Opioid inhibition of reflex urinary bladder contractions: dissociation of supraspinal and spinal mechanisms

The supraspinal and spinal mechanisms of morphine induced inhibition of reflex urinary bladder contractions were studied in the urethane anesthetized rat. The inhibition of bladder contractions by intracerebroventricular (i.c.v.) morphine was abolished by intrathecal (i.t.) naloxone. In addition systemic reserpine (3.5–5.0 mg/kg, i.p.) abolished the inhibitory effect of both systemic morphine and i.c.v. morphine but not that of i.t. morphine. These data support the involvement of separate supraspinal and spinal mechanisms in the effects of morphine on bladder motility. The supraspinal effect appeared to be mediated indirectly via a monoaminergic system whereas the spinal action was mediated independently of monoamines. However, both the indirect supraspinal and the direct spinal mechanism involved a spinal naloxone-sensitive process possibly an endogenous enkephalinergic system.     

Effects of intrathecal nocistatin on the flexor reflex and its interaction with orphanin FQ nociceptin

We studied the effects of intrathecal (i.t.) nocistatin, a peptide identified from the precursor of orphanin FQ/nociceptin (OFQ) on the spinal nociceptive flexor reflex in decerebrate, spinalized, unanesthetized rats and its interaction with i.t. OFQ. Nocistatin induced a moderate, non-dose-dependent facilitation of the flexor reflex without producing reflex depression whereas i.t. OFQ induced a biphasic dose-dependent facilitatory and inhibitory effect. The facilitatory effect of low dose (0.55 pmol) OFQ was significantly increased by nocistatin. On the other hand, the duration, but not magnitude, of reflex depression induced by a high (550 pmol) dose of OFQ was significantly shortened by 5.5 nmol nocistatin. Thus, nocistatin interacts with OFQ in a complex fashion, increasing excitation and reducing inhibition. No evidence was obtained for an antinociceptive effect of nocistatin in rat spinal cord.     

Tonic inhibitory influence of a supraspinal monoaminergic system on recurrent inhibition of an extensor monosynaptic reflex.

Recurrent inhibition of the extensor (quadriceps) monosynaptic reflex (MSR) was antagonized by a 5-hydroxytryptamine (5-HT) precursor, 5-hydroxytryptophan (5-HTP, 75 mg/kg), and a specific 5-HT neuronal uptake blocker, fluoxetine-HC1 (Lilly 110140, 0.25-6 mg/kg), in unanaesthetized decerebrate cats. This inhibition of the flexor (posterior biceps-semitendinosus) MSR was not altered by fluoxetine. Cyproheptadine-HC1 (5 mg/kg) partially reversed the above blocking actions of 5-HTP and fluoxetine and a thoracic "cold block", which eliminates supraspinal inputs to the caudal spinal cord, also eliminated the blockade by fluoxetine on recurrent inhibition. Cyproheptadine (2.5-5 mg/kg) or phenoxybenzamine-HC1 (2.5-5 mg/kg), administered alone, enhanced recurrent inhibition of the extensor but not of the flexor MSR. Since a "cold block" increased recurrent inhibition of the extensor reflex in control animals but failed to alter the inhibition in animals pretreated with either DL-p-chlorophenylalanine (300 mg/kg i.p. for 2 consecutive days) or DL-a-methyl-p-tyrosine methyl ester-HC1 (125 mg/kg i.p. 16 and 4 h prior to experiment), the monoaminergic system would appear to be tonically active. In addition the neuronal uptake blocker, imipramine-HC1 (0.125-4 mg/kg), was more potent in antagonizing recurrent inhibition when injected intra-arterially to the spinal cord than when administered intra-arterially to the brain stem or intravenously, indicating that this agent acts in the spinal cord to block the inhibition. These results support our previous proposal (ref. 18) that a supraspinal system involving 5-HT and noradrenaline antagonizes recurrent inhibition of the quadriceps MSR. This monoaminergic system is tonically active with the 5-HT nerve terminals located in the spinal cord.     

The specific antagonistic effect of intrathecal spantide II on substance P- and C-fiber conditioning stimulation-induced facilitation of the nociceptive flexor reflex in rat.

The effect of intrathecally (i.t.) applied tachykinin antagonist D-NicLys1, 3-Pal3, D-Cl2Phe5, Asn6, D-Trp7.9, Nle11-substance P (SP), spantide II, on the nociceptive flexor reflex was studied in decerebrate, spinalized, unanaesthetized rats over the dose range of 10 ng-10 micrograms. I.t. spantide II usually caused weak facilitation of the flexor reflex, especially at lower doses (10-100 ng) and at higher doses (1-10 micrograms) it sometimes depressed the reflex. Pre-treatment with spantide II (1, 3 or 10 micrograms) effectively antagonized the facilitatory effect of 10 ng i.t. SP on the flexor reflex for about 30 min. The facilitation of the reflex induced by i.t. administration of other neuropeptides present in primary afferents, somatostatin (SOM), vasoactive intestinal peptide (VIP), calcitonin gene-related peptide (CGRP) and galanin (GAL), was not influenced by spantide II. This non-toxic antagonist also effectively blocked facilitation of the flexor reflex induced by C-fiber conditioning stimulation of the sural nerve. The present results indicate that spantide II is an effective and specific tachykinin antagonist in the spinal cord. Furthermore, C-fiber stimulation facilitates the nociceptive flexor reflex through a mechanism involving the release of SP from the central terminals of primary afferents.     

Differential modulation of withdrawal reflexes by a cannabinoid in the rabbit

Inhibition of spinal and trigeminal withdrawal reflexes by morphine and by the cannabinoid agonist HU 210 has been studied in anaesthetized and in decerebrated rabbits. In intact, pentobarbitone-anaesthetized animals, the jaw-depressor reflex (JDR) evoked by stimulation of the tongue, and the reflex elicited in the ankle flexor tibialis anterior (TA) by stimulation of the toes were inhibited to the same extent by morphine (1-30 mg kg(-1) i.v. cumulative). In spinalized, anaesthetized rabbits morphine depressed the JDR to the same level as in non-spinal preparations, but the effect of the opioid on the TA reflex was significantly reduced. All effects of morphine were reversed by naloxone (0.25 mg kg(-1), i.v.). In anaesthetised intact animals, HU 210 depressed the JDR at a dose of 100 nmol kg(-1) i.v. cumulative, reduced reflexes evoked in the knee flexor muscle semitendinosus (ST) by stimulation at the toes at a dose of 30 nmol kg(-1) i.v. cumulative, but had no consistent or significant effects on the TA reflex to toe stimulation. The same results were obtained in spinalized, anaesthetised animals. In decerebrated, spinalized rabbits with no anaesthesia, HU 210 (30 nmol kg(-1)) depressed both ST and TA reflexes evoked by toe stimulation. These data reveal that trigeminal and spinal withdrawal reflexes are equally sensitive to morphine provided the spinal cord is intact, suggesting that at least part of the action of systemic morphine is due to activation of descending inhibition. The present results also show for the first time that cannabinoid agonists can inhibit trigeminal withdrawal reflexes. HU 210 had differential effects on the three reflexes studied depending on the presence or absence of anaesthesia. This is the first occasion on which we have found pharmacological distinctions between withdrawal reflexes, and indicates that spinal sensorimotor processing is more heterogeneous than has been suspected previously.     

Phaclofen antagonizes the depressant effect of baclofen on spinal reflex transmission in rats

The action of phaclofen, the phosphonic acid derivative of baclofen, on baclofen-induced suppression of spinal reflex transmission was tested in anaesthetized rats. Intrathecal (i.th.) injection of phaclofen, 100 nmol, antagonized the depressant effect of baclofen, 2 nmol, on spinal Hoffmann (H)-reflexes and polysynaptic flexor reflexes but ha on the action of muscimol, 20 nmol. The antagonistic effect of phaclofen on baclofen-induced depression of H-reflexes was dose-dependent in doses ranging from 1 to 100 nmol. When administered alone, phaclofen, 100 nmol, was devoid of stimulatory or depressant effects on spinal reflexes. These results indicate that phaclofen specifically antagonizes the reflex suppressant action of baclofen. The lack of intrinsic action of phaclofen suggests that there is no endogenous tonic inhibition mediated by GABAB receptors under the present experimental conditions.     

The effect of systemic cocaine on spinal nociceptive reflex activity in the rat

In the anesthetized rat, cocaine (25 mg/kg i.p.), enhanced the frequency potentiation of nociceptively evoked polysynaptic discharges but did not affect the polysynaptic reflex discharge to single nociceptive stimuli or the habituation of this reflex to repetitive pinch stimuli. The non-nociceptive, short-latency reflex discharge was suppressed for 10-15 min after cocaine administration. The neurogenic extravasation response to antidromic cutaneous C-fiber stimulation was unaffected by cocaine. These findings suggest that systemic cocaine, in doses analgesic for the rat, does not suppress spinal nociceptive reflexes.     

Suppression of the micturition reflex in urethane-anesthetized rats by intracerebroventricular injection of WAY100635, a 5-HT(1A) receptor antagonist

The influence of supraspinal 5-HT_1A receptors on reflex bladder activity was evaluated in anesthetized rats by studying the effects of intracerebroventricular (i.c.v.) administration of WAY100635 (1–100 μg), a selective 5-HT_1A receptor antagonist. The drug dose-dependently decreased the frequency and/or amplitude of isovolumetric reflex bladder contractions. Low doses (1–10 μg) increased the interval between contractions but only slightly reduced the amplitude of the contractions. However, 100 μg of WAY100635 elicited an initial complete block of bladder reflexes followed by a recovery period lasting 10–15 min during which the frequency of reflex contractions was normal but the amplitude was markedly suppressed by 70–80%. Mesulergine (0.1 mg/kg, i.v.), a 5-HT_2C antagonist, which transiently eliminated bladder activity in some rats (five of 11), blocked the inhibitory effect of WAY100635 (10 or 100 μg, i.c.v.) in only two of six rats. Our data coupled with the results of previous studies suggest that spinal and supraspinal 5-HT_1A receptors are involved in multiple inhibitory mechanisms controlling the spinobulbospinal micturition reflex pathway. The regulation of the frequency of bladder reflexes is presumably mediated by a suppression of afferent input to the micturition switching circuitry in the pons, whereas the regulation of bladder contraction amplitude may be related to an inhibition of the output from the pons to the parasympathetic nucleus in the spinal cord.     

The response of the 5-hydroxyindole oxidation current to noxious stimuli in the spinal cord of anesthetized rats: modification by morphine.

The effects of cutaneous noxious heating and of systemic morphine on serotonergic activity in the spinal cord were examined in anesthetized rats. An oxidation current of 5-hydroxyindole signal was seen at 280-300 mV with differential normal pulse voltammetry. Noxious heat stimuli produced a mean signal increase over control values of 15.5 +/- 3.4% at 52 degrees C, and 7.2 +/- 5.5% at 45 degrees C. These increases lasted for 5-10 min. Non-noxious stimuli (37 degrees C) did not affect the 5-hydroxyindole signal. Morphine (0.5, 2.0 and 5.0 mg/kg, i.p.) in the absence of cutaneous stimulation did not change the signal significantly. Systemic morphine alone did not significantly modify the 5-hydroxytryptamine (5-HT) metabolism, as observed in in vivo voltammetry, in the spinal cord of anesthetized rat. However, a low dose of morphine (0.5 mg/kg, i.p.) attenuated the increase in the signal modified by noxious stimuli, and high doses (2.0 or 5.0 mg/kg, i.p.) enhanced it. Both effects of morphine were antagonized by naloxone (0.5 mg/kg, i.v.). It is likely that morphine with noxious stimuli modify the sensitivity of serotonergic descending inhibitory system. It is concluded that noxious heating of the skin increases the 5-HT metabolism in the spinal cord of anesthetized rats and that systemic administration of morphine modulates this 5-HT metabolism.     

Spinal mechanisms underlying A-85380-induced effects on acute thermal pain

Systemic administration of nicotinic receptor (nAChR) agonists is antinociceptive in models of acute pain whereas their intrathecal (i.t.) administration has been reported to be antinociceptive, nociceptive or without effect. It has been hypothesized that the action induced is dependent upon the subtype and location of the nAChR activated. In addition, there is considerable evidence that nAChR ligand-induced antinociception is mediated by other neurotransmitter systems via descending pathways from the brainstem to the spinal cord. The present study investigated the effects of i.t. and systemic administration of A-85380, a novel nAChR agonist, in the paw withdrawal model of acute thermal pain in the rat. Given i.t., A-85380 (1 and 10 nmol/rat) decreased the latency to paw withdrawal by 2–4 s. This pronociception was accompanied by a spontaneous flinching behavior. Both of these effects were differentially blocked by i.t. pretreatment with the nAChR antagonists mecamylamine (10 nmol)>MLA (100 nmol)>DHβE (50% with 1000 nmol) but not by α-bungarotoxin (0% at 0.63 nmol). Given systemically, A-85380 (0.56 μmol/kg, i.p.) induced antinociception as indicated by an increased latency to paw withdrawal, an effect differentially altered by i.t. pretreatment with monoaminergic antagonists (100 nmol/rat). While mecamylamine and prazosin had no effect, scopolamine, methysergide and MDL 72222 partially antagonized and idazoxan completely antagonized A-85380-induced antinociception. Finally, as measured by in vivo microdialysis, levels of 5-HT, but not NE, in the i.t. space of the lumber region of the spinal cord were significantly increased following the systemic administration of A-85380. Together these data suggest that the nociceptive properties of spinally administered nAChR agents are not mediated by either an α₄β₂ or an α₇ subtype nAChR, whereas the antinociceptive properties of systemically-administered nAChR agents are mediated by descending noradrenergic, serotonergic and muscarinic inhibitory pathways.     

Effect of the synthetic polyamine N,N'-bis-(3-aminopropyl) cyclohexane-1,4-diamine (DCD) on rat spinal cord nociceptive transmission

In rats submitted to a C-fiber reflex response paradigm, intravenous (i.v.) administration of 2.5, 5 and 10 mg/kg of the synthetic polyamine N,N'-bis-(3-aminopropyl) cyclohexane-1,4-diamine (DCD) dose-dependently reduced both the integrated C reflex responses and wind-up activity. Inhibitory effects of the polyamine on spinal cord nociceptive transmission are likely to be consequence of blockade by extracellular DCD of NMDA receptor channels localized in dorsal horn neurons, although modulatory actions at supraspinal level and at other ion channels could also be possible.     

The effects of intrathecal galanin and C-fiber stimulation on the flexor reflex in the rat

Galanin (GAL) was applied intrathecally (i.t.) at the lumbar level in decerebrate, spinalized, unanesthetized rats. GAL had no effect on the amplitude of the monosynaptic reflex over a wide concentration range, but at low concentrations if briefly facilitated the flexor reflex and at higher concentrations the facilitation was sometimes followed by a depression. GAL decreased the facilitatory effect of a conditioning stimulus train to C-fibers in the sural nerve. The depressive effect of GAL could be prevented by the i.t. coadministration of calcitonin gene-related peptide (CGRP), but not substance P (SP) and was not reversed by i.t. naloxone or bicuculline. The results illustrate the complex effect of GAL on the spinal cord, possibly exhibiting a biphasic effect. The observed effects on the flexor reflex are probably not due to changes in the excitability of motoneurons. Descending inhibitory pathways or local inhibitory non-GAL interneurons probably are not involved in the depressive effect of GAL. The possibility that the observed effects are related to primary sensory afferents containing not only GAL but also CGRP, and/or to local GAL neurons in the dorsal horn is discussed.     

Effect of ketamine on spinal cord nociceptive transmission in normal and monoarthritic rats.

The effects of systemically and intrathecally administered ketamine on spinal wind-up of normal and monoarthritic rats were studied by using C-fiber reflex responses evoked by repetitive (0.6 Hz) electric stimulation. Both systemic and intrathecal ketamine induced dose-dependent depression of wind-up activity in normal rats, as revealed by the dose-related inhibitory effects of the drug. At the same intraperitoneal doses, ketamine produced a greater inhibitory effect on wind-up activity of monoarthritic rats, compared to normal animals. The intrathecal administration of ketamine also produced wind-up inhibition, the efficacy being higher in the monoarthritic rats. Results indicate that ketamine depresses spinal wind-up, specially in rats submitted to chronic pain, probably due to its antagonistic properties on dorsal horn NMDA receptors, which play a crucial role in the maintenance of chronic pain.     

Depressant and excitant effects of intraspinal microinjections of morphine and methionine-enkephalin in the cat

The effects of intraspinal microinjectins of morphine (10 microgram) and methionine-enkephalin (Met-enkephalin) (5 microgram) on the C-fiber and polysynaptic reflexes in the acute decerebrate low spinal cat were investigated. Microinjected into the dorsal horn, morphine and Met-enkephalin depressed the nociceptive C-fiber reflex (CFR) without altering the short latency polysynaptic reflex. Microinjected into the ventral horn, morphine and Met-enkephalin facilitated the C-fiber and polysynaptic reflexes. Pretreatment of the cats with intravenous naltrexone (2 mg/kg) antagonized the depressant effects produced by dorsal horn intraspinal microinjections of morphine and Met-enkephalin. The excitant effects of ventral horn microinjections of morphine were not antagonized by naltrexone (2 mg/kg). These results support a hypothesis that the analgesic effects of morphine at the spinal cord level are due to interactions with opiate receptors in the dorsal horn.     

Studies of muscarinic receptor subtypes in salivary gland function in anaesthetized rats

The in vivo study aimed to examine whether muscarinic receptor subtypes other than muscarinic M3 receptors exert exocrine functional roles in the rat salivary glands. The effects of pirenzepine, methoctramine and 4-diphenylacetoxy-N-methylpiperidine methiodide (4-DAMP) were examined on secretion from the major salivary glands evoked by acetylcholine (0.001-10 micromol kg(-1) i.v.) in pentobarbitone-anaesthetized rats. Observations were occasionally made on glandular blood flow. 4-DAMP (0.1-100 nmol kg(-1) i.v.) markedly and equipotently inhibited the acetylcholine-evoked fluid responses in all glands. Pirenzepine (0.1 micromol kg(-1) i.v.-10 mmol kg(-1) i.v.) showed significantly lower inhibitory potency than 4-DAMP, most conspicuously in the parotid, while methoctramine (0.1 micromol kg(-1) i.v.-10 mmol kg(-1) i.v.) exerted an even lesser inhibitory effect. Also against acetylcholine-evoked blood flow increases, 4-DAMP showed a conspicuous potency. At 1 and 10 micromol kg(-1) i.v. of pirenzepine, the antagonist reduced the protein concentration in the submandibular saliva, but not in the parotid saliva. While 4-DAMP (1 and 10 nmol kg(-1) i.v.) significantly inhibited acetylcholine-evoked protein secretory responses in the submandibular glands, methoctramine (below 10 micromol kg(-1) i.v.) affected the responses in neither gland. The reduction of the protein concentration in submandibular saliva caused by 4-DAMP and pirenzepine was inhibited by N(omega)-nitro-L-arginine methyl ester (L-NAME; 30 mg kg(-1) i.p.), while L-NAME had no or only minute effects on the parotid protein secretion. Thus, in addition to muscarinic M3 receptors, other muscarinic receptors contribute to in vivo functional responses in rat submandibular and sublingual glands. While these other receptors are muscarinic M1 receptors in the sublingual gland, they may be a different subtype, possibly muscarinic M5 receptors, in the submandibular gland. However, muscarinic M1 receptors may induce indirect effects via nitric oxide in the submandibular gland.     

Supraspinal involvement in the phrenic-to-phrenic inhibitory reflex

Phrenic afferents are capable of attenuating the phrenic motor response elicited by the intercostal-to-phrenic excitatory reflex in decerebrate, paralyzed cats. High spinal transection eliminates the attenuating effect of the bilateral phrenic-to-phrenic inhibitory reflex. These results indicate that although phrenic nerve afferents do exert an inhibitory influence in the cervical spinal cord, some of the inhibitory effects are likely to involve supraspinal mechanisms.     

The effect of glutamate antagonists on c-fos expression induced in spinal neurons by irritation of the lower urinary tract

Chemical irritation of the lower urinary tract (LUT) of the rat increases the expression of c-fos in neurons in the dorsal horn, dorsal commissure and intermediolateral region of the spinal cord. The role of glutamatergic synapses in this response was examined using two glutamate receptor antagonists, MK-801 (an NMDA antagonist) and CNQX (an AMPA antagonist). In rats with an intact spinal cord, MK-801 (3.5 mg/kg, i.v.) administered 15 min before bladder irritation decreased (50–60%) the number of c-fos-positive cells in all regions of the cord. A smaller dose of MK-801 (0.8 mg/kg, i.v.) was ineffective. In spinal transected rats (4–7 days prior to the experiment) MK-801 (3.5 mg/kg, i.v.) decreased c-fos expression only in the medial dorsal horn. CNQX (1.2 mg/kg, i.v.) was ineffective in both preparations. These results indicate that activation of NMDA receptors at glutamate synapses in the central nervous system may play a role in the processing of nociceptive input from the LUT and may also be involved in reflex pathways mediating micturition.     

The effect of morphine on the potassium evoked release of tritiated 5-hydroxytryptamine from spinal cord slices in the rat

The effect of morphine on the potassium (40 mM) evoked release of exogenous [3H]5-HT from slices of the dorsal spinal cord of the rat was studied. The effects of in vitro applied morphine on the slices were compared to those produced by systemic morphine applied to the animals before preparation of the slices. The in vitro application of morphine (10(-6) to 10(-5) M) did not affect the release of [3H]5-HT. By contrast, it was observed that the potassium evoked release of [3H]5-HT from the slices of the spinal cord of rats which had received 10 mg/kg s.c. of morphine 30 min beforehand was significantly increased. The effect of systemic morphine was dose-dependent (in the range of 1.5-10 mg/kg s.c.) and could be blocked by prior administration of naloxone (1 mg/kg i.m.) 2 min before the morphine. The acute administration of 10 mg/kg s.c. of morphine, which did not induce analgesia in rats rendered tolerant to morphine, did not modify the [3H]5-HT release. Higher doses of morphine, which have been shown to restore analgesia in these rats, induced an increase in the release which was significant for a dose of 100 mg/kg s.c. These results demonstrating a specific and dose-dependent increase in the potassium evoked release of [3H]5-HT from spinal dorsal cord slices after systemic administration of morphine, emphasize the role of serotonergic systems in such analgesia. The lack of effect of the drug directly applied in vitro favours a supraspinal site of action of the drug and is in good agreement with recent results in the literature.     

The role of spinal and brainstem adenosine receptors in the modulation of the volume-evoked micturition reflex in the unanesthetized rat

The pharmacology of the spinal, supraspinal and peripheral adenosine receptor subtypes (A1, A2) and their influence on the volume-evoked micturition reflex (VEMR) was studied in a chronic unanesthetised rat model by cystometrography after intrathecal (i.t.), intracerebroventricular (i.c.v.) and intravenous (i.v.) injection. Intrathecally administered A1 adenosine agonist: N6-(L-2-phenylisopropyl)adenosine (R-PIA) and A2 adenosine agonist: 5'-(N-ethylcarboxamido)adenosine (NECA) were equally active with 1.0 nmol reliably producing an increase in the volume necessary to induce the VEMR. At a higher dose (3 nmol), a long-lasting blockade of the VEMR was produced by both agonists. These effects were reversed following intraperitoneal injection of caffeine, an adenosine antagonist. This inhibition of the VEMR outlasted the spinal antinociceptive action which we have previously reported for these two agonists. Contrary to the spinal effect of these agonists, i.c.v. (0.3-3 nmol) and i.v. (100-1000 nmol) injections of R-PIA and NECA resulted in a significant decrease in the volume required to evoke the VEMR. We conclude that at the spinal level a xanthine-sensitive adenosine receptor(s) inhibits the VEMR. Based on several indirect lines of evidence, we speculate that these effects are not mediated by an action on primary afferent input or directly on preganglionic neurons, but on an excitatory interneuronal link.     

Spinal NK1 receptors contribute to the increased excitability of the nociceptive flexor reflex during persistent peripheral inflammation

Hyperalgesia is a characteristic of inflammation and is mediated, in part, by an increase in the excitability of spinal neurons. Although substance P does not appear to mediate fast synaptic events that underlie nociception in the spinal cord, it may contribute to the hyperalgesia and increased excitability of spinal neurons during inflammation induced by complete Freund's adjuvant. We examined the role of endogenous substance P in changes in the excitability of spinal neurons during adjuvant-induced, peripheral inflammation by determining the effect of a selective NK₁ receptor antagonist (RP67580) on the nociceptive flexor reflex in adult rats. Experiments were conducted 2 or 3 days after injection of adjuvant. Animals exhibited moderate thermal hyperalgesia at this time. The flexor reflex was evoked by electrical stimulation of the sural nerve and was recorded in the ipsilateral hamstring muscles. The flexor reflex ipsilateral to the inflamed hindpaw was enhanced approximately two-fold compared to the flexor reflex evoked in untreated animals as determined by the number of potentials and the duration of the reflex. The enhanced reflex in adjuvant-treated animals was most likely due to an increase in the excitability of spinal interneurons because short-latency activity in the hamstring muscles did not differ between untreated animals and adjuvant-treated animals following electrical stimulation of the L₅ dorsal root or the nerve innervating the muscle with a stimulus that was 1.3–1.5 times the threshold for excitation of A-fibers. Intrathecal administration of RP67580 (2.3 and 6.8 nmol) attenuated the flexor reflex evoked in adjuvant-treated animals, but had no effect in untreated animals. Intravenous or intraplantar injection of RP67580 (6.8 nmol) did not affect the flexor reflex in adjuvant-treated animals indicating a spinal action of the drug following intrathecal administration. RP68651, the enantiomer of RP67580, was without effect at doses up to 6.8 nmol, indicating that the effects of comparable doses of RP67580 were due to an action of the drug at NK₁ receptors. However, intrathecal administration of 23 nmol of both drugs attenuated the reflex in adjuvant-treated and control animals indicating that effects of RP67580 at this dose were not mediated entirely by its action at NK₁ receptors. Overall, these data suggest that endogenous substance P has a role in the increased excitability of spinal interneurons observed during persistent inflammation and support the hypothesis that substance P released in the spinal cord contributes to the hyperalgesia that accompanies adjuvant-induced persistent, peripheral inflammation.