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.

Reactive oxygen species (ROS) and nitrogen species (RNS) are involved in cellular signaling processes as a cause of oxidative stress. According to recent studies, ROS and RNS are important signaling molecules involved in pain transmission through spinal mechanisms. In this study, a patch clamp recording was used in spinal slices of rats to investigate the action mechanisms of O2 ⦁- and NO on the excitability of substantia gelatinosa (SG) neuron. The application of xanthine and xanthine oxidase (X/XO) compound, a ROS donor, induced inward currents and increased the frequency of spontaneous excitatory postsynaptic currents (sEPSC) in slice preparation. The application of S-nitroso-N-acetyl-DLpenicillamine (SNAP), a RNS donor, also induced inward currents and increased the frequency of sEPSC. In a single cell preparation, X/XO and SNAP had no effect on the inward currents, revealing the involvement of presynaptic action. X/XO and SNAP induced a membrane depolarization in current clamp conditions which was significantly decreased by the addition of thapsigargin to an external calcium free solution for blocking synaptic transmission. Furthermore, X/XO and SNAP increased the frequency of action potentials evoked by depolarizing current pulses, suggesting the involvement of postsynaptic action. According to these results, it was estblished that elevated ROS and RNS in the spinal cord can sensitize the dorsal horn neurons via pre- and postsynaptic mechanisms. Therefore, ROS and RNS play similar roles in the regulation of the membrane excitability of SG neurons.

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.

Growing evidence suggests that mitochondrial reactive oxygen species (ROS) are involved in various pain states. This study was performed to investigate whether ROS-induced changes in neuronal excitability in trigeminal subnucleus caudalis are related to ROS generation in mitochondria. Confocal scanning laser microscopy was used to measure ROS-induced fluorescence intensity in live rat trigeminal caudalis slices. The ROS level increased during the perfusion of malate, a mitochondrial substrate, after loading of 2′,7′-dichlorofluorescin diacetate (H2DCF-DA), an indicator of the intracellular ROS; the ROS level recovered to the control condition after washout. When pre-treated with phenyl N-tert-butylnitrone (PBN) and 4-hydroxy-2,2,6,6-tetramethylpiperidene-1-oxyl (TEMPOL), malate-induced increase of ROS level was suppressed. To identify the direct relation between elevated ROS levels and mitochondria, we applied the malate after double-loading of H2DCF-DA and chloromethyl-X-rosamine (CMXRos; MitoTracker Red), which is a mitochondria- specific fluorescent probe. As a result, increase of both intracellular ROS and mitochondrial ROS were observed simultaneously. This study demonstrated that elevated ROS in trigeminal subnucleus caudalis neuron can be induced through mitochondrial-ROS pathway, primarily by the leakage of ROS from the mitochondrial electron transport chain.

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.

Using whole cell current- and voltage-clamp recording we investigated the characteristics and pharmacology of group I metabotropic glutamate receptor (mGluR)-mediated responses in rat medial vestibular nucleus (MVN) neurons. In current clamp conditions, activation of mGluR I by application of the group I mGluR agonist (R,S)-3,5-dihydroxyphenylglycine (DHPG) induced a direct excitation of MVN neurons that is characterized by depolarization and increased spontaneous firing frequency. To identify which of mGluR subtypes are responsible for the various actions of DHPG in MVN, we used two subtype-selective antagonists. (S)-(+)- alpha-amino-a-methylbenzeneacetic acid (LY367385) is a potent competitive antagonist that is selective for mGluR1, whereas 2-methyl-6-(phenylethynyl)-pyridine (MPEP) is a potent noncompetitive antagonist that is selective for mGluR5. In voltage clamp conditions, DHPG application increased the frequency of spontaneous and miniature inhibitory postsynaptic currents (IPSCs) but had no effect on amplitude distributions. Antagonism of the DHPG-induced increase of miniature IPSCs required the blockade of both mGluR1 and mGluR5. DHPG application induced an inward current, which can be enhanced under depolarized conditions. DHPG-induced current was blocked by LY367385, but not by MPEP. Both LY367385 and MPEP antagonized the DHPG-induced suppression of the calcium activated potassium current (IAHP). These data suggest that mGluR1 and mGluR5 have similar roles in the regulation of the excitability of MVN neurons, and show a little distinct. Furthermore, mGluR I, via pre- and postsynaptic actions, have the potential to modulate the functions of the MVN.

Recent studies have implicated reactive oxygen species (ROS) as determinants of the pathological pain caused by the activation of peripheral neurons. It has not been elucidated, however, how ROS activate the primary sensory neurons in the pain pathway. In this study, calcium imaging was performed to investigate the effects of NaOCl, a ROS donor, on the intracellular calcium concentration ([Cα2+]i) in acutely dissociated dorsal root ganglion (DRG) neurons. DRG was sequentially treated with 0.2 mg/ml of both protease and thermolysin, and single neurons were then obtained by mechanical dissociation. The administration of NaOCl then caused a reversible increase in the [Cα2+]i], which was inhibited by pretreatment with phenyl-N-tertbuthylnitrone (PBN) and isoascorbate, both ROS scavengers. The NaOCl-induced [Cα2+]i] increase was suppressed both in a calcium free solution and after depletion of the intracellular Cα2+ pool by thapsigargin. Additionally, this increase was predominantly blocked by pretreatment with the transient receptor potential (TRP) antagonists, ruthenium red (50 μM) and capsazepine (10 μM). Collectively, these results suggest that an increase in the intracellular calcium concentration is produced from both extracellular fluid and the intracellular calcium store, and that TRP might be involved in the sensation of pain induced by ROS.

Background : Insect pests damages are increasing on the field of Lycium chinense under eco-friendly organic management and especially, pest damage of Gelechiidae (Ilseopsis parki Povolny) is serious. Currently, various eco-friendly organic materials are registered, but insecticidal activity against Gelechiidae was not verified. Therefore, this experiment was carried out to select eco-friendly organic materials showing insecticidal activity in Gelechiidae. Methods and Results : To test insecticidal activity, the Gelechiidae lavas were collected from the field and Nine eco-friendly organic materials were collected from the market. method. The insecticidal activity assay method was, putting Gelechiidae lavas on the 90mm petridish then water-diluted eco-friendly organic materials were sprayed onto the laver. Result of insecticidal activity was observed with a microscope 10 minutes, 2 hours and 18 hours after the treatment. Microscopic observation results, insecticidal activity was shown only in 5 out of 9 materials. Conclusion : As a result, Insecticidal activity against Gelechiidae was not effective in all tested materials, but the 5 eco-friendly organic materials showed the same effect in repeated experiments. In this study, we selected 5 materials out of 9 eco-friendly organic materials against Gelechiidae, then future studies are planned to select the most suitable materials by field testing the 5 eco-friendly organic materials.

Background : Black Non-Woven Fabric Mulch Culture was knowned increased crop Yield and saved weeding labor in Liriope Pilatyphylla Wang et Tang. But to the removal and planting labor is more needed, So some famers are avoidance that culting method. Methods and Results : So this study was experimented in order to selecting optimun removal time in Liriope Pilatyphylla Wang et Tang mulch culture. Removal time were conventional practices (in April next yesr), September, October and November. In early, Plant length, Root length, Leaf number and number of plants was the long and many by the sooner removal time and also, dry weight was heavier. Black non-woven fabric removal labor was saved by the sooner removal time. The main event of weeds were Cyperus serotinus Rottb and Portulaca oleracea L. In harvest time, Plant length and plants of numbers was the longest and heavier at conventional practice (in April next year) and November removed. Tuberous root number was the more in September removal, Because, the tuber was tall and long. Total1y consideration of the including weeds shooting, weeding labor and Growth and development situation, Black non-woven fabric removal optimum time was September or Conventional practices (in April next year). Conclusion : Black non-woven fabric optimum removal time was the september . In this experiment, increased yield 9, income 16 percent than conventional practices (in April next year).

Background : Natural Mortality Vinyl much culture are increased crop yield and saved weeding workforce. But research is insufficient that of Liriope Pilatyphylla Wang et Tang. Black non-woven fabric mulch culture are increased crop yield and superior to occurrence of weeds, but to the removal and planting more labor needed, so, Farmers are avoidance using that. Methods and Results : So this study designed in order to selecting the best covering material. Using in this study, covering materials were Conventional practices (non covering), Natural mortality vinyl and black non-woven fabric. Soil temperature was continued highly after planting from in mid-may to in august a regular. among them natural mortality vinyl mulch was the highest. Natural mortality vinyl mulch was the fastest that humidity of soil change and Growth and development situation by maintaining proper temperature and humidity. Non mulching was little change in temperature and humidity and the slowest in the Growth and development situation. The main event of weeds were Cyperus serotinus Rottb and Portulaca oleracea L. Weeding labor was saved from 43 to 57 percent in the natural mortality vinyl and black non-woven fabric mulch. Plant length was the shortest in the conventional practices and Root length, Leaf number and number of plants were little changed. Yield was increased from 27 to 29 percent in the natural mortality vinyl and black non-woven fabric mulch than conventional practices. Result of comprehensive economic analysis including weeding labor and yield, the natural mortality vinyl mulch culture was income increased 92 percent than conventional practices. Conclusion : Natural mortality vinyl mulch culture were weeds shooting controled and increased yield 39, income 92 percent than conventional practices (non covering).

Munjangbyeo' is a japonica rice cultivar developed from a cross between Sangsanbyeo and Suweon 397 by Sangju Substation of National Yeongnam Agricultural Experiment Station, RDA in 1999. The cultivar is early matured with heading date of Aug. 2 in ordinar

Sangmibyeo' is a japonica rice cultivar developed from a cross between Sambaegbyeo and Ou 316 by Sangju Substation of National Yeongnam Agricultural Experiment Station, R.D.A. in 1998. The cultivar is early maturing with heading date of August 7 in ordina