• Applied Microscopy
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• v.34 no.4
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• pp.285-294
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• 2004

We investigated the morphology, distribution and synaptic connectivity of cholinergic neurons in the mouse retina by immunocytochemistry, using antisera against choline acetyltransferase (ChAT). ChAT-immunoreactive amacrine cells fall into two groups according to the localization of their somas in the retina: one is situated in the inner nuclear layer (INL), near the border of the inner plexiform layer (IPL), and the other is displaced in the ganglion cell layer (GCL). The dendrites of amacrine cells from the INL ramify in sublamina a and that of the displaced amacrine cells ramify in sublamina b of the IPL. Double labeling with an antisera against ChAT and r-aminobutyric acid (GABA) demonstrated that these labeled cells formed a subpopulation of GABAergic amacrine cells. The synaptic connectivity of ChAT-immunoreactive amacrine cells was identified in the IPL by electron microscopy. The most frequent synaptic input of ChAT-labeled amacrine cells was from bipolar cells in both sublaminae a and b of the IPL, followed by labeled amacrine cells and unlabeled amacrine cells. Their primary output targets were onto ganglion cells in both sublaminae a and b and output onto ganglion cells was more frequently observed in sublamina b of the IPL. Our results suggest that cholinergic amacrine cells in the mouse retina are very similar to their counter parts in other mammals, and they can attribute a major role in the pathway feeding into directionally selective ganglion cells.

• The Korean Journal of Physiology and Pharmacology
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• v.22 no.2
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• pp.173-182
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• 2018

Recent studies have provided several lines of evidence that peripheral administration of oxytocin induces analgesia in human and rodents. However, the exact underlying mechanism of analgesia still remains elusive. In the present study, we aimed to identify which receptor could mediate the analgesic effect of intraperitoneal injection of oxytocin and its cellular mechanisms in thermal pain behavior. We found that oxytocin-induced analgesia could be reversed by $d(CH_2)_5[Tyr(Me)^2,Dab^5]$ AVP, a vasopressin-1a (V1a) receptor antagonist, but not by $desGly-NH_2-d(CH_2)_5[D-Tyr^2,Thr^4]OVT$, an oxytocin receptor antagonist. Single cell RT-PCR analysis revealed that V1a receptor, compared to oxytocin, vasopressin-1b and vasopressin-2 receptors, was more profoundly expressed in dorsal root ganglion (DRG) neurons and the expression of V1a receptor was predominant in transient receptor potential vanilloid 1 (TRPV1)-expressing DRG neurons. Fura-2 based calcium imaging experiments showed that capsaicin-induced calcium transient was significantly inhibited by oxytocin and that such inhibition was reversed by V1a receptor antagonist. Additionally, whole cell patch clamp recording demonstrated that oxytocin significantly increased potassium conductance via V1a receptor in DRG neurons. Taken together, our findings suggest that analgesic effects produced by peripheral administration of oxytocin were attributable to the activation of V1a receptor, resulting in reduction of TRPV1 activity and enhancement of potassium conductance in DRG neurons.

• Proceedings of the Korean Institute of Information and Commucation Sciences Conference
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• 2014.10a
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• pp.989-992
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• 2014

Myelination using Schwann cells and neuron cells was performed in rat. Schwann cells and neuron cells from dorsal root ganglion (DRG) of rat embryos (E16) were cultured, respectively. The embryonic DRG cells purified were cultured and anti-mitotic agents were added. Purified the embryonic Schwann cells were cultured and added to the embryonic DRG cells purified. A purified population of myelination in vitro system was accomplished and identified formation of myelination using antibody of neurofilament protein.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• v.30 no.1
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• pp.1-16
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• 2004

The use of artificial nerve conduit containing viable Schwann cells is one of the most promising strategies to repair the peripheral nerve injury. To fabricate an effective nerve conduit whose microstructure and internal environment are more favorable in the nerve regeneration than existing ones, a new three-dimensional Schwann cell culture technique using $Matrigel^{(R)}$. and dorsal root ganglion (DRG) was developed. Nerve conduit of three-dimensionally arranged Schwann cells was fabricated using direct seeding of freshly harvested DRG into a $Matrigel^{(R)}$ filled silicone tube (I.D. 1.98 mm, 14 mm length) and in vitro rafting culture for 2 weeks. The nerve regeneration efficacy of three-dimensionally cultured Schwann cell conduit (3D conduit group, n=6) was assessed using SD rat sciatic nerve defect of 10 mm, and compared with that of silicone conduit filled with $Matrigel^{(R)}$ and Schwann cells prepared from the conventional plain culture method (2D conduit group, n=6). After 12 weeks, sciatic function was evaluated with sciatic function index (SFI) and gait analysis, and histomorphology of nerve conduit and the innervated tissues of sciatic nerve were examined using image analyzer and electromicroscopic methods. The SFI and ankle stance angle (ASA) in the functional evaluation were $-60.1{\pm}13.9$, $37.9^{\circ}{\pm}5.4^{\circ}$ in 3D conduit group (n=5) and $-87.0{\pm}12.9$, $32.2^{\circ}{\pm}4.8^{\circ}$ in 2D conduit group (n=4), respectively. And the myelinated axon was $44.91%{\pm}0.13%$ in 3D conduit group and $13.05%{\pm}1.95%$ in 2D conduit group to the sham group. In the TEM study, 3D conduit group showed more abundant myelinated nerve fibers with well organized and thickened extracellular collagen than 2D conduit group, and gastrocnemius muscle and biceps femoris tendon in 3D conduit group were less atrophied and showed decreased fibrosis with less fatty infiltration than 2D conduit group. In conclusion, new three-dimensional Schwann cell culture technique was established, and nerve conduit fabricated using this technique showed much improved nerve regeneration capacity than the silicone tube filled with $Matrigel^{(R)}$ and Schwann cells prepared from the conventional plain culture method.

• Journal of Veterinary Clinics
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• v.24 no.4
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• pp.514-521
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• 2007

The sympathetic nerve block improves the blood flow in the innervated regions. For this region, the sympathetic nerve block has been performed in the neural and cerebral disorders. However, the cerebral blood flow regulation of the cranial cervical ganglion block in dogs have not been well defined and the correlation to the changes in the cerebral circulation and the changes in the electroencephalogram is not well defined in dogs yet. Therefore, we investigated the hypothesis that changes in the EEG could be affected by the changes in cerebral blood flow following the cranial cervical ganglion block in dogs. Twenty five beagle dogs were divided into 3 groups; group I(LCCGB, n=10) underwent left sided cranial cervical ganglion block using the 1% lidocaine, group II(L, n=10) injected the 1% lidocaine into the right or left sided digastricus muscle, group III(N/SCCGB, n=5, served as control) underwent the left sided cranial cervical ganglion block using saline. A statistical difference was not found between the control group and the LCCGB group in the 95% spectral edge frequency(SEF) and the median frequency(MF). In the relative band power, the $\delta$ frequency was decreased during 5-25 min, while the $\alpha$ frequency was increased during the same time(p<0.05). But the $\theta$ frequency and the $\beta$ frequency were not shown the significant changes compared with the control group during the same time(p<0.05). These results suggest that the left cranial cervical ganglion block does not induce the change of the cerebral blood flow and its effect is insignificant.

• Journal of Life Science
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• v.19 no.10
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• pp.1346-1351
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• 2009

Recent evidence has shown that many pluripotetic neural crest cells are fate-restricted and that different fate-restricted crest cells emigrate from the neural tube at different times. Jin et al. (2001) identified the expression patterns of Wnts and its antagonists at the time that neural crest cells were being specified and suggested that Wnt signaling was involved in the segregation/differentiation of neural crest cells in the trunk in vitro. In this study, we evaluated the effects of Wnt signaling in avian neural crest lineage segregation. To accomplish this, Wnt signaling was disturbed at the time of neural crest segregation and differentiation by grafting Wnt-3a expressing cells and conducting dominant negative glycogen synthase kinase (dnGSK) electroporation. Stimulation of Wnt signaling induced neural crest lineage segregation and melanoblast specification, and increased the expression levels of genes known to be involved in neural crest development such as cadherin 7 and Slug, which suggests that they are involved in Wnt-induced neural crest lineage differentiation into melanoblasts.

• Advances in pediatric surgery
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• v.7 no.1
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• pp.15-20
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• 2001

The pathophysiology of Hirschsprung's disease (HD) is not fully understood, but recent studies have disclosed that neural cell adhesion molecule (NCAM) and glial cell line-derived neurotrophic factor (GDNF) play important roles in the formation of aganglionic bowel of Hirschsprung's disease. To evaluate the roles of NCAM and GDNF in HD, immunohistochemical analysis was performed using formalin-fixed and paraffin-embedded tissue sections. On the basis of the results, we tried to evaluate them as diagnostic markers. The specimens were obtained from 7 patients with HD who underwent modified Duhamel operation. The diagnosis was based on the clinical findings and the absence of ganglion cells in the nerve plexuses by routine microscopy. NCAM immunoreactivity was found in the nerve plexuses and scattered nerve fibers in the smooth muscle layers of ganglionic segments. In aganglionic segments, the number of NCAM positive nerve fibers in the smooth muscle layers was significantly reduced compared with ganglionic segments. In two cases the nerve plexuses in aganglionic segments, NCAM was negligible. The smooth muscle cells showed diffuse immunoreactivity for GDNF and the staining intensity was not different in the aganglionic and ganglionic segments. However, higher expression of GDNF in the nerve plexus of the ganglionic segments was noted comparing to aganglionic segments. These data suggest that both NCAM and GDNF may play important roles in pathogenesis of Hirschsprung's disease and immunohistochemical staining for NCAM can be used as an ancillary diagnostic tool for HD.

• Applied Microscopy
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• v.30 no.2
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• pp.121-139
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• 2000

The morphogenesis of neuroblasts and plexiform layers, and establishment of its synapses were studied by electron microscopy in human embryos and fetuses ranging from 10 mm to 260 mm crown-rump length ($5\sim30$ weeks of gestational age). At 30 mm fetus the developing retina was composed of outer and inner neuroblastic layers . Cell division of outer neuroblast was occurred until 90 mm fetus. The transient layer of Chievitz was formed by 30 mm fetus, inner plexiform layer by 50 mm fetus, and outer plexiform layer by 150 mm fetus. The cytoplasm of differentiating ganglion cells contained ribosomes, rough endoplasmic reticula, Golgi complexes, microtubules and dense bodies. The processes of $M\ddot{u}ller$ cell penetrated between groups of ganglion cell axons, and formed the cellular component of the inner limiting membrane at 30 mm fetus. At 90 mm fetus radial fibers of M ller cells contained extensive smooth endoplasmic reticula and microtubules. In each specimen , apposing paired membrane specializations were classified as junctions without synaptic vesicles, conventional synapses and ribbon synapses. At 50 mm fetus the processes of neuroblasts in inner plexiform layer were interconnected by junctions without synaptic vesicles. Conventional synapses developed by addition of synaptic vesicles to initially vesicle-free junctions at 90 mm fetus. At 150 mm fetus ribbon synapses were first recognized by the inclusion of a prominent electron-dense material associated with synaptic vesicles. By 260 mm fetus conventional and ribbon synapses and junctions without synaptic vesicles formed similar to those found in the adult.

• The Korean Journal of Physiology and Pharmacology
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• v.2 no.2
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• pp.165-172
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• 1998

Under certain pathophysiological conditions, such as inflammation and ischemia, the concentration of H^+$ ion in the tissue surrounding neurons is changed. Variations in H^+$ concentration are known to alter the conduction and/of the gating properties of several types of ion channels. Several types of K^+$ channels are modulated by pH. In this study, the whole cell configuration of the patch clamp technique has been applied to the recording of the responses of change of external pH on the delayed rectifier K^+$ current of cultured DRG neurons of rat. Outward K^+$ currents were examined in DRG cells, and the Charybdotoxin and Mn^{2+}$ could eliminate Ca^{2+}-dependent$ K^+$ currents from outward K^+$ currents. This outward K^+$ current was activated around -60 mV by step depolarizing pulses from holding potential -70 mV. Outward K^+$ currents were decreased by low external pH. Activation and steady-state inactivation curve were shifted to the right by acidification, while there was small change by alkalization. These results suggest that H^+$ could be alter the sensory modality by changing and modifying voltage-dependent K^+$ currents, which participated in repolarization.

• Biomedical Science Letters
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• v.13 no.2
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• pp.119-125
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• 2007

It is widely known that protein tyrosine kinases (PTKs) are involved in controlling many biological processes such as cell growth, differentiation, proliferation, survival and apoptosis. An $\alpha3\beta4$ subunit combination acts as a major functional acetylcholine receptor (nAChRs) in male rat major pelvic ganglion (MPG) neurons, and their activation induces fast inward currents and intracellular calcium increases. Recently it has been reported that the activity of acetylcholine receptors (AChRs) in some neurons can be negatively regulated by PTKs. However, the exact mechanism of regulation of nAChRs by PTKs is poorly understood. Therefore, we examined the potential role particular in nAChR by PTK using electrophysiology and calcium imaging in male rat MPG neurons. ACh induced inward currents and $(Ca^{2+})_i$ increases in MPG neurons, concomitantly. These responses were inhibited by more than 90% in $Na^+$- or $Ca^{2+}$- free solution. $\alpha$-conotoxin AuIB, a selective $\alpha3\beta4$ nAChR blocket, inhibited ACh-induced inward currents. Genistein (10 $\mu$M), a broad-spectrum tyrosine kinase inhibitor, markedly decreased ACh-induced currents and $Ca^{2+}$ transients, whereas 10 $\mu$M genistin, an inactive analogue, had little effect. Overall these data suggest that the activities of $\alpha3\beta4$ AChRs in MPG neurons are positively regulated by PTK. In conclusion, trosine kinase may be one of the key factors in the regulation of $\alpha3\beta4$ nAChRs in rat MPG neurons, which may play an important roles in the autonomic neuronal function such as synaptic transmission, autonomic reflex, and neuronal plasticity.