Inhibition of trigeminal neurones after intravenous administration of naratriptan through an action at 5-hydroxy-tryptamine (5-HT(1B/1D)) receptors

1. The observation that 5-hydroxytryptamine (5-HT) is effective in treating acute attacks of migraine when administered intravenously resulted in a research effort that led to the discovery of the 5-HT(1B/1D) receptor agonist sumatriptan. 2. Clinical experience has shown sumatriptan to be an effective treatment with some limitations, such as relatively poor bioavailability, which naratriptan was developed to address. Increasing bioavailability has been achieved with greater lipophilicity and thus the potential for greater activity in the central nervous system. 3. In this study the increased access to central sites has been exploited in an attempt to characterize the pharmacology of those central receptors with the newer tools available. Trigeminovascular activation was examined in the model of superior sagittal sinus stimulation. 4. Cats were anaesthetized with alpha-chloralose (60 mg kg(-1), intraperitoneal), paralyzed (gallamine 6 mg kg(-1), intravenously) and ventilated. The superior sagittal sinus was accessed and isolated for electrical stimulation (250 micros pulses, 0.3 Hz, 100 V) by a mid-line circular craniotomy. The region of the dorsal surface of C2 spinal cord was exposed by a laminectomy and an electrode placed for recording evoked activity from sinus stimulation. 5. Stimulation of the superior sagittal sinus resulted in activation of cells in the dorsal horn of C2. Cells fired with a probability of 0.69+/-0.1 at a latency of 9.2+/-0.2 ms. Intravenous (i.v.) administration of naratriptan at clinically relevant doses (30 and 100 microg kg(-1)), inhibited neuronal activity in trigeminal neurones of the C2 dorsal horn, reducing probability of firing without affecting latency. 6. The effect of naratriptan could be reversed by administration of the selective 5-HT(1B/1D) receptor antagonist GR127935 (100 microg kg(-1), i.v.). 7. These data establish that naratriptan acts on central trigeminal neurones since sagittal sinus stimulation activates axons within the tentorial nerve and there are no inhibitory effects mediated within the trigeminal ganglion. Furthermore, given that this inhibition could be reversed by the relatively selective 5-HT(1B/1D) receptor antagonist GR127935, it is highly likely that the anti-migraine effects of drugs of this class with central nervous system access are mediated, at least in part, by 5-HT(1B/1D) receptors within the trigeminal nucleus.     

Omega-agatoxin IVA, a P-type calcium channel antagonist, reduces nociceptive processing in spinal cord neurons with input from the inflamed but not from the normal knee joint--an electrophysiological study in the rat in vivo

High threshold voltage-dependent P- and Q-type calcium channels are involved in neurotransmitter release. In order to investigate the role of P- and Q-type calcium channels in the mechanosensory (nociceptive) processing in the spinal cord, their participation in the responses of spinal wide-dynamic-range neurons to innocuous and noxious mechanical stimulation of the knee and ankle joints was studied in 30 anaesthetized rats. The knee was either normal or acutely inflamed by kaolin/carrageenan. During the topical application of omega-agatoxin IVA (P-type channel antagonist, 0.1 microM) onto the dorsal surface of the spinal cord, the responses to innocuous and noxious pressure applied to the normal knee were increased to respectively 124 +/- 42% and 114 +/- 23% of predrug values (mean +/- SD, P < 0.05, 14 neurons). By contrast, in rats with an inflamed knee, the responses to innocuous and noxious pressure applied to the knee were reduced to respectively 72 +/- 19 and 73 +/- 22% of baseline (mean +/- SD, P < 0.01, 13 neurons). In the same neurons, omega-agatoxin IVA slightly increased the responses to pressure on the non-inflamed ankle whether the knee was normal or inflamed. Thus P-type calcium channels seem to acquire a predominant importance in the excitation of spinal cord neurons by mechanosensory input from inflamed tissue and hence in the generation of inflammatory pain. By contrast, the Q-type channel antagonist, omega-conotoxin MVIIC (1 or 100 microM), had no significant effect upon responses to innocuous or noxious pressure applied to either normal or inflamed knees (25 neurons).     

Actions of kainate and AMPA selective glutamate receptor ligands on nociceptive processing in the spinal cord

Kainate receptors expressing the GluR5 subunit of glutamate receptor are present at high levels on small diameter primary afferent neurones that are considered to mediate nociceptive inputs. This suggests that GluR5 selective ligands could be novel analgesic agents. The role of kainate receptors on C fibre primary afferents has therefore been probed using three compounds that are selective for homomeric GluR5 receptors. The agonist, ATPA, and the antagonists, LY294486 and LY382884, have been tested in four models of nociception: responses evoked by noxious stimulation of the periphery have been recorded electrophysiologically (1) from hemisected spinal cords from neonatal rats in vitro, (2) from single motor units in adult rats in vivo, (3) from dorsal horn neurones in adult rats in vivo, and (4) in hotplate tests with conscious mice. In some protocols comparisons were made with the AMPA selective antagonist GYKI 53655. The agonist ATPA reduced nociceptive reflexes in vitro, but failed to have effects in vivo. In all tests, the GluR5 antagonists reduced nociceptive responses but only at doses that also affected responses to exogenous AMPA. The AMPA antagonist reduced nociceptive responses at doses causing relatively greater reductions of responses to exogenous AMPA. The results indicate that GluR5 selective ligands do reduce spinal nociceptive responses, but they are not strongly analgesic under these conditions of acute nociception.     

Increasing isoflurane from 0.9 to 1.1 minimum alveolar concentration minimally affects dorsal horn cell responses to noxious stimulation

BACKGROUND: The spinal cord appears to be the site at which isoflurane suppresses movement that occurs in response to a noxious stimulus. In an attempt to localize its site of suppressant action, the authors determined the effect of isoflurane on dorsal horn neuronal responses to supramaximal noxious stimulation at end-tidal concentrations that just permitted and just prevented movement. METHODS: Rats (n = 14) were anesthetized with isoflurane, and after lumbar laminectomy, the minimum alveolar concentration (MAC) for each rat was determined using a supramaximal mechanical stimulus. In these same rats, after extracellular microelectrode placement in the lumbar spinal cord, dorsal horn neuronal responses to the supramaximal stimulus were determined at the concentrations of isoflurane that bracketed each rat's MAC (0.1% higher and lower than MAC). The MAC of isoflurane was then re-determined. RESULTS: Dorsal horn neuronal response was 1,757+/-892 impulses/min at 0.9 MAC and 1,508+/-988 impulses/min at 1.1 MAC, a 14% decrease (P < 0.05). Cell responses varied, with some cells increasing their response at the higher concentration of isoflurane. The MAC of isoflurane was 1.38+/-0.2% before and 1.34+/-0.2% after determination of dorsal horn neuronal responses. CONCLUSIONS: Isoflurane, at concentrations that bracket MAC, has a variable and minimal depressant effect on dorsal horn cell responses to noxious mechanical stimulation. These data suggest that the major action of isoflurane to suppress movement evoked by a noxious stimulus might occur primarily at a site other than the dorsal horn.     

Vagal afferent stimulation-produced effects on nociception in capsaicin-treated rats

1. The effects of electrical stimulation of cervical vagal afferent fibers on the nociceptive tail-flick (TF) reflex and responses of spinal dorsal horn neurons to noxious cutaneous stimulation were studied in adult rats treated as neonates with either capsaicin or vehicle. 2. Vagal afferent stimulation (VAS) produced biphasic, intensity-dependent effects on the TF reflex in vehicle-treated and untreated control rats. The TF reflex was facilitated in both groups of rats at lesser intensities of VAS (2.5-50 microA) and fully inhibited at greater intensities of VAS (50-100 microA). In contrast, biphasic effects of VAS on the TF reflex generally were not produced in rats treated as neonates with capsaicin. Facilitation of the TF reflex was produced in these rats by lesser intensities of VAS as well as by typically "inhibitory" intensities of VAS; the TF reflex was never inhibited in 6/12 rats, even at the greatest intensity of VAS tested (1,000 microA). When the TF reflex was inhibited by VAS in capsaicin-treated rats, the intensities required were significantly greater than those required in vehicle-treated or untreated rats. 3. In electrophysiological experiments, 77 neurons were recorded in the lumbar spinal dorsal horn of pentobarbital sodium-anesthetized, paralyzed rats treated as neonates with either vehicle or capsaicin. The neurons had receptive fields on the glabrous skin of the plantar surface of the ipsilateral hind foot, and all responded to mechanical stimuli of both nonnoxious and noxious intensities; 16/77 neurons also responded to noxious thermal stimulation. In vehicle-treated rats, nociceptive responses of 50% of 30 units studied were biphasically modulated by VAS, 33% were only inhibited, and 17% were only facilitated by VAS at the intensities tested (5-500 microA). In capsaicin-treated rats, nociceptive responses of 32% of 47 units studied were biphasically modulated by VAS, 15% were only inhibited, and 34% were only facilitated by VAS at the intensities tested (5-500 microA). In addition, nociceptive responses of neurons facilitated at lesser intensities of VAS and not affected at greater intensities of VAS were observed in capsaicin-treated rats (19% of the 47-unit sample). Overall, the proportion of the neuronal sample inhibited by VAS was less, and the proportion of the sample facilitated by VAS was greater in capsaicin-treated rats compared with vehicle-treated rats. 4. The efficacy of the capsaicin treatment was evaluated immunocytochemically.(ABSTRACT TRUNCATED AT 400 WORDS)     

Biphasic modulation of spinal nociceptive transmission from the medullary raphe nuclei in the rat

The modulatory effects of electrical and chemical (glutamate) stimulation in the rostral ventromedial medulla (RVM) on spinal nociceptive transmission and a spinal nociceptive reflex were studied in rats. Electrical stimulation at a total 86 sites in the RVM in the medial raphe nuclei (n = 54) and adjacent gigantocellular areas (n = 32) produced biphasic (facilitatory and inhibitory, n = 43) or only inhibitory (n = 43) modulation of the tail-flick (TF) reflex. At these 43 biphasic sites in the RVM, facilitation of the TF reflex was produced at low intensities of stimulation (5-25 microA) and inhibition was produced at greater intensities of stimulation (50-200 microA). At 43 sites in the RVM, electrical stimulation only produced intensity-dependent inhibition of the TF reflex. Activation of cell bodies in the RVM by glutamate microinjection reproduced the biphasic modulatory effects of electrical stimulation. At biphasic sites previously characterized by electrical stimulation, glutamate at a low concentration (5 nmol) produced facilitation of the TF reflex; a greater concentration (50 nmol) only inhibited the TF reflex. In electrophysiological experiments, electrical stimulation at 62 sites in the RVM produced biphasic (n = 26), only inhibitory (n = 26), or only facilitatory (n = 10) modulation of responses of lumbar spinal dorsal horn neurons to noxious cutaneous thermal (50 degrees C) or mechanical (75.9 g) stimulation. Facilitatory effects were produced at lesser intensities of stimulation and inhibitory effects were produced at greater intensities of stimulation. The apparent latencies to stimulation-produced facilitation and inhibition, determined with the use of a cumulative sum method and bin-by-bin analysis of spinal neuron responses to noxious thermal stimulation of the skin, were 231 and 90 ms, respectively. The spinal pathways conveying descending facilitatory and inhibitory influences were found to be different. Descending facilitatory influences on the TF reflex were conveyed in ventral/ventrolateral funiculi, whereas inhibitory influences were conveyed in dorsolateral funiculi. The results indicate that descending inhibitory and facilitatory influences can be simultaneously engaged throughout the RVM, including nucleus raphe magnus, and that such influences are conveyed in different spinal funiculi.     

Inhibition of trigeminal neurons by intravenous administration of the serotonin (5HT)1B/D receptor agonist zolmitriptan (311C90): are brain stem sites therapeutic target in migraine?

Migraine is a common and debilitating condition. Its treatment has received considerable attention in recent times with the introduction into clinical use of the serotonin (5HT)1B/D-like agonist sumatriptan. It is known from human studies that the intracranial blood vessels and dura mater are important pain-sensitive structures since mechanical or electrical stimulation of these vessels, such as the superior sagittal sinus, causes pain. We have developed a model of craniovascular pain by stimulating the superior sagittal sinus and monitoring trigeminal neuronal activity using electrophysiological techniques. In this study we determined the effect of intravenous administration of the novel anti-migraine compound zolmitriptan (311C90) upon evoked neuronal activity in trigeminal neurons. Nine adult cats were anaesthetised with alpha-chloralose (60 mg/kg, i.p.; 20 mg/kg, i.v., 2-hourly) with all surgery being conducted under halothane (1-3%). The superior sagittal sinus was isolated for electrical stimulation. Recordings were made from caudal trigeminal neurons at the C2 level of the cervical spinal cord with tungsten-in-glass microelectrodes. Signals were amplified and analysed by a custom-written program that enabled software filtering and extraction of both evoked potential and single cell data. Data were collected before and after administration of zolmitriptan. Electrical stimulation of the superior sagittal sinus resulted in activation of neuronal elements within the trigeminal nucleus that could be monitored as single unit activity or as evoked potentials, the latter reflecting both primary afferent and trigeminal cell body activity. The evoked potential recorded from the trigeminal nucleus was 207 +/- 14 microV and was reduced by zolmitriptan (100 micrograms/kg, i.v.) to a mean of 98 +/- 17 microV. Similarly, the probability of firing for trigeminal neurons was reduced from a control level of 0.63 +/- 0.1 to 0.13 +/- 0.05 after a dose of 100 micrograms/kg intravenously. These effects were dose-dependent and were significantly different from the effect of vehicle (P < 0.05). These data demonstrate that systemically administered zolmitriptan can inhibit evoked trigeminovascular activity within the trigeminal nucleus. This inhibition of trigeminal activity may play a role in the anti-migraine actions of this compound and offers the prospect of a third pathophysiologically consistent target site for anti-migraine drug effects.     

Predictable deuteration of recombinant proteins expressed in Escherichia coli

Carrageenan was used to study inflammation-induced changes in spinal nociception and its brain stem modulation in the pentobarbitone-anesthetized rat. Carrageenan was administered intraplantarly into one hindpaw 2 h before the start of electrophysiological single unit recordings of wide-dynamic range (WDR) neurons of the spinal dorsal horn. Carrageenan produced a significant leftward shift in the stimulus-response function for mechanical stimuli, whereas that for noxious heat stimuli was short of statistical significance. Conditioning electrical stimulation in the rostroventromedial medulla (RVM) significantly attenuated noxious heat-evoked, but not mechanically evoked, responses to spinal dorsal horn WDR neurons in the control (contralateral) side. However, in the carrageenan-treated side RVM stimulation had no significant effect on mechanically or noxious heat-evoked responses. Following direct spinal administration of neuropeptide FF (NPFF), noxious heat-evoked responses, but not mechanically evoked responses, were attenuated by RVM-stimulation also in the carrageenan-treated side. This selective NPFF-induced enhancement of brain stem-spinal inhibition was not reversed by naloxone. The results indicate that carrageenan-induced inflammation significantly changes the response properties of spinal nociceptive neurons and their brain stem-spinal modulation. During inflammation, NPFF in the spinal cord produces a submodality-selective potentiation of the antinociceptive effect induced by brain stem-spinal pathways, independent of naloxone-sensitive opioid receptors.     

Somatotopic and laminar organization of fos-like immunoreactivity in the medullary and upper cervical dorsal horn induced by noxious facial stimulation in the rat

The distribution of fos-like-immunoreactivity (fos-LI) in the medullary and upper cervical dorsal horn was examined following noxious facial stimulation, in order to evaluate the use of fos as a marker for neuronal activation in trigeminal nociceptive pathways. Control animals that received urethane anesthesia and no facial stimulation showed substantial bilateral labeling in the trigeminal complex that was restricted to one rostrocaudal level, at the transition between the medullary dorsal horn (nucleus caudalis) and nucleus interpolaris. Noxious mechanical stimulation (pinch) of different facial sites produced labeling in the ipsilateral dorsal horn whose distribution varied predictably with the rostrocaudal and dorsoventral position of the facial stimulation site, such that rostral facial sites were represented rostrally in the dorsal horn and dorsal sites were represented ventrolaterally. The cornea was exceptional among the facial stimulation sites in that it had a specific representation at two distinct rostrocaudal levels, in C1 and the interpolaris-caudalis transition region; the position of the rostral peak was somatotopically inappropriate, based on the representation of other facial sites. The proportion of labelling in laminae III-IV relative to laminae I-II was higher with noxious mechanical stimulation than with noxious thermal (55 degrees C) or chemical (subcutaneous injection of capsaicin) stimulation. The proportion of labelling in laminae III-IV produced by electrical stimulation of the infraorbital nerve was no greater than that produced by pinch. The results suggest that fos-LI mapping can be a useful method for the investigation of somatotopy but is subject to serious limitations when used for the investigation of laminar organization. The results also suggest that the interpolaris-caudalis transition region may have properties that are distinct from those of the rest of the trigeminal complex, possibly related to an involvement in autonomic function.     

Involvement of the caudal medulla in negative feedback mechanisms triggered by spatial summation of nociceptive inputs

In the rat, applying noxious heat stimuli to the excitatory receptive fields and simultaneously to adjacent, much larger, areas of the body results in a surface-related reduction in the responses of lumbar dorsal horn convergent neurons. These inhibitory effects induced by spatial summation of nociceptive inputs have been shown to involve a supraspinally mediated negative feedback loop. The aim of the present study was to determine the anatomic level of integration of these controls and hence to ascertain what relationships they might share with other descending controls modulating the transmission of nociceptive signals. The responses of lumbar convergent neurons to noxious stimulation (15-s immersion in a 48 degrees C water bath) applied to increasing areas of the ipsilateral hindlimb were examined in several anesthetized preparations: sham-operated rats, rats with acute transections performed at various levels of the brain stem, and spinal rats. The effects of heterotopic noxious heat stimulation (tail immersion in a 52 degrees C water bath) on the C-fiber responses of these neurons also were analyzed. The electrophysiological properties of dorsal horn convergent neurons, including their responses to increasing stimulus surface areas, were not different in sham-operated animals and in animals the brain stems of which had been transected completely rostral to a plane -2. 8 mm remote from interaural line (200 micron caudal to the caudal end of the rostral ventromedial medulla). In these animals, increasing the stimulated area size from 4.8 to 18 cm2 resulted in a 35-45% reduction in the responses. In contrast, relative to responses elicited by 4.8 cm2 stimuli, responses to 18 cm2 were unchanged or even increased in animals with transections at more caudal level and in spinal animals. Inhibitions of the C-fiber responses elicited by heterotopic noxious heat stimulation were in the 70-80% range during conditioning in sham-operated animals and in animals with rostral brain stem transections. Such effects were reduced significantly (residual inhibitions in the 10-20% range) in animals with transections >500 micron caudal to the caudal end of the rostral ventromedial medulla and in spinal animals. It is concluded that the caudal medulla constitutes a key region for the expression of negative feed-back mechanisms triggered by both spatial summation of noxious inputs and heterotopic noxious inputs.     

Chronic spinal nerve ligation induces changes in response characteristics of nociceptive spinal dorsal horn neurons and in their descending regulation originating in the periaqueductal gray in the rat

We studied whether a chronic neuropathy induced by unilateral spinal nerve ligation changes the response characteristics of spinal dorsal horn wide-dynamic range (WDR) neurons or their periaqueductal gray (PAG)-induced descending modulation. Experiments were performed in rats with behaviorally demonstrated allodynia induced by spinal nerve ligation and in a group of nonneuropathic control rats. The stimulus-response functions of WDR neurons for mechanical and thermal stimuli and the modulation of their peripherally evoked responses by electrical stimulation of the PAG were determined under pentobarbital anesthesia. The results showed that neuropathy caused a significant leftward shift in stimulus-response functions for mechanical stimuli. In contrast, stimulus-response functions for noxious heat stimuli in the neuropathic limb were, if anything, shifted rightward, although this shift was short of statistical significance. In neuropathic rats, PAG stimulation produced a significantly stronger attenuation of spinal neuronal responses induced by noxious heat in the unoperated than in the operated side. At the intensity that produced attenuation of noxious heat stimuli, PAG stimulation did not produce any significant change in spinal neuronal responses evoked by mechanical stimuli either from the operated or the nonoperated hindlimb of the neuropathic rats. Spontaneous activity of WDR neurons was higher in the operated side of neuropathic rats than in control rats. Afterdischarges evoked by peripheral stimuli were observed in 1/16 of the WDR neurons ipsilateral to spinal nerve ligation and not at all in other experimental groups. The WDR neurons studied were not activated by innocuous or noxious cold stimuli. The results indicate that spinal nerve ligation induces increased spontaneous activity and enhanced responses to mechanical stimuli in the spinal dorsal horn WDR neurons, whereas noxious heat-evoked responses are not significantly changed or if anything, attenuated. Moreover, the inhibition of noxious heat stimuli by PAG stimulation is attenuated in the neuropathic side. It is proposed that the observed changes in the response characteristics of the spinal dorsal horn WDR neurons and in their descending modulation may contribute to the neuropathic symptoms in these animals.     

Synthesis of thiazole-4-carboxamide-adenine difluoromethylenediphosphonates substituted with fluorine at C-2'' of the adenosine

Stimulation in the nucleus raphe magnus (NRM) inhibits transmission of nociceptive information within the spinal cord through activation of bulbospinal pathways. This study used microdialysis in combination with high pressure liquid chromatography to measure the release of serotonin (5HT) and several amino acids, including glutamate, aspartate and glycine, from the lumbar dorsal horn during electrical stimulation within the NRM in the alpha-chloralose anesthetized cat. Observed release of putative neurotransmitters was correlated with inhibition of nociceptive projection neurons recorded from sites within 800 microns rostral or caudal to the dialysis fiber. NRM stimulus parameters considered to preferentially activate myelinated fibers caused inhibition of nociceptive evoked activity, and increased the release of excitatory amino acids and glycine within the spinal cord, with no detectable release of 5HT. When pulse widths were lengthened and unmyelinated fibers were also activated, increases in 5HT in the spinal dialysate were observed as well. Strychnine administered through the dialysis fiber (0.02-1 mM) antagonized NRM-induced inhibition when 5HT release was not detected. Inhibition produced by stimulation that increased 5HT concentrations was relatively strychnine resistant. These results point to a raphe-spinal inhibitory pathway that is not dependent on 5HT, the activation of which results in the spinal release of glycine.     

5-HT autoreceptors in the regulation of 5-HT release from guinea pig raphe nucleus and hypothalamus

5-HT autoreceptors involved in the regulation of 5-HT release in the guinea pig dorsal raphe nucleus have been studied in comparison with those in the hypothalamus. In vitro release was measured in slices of raphe and hypothalamus prelabelled with [3H]5-HT, superfused with Krebs solution and depolarized electrically. The non-selective 5-HT receptor agonist, 5-carboxamidotryptamine (5-CT) (0.1-10 nM for raphe: 1-100 nM for hypothalamus) and antagonist, methiothepin (10-1000nM), decreased and increased, respectively, the release of [3H]5-HT evoked by electrical stimulation in either of these regions when given alone. The selective 5-HT1B/D receptor antagonist, GR127935 (100-1000 nM), and the 5-HT1D receptor antagonist, ketanserin (300-1000 nM), had no significant effect on this release in either of these regions. Methiothepin and GR127935 (100-1000 nM) shifted to the right the concentration-effect curve of 5-CT in both the raphe and the hypothalamus. At 300 nM, ketanserin shifted to the right the concentration-effect curve of 5-CT in the raphe but did not modify the 5-CT curve in the hypothalamus. In microdialysis experiments ketanserin, applied locally at 10 microM, increased the extracellular levels of 5-HT in the dorsal raphe nucleus of the freely moving guinea pig, whereas 5-HT levels were unchanged in the hypothalamus. Ketanserin at 1 microM did not affect the decrease in 5-HT output induced by the selective 5-HT1B/D receptor agonist, naratriptan (used at 10 microM in raphe and 0.1 microM in hypothalamus), in the raphe or the hypothalamus. In the raphe, WAY100635, a 5-HT1A receptor antagonist, at 1 microM, did not prevent naratriptan (10 microM) from reducing the extracellular levels of 5-HT. These results suggest that, in the conditions used in this study, the release of 5-HT in the dorsal raphe nucleus is possibly modulated in part by 5-HT1B receptors but essentially the control is through 5-HT receptors whose subtype is still to be determined. In the hypothalamus, however, it is clear that only 5-HT1B receptors are involved in the modulation of 5-HT neurotransmission.     

Codon bias evolution in Drosophila. Population genetics of mutation-selection drift

Reflex cardiovascular responses elicited by noxious oro-facial stimulation are well known but the neural pathways that underlie trigeminal cardiovascular reflex reactions remain to be elucidated. In previous studies, we have shown that noxious electrical stimulation of the mandibular incisor in the anesthetized rat elicits increases in mean arterial blood pressure and heart rate (Allen, G.V., Barbrick, B. and Esser, M.J., Trigeminal parabrachial connections: possible pathway for nociception-induced cardiovascular reflex responses, Brain Res., 715 (1996) 125-135). In this study, microinjections of the presynaptic blocker, cobalt chloride, or the anesthetic agent, lidocaine, were made into selected brainstem sites to identify neural pathways that are involved in mediation of the reflex pressor responses. Ipsilateral and bilateral injections of chemical blocker into the dorsomedial spinal trigeminal nucleus, pars caudalis, lateral parabrachial nucleus and the rostral ventral lateral medulla/caudal A5 region attenuated the reflex cardiovascular response. Bilateral injections of cobalt chloride into the dorsomedial subnucleus caudalis resulted in 70-100% attenuation of the reflex pressor response. Bilateral injections of cobalt chloride and/or lidocaine into the lateral parabrachial nucleus or the rostral ventral lateral medulla/A5 region resulted in 43-57% and 44-100% attenuation of the reflex pressor response, respectively. There were no significant differences in the degree or duration of attenuation of the reflex pressor responses produced by cobalt chloride compared to that produced by lidocaine injections. The reflex pressor responses usually returned to baseline levels approximately 60 min following injection of the chemical blocker substance. The results indicate that noxious electrical stimulation of the mandibular incisor elicits a reflex increase in mean arterial blood pressure which is initially mediated in the dorsomedial spinal trigeminal nucleus, pars caudalis and is subsequently mediated in the lateral parabrachial nucleus and the rostral ventral lateral medulla/caudal A5 region.     

Release of immunoreactive substance P in the trigeminal brain stem nuclear complex evoked by chemical stimulation of the nasal mucosa and the dura mater encephali--a study with antibody microprobes

In order to study a possible involvement of substance P in the processing of chemonociceptive input from the nasal mucosa and the dura mater encephali in the spinal trigeminal, the release of immunoreactive substance P was measured in the trigeminal brain stem nuclear complex in anaesthetized rats. Microprobes coated with antibody to substance P were inserted into the lateral area of the brain stem up to 1 mm posterior to the obex corresponding to the trigeminal subnucleus caudalis. When the nasal mucosa was stimulated by topical administration of mustard oil (1% and 5%) into the nostrils, immunoreactive substance P was mainly detected in the dorsal region of the trigeminal brain stem nuclear complex with a maximum in the superficial gray matter. When the dura mater encephali was stimulated by topical administration of Tyrode's solution (pH 6.2), immunoreactive substance P was mainly released in the ventral region of the trigeminal brain stem nuclear complex; with pH 5.5 the release was more diffuse extending from the ventral to the dorsal part of the spinal trigeminal nucleus. Release was maximal rather after than during the administration of the stimuli, and it considerably outlasted the stimulation periods. These data suggest that substance P plays an important role in the processing of chemonociceptive inputs from the nasal mucosa and the dura mater encephali in the trigeminal brain stem nuclear complex. Substance P may be important, therefore, in the generation of those headaches that are caused by affections of the nasal mucosa and the dura mater encephali. Since enhanced levels of immunoreactive substance P were present for considerable time periods beyond the administration of the stimuli, substance P and neurokinin-1 receptors may be involved in long-lasting neuronal events following noxious stimulation.     

The NSAID dexketoprofen trometamol is as potent as mu-opioids in the depression of wind-up and spinal cord nociceptive reflexes in normal rats

The aim of this study was to examine the potency of the antinociceptive effects of the non-steroidal antiinflammatory drug (NSAID), Dexketoprofen Trometamol (the active enantiomer of ketoprofen) on spinal cord nociceptive reflexes. These effects were compared with those of the mu-opioid receptor agonist fentanyl in normal animals. The experiments were performed in male Wistar rats anaesthetised with alpha-chloralose. The nociceptive reflexes were recorded as single motor units in peripheral muscles, activated by mechanical and electrical stimulation. Both dexketoprofen and fentanyl inhibited responses evoked by mechanical and electrical stimulation with doses in the same nanomolar range (dexketoprofen ID50s: 100 and 762 nmol kg-1 and fentanyl: 40 and 51 nmol kg-1, respectively). Dexketoprofen and fentanyl also significantly inhibited wind-up. Since fentanyl has been shown to be some 1000 times more potent than morphine in this type of experiments, we conclude that dexketoprofen has central analgesic actions in normal animals and depresses nociceptive responses with a potency similar to that of mu-opioid agonists.     

ATP P2x receptors and sensory synaptic transmission between primary afferent fibers and spinal dorsal horn neurons in rats

ATP P2x receptors and sensory synaptic transmission between primary afferent fibers and spinal dorsal horn neurons in rats. J. Neurophysiol. 80: 3356-3360, 1998. Glutamate is a major fast transmitter between primary afferent fibers and dorsal horn neurons in the spinal cord. Recent evidence indicates that ATP acts as another fast transmitter at the rat cervical spinal cord and is proposed to serve as a transmitter for nociception and pain. Sensory synaptic transmission between dorsal root afferent fibers and neurons in the superficial dorsal horn of the lumbar spinal cord were examined by whole cell patch-clamp recording techniques. Experiments were designed to test if ATP could serve as a transmitter at the lumbar spinal cord. Monosynaptic excitatory postsynaptic currents (EPSCs) were completely abolished after the blockade of both glutamatergic alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid/kainate and N-methyl--aspartate receptors. No residual current was detected, indicating that glutamate but not ATP is a fast transmitter at the dorsal horn of the lumbar spinal cord. Pyridoxal-phosphate-6-azophenyl-2',4'-disulfonic acid (PPADS), a selective P2x receptor antagonist, produced an inhibitory modulatory effect on fast EPSCs and altered responses to paired-pulse stimulation, suggesting the involvement of a presynaptic mechanism. Intrathecal administration of PPADS did not produce any antinociceptive effect in two different types of behavioral nociceptive tests. The present results suggest that ATP P2x2 receptors modulate excitatory synaptic transmission in the superficial dorsal horn of the lumbar spinal cord by a presynaptic mechanism, and such a mechanism does not play an important role in behavioral responses to noxious heating. The involvement of other P2x subtype receptors, which is are less sensitive to PPADS, in acute nociceptive modulation and persistent pain remains to be investigated.     

Central mechanism of an extensive analgesic effect due to strong electroacupuncture of acupoint on spinal dorsal horn neurons]

Experiments were performed on male rats. The responses of dorsal horn convergent neurons in spinal cord (T12-L1) to noxious stimulation of hind paw were recorded extracellularly with glass microelectrode. When low intensity (2 V) electroacupuncture (EA) was used, the nociceptive responses of convergent neurons were inhibited by EA at "Zusanli" near noxious stimulation area, but not at "Xiaguan" far from the area. When intensity (18 V) high than the threshold of C fibers EA was applied at the far acupoint "Xiaguan", obvious analgesic effects on convergent neurons were also produced, showing an extensive analgesic effect of strong EA at acupoint. This extensive analgesic effect was abdicated by lesion of nucleus raphe magnus (NRM), but still persisted to some extent by EA at the same segment acupoint "Zusanli" with 18 V or 2 V intensity. The results suggest that, the extensive analgesia of strong EA at far segment acupoint may be mainly mediated by noxious stimulation through NRM, a negative feedback mechanism modulating pain of supraspinal cord. The analgesia due to 2 V EA at the same segment acupoint may be mainly produced by gate control in spinal cord, but also to some extend by supraspinal cord mechanism.     

Antinociception produced by microinjection of morphine in the rat periaqueductal gray is enhanced in the foot, but not the tail, by intrathecal injection of alpha1-adrenoceptor antagonists

Antinociception produced by microinjection of morphine in the ventrolateral periaqueductal gray is mediated in part by alpha2-adrenoceptors in the spinal cord dorsal horn. However, several recent reports demonstrate that microinjection of morphine in the ventrolateral periaqueductal gray inhibits nociceptive responses to noxious heating of the tail by activating descending neuronal systems that are different from those that inhibit the nociceptive responses to noxious heating of the feet. More specifically, alpha2-adrenoceptors appear to mediate the antinociception produced by morphine using the tail-flick test, but not that using the foot-withdrawal or hot-plate tests. The present study extended these findings and determined the role of alpha1-adrenoceptors in mediating the antinociceptive effects of morphine microinjected into the ventrolateral periaqueductal gray using both the foot-withdrawal and the tail-flick responses to noxious radiant heating in lightly anesthetized rats. Intrathecal injection of selective antagonists was used to determine whether the antinociceptive effects of morphine were modulated by alpha1-adrenoceptors. Injection of the selective alpha1-adrenoceptor antagonists prazosin or WB4101 potentiated the increase in the foot-withdrawal response latency produced by microinjection of morphine in the ventrolateral periaqueductal gray. In contrast, either prazosin or WB4101 partially reversed the increase in the tail-flick response latency produced by morphine. These results indicate that microinjection of morphine in the ventrolateral periaqueductal gray modulates nociceptive responses to noxious heating of the feet by activating descending neuronal systems that are different from those that inhibit the nociceptive responses to noxious heating of the tail. More specifically, alpha1-adrenoceptors mediate a pro-nociceptive action of morphine using the foot-withdrawal response, but in contrast, alpha1-adrenoceptors appear to mediate part of the antinociceptive effect of morphine determined using the tail-flick test.     

Distribution of trigeminohypothalamic and spinohypothalamic tract neurons displaying substance P receptor-like immunoreactivity in the rat

Primary afferent neurons containing substance P (SP) are apparently implicated in the transmission of noxious information from the periphery to the central nervous system, and SP released from primary afferent neurons acts on second-order neurons with the SP receptor (SPR). In the rat, nociceptive information reached the hypothalamus not only through indirect pathways but also directly through trigeminohypothalamic and spinohypothalamic pathways. Thus, in the present study, the distribution pattern of trigeminohypothalamic and spinohypothalamic tract neurons showing SPR-like immunoreactivity (SPR-LI) was examined in the rat by a retrograde tract-tracing method combined with immunofluorescence histochemistry for SPR. A substantial number of trigeminal and spinal neurons with SPR-LI were retrogradely labeled with Fluoro-Gold (FG) injected into the hypothalamic regions. These neurons were distributed mainly in lamina I of the medullary and spinal dorsal horns, lateral spinal nucleus, regions around the central canal of the spinal cord, and the lateral aspect of the deep part of the spinal dorsal horn. A number of SPR-LI neurons in the spinal parasympathetic nucleus were labeled with FG injected into the area around the paraventricular hypothalamic nucleus. Some SPR-LI neurons in the lateral spinal nucleus and the lateral aspect of the deep part of the spinal dorsal horn were also labeled with FG injected into the septal region. On the basis of the distribution areas of SPR-LI trigeminal and spinal neurons projecting to the hypothalamic and septal regions, it is likely that these neurons are involved in the transmission of somatic and/or visceral noxious information.