• The Korean Journal of Physiology and Pharmacology
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• v.13 no.5
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• pp.373-378
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• 2009

Cerebellar Purkinje cells (PCs) play a crucial role in motor functions and their progressive degeneration is closely associated with spinocerebellar ataxias. Although immunohistochemical (IHC) analysis can provide a valuable tool for understanding the pathophysiology of PC disorders, the method validation of IHC analysis with cerebellar tissue specimens is unclear. Here we present an optimized and validated IHC method using antibodies to calbindin D28k, a specific PC marker in the cerebellum. To achieve the desired sensitivity, specificity, and reproducibility, we modified IHC analysis procedures for cerebellar tissues. We found that the sensitivity of staining varies depending on the commercial source of primary antibody. In addition, we showed that a biotin-free signal amplification method using a horseradish peroxidase polymer-conjugated secondary antibody increases both the sensitivity and specificity of ICH analysis. Furthermore, we demonstrated that dye filtration using a $0.22\;{\mu}m$ filter eliminates or minimizes nonspecific staining while preserving the analytical sensitivity. These results suggest that our protocol can be adapted for future investigations aiming to understand the pathophysiology of cerebellar PC disorders and to evaluate the efficacy of therapeutic strategies for treating' these diseases.

• Journal of Life Science
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• v.15 no.4 s.71
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• pp.607-612
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• 2005

Heat shock protein 70 (HSP70) is a multigene family composed of constitutively expressed members(Hsc70) and stress-inducible members (Hsp70). In the mammalian nervous system, a considerable amount of HSPs is also synthesized under normal conditions suggesting that they play an important role in the metabolism of unstressed cells. In this study we examined the expression of Hsp70 in the synapses of rat cerebellar neurons. Immunohistochemistry using specific antibodies revealed that both Hsp70 and Hsc70 are expressed in the cerebellar tissue, with strongest expression in Purkinje cells followed by granule cells. Neurons in deep cerebellar nuclei were also intensely stained by Hsp70 antibody. Immunocytochemical stainings of cultured cerebellar cells showed that Hsp70 is expressed in both Purkinje and granule cells. The expression was punctate in the soma and along dendritic trees, and the punctae were colocalized with those of PSD95, a postsynaptic marker. Immunoblotting also indicates that Hsp70 is associated with the postsynaptic density fraction. Taken together, our results indicate that the Hsp70 is expressed in cerebellar neurons in normal conditions, and that some are localized in the synapses.

• The Korean Journal of Zoology
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• v.33 no.2
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• pp.141-151
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• 1990

Raf protein kinases and protein kinase C belong to serine/threonine-specific proteins in the cytoplasin, and are similar to each other in functional structure and the aspect of the distribution of celI. The distribution of raf protein kinase in the cerebrum of Geoclemys reevesfi as studied by using the antibodies against a-raf and c-raf protein kinase which induce the expression of raf fainily oncogenes. In general, raf protein kinases were distributed in such restricted regions as the general pallium, hippocampal formation, pdmordiuin hippocampi,nucleus of lateral olfactory tract, basal amygdaloid nucleus, and bed of stria terminalis. Immunological labeling of c-raf protein kinase was more widespread than that of a-raf. However, the intensity of the labeling of c-raf was lower than that of a-raf. The spherical cells of basal amygdaloid nucleus is a ring-like form, because only the cytoplasm was imunolabeled. Especially, c-raf protein kinase occurred in the cells which contained protein kinase C abundandy such as pyramidal cells and Purkinje cells. This suggests that a- and e-raf protein kinases may synegistically induce carclnoma with myc gene which is activated by protein kinase C.

• 한국체육학회지인문사회과학편
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• v.58 no.4
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• pp.481-492
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• 2019

The purpose of this study was to investigate the effects of treadmill exercise on cerebellar astrocyte activation and purkinje cells, neurotrophic factors expression, and motor function in aged rats. Sprague-Dawley (SD) rats were used and divided into three groups; (1) Young Control Group (YCG; 3months aged, n=10); (2) Old Control Group; (OCG; 24months aged, n=10); (3) Old Exercise Group (OEG; 24months aged, n=10). Rats were then subjected to treadmill exercise for 5 days per week for 12 weeks during which time the speed of the treadmill was gradually increased. The results revealed that in the rota-rod test, motor function was significantly increased in the OEG compared to the OCG (p<.05), and similarly YCG. Number of calbindin-positive purkinje cell expression significantly increased in the cerebellar vermis of OEG compared to the OCG (p<.05), and similarly YCG. GFAP-, NMDAR-positive cell expression significantly increased in the OEG (respectively p<.001), GFAP and GLAST protein levels were significantly increased in the cerebellum of OEG compared to the OCG (p<.05, p<.001) and similarly YCG. BDNF and NGF protein levels were highest in the YCG, increased in the OEG compared to OCG (p<.001, p<.05). These result show that regular exercise not only improved astrocyte activation, but also increased purkinje cell expression in the cerebellum and motor function by increasing the neurotrophic factors in aged rats.

• Korean Journal of Veterinary Pathology
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• v.3 no.2
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• pp.73-76
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• 1999

The tissue distribution and cellular localization of viral antigen in the brain of aborted fetus with bovine viral diarrhea virus(BVDV) infection was studied; BVDV antigens was detected in spleen, kidney, lung, eyelid as well as brain. In the brain, the virus was recognized in neurons and non-neuronal cells in the cerebellum and cerebrum. Many cells in the superficial layer and occasional Purkinje cells had BVDV antigens. As well, BVDV was also found in the perivascular cells, vascular endothelial cells and smooth muscle cells in the vessels and neuroglial cells in the white matter. This finding suggests that BVD virus favors infect progenitor cells in the brain, notably in the superficial layer of cerebellum, and damage normal development of cerebellum, which leads to cerebellar hypoplasia.

• Proceedings of the Korean Society of Applied Pharmacology
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• 2003.11a
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• pp.112-112
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• 2003

Voltage-gated $K^{+}$ (Kv) channels represent a structurally and functionally diverse group of membrane proteins. These channels play an important role in determining the length of the cardiac action potential and are the targets for antiarrhythmic drugs. Many $K^{+}$ channel genes have been cloned from human myocardium and functionally contribute to its electrical activity. One of these channels, Kv1.5, is one of the more cardiovascular-specific $K^{+}$ channel isoforms identified to date and forms the molecular basis for an ultra-rapid delayed rectifier $K^{＋}$ current found in human atrium. Thus, the blocker of hKv1.5 is expected to be an ideal antiarrhythmic drug for atrial fibrillation. Chelidonine was isolated from Chelidonium majus L. We examined the effect of chelidonine on the hKv1.5 current expressed in Ltk-cells using whole cell mode of patch clamp techniques. Chelidonine selectively inhibited the hKv1.5 current expressed in Ltk-cells in a concentration-dependent manner, whereas did not affect the HERG current expressed in HEK-293 cells. Additionally, chelidonine reduced the tail current amplitude recorded at -50 mV after 250 ms depolarizing pulses to +60 mV, and slowed the deactivation time course resulting in a 'crossover' phenomenon when the tail currents recorded under control conditions and in the presence of chelidonine were superimposed. We found that chelidonine also inhibited the $K^{+}$ current in isolated human atrial myocytes where hKv1.5 channels were predominantly expressed. Furthermore, we examined the effects of chelidonine on the action potentials in rabbit hearts using conventional microelectrode technique. Chelidonine prolonged the action potential durations (APD) of atrial, ventricular myocytes and Purkinje fibers in a dose-dependent manner. However, the effect of chelidonine on atrial APD was frequency-dependent whereas the effect of chelidonine on the APDs of ventricular myocytes and Purkinje fibers was not frequency- dependent. Also, the selective action of chelidonine on heart was more potent than dofetilide, $K^{+}$ channel blocker.

• Journal of Environmental Health Sciences
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• v.12 no.2
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• pp.1-9
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• 1986

The purpose of this study is to determine the antidotal effects of thiamine in phenylmercury poisoning rats. Sixty-six rats were divided into six groups the control group, the $40{\gamma}$ thiamine-only-dosed group, the 6 ppm phenylmercury-only-dosed group, the simultaneously-dosed-group with 6 ppm mercury & $20{\gamma}$ thiamine, and with 6 ppm mercury & $40{\gamma}$ thiamine, and with 6 ppm mercury & $80{\gamma}$ thiamine. The thiamine was put into pellet by various concentrations, and phenylmercury was mixed in drinking water by 6 ppm concentration. The rats were sacrificed for observing the histopathological changes of brain, liver and kidney. The remits summarized are as follows 1. In the group dosed with only $40{\gamma}$ thiamine, the tissues of brain, liver and kidney did not show any abnormal architecture. 2. The phenyhnercury-only-dosed group and the simultaneoulsy-dosed group with mercury and $20{\gamma}$ thiamine showed remarkable degenerative or necrotic hepatic cells. In addition, a remarkable swelling and necrosis on epithelium of proximal tubules in kidney were found. 3. The simultaneously-dosed group with mercury and $40{\gamma}$ thiamine showed moderate degeneration and necrosis of Purkinje cells in cerebellum. A moderate necrosis and swelling on epithelium of proximal tubules and a large amount of tubular casts were found as well. 4. The simultaneously-dosed group with mercury and $80{\gamma}$ thiamine showed a slight degenerative change of Purkinje cells. A slight degenerative change on epithelium of proximal tubules and a small amount of tubular casts were also found.

• Applied Microscopy
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• v.31 no.1
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• pp.1-8
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• 2001

The morphological features of neuronal dendritic spines are changed their shapes, sizes and density in response to physiological or pathological conditions . Therefore, exact analysis of spines warrants understanding of neuronal function. The size of the spine is at the borderline of resolution with light microscopy. High voltage electron microscopy Provide excellent resolution of the spines with proper stain techniques thanks to its higher resolution and penetration power. We evaluated more effective staining method for observing dendritic spines after labeling Purkinje cells with anti-calbindin 28 kD immunohistochemistry or Golgi staining methods. 4 fm thickness sections were observed with high voltage electron microscopy and some morphometric analyses were performed. Both Golgi staining and immunohistochemistry revealed the detail structures of the Purkinje cell such as soma, dendrites, and dendritic spines. High voltage electron micrographs with Golgi staining provide more precise morphology and are easy to measure. Average density of spine is $24.5{\pm}3.6/10{\mu}m$ and its length is $1.12{\pm}0.22{\mu}m$. For quantitative analysis of the spines, high voltage electron, micrographs with Golgi staining are more effective. This preliminary result is expected to be useful for further study of spine plasticity in various conditions.

• Molecules and Cells
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• v.26 no.2
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• pp.186-192
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• 2008

NELL2, a neural tissue-enriched protein, is produced in the embryo, and postembryonically in the mammalian brain, with a broad distribution. Although its synthesis is required for neuronal differentiation in chicks, not much is known about its function in the adult mammalian brain. We investigated the distribution of NELL2 in various regions of the adult rat brain to study its potential functions in brain physiology. Consistent with previous reports, NELL2-immunoreactivity (ir) was found in the cytoplasm of neurons, but not in glial fibrillary acidic protein (GFAP)-positive glial cells. The highest levels of NELL2 were detected in the hippocampus and the cerebellum. Interestingly, in the cerebellar cortex NELL2 was observed only in the GABAergic Purkinje cells not in the excitatory granular cells. In contrast, it was found mainly in the hippocampal dentate gyrus and pyramidal cell layer that contains mainly glutamatergic neurons. In the dentate gyrus, NELL2 was not detected in the GFAP-positive neural precursor cells, but was generally present in mature neurons of the subgranular zone, suggesting a role in this region restricted to mature neurons.

• Proceedings of the Korean Biophysical Society Conference
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• 2002.06b
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• pp.37-37
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• 2002

The Na$^{＋}$-Ca$^{2＋}$ exchanger (NCX) is known to playa critical role in the regulation of intracellular $Ca^{2＋}$ in many tissues and cells. Three isoforms have been cloned (NCXl, NCX2, NCX3). Among the isoforms, NCX2 and NCX3 are expressed at high levels in brain and in a few other tissues. But the differential properties of the isoforms are not yet clearly established.(omitted)