Yoshiyuki Tsuboi

Astroglia in Medullary Dorsal Horn (Trigeminal Spinal Subnucleus Caudalis) Are Involved in Trigeminal Neuropathic Pain Mechanisms

The aim of this study was to investigate whether astroglia in the medullary dorsal horn (trigeminal spinal subnucleus caudalis; Vc) may be involved in orofacial neuropathic pain following trigeminal nerve injury. The effects of intrathecal administration of the astroglial aconitase inhibitor sodium fluoroacetate (FA) were tested on Vc astroglial hyperactivity [as revealed by glial fibrillary acid protein (GFAP) labeling], nocifensive behavior, Vc extracellular signal-regulated kinase phosphorylation (pERK), and Vc neuronal activity in inferior alveolar nerve-transected (IANX) rats. Compared with sham-control rats, a significant increase occurred in GFAP-positive cells in ipsilateral Vc at postoperative day 7 in IANX rats, which was prevented following FA administration. FA significantly increased the reduced head withdrawal latency to high-intensity heat stimulation of the maxillary whisker pad skin in IANX rats, although it did not significantly affect the reduced escape threshold to low-intensity mechanical stimulation of the whisker skin in IANX rats. FA also significantly reduced the increased number of pERK-like immunoreactive cells in Vc and the enhanced Vc nociceptive neuronal responses following high-intensity skin stimulation that were documented in IANX rats, and glutamine administration restored the enhanced responses. These various findings provide the first documentation that astroglia is involved in the enhanced nociceptive responses of functionally identified Vc nociceptive neurons and in the associated orofacial hyperalgesia following trigeminal nerve injury.

Morphological and electrophysiological properties of ACCx nociceptive neurons in rats.

A total of 33 neurons with cutaneous receptive fields were recorded from the anterior cingulate cortex (ACCx) and successfully injected with neurobiotin. All neurons were in area 24 of the ACCx. Neurons from the ACCx had large receptive fields (RFs), usually bilateral, and some had RFs covering the whole body surface. Most of the neurons were in the deep laminae and had a pyramidal soma with thick apical dendrites and many spines. Thirteen of 33 neurons were classified as pyramidal nociceptive specific (NS) neurons and 12 as noxious-tap neurons, 3 neurons received inhibitory input and were in lamina V. Two non-pyramidal noxious-tap neurons were located in lamina V and 1 pyramidal noxious-tap neuron was located in lamina VI, and 2 pyramidal NS neurons were in lamina III. Axon collaterals of NS neurons were mainly distributed around the soma, whereas those of noxious-tap neurons were also distributed far from the soma. A large number of varicosities were observed on the axon collaterals of both NS and noxious-tap neurons. Our results suggest that NS neurons in the ACCx send information locally to the vicinity of the soma, while noxious-tap neurons send information to a wider area of the ACCx.

Mechanisms involved in modulation of trigeminal primary afferent activity in rats with peripheral mononeuropathy

In order to clarify the mechanisms underlying the changes in primary afferent neurons in trigeminal neuropathic pain, a chronic constriction nerve injury model of the infraorbital nerve (ION‐CCI) was developed in rats. Mechanical allodynia was observed at 3 days after ION‐CCI and lasted more than 14 days. Single‐unit activities were recorded from the ION of anesthetized rats. C‐, Aβ‐ and Aδ‐units were identified on the basis of their conduction velocity. Aδ‐units were frequently encountered at a later period after ION‐CCI. The highest Aδ‐spontaneous activity was recorded at 3 days after ION‐CCI and progressively decreased after that, but spontaneous activity was still higher at 14 days after ION‐CCI than that of naïve rats. Mechanical‐evoked responses of Aδ‐units were also highest at 3 days after ION‐CCI and then gradually decreased. In consideration of these data, patch‐clamp recordings were performed on medium to large size neurons of the dissociated trigeminal ganglion (TRG). Patch‐clamp recordings revealed that the IK (sustained) and IA (transient) in rats with ION‐CCI were significantly smaller than those of naïve rats, and correlated with an increase in duration of repolarization phase and a decrease in duration of depolarization phase, respectively. The hyperpolarization‐activated current (Ih) was significantly larger in TRG neurons of rats with ION‐CCI as compared with those of naïve rats. The present results suggest that Ih, IK and IA in Aδ‐afferent neurons in TRG are significantly involved in the changes in afferent spontaneous activity and mechanically evoked activity that accompany mechanical allodynia produced by trigeminal nerve injury.

Alteration of the Second Branch of the Trigeminal Nerve Activity Following Inferior Alveolar Nerve Transection in Rats

Abstract After transection of the inferior alveolar nerve (IAN), the whisker pad area, which is innervated by the infraorbital nerve (ION) that was not injured, showed hypersensitivity to mechanical stimulation. Two days after IAN transection, threshold intensity for escape behavior to mechanical stimulation of the ipsilateral whisker pad area was less than 4.0 g, indicating mechanical allodynia. A total of 68 single fiber discharges were recorded from ION fibers at 3 days after IAN transection. The responses of C‐ and A‐fibers were classified according to their conduction velocity. The C‐fiber activities were not affected by IAN transection, whereas A‐fiber activities were significantly enhanced by IAN transection as indicated by an increase in background activity and mechanically evoked response. Since the A‐fiber responses were significantly affected by IAN transection, patch clamp recording was performed from middle to large diameter retrogradely labeled and acutely dissociated trigeminal ganglion (TRG) neurons. The IK (sustained) and IA (transient) currents were significantly smaller and hyperpolarization‐activated current (Ih) was significantly larger in TRG neurons of rats with IAN transection as compared to those of naive rats. Furthermore, current injection into TRG neurons induced high frequency spike discharges in rats with IAN transection. These data suggest that changes in K+ current and Ih observed in the uninjured TRG neurons reflect an increase in excitability of TRG neurons innervated by the ION after IAN transection, resulting in the development of mechano‐allodynia in the area adjacent to the injured IAN innervated region.

Potassium channels as a potential therapeutic target for trigeminal neuropathic and inflammatory pain.

Previous studies in several different trigeminal nerve injury/inflammation models indicated that the hyperexcitability of primary afferent neurons contributes to the pain pathway underlying mechanical allodynia. Although multiple types of voltage-gated ion channels are associated with neuronal hyperexcitability, voltage-gated K+ channels (Kv) are one of the important physiological regulators of membrane potentials in excitable tissues, including nociceptive sensory neurons. Since the opening of K+ channels leads to hyperpolarization of cell membrane and a consequent decrease in cell excitability, several Kv channels have been proposed as potential target candidates for pain therapy. In this review, we focus on common changes measured in the Kv channels of several different trigeminal neuropathic/inflammatory pain animal models, particularly the relationship between changes in Kv channels and the excitability of trigeminal ganglion (TRG) neurons. We also discuss the potential of Kv channel openers as therapeutic agents for trigeminal neuropathic/inflammatory pain, such as mechanical allodynia.

Effects of intrastriatal injections of selective dopamine D-1 and D-2 agonists and antagonists on jaw movements of rats

The effects of bilateral intrastriatal injections of the selective D-1 and D-2 antagonists, SCH23390 and sulpiride on apomorphine-induced jaw movements were studied in ketamine-anaesthetized rats after C1 spinal transection. A photo-transducer attached to the lower mandible automatically detected jaw movements. Apomorphine (0.2, 0.5 and 1.0 mg/kg i.v.) dose dependently increased jaw movements, an effect prevented by prior administration into the ventral striatum of either SCH23390 (0.1, 0.5 and 1 microgram) or sulpiride (125 ng). To be effective, SCH23390 had to be given less than 30 min before apomorphine whereas sulpiride had to be given earlier. Sulpiride injected into the dorsal striatum potentiated the effects of apomorphine, an action prevented by administering the sulpiride with SCH23390. Local application of the selective D-1 and D-2 agonists, SKF38393 (5 micrograms) and quinpirole (10 micrograms) into sites within the ventral striatum from which repeated jaw movements could be obtained by electrical stimulation, also evoked jaw movements; the effects of combining the two drugs were much greater than the effects of either drug alone.

Organization of pERK-immunoreactive cells in trigeminal spinal nucleus caudalis and upper cervical cord following capsaicin injection into oral and craniofacial regions in rats

To define the somatotopic arrangement of neurons in the trigeminal spinal subnucleus caudalis and upper cervical cord activated by acute noxious stimulation of various orofacial sites, pERK expression was analyzed in these neurons. After capsaicin injection into the tongue, lower gum, upper and lower lips, or mental region, pERK‐like immunoreactive (pERK‐LI) cells were distributed mainly in the dorsal half of the trigeminal spinal nucleus interporalis (Vi) and caudalis (Vc) transition zone (Vi/Vc zone), middle Vc, and Vc and upper cervical cord transition zone (Vc/C2 zone). pERK‐LI cells were distributed throughout the dorsal to ventral portion of the Vi/Vc zone, middle Vc, and Vc/C2 zone following capsaicin injection into the anterior hard palate, upper gum, buccal mucosa, or vibrissal pad and in the ventral portion of the Vi/Vc zone, middle Vc, and Vc/C2 zone following snout, ophthalmic, or ocular injection of capsaicin. The rostrocaudal distribution area of pERK‐LI cells was more extensive from the Vi/Vc zone to the Vc/C2 zone after intraoral injection than that after facial injection, and the rostrocaudal distribution of pERK‐LI cells from the Vi/Vc zone to the Vc/C2 zone had a somatotopic arrangement, with the snout being represented most rostrally and ophthalmic, ocular, or mental regions represented most caudally. These findings suggest that the pERK‐LI cells expressed from the Vi/Vc zone to the Vc/C2 zone following injection of capsaicin in facial and intraoral structures may be differentially involved in pain perception in facial and intraoral sites. J. Comp. Neurol. 507:1428–1440, 2008.

Dorsal column-thalamic pathway is involved in thalamic hyperexcitability following peripheral nerve injury: a lesion study in rats with experimental mononeuropathy.

Abstract A total of 68 neurons were recorded from the ventro‐postero‐lateral nucleus of thalamus (VPL) in rats with a unilateral chronic constriction injury (CCI) of the sciatic nerve (n=20), sham operation (n=24) and naive rats (n=24), and effects of the lesion of dorsal column (DC) pathway [DC lesion or DC+gracile nucleus lesions] on VPL nucleus neuronal activities were studied. In the VPL nucleus contralateral to the CCI (receiving input from the injured nerve), response latencies of low threshold mechanoreceptive (LTM) and wide dynamic range (WDR) neurons to electrical stimulation of the sciatic nerve were significantly longer than that in the contralateral VPL nucleus receiving input from the sham‐operated side (P<0.05). In contrast, response latencies of LTM and WDR neurons to DC stimulation were not different between the sham operated and CCI sides (P>0.05). Background activity of WDR neurons was significantly higher in the VPL nucleus contralateral to the CCI side when compared to neurons in the VPL nucleus contralateral to the sham operated side and in naive animals. Responses of LTM and WDR neurons to innocuous mechanical stimulation of the receptive fields were significantly decreased after DC and DC+gracile nucleus lesions in all animals. However, the responses of WDR neurons to noxious stimuli were selectively reduced only in rats with CCI by DC and DC+gracile nucleus lesions (P<0.05). The decrease in noxious stimulus‐evoked responses of WDR neurons in the VPL nucleus contralateral to the CCI side after DC and DC+gracile nucleus lesions was greater than that in the VPL nucleus contralateral to the sham operated side and naive animals. These results indicated that DC and DC+gracile nucleus lesions produced selective and stronger effect on noxious responses of VPL nucleus WDR neurons receiving input from the site of nerve injury. The findings suggest that the gracile nucleus‐thalamic pathway conveys, or modulates, nociceptive information to the VPL nucleus following peripheral nerve injury, resulting in an increase in VPL nucleus response to noxious stimuli that contributes to the development of mechanical hyperalgesia.

Responses of dorsal column nuclei neurons in rats with experimental mononeuropathy

&NA; To examine the functional role of dorsal column nuclei in neuropathic pain, electrophysiological properties of low‐ and high‐threshold dorsal column nuclei neurons in neuropathic and normal rats were examined. Single‐neuronal activities were recorded from the gracile nucleus (GN) in rats at 10–14 days after application of four loose ligatures around the sciatic nerve (chronic constriction nerve injury; CCI). A total of 190 units were recorded from the GN in naive and CCI rats. The largest population of low‐threshold mechanoreceptive (LTM) neurons recorded from the GN of naive rats were classified as rapidly‐adapting (RA) LTM neurons, whereas those from CCI rats were slowly‐adapting (SA) neurons. Mean orthodromic latencies of GN neurons ipsilateral to the CCI after sciatic nerve stimulation were significantly longer than those of naive animals and those of GN neurons without receptive fields were significantly longer than any other type of neurons. One hundred and eight of 190 GN neurons were also antidromically activated following electrical stimulation of the ventro‐lateral posterior nucleus of the thalamus. Furthermore, when stronger stimuli were applied to the sciatic nerve, some GN neurons also responded with long latencies. GN neurons of sham‐operated naive rats and those contralateral to the CCI had mechanical receptive fields on the paw, whereas 8.0% of the GN neurons in the CCI side did not have any detectable mechanical receptive field. Receptive field size was not significantly different between neurons ipsilateral or contralateral to the CCI and those of naive rats. Spontaneous activity of GN neurons from the ipsilateral side was significantly higher than those from the contralateral side. On the other hand, spontaneous activity of GN neurons both ipsilateral and contralateral to the CCI were significantly higher than those from naive rats. Furthermore, afterdischarges of GN neurons ipsilateral and contralateral to the CCI were significantly higher than those of naive rats. The present data suggest that the dorsal column pathway is involved in CCI‐produced sensory abnormalities by conveying their hyperactivity to thalamic neurons.

Mechanisms involved in an increment of multimodal excitability of medullary and upper cervical dorsal horn neurons following cutaneous capsaicin treatment

BackgroundIn order to evaluate mechanisms that may underlie the sensitization of trigeminal spinal subnucleus caudalis (Vc; the medullary dorsal horn) and upper cervical spinal cord (C1-C2) nociceptive neurons to heat, cold and mechanical stimuli following topical capsaicin treatment of the facial skin, nocifensive behaviors as well as phosphorylation of extracellular regulated-kinase (pERK) in Vc and C1-C2 neurons were studied in rats.ResultsCompared to vehicle application, capsaicin application to the lateral facial skin produced 1 hour later a flare in the skin, and also induced significantly greater nocifensive behaviors to heat, cold or mechanical stimulus of the lateral facial skin. The intrathecal (i.t.) injection of the MEK inhibitor PD98059 markedly attenuated the nocifensive behaviors to these stimuli in capsaicin-treated rats. Moreover, the number of pERK-like immunoreactive (pERK-LI) cells in Vc and C1-C2 was significantly larger following the heat, cold and mechanical stimuli in capsaicin-treated rats compared with vehicle-treated rats. The number of pERK-LI cells gradually increased following progressive increases in the heat or mechanical stimulus intensity and following progressive decrease in the cold stimulus. The ERK phosphorylation in Vc and C1-C2 neurons was strongly inhibited after subcutaneous injection of the capsaicin antagonist capsazepine in capsaicin-treated rats.ConclusionThe present findings revealed that capsaicin treatment of the lateral facial skin causes an enhancement of ERK phosphorylation in Vc and C1-C2 neurons as well as induces nocifensive behavior to heat, cold and mechanical simulation of the capsaicin-treated skin. The findings suggest that TRPV1 receptor mechanisms in rat facial skin influence nociceptive responses to noxious cutaneous thermal and mechanical stimuli by inducing neuroplastic changes in Vc and C1-C2 neurons that involve in the MAP kinase cascade.