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1 KISEP Otology Korean J Otolaryngol 2002;45:214-8 살리실산투여후흰쥐뇌간에서 c-fos 발현의변화 김상철 1 전병훈 1 최익수 1 정범조 1 김동석 2 C-fos Expression in Brain Stem of Rats after Salicylic Acid Treatment Sang Chul Kim, MD 1, Byung Hoon Jun, MD 1, Ick Soo Choi, MD 1, Bum Jo Jung, MD 1 and Dong Suk Kim, MD 2 1 Department of Otolaryngology, Inje University College of Medicine, Seoul Paik Hospital, Seoul 2 Dong Suk Kim ENT Clinic, Incheon, Korea ABSTRACT Background and ObjectivesSubjective tinnitus, a distracting internal noise, is experienced by humans. Tinnitus is evoked by salicylic acid treatment in rats as confirmed by Jastreboff in 1994 in an animal behavior model of tinnitus with salicylic acid. The objective of this study is to evaluate c-fos expression in the brain stem of rats after salicylic acid treatment. Materials and MethodAfter salicylic acid 450 mg/kg and saline treatment 450 mg/kg, c-fos immunohistochemical staining expression in the auditory and nonauditory brain stem nuclei were observed. ResultsMany immunoreactive cells were observed in the Locus Ceruleus of the salicylic acid treated animals, but not in the saline treated animals. No immunoreactive cells were found in the auditory brain stem nuclei. ConclusionThe Locus Ceruleus is the nucleus of the brain stem and produce norepinephrine which results in arousal of the neuronal activity for stress. These results suggest that salicylic acid may evoke tinnitus through a combined effect on the auditory and nonauditory brain nuclei. It seems possible that the interaction of these effects at particular locations of the brain causes tinnitus. Korean J Otolaryngol 2002;45:214-8 KEY WORDSTinnitus Salicylic acid C-fos protein Locus ceruleus. 214 실험동물
2 이명의유도 조직준비 215
3 살리실산을 이용한 흰쥐 이명 모델에서 c-fos의 발현 비교 c-fos에 대한 면역조직화학반응 양성세포의 발현이 LC에서 응 양성세포 발현양상을 비교하였다. 실험군과 대조군 모두 나타났고, 생리식염수를 주입한 대조군에서는 동일한 영역에 에서 CN는 항상 c-fos 면역조직화학반응 양성세포들이 발 서 c-fos 면역조직화학반응 양성세포가 관찰되지 않았다. 견되며, 복측 핵보다는 배측 핵에서 c-fos 면역조직화학반 c-fos 면역조직화학반응 양성세포의 발현은 뇌 반구별로 차 응 양성세포들이 더 밀집된 양상으로 나타난다(Fig. 1). IC 이를 나타내지 않았다. 다른 청각 뇌간 신경핵들은 처치 효 는 실험군과 대조군에서 모두 c-fos 면역조직화학반응 양 과에 따른 c-fos 면역조직화학반응 양성세포에 있어서 차이 성세포가 관찰되지 않았다. 그외 상행성 청각 중추 신경경로 를 보이지 않았다. 소뇌에서는 c-fos 면역조직화학반응 양 인 SOC, LL, MGB, AC에서도 실험군과 대조군에서 c-fos 성세포가 실험군과 대조군 모두에서 나타났다. 실험결과는 각 면역조직화학반응 양성세포가 발현되지 않았다. 각의 실험에서 모두 일치했다. 비청각 중추 신경핵 흰쥐에서 LC는 뇌교 부위에서 PeriVenricular Grey 외측 청각 중추 신경핵 청각 중추 신경핵에서는 실험군과 대조군 사이의 처치에 벽의 세포들이 길어져서 구성되어 있는 중간크기의 신경핵으 따른 변화가 거의 없었다. 하부 청각 중추 신경핵부터 상부 로 LC에서 살리실산 처치 세시간 경과 후 관찰한 결과, c- 청각 신경핵까지 따라 올라 가면서 살리실산을 처치한 실험 fos 면역조직화학반응 양성세포가 많이 밀집되어 있었다. 실 군과 생리식염수를 처치한 대조군의 c-fos 면역조직화학반 험군에서 c-fos 면역조직화학반응 양성세포들은 LC의 배측 Fig. 1. The immunohistochemical staining pattern of c-fos on the right cochlear nucleus (CN) salicylic acid treated (A) and Saline control group (B). The expression of c-fos immuno-histochemical staining on CN was similar in both group, although the dorsal nucleus of CN (arrow) was more densely stained than ventral nucleus. round circle was airbubble (arrowhead) ( 400). A A B B Fig. 2. Immunoreactivity in LC. A Many c-fos immunoreactive cells are found in Locus Ceruleus (LC) after salicylate treatment (arrow). B LC lacks c-fos immunoreactivity after saline treatment (arrowhead). MeV mesencephalic trigeminal nucleus (MMTN), Cb cerebellum. 216 Korean J Otolaryngol 2002 ;45 :214-8
4 Fig. 3. Anatomy of LC. ASchematic diagram for location of the locus Ceruleus LC in the coronal section of rat brain. BSagittal section of the rat brain showing the course of the dorsal and ventral noradrenergic bundles from the LC. Solid lines indicate the major projections of the LC, including the dorsal noradrenergic bundle and central gray longitudinal fasciculus thick line and ventral tegmental bundle thin line. Dotted lines indicate the projections of the cell groups comprising the ventral noradrenergic bundle The Rat brain in stereotaxic coordinates 3rd editiongeorge Paxinos & Charles Watson academic press,
5 REFERENCES 1) Jastreboff PJ, Sasaki CT. An animal model of tinnitus: A decade of development. Am J Otolayngol 1994;15: ) Bauer CA, Brozoski TJ, Rojas R, Boley J, Wyder M. Behavioral model of chronic tinnitus in rats. Otolaryngol Head Neck Surg 1999;121: ) Boettcher FA, Salvi RJ. Salicylate ototoxicity: Review and syn- thesis. Am J Otolaryngol 1991;12: ) Jung TT, Rhee CK, Lee CS, Park YS, Choi DC. Ototoxicity of salicylate, nonsteroidal antiinflammatory drugs, and quinine. Otolaryngol Clin North Am 1993;26: ) Morgan JI, Curran T. Stimulus-transcription coupling in the nervous system: Involvement of the inducible proto-oncogenes fos and jun. Annu Rev Neurosci 1991;14: ) Senba E, Matsunaga K, Tohyama M, Noguchi K. Stress-induced c-fos expression in the rat brain: Activation mechanism of sympathetic pathway. Brain Res Bull 1993;31: ) Paxinos G, Watson C. The Rat Brain in Stereotaxic Coordinates, fourth edition;1997. p ) Ehret G, Fischer R. Neuronal activity and tonotopy in the auditory system visualized by c-fos gene expression. Brain Res 1991; 567: ) Sandner G, Oberling P, Silveira MC, Di Scala G, Rocha B, Bargri A, et al. What brain structures are active during emotions? Effects of brain stimulation elicited aversion on c-fos immunoreactivity and behavior. Behav Brain Res 1993;58: ) Keilmann A, Herdegen T. The c-fos transcription factor in the auditory pathway of the juvenile rat: Effects of acoustic deprivation and repetitive stimulation. Brain Res 1997;753: ) Wallhausser-Franke E, Braun S, Langner G. Salicylate alters 2- DG uptake in the auditory system: A model for tinnitus? Neuroreport 1996;7: ) Wallhausser-Franke E. Salicylate evokes c-fos expression in the brain stem: Implications for tinnitus. Neuroreport 1997;8: ) Martin RL, Webster WR, Serviere J. The frequency organizaion of the inferior colliculus of the guinea pig: A [14C]-2-deoxyglucose study. Hear Res 1988;33: ) Klepper A, Herbert H. Distribution and origin of noradrenergic and serotonergic fibers in the cochlear nucleus and inferior colliculus of the rat. Brain Res 1991;557: ) Swanson LW. The locus Ceruleus: A cytoarchitectonic, Golgi and immu-nohistochemical study in the albino rat. Brain Res 1976; 110: ) Foote SL, Bloom FE, Aston-Jones G. Nucleus locus ceruleus: New evidence of anatomical and physiological specificity. Physiol Rev 1983;63: ) Chan RK, Sauchenko PE. Spatially and temporally differentiated patterns of c-fos expression in brainstem catecholaminergic cell groups induced by cardiovascular challenges in the rat. J Comp Neurol 1994;348: ) Singewald N, Zhou GY, Schneider C. Release of excitatory and inhibitory amino acids from the locus Ceruleus of conscious rats by cardiovascular stimuli and various forms of acute stress. Brain Res 1995;704: ) Krukoff TL, Morton TL, Harris KH, Jhamandas JH. Expression of c-fos protein in rat brain elicited by electrical stimulation of the pontine parabrachial nucleus. J Neurosci 1992;12: Korean J Otolaryngol 2002;45:214-8
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