Glial cells, including astrocytes and microglia, interact closely with neurons and modulate pain transmission, particularly under pathological conditions. In this study, we examined the excitability of substantia gelatinosa (SG) neurons of the spinal dorsal horn using a patch clamp recording to investigate the roles of microglial activation in the nociceptive processes of rats. We used xanthine/xanthine oxidase (X/XO), a generator of superoxide anion (O2∙–), to induce a pathological pain condition. X/XO treatment induced an inward current and membrane depolarization. The inward current was significantly inhibited by minocycline, a microglial inhibitor, and fluorocitrate, an astrocyte inhibitor. To examine whether toll-like receptor 4 (TLR4) in microglia was involved in the inward current, we used lipopolysaccharide (LPS), a highly specific TLR4 agonist. The LPS induced inward current, which was decreased by pretreatment with Tak-242, a TLR4-specific inhibitor, and phenyl N-t-butylnitrone, a reactive oxygen species scavenger. The X/XO-induced inward current was also inhibited by pretreatment with Tak-242. These results indicate that the X/XO-induced inward current of SG neurons occurs through activation of TLR4 in microglial cells, suggesting that neuroglial cells modulate the nociceptive process through central sensitization.

Background: Lumbar disc herniation (LDH) causes neurological symptoms by compression of the dura mater and nerve roots. Due to the changed in proprioception inputs that can result in abnormal postural pattern, delayed reaction time, and changed in deep tendon reflex. Objective: To investigate the effects of lumbar stabilization exercises on motor neuron excitability and neurological symptoms in patients with LDH. Design: Randomized Controlled Trial (single blind) Methods: Thirty patients with LDH were recruited; they were randomly divided into the balance center stabilization resistance exercise group (n=15) and the Nordic walking group (n=15). Each group underwent their corresponding 20- minute intervention once a day, four times a week, for four weeks. Participants’ motor neuron excitability and low back pain were assessed before and after the four-week intervention. Results: There were significant differences in all variables within each group (p<.05). There were significant differences between the experimental and control groups in the changes of upper motor neuron excitability and pain (p<.05), but not in the changes of lower motor neuron excitability and Korean Oswestry Disability Index. Conclusion: Lumbar stabilization exercises utilizing concurrent contraction of deep and superficial muscles improved low back function in patients with LDH by lowering upper motor neuron excitability than compared to exercises actively moving the limbs. Lumbar stabilization exercises without pain have a positive impact on improving motor neuron excitability.

Recent studies indicate that mitochondria are an important source of reactive oxygen species (ROS) in the spinal dorsal horn. In our previous study, application of malate, a mitochondrial electron transport complex I substrate, induced a membrane depolarization, which was inhibited by pretreatment with ROS scavengers. In the present study, we used patch clamp recording in the substantia geletinosa (SG) neurons of spinal slices, to investigate the cellular mechanism of mitochondrial ROS on neuronal excitability. DNQX (an AMPA receptor antagonist) and AP5 (an NMDA receptor antagonist) decreased the malate-induced depolarization. In an external calcium free solution and addition of tetrodotoxin (TTX) for blockade of synaptic transmission, the malateinduced depolarization remained unchanged. In the presence of DNQX, AP5 and AP3 (a groupⅠ metabotropic glutamate receptor (mGluR) antagonist), glutamate depolarized the membrane potential, which was suppressed by PBN. However, oligomycin (a mitochondrial ATP synthase inhibitor) or PPADS (a P2 receptor inhibitor) did not affect the substrates-induced depolarization. These results suggest that mitochondrial substrate-induced ROS in SG neuron directly acts on the postsynaptic neuron, therefore increasing the ion influx via glutamate receptors.

Reactive oxygen species (ROS) and nitrogen species (RNS) are both important signaling molecules involved in pain transmission in the dorsal horn of the spinal cord. Xanthine oxidase (XO) is a well-known enzyme for the generation of superoxide anions (O2 ⦁-), while S-nitroso-N-acetyl-DLpenicillamine (SNAP) is a representative nitric oxide (NO) donor. In this study, we used patch clamp recording in spinal slices of rats to investigate the effects of O2 ⦁- and NO on the excitability of substantia gelatinosa (SG) neurons. We also used confocal scanning laser microscopy to measure XO- and SNAP-induced ROS and RNS production in live slices. We observed that the ROS level increased during the perfusion of xanthine and xanthine oxidase (X/XO) compound and SNAP after the loading of 2′,7′-dichlorofluorescin diacetate (H2DCF-DA), which is an indicator of intracellular ROS and RNS. Application of ROS donors such as X/XO, β -nicotinamide adenine dinucleotide phosphate (NADPH), and 3-morpholinosydnomimine (SIN-1) induced a membrane depolarization and inward currents. SNAP, an RNS donor, also induced membrane depolarization and inward currents. X/XO-induced inward currents were significantly decreased by pretreatment with phenyl N-tert-butylnitrone (PBN; nonspecific ROS and RNS scavenger) and manganese(III) tetrakis(4-benzoic acid) porphyrin (MnTBAP; superoxide dismutase mimetics). Nitro-L-arginine methyl ester (NAME; NO scavenger) also slightly decreased X/XO-induced inward currents, suggesting that X/XO-induced responses can be involved in the generation of peroxynitrite (ONOO-). Our data suggest that elevated ROS, especially O2 ⦁-, NO and ONOO-, in the spinal cord can increase the excitability of the SG neurons related to pain transmission.

Nitric Oxide (NO) is an important signaling molecule in the nociceptive process. Our previous study suggested that high concentrations of sodium nitroprusside (SNP), a NO donor, induce a membrane hyperpolarization and outward current through large conductances calcium-activated potassium (BKca) channels in substantia gelatinosa (SG) neurons. In this study, patch clamp recording in spinal slices was used to investigate the sources of Ca²+ that induces Ca²+-activated potassium currents. Application of SNP induced a membrane hyperpolarization, which was significantly inhibited by hemoglobin and 2-(4-carboxyphenyl) -4,4,5,5- tetramethylimidazoline-1-oxyl-3-oxide potassium salt (c-PTIO), NO scavengers. SNP-induced hyperpolarization was decreased in the presence of charybdotoxin, a selective BKCa channel blocker. In addition, SNP-induced response was significantly blocked by pretreatment of thapsigargin which can remove Ca²+ in endoplasmic reticulum, and decreased by pretreatment of dentrolene, a ryanodine receptors (RyR) blocker. These data suggested that NO induces a membrane hyperpolarization through BKca channels, which are activated by intracellular Ca²+ increase via activation of RyR of Ca²+ stores.

Reactive oxygen species (ROS) and nitrogen species (RNS) are implicated in cellular signaling processes and as a cause of oxidative stress. Recent studies indicate that ROS and RNS are important signaling molecules involved in nociceptive transmission. Xanthine oxidase (XO) system is a well-known system for superoxide anions (O2˙-) generation, and sodium nitroprusside (SNP) is a representative nitric oxide (NO) donor. Patch clamp recording in spinal slices was used to investigate the role of O2˙- and NO on substantia gelatinosa (SG) neuronal excitability. Application of xanthine and xanthine oxidase (X/XO) compound induced membrane depolarization. Low concentration SNP (10μM) induced depolarization of the membrane, whereas high concentration SNP (1 mM) evoked membrane hyperpolarization. These responses were significantly decreased by pretreatment with phenyl N-tert-butylnitrone (PBN; nonspecific ROS and RNS scavenger). Addition of thapsigargin to an external calcium free solution for blocking synaptic transmission, led to significantly decreased X/XO-induced responses. Additionally, X/XO and SNP-induced responses were unchanged in the presence of intracellular applied PBN, indicative of the involvement of presynaptic action. Inclusion of GDP-β-S or suramin (G protein inhibitors) in the patch pipette decreased SNP-induced responses, whereas it failed to decrease X/XO-induced responses. Pretreatment with n-ethylmaleimide (NEM; thiol-alkylating agent) decreased the effects of SNP, suggesting that these responses were mediated by direct oxidation of channel protein, whereas X/XO-induced responses were unchanged. These data suggested that ROS and RNS play distinct roles in the regulation of the membrane excitability of SG neurons related to the pain transmission.

Recent studies indicate that reactive oxygen species (ROS) can act as modulators of neuronal activity, and are critically involved in persistent pain primarily through spinal mechanisms. In this study, we investigated the effects of NaOCl, a ROS donor, on neuronal excitability and the intracellular calcium concentration ([Ca²⁺]_i) in spinal substantia gelatinosa (SG) neurons. In current clamp conditions, the application of NaOCl caused a membrane depolarization, which was inhibited by pretreatment with phenyl-N-tert-buthylnitrone (PBN), a ROS scavenger. The NaOCl-induced depolarization was not blocked however by pretreatment with dithiothreitol, a sulfhydrylreducing agent. Confocal scanning laser microscopy was used to confirm whether NaOCl increases the intracellular ROS level. ROS-induced fluorescence intensity was found to be increased during perfusion of NaOCl after the loading of 2′,7′-dichlorofluorescin diacetate (H₂DCF-DA). NaOCl-induced depolarization was not blocked by pretreatment with external Ca²⁺ free solution or by the addition of nifedifine. However, when slices were pretreated with the Ca²⁺ ATPase inhibitor thapsigargin, NaOCl failed to induce membrane depolarization. In a calcium imaging technique using the Ca²⁺-sensitive fluorescence dye fura-2, the [Ca²⁺]_i was found to be increased by NaOCl. These results indicate that NaOCl activates the excitability of SG neurons via the modulation of the intracellular calcium concentration, and suggest that ROS induces nociception through a central sensitization.

Recent studies indicate that reactive oxygen species (ROS) are critically involved in persistent pain primarily through spinal mechanisms, and that mitochondria are the main source of ROS in the spinal dorsal horn. To investigate whether mitochondrial ROS can induce changes in mem¬brane excitability on spinal substantia gelatonosa (SG) neurons, we examined the effects of mitochondrial electron transport complex (ETC) substrates and inhibitors on the membrane potential of SG neurons in spinal slices. Application of ETC inhibitors, rotenone or antimycin A, resulted in a slowly developing and slight membrane depolarization in SG neurons. Also, application of both malate, a complex I substrate, and succinate, a complex II substrate, caused reversible membrane depolarization and enhanced firing activity. Changes in membrane potential after malate exposure were more prominent than succinate exposure. When slices were pretreated with ROS scavengers such as phenyl-N-tert-buthylnitrone (PBN), catalase and 4-hydroxy-2,2,6,6-tetramethylpiperidine-1-oxyl (TEMPOL), malate-induced depolarization was significantly decreased. Intracellular calcium above 100 µM increased malate-induced depolarization, witch was suppressed by cyclosporin A, a mitochondrial permeability transition (MPT) inhibitor. These results suggest that enhanced production of spinal mitochondrial ROS can induce nociception through central sensitization.

The superficial dorsal horn, particularly substantia gelatinosa (SG) in the spinal cord, receives inputs from small-diameter primary afferents that predominantly convey noxious sensation. Reactive oxygen species (ROS) are toxic agents that may be involved in various neurodegenerative diseases. Recent studies indicate that ROS are also involved in persistent pain through a spinal mechanism. In the present study, whole cell patch clamp recordings were carried out on SG neurons in spinal cord slice of young rats to investigate the effects of hydrogen peroxide on neuronal excitability and excitatory synaptic transmission. In current clamp condition, tert-buthyl hydroperoxide (t-BuOOH), an ROS donor, depolarized membrane potential of SG neurons and increased the neuronal firing frequencies evoked by depolarizing current pulses. When slices were pretreated with phenyl-N-tert-buthylnitrone (PBN) or ascorbate, ROS scavengers, t-BuOOH did not induce hyperexcitability. In voltage clamp condition, t-BuOOH increased the frequency and amplitude of spontaneous excitatory postsynaptic currents (sEPSCs), and monosynaptically evoked excitatory postsynaptic currents (eEPSCs) by electrical stimulation of the ipsilateral dorsal root. These data suggest that ROS generated by peripheral nerve injury can modulate the excitability of the SG neurons via pre- and postsynaptic actions.

본 연구의 목적은 ``bi-directional(양방향성)`` 복합자극에 의한 대뇌 운동피질의 흥분성 조절의 효과를 살펴보는 것이다. 구체적으로 Pitcher 등(2003a)이 제안한 복합 자극이 정상인의 운동피질 흥분성에 미치는 효과와 Wherhahn 등(2002)의 말초 구심성 차단으로 대칭측 대뇌반구의 흥분성 억제가 동측 대뇌반구의 운동피질 흥분성을 촉진시킨다는 연구를 양측성 복합 자극 훈련에 적용하여 운동피질 흥분성에 미치는 효과를 검증하였다. 노인 10명과 성인 15명이 참여하였으며, 피험자들은 단측성 복합 자극을 받고 1주후 양측성 복합 자극을 수행하였다. 복합 자극은 말초신경자극(양측성: 저빈도 3Hz와 고빈도 30Hz; 단측성: 고빈도 30Hz)과 경두개자기자극기로 운동역치의 120% 강도로 대뇌에 직접 자극하였다. 그 결과, 성인과 노인집단 모두 단측성과 양측성 복합자극을 통해 운동유발전위 크기가 증가하였으며, 양측성 복합 자극이 단측성 복합 자극에 비해 운동유발전위 크기가 더 큰 것으로 나타났다. 이러한 연구결과를 통해 인간의 대뇌에 양방향성으로 조절 가능한 시냅스가 존재 할 수 있다는 가능성을 제시하고 있다.

본 연구는 정상 성인과 만성 뇌졸중 환자를 대상으로 하여 활동관찰, 운동심상, 그리고 거울을 활용한 시각적 착시가 운동피질 활성화에 어떠한 영향을 주는지 신경생리적 측면에서 검증하고자 하였다. 피험자는 정상 성인 10명, 만성 뇌졸중 환자 10명, 총 20명이 참여하여, 다섯 가지의 실험 조건(통제조건, 비활동조건, 활동조건, 심상조건, 거울조건)에서 오른손 엄지와 검지를 메트로놈 1 Hz 신호에 따라 반복적으로 움직이는 과제를 수행하였다. 실험 도구는 경두개자기자극기, 자극 코일, 근전도 시스템, 거울 등이 사용되었다. 실험 결과, 정상 성인과 뇌졸중 환자 모두 거울조건에서 운동유발전위가 가장 큰 것으로 나타났으며, 두 집단 간에는 차이가 없는 것으로 나타났다. 정상 성인의 경우에는 심상조건, 활동조건, 비활동조건 순으로 운동유발전위가 큰 것으로 나타났으며, 뇌졸중 환자의 경우 심상조건과 활동조건은 통계적으로 유의한 차이가 없었으며, 비활동조건에서 가장 낮은 운동유발전위가 발생하였다.

본 연구의 목적은 양측성 단일 운동 훈련이 만성 뇌졸중 노인의 상지 운동기능회복에 어떠한 영향을 주는지 살펴보고 손상측 대뇌 운동피질 활성화에 미치는 영향에 대해 검증하고자 하였다. 이러한 연구 목적을 달성하기 위해 총 10명의 만성 뇌졸중 노인들이 양측성 훈련과 단측성 훈련에 각각 5명씩 참여하였으며, 실험을 위해 동작분석 시스템, 경두개자기자극기, 근전도 기기, 재활훈련도구가 사용되었다. 훈련과제는 나무 말뚝 옮기기, 나무 팩 맞추기, 그리고 물 마시기 모방 과제로 구성하여 주 2회 총 16회의 훈련을 실시하였다. 그 결과, 양측성 훈련 집단은 기능적 뻗기 검사에서 최대속도가 증가하였으며, 운동시간이 감소한 것으로 나타났다. 기능적 검사에서도 씨앗 옮기기, 무거운 물건 옯기기 과제에서 수행시간이 감소한 것으로 나타났다. 또한 경두개자기자극기를 활용한 신경생리적 검사에서도 130%, 140%, 그리고 150% 자극 강도에서 운동유발전위의 크기가 증가한 것으로 나타났다. 이러한 결과를 통해 양측성 단일 운동 훈련이 손상측 상지의 운동기능회복과 대뇌 운동피질 활성화에 긍정적인 효과가 있는 것으로 알 수 있다.