• Title/Summary/Keyword: spinal cord slice

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Effects of Hydrogen Peroxide on Neuronal Excitability and Synaptic Transmission in Rat Substantia Gelatinosa Neurons

  • Son, Yong;Chun, Sang-Woo
    • International Journal of Oral Biology
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    • v.32 no.4
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    • pp.153-160
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    • 2007
  • 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.

Isolation and Electrical Characterization of the Rat Spinal Dorsal Horn Neurons (랫드 척수후각 단일세포 분리 및 특성에 관한 연구)

  • Han, Seong-Kyu;Ryu, Pan-Dong
    • The Korean Journal of Pharmacology
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    • v.32 no.2
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    • pp.283-292
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    • 1996
  • The spinal dorsal horn is the area where primary afferent fibers terminate and cutaneous sensory information is processed. A number of putative neurotransmitter substances, including excitatory and inhibitory amino acids and peptides, are present in this region. In this study, single neurons of the spinal dorsal horn were acutely isolated and the properties of whole cell current and responses to excitatory and inhibitory neurotransmitters were studied by patch clamp technique. Transverse slice ($(300{\mu}m$) of lumbar spinal cords from young rats$(7{\sim}14\;days)$ were sequentially treated with two pretenses(pronase 0.75 mg/ml and thermolysin 0.75 mg/ml), then single neurons were mechanically dissociated. These neurons showed near-intact morphology such as multipolar, ellipsoidal and bipolar, and pyramidal cells and we recorded the typical whole cell currents of $K^+$, $Ca^{2+}$ and ligand-operated channels from these neurons. Glutamate $(30{\mu}M)$ and N-methyl-D-aspartate(NMDA, $30{\mu}M)$ induced inward currents of $117{\pm}12.4$ pA(n=5) and $49{\pm}6.9$ pA(n=3), respectively. Glycine $(1{\mu}M)$ potentiated glutamate-induced currents $4{\sim}5$ times and NMDA-induced currents $8{\sim}10$ times. In addition, glycine $(30{\mu}M)$ induced Inward current ($31{\pm}6.1$ nA, n=2), which was rapidly desensitized after the peak to a new steady-state level. However, the inward currents induced by ${\gamma}-amino$ butyric acid(GABA, $1{\mu}M$) decreased continuously after the peak($226{\pm}41.6$ pA, n=3) under the similar experimental condition. The ionic currents and pharmacological responses of isolated neurons in this work were similar to those observed in vivo or in vitro spinal cord slice, indicating that acutely isolated neurons could be effectively used for further pharmacological studies.

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Roles of Reactive Oxygen Species on Neuronal Excitability in Rat Substantia Gelatinosa Neurons (척수 아교질 신경세포의 흥분성에 대한 활성산소종의 역할)

  • Choi, Jeong-Hee;Kim, Jae-Hyo;Lim, Sung-Jun;Park, Byung-Rim;Kwon, Kang-Beom
    • Journal of Physiology & Pathology in Korean Medicine
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    • v.21 no.2
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    • pp.432-437
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    • 2007
  • 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 substantia gelatinosa (SG) neurons in spinal cord slice of neonatal rats to investigate the effects of ROS on neuronal excitability and excitatory synaptic transmission. In current clamp condition, tert-buthyl hydroperoxide (t-BuOOH), an ROS donor, induced a electrical hyperexcitability during t-BuOOH wash-out followed by a brief inhibition of excitability in SG neurons. Application of t-BuOOH depolarized membrane potential of SG neurons and increased the neuronal firing frequencies evoked by depolarizing current pulses. Phenyl-N-tert-buthylnitrone (PBN), an ROS scavenger, antagonized t-BuOOH induced hyperexcitability. IN voltage clamp conditions, t-BuOOH increased the frequency and amplitude of spontaneous excitatory postsynaptic currents (sEPSCs). In order to determine the site of action of t-BuOOH, miniature excitatory postsynaptic currents (mEPSCs) were recorded. t-BuOOH increased the frequency and amplitude of mEPSCs, indicating that it may modulate the excitability of the SG neurons via pre- and postsynaptic actions. These data suggest that ROS generated by peripheral nerve injury can induce central sensitization in spinal cord.

DAMGO modulates two-pore domain K+ channels in the substantia gelatinosa neurons of rat spinal cord

  • Cho, Pyung Sun;Lee, Han Kyu;Lee, Sang Hoon;Im, Jay Zoon;Jung, Sung Jun
    • The Korean Journal of Physiology and Pharmacology
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    • v.20 no.5
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    • pp.525-531
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    • 2016
  • The analgesic mechanism of opioids is known to decrease the excitability of substantia gelatinosa (SG) neurons receiving the synaptic inputs from primary nociceptive afferent fiber by increasing inwardly rectifying $K^+$ current. In this study, we examined whether a ${\mu}$-opioid agonist, [D-Ala2,N-Me-Phe4, Gly5-ol]-enkephalin (DAMGO), affects the two-pore domain $K^+$ channel (K2P) current in rat SG neurons using a slice whole-cell patch clamp technique. Also we confirmed which subtypes of K2P channels were associated with DAMGO-induced currents, measuring the expression of K2P channel in whole spinal cord and SG region. DAMGO caused a robust hyperpolarization and outward current in the SG neurons, which developed almost instantaneously and did not show any time-dependent inactivation. Half of the SG neurons exhibited a linear I~V relationship of the DAMGO-induced current, whereas rest of the neurons displayed inward rectification. In SG neurons with a linear I~V relationship of DAMGO-induced current, the reversal potential was close to the $K^+$ equilibrium potentials. The mRNA expression of TWIK (tandem of pore domains in a weak inwardly rectifying $K^+$ channel) related acid-sensitive $K^+$ channel (TASK) 1 and 3 was found in the SG region and a low pH (6.4) significantly blocked the DAMGO-induced $K^+$ current. Taken together, the DAMGO-induced hyperpolarization at resting membrane potential and subsequent decrease in excitability of SG neurons can be carried by the two-pore domain $K^+$ channel (TASK1 and 3) in addition to inwardly rectifying $K^+$ channel.

Pre- and Postsynaptic Actions of Reactive Oxygen Species and Nitrogen Species in Spinal Substantia Gelatinosa Neurons

  • Park, Areum;Chun, Sang Woo
    • International Journal of Oral Biology
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    • v.43 no.4
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    • pp.209-216
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    • 2018
  • 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 $O_2{^{{\bullet}_-}}$ 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-DL-penicillamine (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.

Involvement of Crosstalk Between cAMP and cGMP in Synaptic Plasticity in the Substantia Gelatinosa Neurons

  • Kim, Tae-Hyung;Chung, Ge-Hoon;Park, Seok-Beom;Chey, Won-Young;Jun, Sung-Jun;Kim, Joong-Soo;Oh, Seog-Bae
    • International Journal of Oral Biology
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    • v.36 no.2
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    • pp.83-89
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    • 2011
  • Substantia gelatinosa (SG) neurons receive synaptic inputs from primary afferent $A{\delta}$- and C-fibers, where nociceptive information is integrated and modulated by numerous neurotransmitters or neuromodulators. A number of studies were dedicated to the molecular mechanism underlying the modulation of excitability or synaptic plasticity in SG neurons and revealed that second messengers, such as cAMP and cGMP, play an important role. Recently, cAMP and cGMP were shown to downregulate each other in heart muscle cells. However, involvement of the crosstalk between cAMP and cGMP in neurons is yet to be addressed. Therefore, we investigated whether interaction between cAMP and cGMP modulates synaptic plasticity in SG neurons using slice patchclamp recording from rats. Synaptic activity was measured by excitatory post-synaptic currents (EPSCs) elicited by stimulation onto dorsal root entry zone. Application of 1 mM of 8-bromoadenosine 3,5-cyclic monophosphate (8-Br-cAMP) or 8-bromoguanosine 3,5-cyclic monophosphate (8-Br-cGMP) for 15 minutes increased EPSCs, which were maintained for 30 minutes. However, simultaneous application of 8-BrcAMP and 8-Br-cGMP failed to increase EPSCs, which suggested antagonistic cross-talk between two second messengers. Application of 3-isobutyl-1-methylxanthine (IBMX) that prevents degradation of cAMP and cGMP by blocking phosphodiesterase (PDE) increased EPSCs. Co-application of cAMP/cGMP along with IBMX induced additional increase in EPSCs. These results suggest that second messengers, cAMP and cGMP, might contribute to development of chronic pain through the mutual regulation of the signal transduction.

The Usability Analysis of 3D-CRT, IMRT, Tomotherpy Radiation Therapy on Nasopharyngeal Cancer (NPC의 방사선치료시 3D-CRT, IMRT, Tomotherapy의 유용성 분석)

  • Song, Jong-Nam;Kim, Young-Jae;Hong, Seung-Il
    • Journal of the Korean Society of Radiology
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    • v.6 no.5
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    • pp.365-371
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    • 2012
  • The radiation therapy treatment technique is developed from 3D-CRT, IMRT to Tomotherapy. and these three technique was most widely using methods. We find out a comparison normal tissue doses and tumor dose of 3D-CRT, IMRT(Linac Based), and Tomotherapy on Head and Neck Cancer. We achieved radiological image used the Human model phantom (Anthropomorphic Phantom) and it was taken CT simulation (Slice Thickness : 3mm) and GTV was nasopharngeal region and PTV(including set-up margin) was GTV plus 2mm area. and transfer those images to the radiation planning system (3D-CRT - ADAC-Pinnacle3, Tomotherapy - Tomotherapy Hi-Art System). The prescription dose was 7020 cGy and measuring PTV's dose and nomal tissue (parotid gland, oral cavity, spinal cord). The PTV's doses was Tomotherapy, Linac Based - IMRT, 3D-CRT was 6923 cGy, 6901 cGy and 6718 cGy its dose value was meet TCP because its value was up to the 95% based on 7020 cGy, Nomal tissue (parotid gland, oral cavity, spinal cord) was 1966 cGy(Tomotherapy), 2405 cGy(IMRT), 2468 cGy(3D-CRT)[parotid gland], 2991 cGy(Tomotherapy), 3062 cGy(IMRT), 3684 cGy (3D-CRT)[oral cavity], 1768 cGy(Tomotherapy), 2151 cGy(IMRT), 4031 cGy(3D-CRT)[spinal cord] its value did not exceeded NTCP. All the treatment techniques are equated with tumor and nomal tissue doses. The 3D-CRT was worse than other techniques on dose distribution, but it is reasonable in terms of TCP and NTCP baseline Tomotherapy, IMRT -dose distribution was relatively superior- was hard to therapy to claustrophobic patients and patients with respiratory failure. Particularly, in case on Tomotherapy, it take MVCT before treatment so dose measurement will be unnecessary radiation exposure to patients. Conclusion, Tomotherapy was the best treatment technique and 2nd was IMRT, and 3rd 3D-CRT. But applicable differently depending on the the patient's condition even though dose not matter.

Mechanism of Glutamate-induced $[Ca^{2+}]i$ Increase in Substantia Gelatinosa Neurons of Juvenile Rats

  • Jung, Sung-Jun;Choi, Jeong-Sook;Kwak, Ji-Yeon;Kim, Jun;Kim, Jong-Whan;Kim, Sang-Jeong
    • The Korean Journal of Physiology and Pharmacology
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    • v.7 no.2
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    • pp.53-57
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    • 2003
  • The glutamate receptors (GluRs) are key receptors for modulatory synaptic events in the central nervous system. It has been reported that glutamate increases the intracellular $Ca^{2+}$ concentration ($[Ca^{2+}]_i$) and induces cytotoxicity. In the present study, we investigated whether the glutamate-induced $[Ca^{2+}]_i$ increase was associated with the activation of ionotropic (iGluR) and metabotropic GluRs (mGluR) in substantia gelatinosa neurons, using spinal cord slice of juvenile rats (10${\sim}21 day). $[Ca^{2+}]_i$ was measured using conventional imaging techniques, which was combined with whole-cell patch clamp recording by incorporating fura-2 in the patch pipette. At physiological concentration of extracellular $Ca^{2+}$, the inward current and $[Ca^{2+}]_i$ increase were induced by membrane depolarization and application of glutamate. Dose-response relationship with glutamate was observed in both $Ca^{2+}$ signal and inward current. The glutamate-induced $[Ca^{2+}]_i$ increase at holding potential of -70 mV was blocked by CNQX, an AMPA receptor blocker, but not by AP-5, a NMDA receptor blocker. The glutamate-induced $[Ca^{2+}]_i$ increase in $Ca^{2+}$ free condition was not affected by iGluR blockers. A selective mGluR (group I) agonist, RS-3,5-dihydroxyphenylglycine (DHPG), induced $[Ca^{2+}]_i$ increase at holding potential of -70 mV in SG neurons. These findings suggest that the glutamate-induced $[Ca^{2+}]_i$ increase is associated with AMPA-sensitive iGluR and group I mGluR in SG neurons of rats.

A Study of Energy Dependency in Intensity Modulated Radiation Therapy of Lung Cancer (폐암환자의 세기조절방사선치료에서 에너지에 따른 선량분포 특성 비교)

  • Kim, Sung-Kyu;Kim, Myung-Se;Yun, Sang-Mo
    • Progress in Medical Physics
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    • v.19 no.3
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    • pp.191-199
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    • 2008
  • PTV considered for the energy, dose distribution exposed to lung and spinal cord, and the characteristic of DVH(Dose Volume Histogram) were compared and investigated by planning the intensity modulated radiation therapy (IMRT) using the photon energies of 6 MV and 10 MV according to tumor location like as the anterior, middle, and posterior regions of lung, and the mediastinum region in lung cancer patients. Our institution installed the linear accelerator (Varian 21 EX-s, USA) equipped with 120 multileaf collimator for lung cancer patients, which is producing the photon energies of 6 MV and 10 MV, and radiation therapy planning was performed with ECLIPSE system (Varian, SomaVision 6.5, USA), which support inverse treatment planning. The tomographic images of 3 mm slice thickness for lung cancer patients were acquired using planning CT, and acquired tomographic images were sent to the Varis system, and then treatment planning was performed in the ECLIPSE system. The radiation treatment planning of the IMRT was processed from various angles according to the regions of the tumor, and using various beam lines according to the size and location of the tumor. The investigation of the characteristic of dose distributions for the energy of 6 MV and 10 MV according to tumor locations in lung cancer patients resulted that the maximum dose of 10 MV energy was 1.2% less than that of 6 MV energy without depending on the tumor location of lung cancer, and the reduction effects of MU were occurred from 10 to 25 MU. Radiation dose exposed to the lung satisfied the less 30% of V20, however radiation dose in 6 MV energy was from 0.1% to 0.5% less than that in 10 MV energy. Radiation dose exposed to the spinal cord for 6 MV energy was from 0.6% to 2.1% less than that for 6 MV energy.

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THE CURRENT STATUS OF BIOMEDICAL ENGINEERING IN THE USA

  • Webster, John G.
    • Proceedings of the KOSOMBE Conference
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    • v.1992 no.05
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    • pp.27-47
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    • 1992
  • Engineers have developed new instruments that aid in diagnosis and therapy Ultrasonic imaging has provided a nondamaging method of imaging internal organs. A complex transducer emits ultrasonic waves at many angles and reconstructs a map of internal anatomy and also velocities of blood in vessels. Fast computed tomography permits reconstruction of the 3-dimensional anatomy and perfusion of the heart at 20-Hz rates. Positron emission tomography uses certain isotopes that produce positrons that react with electrons to simultaneously emit two gamma rays in opposite directions. It locates the region of origin by using a ring of discrete scintillation detectors, each in electronic coincidence with an opposing detector. In magnetic resonance imaging, the patient is placed in a very strong magnetic field. The precessing of the hydrogen atoms is perturbed by an interrogating field to yield two-dimensional images of soft tissue having exceptional clarity. As an alternative to radiology image processing, film archiving, and retrieval, picture archiving and communication systems (PACS) are being implemented. Images from computed radiography, magnetic resonance imaging (MRI), nuclear medicine, and ultrasound are digitized, transmitted, and stored in computers for retrieval at distributed work stations. In electrical impedance tomography, electrodes are placed around the thorax. 50-kHz current is injected between two electrodes and voltages are measured on all other electrodes. A computer processes the data to yield an image of the resistivity of a 2-dimensional slice of the thorax. During fetal monitoring, a corkscrew electrode is screwed into the fetal scalp to measure the fetal electrocardiogram. Correlations with uterine contractions yield information on the status of the fetus during delivery To measure cardiac output by thermodilution, cold saline is injected into the right atrium. A thermistor in the right pulmonary artery yields temperature measurements, from which we can calculate cardiac output. In impedance cardiography, we measure the changes in electrical impedance as the heart ejects blood into the arteries. Motion artifacts are large, so signal averaging is useful during monitoring. An intraarterial blood gas monitoring system permits monitoring in real time. Light is sent down optical fibers inserted into the radial artery, where it is absorbed by dyes, which reemit the light at a different wavelength. The emitted light travels up optical fibers where an external instrument determines O2, CO2, and pH. Therapeutic devices include the electrosurgical unit. A high-frequency electric arc is drawn between the knife and the tissue. The arc cuts and the heat coagulates, thus preventing blood loss. Hyperthermia has demonstrated antitumor effects in patients in whom all conventional modes of therapy have failed. Methods of raising tumor temperature include focused ultrasound, radio-frequency power through needles, or microwaves. When the heart stops pumping, we use the defibrillator to restore normal pumping. A brief, high-current pulse through the heart synchronizes all cardiac fibers to restore normal rhythm. When the cardiac rhythm is too slow, we implant the cardiac pacemaker. An electrode within the heart stimulates the cardiac muscle to contract at the normal rate. When the cardiac valves are narrowed or leak, we implant an artificial valve. Silicone rubber and Teflon are used for biocompatibility. Artificial hearts powered by pneumatic hoses have been implanted in humans. However, the quality of life gradually degrades, and death ensues. When kidney stones develop, lithotripsy is used. A spark creates a pressure wave, which is focused on the stone and fragments it. The pieces pass out normally. When kidneys fail, the blood is cleansed during hemodialysis. Urea passes through a porous membrane to a dialysate bath to lower its concentration in the blood. The blind are able to read by scanning the Optacon with their fingertips. A camera scans letters and converts them to an array of vibrating pins. The deaf are able to hear using a cochlear implant. A microphone detects sound and divides it into frequency bands. 22 electrodes within the cochlea stimulate the acoustic the acoustic nerve to provide sound patterns. For those who have lost muscle function in the limbs, researchers are implanting electrodes to stimulate the muscle. Sensors in the legs and arms feed back signals to a computer that coordinates the stimulators to provide limb motion. For those with high spinal cord injury, a puff and sip switch can control a computer and permit the disabled person operate the computer and communicate with the outside world.

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