Gabapentin and pregabalin ameliorate mechanical hypersensitivity after spinal cord injury in mice.

The antiepileptic drugs gabapentin and pregabalin exhibit well-established analgesic effects in patients with several neuropathic conditions. In the present study, we examined their effects on mechanical hypersensitivity in mice subjected to weight-drop spinal cord injury. Hindlimb motor function and mechanical hypersensitivity were evaluated using the Basso-Beattie-Bresnahan (BBB) locomotor rating scale and the von Frey test, respectively, for 4 weeks after spinal cord injury. Despite gradual recovery of hindlimb motor function after spinal cord injury, mice exhibited continuous development of mechanical hypersensitivity. Gabapentin (30 and 100 mg/kg) and pregabalin (10 and 30 mg/kg), administered intraperitoneally on the 28th day after spinal cord injury, reduced mechanical hypersensitivity in a dose-dependent manner. These results suggest that gabapentin and pregabalin could be useful therapeutic tools for patients with neuropathic pain after spinal cord injury.

Glycine transporter blockade ameliorates motor ataxia in a mouse model of spinocerebellar atrophy.

Ataxic movement, the common major symptom of spinocerebellar atrophy, has been considered to involve impaired glutamatergic excitatory neurotransmission in the cerebellum. Considering the therapeutic importance of ataxia control, we assessed the effectiveness of increasing the extracellular concentration of glycine by administering it exogenously or via blockade of glycine transporter 1, using its selective inhibitors sarcosine and N-[3-(4'-fluorophenyl)-3-(4'-phenylphenoxy)propyl]sarcosine (NFPS), for amelioration of motor ataxia in a mouse model of spinocerebellar atrophy developing after neonatal treatment with cytosine beta-D-arabinofuranoside. Intracerebroventricular (i.c.v.) injection of sarcosine (3, 10, and 30 microg) and NFPS (0.01 and 0.03 microg) reduced the number of falls without affecting spontaneous motor activity, and therefore the falling index [(number of falls / spontaneous motor activity) x 100], and dose-dependently ameliorated ataxic movements. Similar effects were observed upon i.c.v. injection of D-serine (1 and 10 microg), an agonist of the glycine-recognition site of the N-methyl-D-aspartate (NMDA) receptor. However, exogenously injected glycine (1, 3, and 10 microg, i.c.v.) only weakly ameliorated the ataxic movements at 3 microg. These results suggest the therapeutic relevance of GlyT1 inhibitors for amelioration of motor ataxia in spinocerebellar atrophy by increasing the endogenous concentration of glycine near the glycine-recognition site of the NMDA receptor.

Neurochemical evidence that supraspinally administered gabapentin activates the descending noradrenergic system after peripheral nerve injury.

We have previously demonstrated that gabapentin supraspinally activates the descending noradrenergic system to produce analgesic effects after peripheral nerve injury. To further establish the neurochemical basis for its supraspinally mediated analgesic action, concentrations of spinal noradrenaline, 4-hydroxy-3-methoxyphenylglycol (MHPG), serotonin (5-HT), 5-hydroxyindoleacetic acid (5-HIAA) and dopamine were measured using high-performance liquid chromatography in a murine neuropathic pain model that was prepared by partial ligation of the sciatic nerve (the Seltzer model). Intracerebroventricularly (i.c.v.) administered gabapentin (100 and 300 microg) increased the spinal MHPG concentration and the MHPG/noradrenaline ratio and alleviated mechanical hypersensitivity, whereas the concentrations of noradrenaline, 5-HT, 5-HIAA and dopamine were unchanged. By contrast, i.c.v. gabapentin neither affected the spinal MHPG concentration and MHPG/noradrenaline ratio nor exhibited analgesic effects in animals subjected to a sham operation. In addition, spinal monoamine levels in ligated animals were not changed after intrathecal administration of gabapentin which however generated analgesic effects. Thus, the supraspinally mediated analgesic effects of gabapentin are correlated with an increase in spinal noradrenaline turnover.

Phenytoin and carbamazepine delay the initial depression of the population spike upon exposure to in vitro ischemia and promote its post-ischemic functional recovery in rat hippocampal slices.

Antiepileptic drugs have been shown to reduce the severity of neurodegeneration resulting from stroke or brain injury. In the present study, we evaluated the effects of the antiepileptic drugs phenytoin and carbamazepine on the time course of changes in the population spike (PS) during brief oxygen/glucose deprivation (OGD) in the CA1 pyramidal region of rat hippocampal slices in vitro. After introducing simulated ischemia by OGD, the PS was initially inhibited, followed by transient recovery and subsequent reinhibition again concomitantly with disappearance of the presynaptic volley (PV). The slices were then reperfused with oxygen/glucose-containing solution. Both phenytoin and carbamazepine (30 and 100 muM each) concentration-dependently delayed the initial inhibition and the time to transient recovery of the PS during OGD, thus prolonging the time until disappearance of the PV. However, they significantly promoted restoration of the PS after reperfusion. These results suggest that treatment with phenytoin and carbamazepine increases the resistance of tissue to energy deprivation, as evidenced by the facilitated post-ischemic recovery of the PS, despite prolonged ischemia.

Gabapentin depresses C-fiber-evoked field potentials in rat spinal dorsal horn only after induction of long-term potentiation.

C-fiber-evoked field potentials in response to electrical stimulation of the sciatic nerve were recorded in the dorsal horn of the rat lumbar spinal cord, and their long-term potentiation (LTP) was induced by high-frequency stimulation applied on the sciatic nerve as a synaptic model of hypersensitivity underlying an increased efficacy of nociceptive transmission. We evaluated the effect of gabapentin on the basal C-fiber-evoked field potentials and their established LTP. Intravenously administered gabapentin (10 and 30 mg/kg, i.v.) reduced the LTP of C-fiber-evoked field potentials in a dose-dependent manner when applied 60 min after establishment of the LTP. However, gabapentin did not affect the basal C-fiber-evoked field potentials or induction of the LTP. Thus, gabapentin was effective only in sensitized conditions. By contrast, morphine HCl (1 and 3 or 10 mg/kg, i.v.) reduced both the basal responses and their established LTP. The combination of gabapentin and morphine at lower doses of each drug appeared to result in a stronger reduction on the established LTP than that of each drug alone, suggesting that combination therapy can generate better analgesia in the treatment of chronic pain.

Fluvoxamine, a selective serotonin reuptake inhibitor, exerts its antiallodynic effects on neuropathic pain in mice via 5-HT2A/2C receptors.

There is an association between depression and chronic pain, and some antidepressants exert antinociceptive effects in humans and laboratory animals. We examined the effects of fluvoxamine, a selective serotonin reuptake inhibitor, on mechanical allodynia and its mechanism of action in the mouse chronic pain model, which was prepared by partially ligating the sciatic nerve. The antiallodynic effect was measured using the von Frey test. Fluvoxamine produced antiallodynic effects following both systemic and intrathecal administration. In 5-hydroxytryptamine (5-HT)-depleted mice, prepared by intracerebroventricular injection of 5,7-dihyroxytryptamine, the fluvoxamine-induced antiallodynic effect was significantly attenuated. The antiallodynic effects of systemic fluvoxamine were also reduced by both systemic and intrathecal administration of ketanserin, a 5-HT2A/2C receptor antagonist. In addition, fluvoxamine also induced antinociceptive effect in the acute paw pressure test, and this effect was antagonized by the 5-HT3 receptor antagonist granisetron. These results indicate that fluvoxamine exerts its antiallodynic effects on neuropathic pain via descending 5-HT fibers and spinal 5-HT2A or 5-HT2C receptors, and the antinociception on acute mechanical pain via 5-HT3 receptors.

Spinal cord injury-specific depression of monosynaptic spinal reflex transmission by l-5-hydroxytryptophan results from loss of the 5-HT uptake system and not 5-HT receptor supersensitivity.

We studied changes in the spinal segmental reflex and serotonergic (5-HT) responses in rats after spinal cord injury (SCI) produced by the weight-dropping method at the T8 level. The spinal monosynaptic reflex amplitude (MSR) was recorded from the L5 ventral root following stimulation of the ipsilateral L5 dorsal root. The 5-HT precursor l-5-hydroxytryptophan (L-5-HTP) depressed MSR in the spinal cord injured rats but not in normal rats. We investigated whether the SCI-specific depression of MSR by L-5-HTP was attributable to postsynaptic supersensitivity of 5-HT receptors or presynaptic loss of the 5-HT uptake system. Sumatriptan, a selective 5-HT(1B/1D) receptor agonist that is not taken up by 5-HT transporters, depressed the MSR similarly in both SCI and normal rats, suggesting that SCI resulted in the loss of 5-HT terminals and not postsynaptic supersensitivity of 5-HT receptors.

Spinal alpha(2)-adrenergic and muscarinic receptors and the NO release cascade mediate supraspinally produced effectiveness of gabapentin at decreasing mechanical hypersensitivity in mice after partial nerve injury.

After partial nerve injury, the central analgesic effect of systemically administered gabapentin is mediated by both supraspinal and spinal actions. We further evaluate the mechanisms related to the supraspinally mediated analgesic actions of gabapentin involving the descending noradrenergic system. Intracerebroventricularly (i.c.v.) administered gabapentin (100 microg) decreased thermal and mechanical hypersensitivity in a murine chronic pain model that was prepared by partial ligation of the sciatic nerve. These effects were abolished by intrathecal (i.t.) injection of either yohimbine (3 microg) or idazoxan (3 microg), alpha(2)-adrenergic receptor antagonists. Pretreatment with atropine (0.3 mg kg(-1), i.p. or 0.1 microg, i.t.), a muscarinic receptor antagonist, completely suppressed the effect of i.c.v.-injected gabapentin on mechanical hypersensitivity, whereas its effect on thermal hypersensitivity remained unchanged. Similar effects were obtained with pirenzepine (0.1 microg, i.t.), a selective M(1)-muscarinic receptor antagonist, but not with methoctramine (0.1 and 0.3 microg, i.t.), a selective M(2)-muscarinic receptor antagonist. The cholinesterase inhibitor neostigmine (0.3 ng, i.t.) potentiated only the analgesic effect of i.c.v. gabapentin on mechanical hypersensitivity, confirming spinal acetylcholine release downstream of the supraspinal action of gabapentin. Moreover, the effect of i.c.v. gabapentin on mechanical but not thermal hypersensitivity was reduced by i.t. injection of L-NAME (3 microg) or L-NMMA (10 microg), both of which are nitric oxide (NO) synthase inhibitors. Systemically administered naloxone (10 mg kg(-1), i.p.), an opioid receptor antagonist, failed to suppress the analgesic actions of i.c.v. gabapentin, indicating that opioid receptors are not involved in activation of the descending noradrenergic system by gabapentin. Thus, the supraspinally mediated effect of gabapentin on mechanical hypersensitivity involves activation of spinal alpha(2)-adrenergic receptors followed by muscarinic receptors (most likely M(1)) and the NO cascade. In contrast, the effect of supraspinal gabapentin on thermal hypersensitivity is independent of the spinal cholinergic-NO system.

Spinal ventral root after-discharges as a pain index: involvement of NK-1 and NMDA receptors.

Nociceptive signals are transmitted to the spinal dorsal horn via primary afferent fibers, and the signals induce withdrawal reflexes by activating spinal motoneurons in the ventral horn. Therefore, nociceptive stimuli increase motoneuronal firing and ventral root discharges. This study was aimed to develop a method for the study of pain mechanisms and analgesics by recording ventral root discharges. Spinalized rats were laminectomized in the lumbo-sacral region. The fifth lumbar ventral root was sectioned and placed on a pair of wire electrodes. Multi unit efferent discharges from the ventral root were increased by mechanical stimulation using a von Frey hair applied to the plantar surface of the hindpaw. The low-intensity mechanical stimuli increased the discharges during stimulation (during-discharges) without increasing the discharges after cessation of stimulation (after-discharges), and the high-intensity mechanical stimuli increased both during- and after-discharges. Pretreatment with resiniferatoxin, an ultrapotent analogue of capsaicin, halved during-discharges and eliminated after-discharges, suggesting that after-discharges are generated by heat- and mechanosensitive polymodal nociceptors. Ezlopitant, a neurokinin-1 (NK-1) receptor antagonist, but not its inactive enantiomer, selectively reduced the after-discharges. Ketamine, an N-methyl-D-aspartate (NMDA) receptor antagonist, preferentially reduced the after-discharges, demonstrating that NK-1 and NMDA receptors mediate the after-discharges. Morphine reduced the after-discharges without affecting during-discharges. By contrast, mephenesin, a centrally acting muscle relaxant, reduced both during- and after-discharges. There results suggest that simultaneous recordings of during- and after-discharges are useful to study pain mechanisms and analgesics as well as to discriminate the analgesic effects from the side effects such as muscle relaxant effects.

Presynaptic I1-imidazoline receptors reduce GABAergic synaptic transmission in striatal medium spiny neurons.

Imidazoline receptors are expressed widely in the CNS. In the present study, whole-cell patch-clamp recordings were made from medium spiny neurons in dorsal striatum slices from the rat brain, and the roles of I1-imidazoline receptors in the modulation of synaptic transmission were studied. Moxonidine, an I1-imidazoline receptor agonist, decreased the GABAA receptor-mediated IPSCs in a concentration-dependent manner. However, glutamate-mediated EPSCs were hardly affected. The depression of IPSCs by moxonidine was antagonized by either idazoxan or efaroxan, which are both imidazoline receptor antagonists containing an imidazoline moiety. In contrast, yohimbine and SKF86466 (6-chloro-2,3,4,5-tetrahydro-3-methyl-1H-3-benzazepine), which are alpha2-adrenergic receptor antagonists with no affinity for imidazoline receptors, did not affect the moxonidine-induced inhibition of IPSCs. Moxonidine increased the paired-pulse ratio and reduced the frequency of miniature IPSCs without affecting their amplitude, indicating that this agent inhibits IPSCs via presynaptic mechanisms. Moreover, the sulfhydryl alkylating agent N-ethylmaleimide (NEM) significantly reduced the moxonidine-induced inhibition of IPSCs. Thus, the activation of presynaptic I1-imidazoline receptors decreases GABA-mediated inhibition of medium spiny neurons in the striatum, in which NEM-sensitive proteins such as G(i/o)-type G-proteins play an essential role. The adenylate cyclase activator forskolin partly opposed IPSC inhibition elicited by subsequently applied moxonidine. Furthermore, the protein kinase C (PKC) activator phorbol 12,13-dibutyrate attenuated and the PKC inhibitor chelerythrine potentiated the moxonidine-induced inhibition of IPSCs. These results suggest that IPSC inhibition via presynaptic I1-imidazoline receptors involves intracellular adenylate cyclase activity and is influenced by static PKC activity in the striatum.

Taltirelin improves motor ataxia independently of monoamine levels in rolling mouse nagoya, a model of spinocerebellar atrophy.

To examine the relationship between motor ataxia and monoamine levels in the central nervous system, the contents and concentrations of noradrenaline (NA), dopamine (DA) and serotonin (5-HT) in the cerebellum, brain stem and spinal cord were measured in rolling mouse Nagoya (RMN), a murine model of spinocerebellar atrophy. The tissue weight of the cerebellum and spinal cord, but not that of the brain stem was significantly lower in RMN than in the control group. In RMN, the NA content of the brain stem and spinal cord, but not the cerebellum were decreased relative to the control, and the concentration of NA in the spinal cord was also lower, but not significant. The DA and 5-HT contents in each tissue did not differ from those of the control, but the concentrations of monoamines, except for DA, were elevated in the brain stem and spinal cord in RMN. In particular, the concentrations of NA, DA and 5-HT in the cerebellum were significantly increased in RMN. Repeated administration of tartilerin hydrate, an analog of thyrotropin-releasing hormone, improved the ataxia of RMN, and elicited no obvious changes in either monoamine content or concentration of cerebellum, brain stem and spinal cord. These results indicate that the concentration of DA, as well as NA and 5-HT, increased in the RMN cerebellum, and that tartilerin improves the motor function of these mice via mechanisms other than changes in the levels of NA, DA and 5-HT in the central nervous system.

Centrally mediated antihyperalgesic and antiallodynic effects of zonisamide following partial nerve injury in the mouse.

Some antiepileptic drugs are used clinically to relieve neuropathic pain. We have evaluated the effects and investigated the possible mechanisms of action of zonisamide, an antiepileptic drug, on thermal hyperalgesia and tactile allodynia in a murine chronic pain model that was prepared by partial ligation of the sciatic nerve. Subcutaneously administered zonisamide (10 and 30 mg/kg) produced antihyperalgesic and antiallodynic effects in a dose-dependent manner; these effects were manifested by elevation of the withdrawal threshold in response to a thermal (plantar test) or mechanical (von Frey) stimulus, respectively. Similar analgesic effects were obtained in both the plantar and von Frey tests when zonisamide was injected either intracerebroventricularly (i.c.v., 10 and 30 microg) or intrathecally (i.t., 10 and 30 microg). It is thought that this elevation of the thermal and mechanical withdrawal thresholds after local injection of zonisamide is not generated secondarily via impaired motor activity, since zonisamide (30 microg, i.c.v. or i.t.) did not affect locomotor activity, as assessed in sciatic-nerve-ligated mice. Moreover, the nitric oxide synthase inhibitor L-NAME, when injected either i.c.v. or i.t., potentiated the analgesic effects of zonisamide. In contrast, neither i.c.v. nor i.t. zonisamide produced antinociceptive effects against acute thermal and mechanical nociception in non-ligated mice. Together, following peripheral nerve injury, it appears that zonisamide produces centrally mediated antihyperalgesic and antiallodynic effects partly via the blockade of nitric oxide synthesis.

Functional alteration of inhibitory influences on spinal motor output in painful diabetic neuropathy in rats.

Diabetes is frequently accompanied by painful polyneuropathies that are mediated by enhanced neuronal excitability in the spinal cord, partly because of decrease in spinal intrinsic inhibitory influences. Changes in spinal excitatory-inhibitory balance may alter spinal segmental motor output. In the study presented here, the mono- and disynaptic (the fastest polysynaptic) reflexes (MSR and DSR, respectively) were recorded from L5 ventral roots in response to stimulation of the ipsilateral L5 dorsal root in spinalized streptozotocin (STZ)-induced diabetic rats with a reduced withdrawal threshold to mechanical stimuli. The diabetic rats generally exhibited larger spinal reflex amplitudes, the DSR being influenced in particular. We addressed whether recurrent and presynaptic inhibition of the spinal reflexes were altered in STZ-treated animals. The recurrent inhibition of the MSR and DSR elicited by preceding antidromic conditioning stimulation delivered to the recorded L5 ventral root was markedly suppressed in diabetic rats. By contrast, the presynaptic inhibition of the MSR and DSR elicited by preceding conditioning stimulation to the ipsilateral L4 dorsal root was not impaired. Thus, in diabetic painful neuropathy, reduced spinal intrinsic inhibition in the ventral horn contributes to an enhanced spinal segmental motor output.

Involvement of supraspinal imidazoline receptors and descending monoaminergic pathways in tizanidine-induced inhibition of rat spinal reflexes.

The neuronal pathways involved in the muscle relaxant effect of tizanidine were examined by measurement of spinal reflexes in rats. Tizanidine (i.v. and intra-4th ventricular injection) decreased the mono- and disynaptic (the fastest polysynaptic) reflexes (MSR and DSR, respectively) in non-spinalized rats. Depletion of central noradrenaline by 6-hydroxydopamine abolished the depressant effect of tizanidine on the MSR almost completely and attenuated the effect on the DSR. Co-depletion of serotonin by 5,6-dihydroxytryptamine and noradrenaline resulted in more prominent attenuation of tizanidine-induced inhibition of the DSR. Supraspinal receptors were then studied using yohimbine- and some imidazoline-receptor ligands containing an imidazoline moiety. Idazoxan (I1, I2, I3, and alpha2), efaroxan (I1, I3, and alpha2), and RX821002 (I3 and alpha2), but not yohimbine, an alpha2-adrenergic receptor antagonist with no affinity for I receptors, antagonized the inhibitory effects of tizanidine. Thus, supraspinal I receptors (most likely I3) and descending monoaminergic influences are necessary for tizanidine-induced inhibition of spinal segmental reflexes.

Enhanced wind-up of the C-fiber-mediated nociceptive flexor reflex movement following painful diabetic neuropathy in mice.

We examined wind-up of the nociceptive flexor withdrawal responses in diabetic mice that had developed tactile allodynia after treatment with streptozotocin (STZ). In control and STZ-treated mice, simultaneous activation of Adelta- and C-fibers by electrical stimuli at C-fiber intensity delivered to the ventral aspect of the toe elicited a biphasic withdrawal reflex composed of short- and long-latency movements of the ipsilateral hind paw that were respectively mediated by activation of Adelta- and C-fibers. There were no significant differences between control and diabetic mice in the activation threshold of each reflex movement or the amplitude of reflexes elicited by various stimulus intensities. However, a repetitive conditioning stimulus (CS) elicited significantly greater wind-up of the C-fiber-mediated movement and early saturation of wind-up in diabetic mice. In both control and diabetic mice, the CS elicited no or occasionally slight wind-up of the A delta-fiber-mediated movement. Moreover, post-CS facilitation, which reflects the prolonged excitability increase, was observed in both Adelta-fiber- and C-fiber-mediated movements of control mice, whereas significant post-CS facilitation was only obtained in the C-fiber-mediated movement of diabetic mice, which may reflect supraspinal descending influences. Such changes in the excitability of spinal neurons in diabetic mice may represent some aspect of painful diabetic neuropathy.

Role of descending noradrenergic system and spinal alpha2-adrenergic receptors in the effects of gabapentin on thermal and mechanical nociception after partial nerve injury in the mouse.

1. To gain further insight into the mechanisms underlying the antihyperalgesic and antiallodynic actions of gabapentin, a chronic pain model was prepared by partially ligating the sciatic nerve in mice. The mice then received systemic or local injections of gabapentin combined with either central noradrenaline (NA) depletion by 6-hydroxydopamine (6-OHDA) or alpha-adrenergic receptor blockade. 2. Intraperitoneally (i.p.) administered gabapentin produced antihyperalgesic and antiallodynic effects that were manifested by elevation of the withdrawal threshold to a thermal (plantar test) or mechanical (von Frey test) stimulus, respectively. 3. Similar effects were obtained in both the plantar and von Frey tests when gabapentin was injected intracerebroventricularly (i.c.v.) or intrathecally (i.t.), suggesting that it acts at both supraspinal and spinal loci. This novel supraspinal analgesic action of gabapentin was only obtained in ligated neuropathic mice, and gabapentin (i.p. and i.c.v.) did not affect acute thermal and mechanical nociception. 4. In mice in which central NA levels were depleted by 6-OHDA, the antihyperalgesic and antiallodynic effects of i.p. and i.c.v. gabapentin were strongly suppressed. 5. The antihyperalgesic and antiallodynic effects of systemic gabapentin were reduced by both systemic and i.t. administration of yohimbine, an alpha2-adrenergic receptor antagonist. By contrast, prazosin (i.p. or i.t.), an alpha1-adrenergic receptor antagonist, did not alter the effects of gabapentin. 6. It was concluded that the antihyperalgesic and antiallodynic effects of gabapentin are mediated substantially by the descending noradrenergic system, resulting in the activation of spinal alpha2-adrenergic receptors.

GABAB receptors do not mediate the inhibitory actions of gabapentin on the spinal reflex in rats.

The clinical effectiveness of gabapentin for the treatment of epilepsy, spasticity, and neuropathic pain has been established. The mechanisms responsible for those actions, however, are still not clearly understood. We have recently demonstrated that gabapentin reduces the spinal reflex in rats via mechanisms that do not involve gamma-aminobutyric acid (GABA)A receptors. In the study, we attempted to explore the involvement of GABAB receptors in gabapentin-induced inhibition of the spinal reflexes in spinalized rats. Stimulation of the dorsal root at L5 elicited the segmental mono-(MSR) and polysynaptic reflex (PSR) in the ipsilateral ventral root. The microinjection of gabapentin (1.5 and 5 nmol) into the ventral horn reduced both MSR and PSR, whereas the injection into the dorsal horn only inhibited the PSR, indicating that systemic gabapentin inhibits the MSR at the ventral horn and it inhibits the PSR at both the ventral and dorsal horns. The GABAB-receptor antagonist CGP35348 (0.5 nmol) injected into the ventral horn antagonized the inhibition of the spinal reflexes by the GABAB-receptor agonist baclofen (i.v.) but not by gabapentin (i.v.). Thus, GABAB receptors do not appear to contribute to the gabapentin-induced inhibition of the spinal reflex.

Endogenously released 5-hydroxytryptamine depresses the spinal monosynaptic reflex via 5-HT1D receptors.

In the spinal cord, various 5-hydroxytryptamine (5-HT) receptor subtypes are involved in the modulation of motor output. Previously, we have shown that 5-HT1B receptors mediate the monosynaptic reflex depression induced by exogenously applied 5-HT that was formed from the precursor L-5-hydroxytryptophan in spinalized rats. In this study, we determined the effects of endogenous 5-HT, which was released from serotonergic terminals by DL-p-chloroamphetamine, on spinal reflexes. DL-p-chloroamphetamine depressed the monosynaptic reflex and increased the polysynaptic reflex. The depletion of 5-HT abolished the monosynaptic reflex depression, but the increase in polysynaptic reflexes was maintained, suggesting that endogenous 5-HT released by DL-p-chloroamphetamine mediates depression of the monosynaptic reflex in the spinal cord. The depression of the monosynaptic reflex was antagonized by GR127935 (N-[methoxy-3-(4-methyl-l-piperazinyl)phenyl]-2'-methyl-4'-(5-methyl-1,2,4-oxadiazol-3-yl)[1,1-biphenyl]-4-carboxamide; 5-HT1B/1D receptor antagonist) and BRL15572 (3-[4-(4-chlorophenyl)piperazin-1-yl]-1,1-diphenyl-2-propanol; 5-HT1D receptor antagonist) but not by isamoltane (5-HT(1B) receptor antagonist). These results suggest that 5-HT released from serotonergic terminals depresses monosynaptic reflex transmission via 5-HT1D receptors.

Peritumoral CpG oligodeoxynucleotide treatment inhibits tumor growth and metastasis of B16F10 melanoma cells.

Although melanoma mostly affects the skin, it is notorious for its propensity to easily develop metastasis. Metastatic melanoma is highly resistant to a variety of therapies. We examined the anti-metastatic potential of peritumoral monotherapy against murine cutaneous B16F10 melanoma with synthetic oligodeoxynucleotides (ODN) containing unmethylated CpG motifs. We demonstrated that repeated peritumoral injections of CpG ODN significantly reduced skin tumor size. Peritumoral CpG ODN-treatment of skin tumors prevented the development of pulmonary B16F10 colonies. Adoptive transfer of splenocytes obtained from CpG ODN-treated mice markedly reduced the number of previously established pulmonary colonies in recipient naïve mice. T-lymphocyte depletion studies indicated that the anti-metastatic effect was dependent on both CD4+ and CD8+ T cells. These results suggest that CpG ODN are promising as a preventive and therapeutic anti-metastatic measure against melanoma.

Antinociceptive effects of sodium channel-blocking agents on acute pain in mice.

The effects of various sodium channel blocking agents on acute thermal and mechanical nociception, as assessed using the plantar and tail pressure tests, respectively, were compared with the effects of morphine. The drugs used were mexiletine, lidocaine, carbamazepine, phenytoin, eperisone, tolperisone, and zonisamide. The sodium channel blocking agents exhibited a rather preferential elevation of the threshold for thermal nociception. By contrast, morphine produced similar analgesic effects on thermal and mechanical nociception. In the sciatic nerve isolated from mice, mexiletine, lidocaine, eperisone, and tolperisone impaired the propagation of low frequency action potentials (evoked at 0.2 Hz). Carbamazepine, phenytoin, and zonisamide generated a more frequency-dependent local anesthetic action with their obvious effects on higher frequency action potentials (evoked at 5 and/or 10 Hz). Our results show that sodium channel blocking agents have a preferential antinociceptive action against thermal stimulation that is likely to be attributed to their local anesthetic action.