All C-terminal fragments of substance P were less potent than substance P in stimulating amylase release. The C-terminal hexapeptide of substance P was the minimum structure for potent activity in this system, having 1/3 to 1/8 the potency of substance P. There was a dramatic drop in potency for the C-terminal pentapeptide of substance P or substance P free acid. Physalaemin was more potent than substance P (ED50 = 7 nM), eledoisin was about equipotent with substance P (ED50 = 17 nM), and kassinin less potent that substance P (ED50 = 150 nM). 6 The structure-activity profile observed is very similar to that for stimulation of salivation in vivo, indicating that the same receptors are involved in mediating these responses. 7 All the fragments of substance P tested were capable of eliciting a full amylase release response. This indicates that the apparent partial agonist action of the C-terminal nonapeptide fragment on in vivo salivation is not explicable at the receptor level.     

Differential effects of wortmannin on the release of substance P and amino acids from the isolated spinal cord of the neonatal rat

1. Effects of wortmannin, an inhibitor of myosin light chain kinase, on the release of substance P and amino acids, GABA and glutamate, were investigated in the isolated spinal cord preparation of the neonatal rat.
2. Wortmannin at 0.5–10 μM depressed the release of substance P evoked by high-K⁺ (90 mM) medium from the spinal cord (IC₅₀=1.1 μM). Wortmannin also depressed the high-K⁺ (70 mM)-evoked release of substance P from cultured dorsal root ganglion neurons of neonatal rats. In contrast, the high-K⁺ (90 mM)-evoked release of GABA and glutamate from the spinal cord was not affected by wortmannin (0.1–10 μM).
3. Upon stimulation of a dorsal root, a monosynaptic reflex and a subsequent slow ventral root depolarization were evoked in the ipsilateral ventral root of the same segment in the isolated spinal cord preparation. The magnitude of the slow ventral root depolarization was depressed gradually to about 70% of the control during the course of 30 min under wortmannin (1 μM). In contrast, the monosynaptic reflex was unaffected by wortmannin.
4. Immunofluorescent staining revealed that immunoreactivities of substance P and myosin II were colocalized at presynaptic terminals in the dorsal horn of the neonatal rat spinal cord.
5. The present results suggest that myosin phosphorylation by myosin light chain kinase may play a crucial role in the release of substance P, but not in the release of GABA and glutamate in the neonatal rat spinal cord. This may reflect a difference in the exocytic mechanisms of substance P-containing large dense core vesicles and amino acid-containing small clear vesicles.

     

Epileptiform activity induced by magnesium-free solution in slices of rat amygdala: antagonism by N-methyl-D-aspartate receptor antagonists

The effects of the N-methyl-D-aspartate (NMDA) receptor antagonists, D-2-amino-5-phosphonovalerate (D-APV) and phencyclidine (PCP), were studied in vitro on epileptiform activity induced in magnesium-free solution in neurons of the basolateral amygdala of the rat, using intracellular recording techniques. Twenty to 30 min after switching to magnesium-free medium, spontaneous interictal-like events were observed in 33 out of 37 amygdala slices. The spontaneous interictal-like events consisted of an initial burst followed by a number of afterdischarges. Superfusion with D-APV, a competitive NMDA receptor antagonist, reversibly reduced the duration of the events was also reduced. The IC50, estimated from the graph of the concentration-response relationship, was approximately 10 microM which is close to the IC50 for the binding of D-AVP to the NMDA receptor in other regions of the brain. The effect of phencyclidine, a noncompetitive NMDA receptor antagonist, was similar to that of D-APV. These results suggest that activation of NMDA receptors plays an intrinsic role in the induction or propagation of epileptiform activity seen in magnesium-free solution in the neurons of the amygdala.     

Bombesin receptor antagonists. 2. Analogues based on substance P antagonists.

Although bombesin (BN) and substance P share only the C-terminal dipeptide amide, some substance P receptor antagonists are also weak bombesin receptor antagonists. In order to increase the selectivity of the antagonism for the BN receptor, a series of hybrid peptides were synthesized by the solid-phase methodology, and screened on 3T3 fibroblasts for binding and mitogenic activity. The analogues inhibiting BN-induced thymidine incorporation were further tested for peripheral (amylase release and urinary bladder contraction) and central activity (grooming behaviour).     

In vitro prostanoid release from spinal cord following peripheral inflammation: effects of substance P, NMDA and capsaicin

1. Spinal prostanoids are implicated in the development of thermal hyperalgesia after peripheral injury, but the specific prostanoid species that are involved are presently unknown. The current study used an in vitro spinal superfusion model to investigate the effect of substance P (SP), N-methyl-d-aspartate (NMDA), and capsaicin on multiple prostanoid release from dorsal spinal cord of naive rats as well as rats that underwent peripheral injury and inflammation (knee joint kaolin/carrageenan). 2. In naive rat spinal cords, PGE2 and 6-keto-PGF1alpha, but not TxB2, levels were increased after inclusion of SP, NMDA, or capsaicin in the perfusion medium. 3. Basal PGE2 levels from spinal cords of animals that underwent 5-72 h of peripheral inflammation were elevated relative to age-matched naive cohorts. The time course of this increase in basal PGE2 levels coincided with peripheral inflammation, as assessed by knee joint circumference. Basal 6-keto-PGF1alpha levels were not elevated after injury. 4. From this inflammation-evoked increase in basal PGE2 levels, SP and capsaicin significantly increased spinal PGE2 release in a dose-dependent fashion. Capsaicin-evoked increases were blocked dose-dependently by inclusion of S(+) ibuprofen in the capsaicin-containing perfusate. 5. These data suggest a role for spinal PGE2 and NK-1 receptor activation in the development of hyperalgesia after injury and demonstrate that this relationship is upregulated in response to peripheral tissue injury and inflammation.     

Clonidine reduces substance P binding in rat spinal cord membrane preparation.

The effects of clonidine on substance P (SP) binding was investigated using rat brain and spinal cord membrane preparations preincubated with various concentrations of clonidine. [3H]SP specific binding in the spinal cord was significantly decreased with 10(-4) M clonidine, but no effect on binding was seen in the brain. Scatchard analysis of SP binding indicated that Bmax was significantly depressed without changing the affinity. The mechanism of clonidine-induced analgesia includes a spinal neural component and action on the SP receptors.     

The effects of cholinoceptor agonists and antagonists on C-fibre evoked responses in the substantia gelatinosa of neonatal rat spinal cord slices.

1. The effects of cholinoceptor agonists and antagonists were studied on neurones in the substantia gelatinosa (SG) of an in vitro spinal cord slice and nerve preparation from neonatal rats. 2. Bath application of carbachol (1-50 microM) reduced, in a dose-related manner, the amplitude and duration of the excitatory postsynaptic potentials (e.p.s.ps) evoked in response to nerve stimulation. 3. The latencies and stimulation thresholds required to evoke these e.p.s.ps suggested that the majority were due to C-fibre activation. 4. The reduction in e.p.s.p. amplitude and duration produced by carbachol was reversed by the muscarinic antagonists, atropine (in 8 out of 11 cells), pirenzepine (in 7 out of 9 cells) and methoctramine (in 8 out of 9 cells) and by the nicotinic antagonist mecamylamine (in 3 out of 7 cells). 5. Injection of small hyperpolarizing or depolarizing pulses was associated with no change in conductance in 19 out of 26 (73%) of cells tested, suggesting that an action at a site presynaptic to the neurone studied could account for part of the effect of carbachol. 6. It is proposed that some of the cholinoceptors associated with the e.p.s.p. depression are located on C-fibres.     

NK3 receptor blockade prevents hyperalgesia and the associated spinal cord substance P release in monoarthritic rats

Previous studies in vitro have shown that NK3 receptors exist on primary afferent terminals in rat spinal cord and mediate potentiation of the depolarisation-evoked substance P (SP) release. In the present study we have investigated the role of the NK3 receptor-mediated SP release system in a model of inflammatory pain. Monoarthritis was induced in rats by unilateral injection of complete Freund's adjuvant (CFA); withdrawal latencies to a thermal stimulus were subsequently measured at various times following CFA. The CFA-treated paw displayed hyperalgesia as early as 4 h after CFA injection and hyperalgesia was maintained until day 4 but had disappeared by day 21. The thermal hyperalgesia was associated with an increase in basal SP release from spinal cord synaptosomes. The possible involvement of endogenous neurokinin B acting at NK3 receptors was tested by using SB 223412-A [(S)-(-)-N-(alpha-ethylbenzyl)-3-hydroxy-2-phenylquinoline-4-carbo xamide hydrochloride], a novel, potent (Ki=30 nM) and selective (Ki>10,000 nM for NK1 and NK2 receptors), non-peptidic NK3 receptor antagonist. In vitro SB 223412-A antagonised the potentiation of SP release produced by senktide in spinal cord synaptosomes. Administered systemically to monoarthritic rats (50 mg/kg, p.o., b.i.d., for 4 days), the NK3 receptor antagonist SB 223412-A significantly reduced thermal hyperalgesia and normalised the basal release of SP from spinal cord synaptosomes. The data suggest that neurokinin B acting at NK3 receptors that mediate SP release within the spinal cord play a role in inflammation. These NK3 receptors may represent, therefore, appropriate targets in the therapy of inflammatory pain.     

Lack of effect of baclofen on substance P and somatostatin release from the spinal cord in vitro

Summary High concentrations of K⁺ increase the release of substance P (SP) and somatostatin (SRIF) from superfused slices of rat spinal cord. This increase is Ca-dependent. Baclofen (100–500 M) does not significantly alter the K⁺-evoked release of SP or SRIF. Stereoisomers of baclofen and GABA, similarly, are without effect. The spinal analgesic action of baclofen does not appear to be due to alterations in the release of SP or SRIF.     

Serotonin type-2 receptor mediated regulation of substance P release in the ventral spinal cord and the effects of chronic antidepressant treatment

Summary Regulation of the release of substance P (SP) by the coexisting neurotransmitter serotonin (5-hydroxytryptamine, 5-HT) in the ventral spinal cord and the effects of chronic antidepressant treatment mediated changes in serotonin metabolism on the regulation, were examined.The K⁺ (40 mmol/l) evoked release of (SP) from slices of the ventral spinal cord of the rat was potentiated by (5-HT) applied to 100 mol/l concentration. This effect was blocked by the serotoninergic antagonists methysergide (10 mol/l), methiotepin (10 mol/l) and fully blocked by ketanserin (10 mol/l). Thus the 5-HT receptor which regulates the release of SP appears to belong to the type-2 5-HT receptors. Chronic treatment with the selective serotonin uptake inhibitor zimelidine (14 days, 2×10 mol/kg/day, p.o.) lowered the tissue levels of the 5-HT metabolite: 5-hydroxyindol acetic acid (5-HIAA) and elevated the tissue levels of SP in both the ventral and dorsal spinal cord as compared to that in the vehicle treated group (14 days, 2×5 ml saline/kg/day, p.o.). The decrease in the 5-HIAA levels after chronic zimelidine treatment was quantitatively similar in the dorsal (33%,p<0.01) and ventral (31%,p<0.05) spinal cord. The increase in SP levels after chronic zimelidine treatment was more pronounced in the ventral cord (80%,p<0.01) where the majority of the SP containing nerve endings also contain 5-HT, than in the dorsal spinal cord (22% increase in SP,p<0.05), where only a minor fraction of the SP-containing nerve endings shows a 5-HT/SP coexistence. Chronic treatment with imipramine (14 days, 2×10 mol/kg/day, p.o.) gave qualitatively similar results to those obtained by zimelidine treatment, but increases in SP levels, and decreases in 5-HT and 5-HIAA levels in ventral and dorsal spinal cord, were less pronounced. The K⁺ (40 mmol/l) evoked release of SP was studied in a slice preparation of the ventral spinal cord, from rats chronically treated with imipramine, zimelidine and saline. In the zimelidine treated group the amount of SP released (pmol/g tissue) and the fractional SP release upon K⁺ (40 mmol/l) stimulation was increased by 53% (p<0.01) and 42% (p<0.01) respectively, when compared to the control group. No significant changes in the amount of SP released or fractional SP release were observed when tissue preparations from rats treated with imipramine were used. Thus, it seems that treatment with specific serotoninergic or monoaminergic antidepressant drugs does not only change the tissue levels of the monoamine and its metabolite but also affects the coexisting peptidergic transmitter; SP in the ventral spinal cord. This change is also reflected in the size of the releaseable pool of SP.     

Baclofen and phaclofen modulate GABA release from slices of rat cerebral cortex and spinal cord but not from retina.

1. The effects of (-)-baclofen, muscimol and phaclofen on endogenous gamma-aminobutyric acid (GABA) release from rat cortical slices, spinal cord slices and entire retinas were studied. 2. The spontaneous resting release of GABA from the three tissues was 3 to 6 pmol mg-1 wet wt 10 min-1. Depolarization of cortical slices with KCl (50 mM) (high-K) produced an 8 fold increase in GABA release but high-K did not evoke an increased release of GABA from spinal slices or retinas. 3. When rats were injected with gamma-vinyl-GABA (250 mg kg-1 i.p.) (GVG) 18 h before death, the tissue GABA stores were increased 3 to 6 fold and high-K then evoked striking Ca-dependent releases of GABA from all three tissues. Thus, in subsequent experiments, unless otherwise stated, the nervous tissues were taken from GVG-treated rats. 4. (-)-Baclofen (10 microM) significantly reduced the K-evoked release of GABA from cortical and spinal slices but retinal release was not affected, even at a concentration of (+/-)-baclofen of 1 mM. For cortical slices, the IC50 for baclofen was approximately 5.2 microM. The inhibitory effect of baclofen on GABA release from cortical slices also occurred in slices prepared from saline-injected rats, indicating that GVG treatment did not qualitatively affect the results. 5. The inhibitory effect of (-)-baclofen on the K-evoked release of GABA from cortical and spinal slices was antagonised by phaclofen (500 microM), confirming that baclofen was producing its effects by acting at the GABAB-receptor.(ABSTRACT TRUNCATED AT 250 WORDS)     

NK-3 receptors mediate enhancement of substance P release from capsaicin-sensitive spinal cord afferent terminals

1. The effects of NK-3 receptor agonists on the release of substance P-immunoreactivity (SP-LI) have been investigated using superfused rat spinal cord synaptosomes.
2. The Ca²⁺-dependent overflow of SP-LI evoked by 35 mM KCl was concentration-dependently enhanced by senktide (EC₅₀=52 nM; maximal effect=70%) or [MePhe⁷]NKB (EC₅₀=5.5 nM; maximal effect=125%), both selective agonists at receptors of the NK-3 type.
3. The potentiation of the SP-LI overflow elicited by 100 nM senktide or [MePhe⁷]NKB was prevented by the NK-3 receptor antagonist (+)-SR142801. The antagonist halved, at 10 nM, and almost abolished, at 100 nM, the effect of both agonists. The effect of senktide or [MePhe⁷]NKB was insensitive to antagonists at NK-1 or NK-2 receptors.
4. Capsaicin (0.1–1 μM) stimulated SP-LI release in a concentration-dependent manner from spinal cord synaptosomes. The SP-LI overflow elicited by 1 μM capsaicin was completely dependent on external Ca²⁺. Senktide could not affect the capsaicin-evoked release of SP-LI.
5. Senktide failed to potentiate the K⁺-evoked overflow of SP-LI from synaptosomes previously exposed for 15 min in superfusion to capsaicin.
6. The results show that release-enhancing NK-3 receptors are located on axon terminals of capsaicin-sensitive primary afferent neurones in the spinal cord. Antagonists at NK-3 receptors might help controlling pain transmission.

     

Regulation of cardiovascular sympathetic neurons by substance P and gamma-aminobutyric acid in the rat spinal cord.

The spinal regulation of cardiovascular sympathetic preganglionic neurons by substance P (SP) and gamma-aminobutyric acid (GABA) was investigated in conscious rats. Intrathecal injection at the T-9 spinal level of bicuculline, a GABAA receptor antagonist, evoked increases in mean arterial pressure (MAP) and heart rate (HR) which were maximal at 5.0 and 0.5 nmol, respectively. Phaclofen, a GABAB receptor antagonist, produced no cardiovascular changes up to 2 mumol while 10 mumol evoked a rise in MAP and HR. Muscimol, a GABAA receptor agonist, produced a decrease in MAP which was maximal at 5.0 nmol and had no effect on HR. Baclofen, a GABAB receptor agonist, was without cardiovascular effects up to 5.0 nmol, while 50 and 100 nmol evoked a fall in MAP and HR. The pressor response to SP (16.25 nmol, T-9) was antagonised by 0.5-50 nmol muscimol or baclofen in a dose-related manner and the pressor response to SP was still inhibited by 40 nmol GABA in capsaicin-treated animals. However, when SP was injected at T-2, the rise in both MAP and HR was blocked by 50 nmol baclofen. Similarly, 50 nmol muscimol blocked the rise in both MAP and HR induced by 15 nmol thyrotropin-releasing hormone. In contrast, 50 nmol glycine failed to alter the cardiovascular response to SP co-injected either at T-9 or T-2. Baclofen was found to reduce significantly the basal release of epinephrine when injected at the T-9 level. These results provide pharmacological evidence for a possible tonic GABAergic inhibitory input onto cardiovascular sympathetic preganglionic neurons mediated by GABAA and GABAB receptors.     

The serotonin receptor agonist 5-methoxy-N,N-dimethyltryptamine facilitates noradrenaline release from rat spinal cord slices and inhibits monoamine oxidase activity

1. The influences of the purported serotonergic agonist 5-methoxy-N,N-dimethyltryptamine (MeODMT) on noradrenaline release and metabolism were investigated in a rat spinal cord release model and a monoamine oxidase (MAO) assay.2. MeODMT inhibited the basal outflow of tritium from rat spinal cord slices preincubated with [³H]noradrenaline and enhanced the electrically-evoked overflow.3. Effects on basal outflow were not observed, when monoamine oxidase (MAO) was inhibited by pargyline. Effects on the evoked overflow were not observed in the presence of metitepine or phentolamine.4. Preferential inhibition by MeODMT of MAO A-type enzyme activity was found in a direct assay.5. The results provide evidence for two different effects by which MeODMT reinforces noradrenergic neurotransmission in the rat spinal cord: facilitation of stimulation-evoked noradrenaline release and inhibition of noradrenaline metabolism by MAO inhibition.     

Substance P enhances the release of endogenous serotonin from rat ventral spinal cord

The effect of the tachykinin neuropeptides, substance P (SP), neurokinin A (NKA) and the neurokinin B (NKB) receptor agonist, senktide, on the potassium-evoked release of endogenous serotonin (5-hydroxytryptamine, 5-HT) was investigated in superfused tissue slices of rat ventral spinal cord, where 5-HT is known to coexist with SP. Endogenous 5-HT was assayed by HPLC with electrochemical detection. The evoked release of 5-HT was significantly enhanced by 10(-4) M SP (190% increase) and 10(-5) M SP (74% increase) but not by 10(-6) M SP, NKA (10(-5) and 10(-4) M) and senktide (10(-5) and 10(-4) M) had no significant effect on the 5-HT release. The results suggest that, in the rat ventral spinal cord, where most of the 5-HT and SP is stored in the same nerve endings. SP but not NKA nor NKB potentiates the evoked release of 5-HT in a dose-dependent manner.