笔者要感谢保罗·维悌希博士和吉纳维芙方女士的mRNA的提取和PCR援助，以及加内特凋谢和罗德尼·威廉斯纪念基金会的支持。

伦理学声明:下面列出的所有程序批准了悉尼动物伦理委员会的大学。 1.动物注:1和8 - 9个月大的小鼠（C57 / BL6）是从动物资源中心（珀斯，澳大利亚）中获得。老鼠被安置在博世灭鼠设施，在悉尼大学。 <ol><li>家鼠在上一个12/12小时亮/暗循环以获得食物和水随意标准鼠标笼中。 </li><li>鸿沟小鼠各年龄组为近红外（NIR）治疗，或假处理（对照）组进行比较。 </li></ol> 2.近红外（NIR）照射及深水治疗<ol><li>剃鼠标的头部和颈部区域的毛皮用电动剃刀尽可能防止头发快速再生完成治疗方案的前。为了保持小鼠的无病原体状态，使用70％的乙醇剃须动物，并从不同的笼子里的动物之间的F10兽用消毒剂清洗之间的乐器。这样做2 - 3天前开始治疗等动物不是过度处理。 </li><li>抑制小鼠握住尾部的近端，并允许它在用一只手或台式手掌放松，以便尽量减少应力，这可能会导致错误的结果。 </li><li>持有NIR（670 nm）的LED（发光二极管）器件1 - 2厘米的距离暴露（光头）区和开关设备上的90秒（ 图1A）。 注意:温度变化的90秒曝光结果测定为&lt;0.2℃下在100毫升的水中。 </li><li>对于假治疗组动物的2.3，但离开该装置关闭（ 图1B） -重复步骤2.2。 </li><li>重复步骤2.2 - 2.3对NIR阻断治疗组的动物，但覆盖器件有铝箔。 </li><li>每天approximat 2.5 - 重复步骤2.2E 24小时的间隔，连续5天。 </li><li>提取两个耳朵上的第五天后处理前庭感觉上皮（3嵴和1个椭圆囊斑）（见下文第3节 ）。 </li></ol> 3.组织提取20 <ol><li>制备300毫升基于甘油的人工脑脊液（ACSF）的组成（以mM计）:26的NaHCO 3，11葡萄糖，250甘油，2.5氯化钾，1.2的NaH 2 PO 4，1.2的MgCl 2，和2.5 氯化钙 。在加入氯化钙 ，燃气与卡波金（95％O 2和5％CO 2）的溶液来建立的pH为7.4，并避免钙沉淀（混浊）。寒意在-80℃冷冻该溶液45分钟，以使冰浆料形成。 </li><li>准备在标螺杆顶端微管的RNA分离裂解缓冲液（停止盖子的爆裂时冻结和解冻在液氮之间管压力的变化），根据制造商或通常的做法实验室的指示。有液氮准备在铝杜瓦瓶中。 注:液氮操作时，使用的防护服和eyeware。确保使用液氮在一个通风良好的房间作为其气体形式的体积大约是700倍，它的液体形式，并可能导致窒息。 </li><li>通过腹膜内注射深深地麻醉小鼠用氯胺酮（400毫克/千克）。允许后肢反射完全消退作为指示小鼠完全麻醉。 </li><li>杀头的小鼠用锋利的不锈钢剪刀，并沿着用刀片头骨的矢状皮肤切口（四舍五入＃22）。在这一点上和整个步骤3.5 - 3.9，保持颅，脑和潜在的前庭器官尽可能凉爽通过在组织常规应用冰冷学联的。 </li><li>使用标准模式剪刀尖手臂做出SMaL公司升切口头骨的Lambda和沿矢状缝切割。 </li><li>轻轻滑动的浅表骨钳一个臂顶骨保持刀片尽可能接近到骨头的下表面，而不拖动大脑下方。安全和扯远了顶骨横向和枕骨后部，直到大脑暴露。 </li><li>用一个不锈钢小铲子和解除大脑远离前部和中颅窝暴露前庭耳蜗神经（CN八）。横切神经以防止在该初级传入轴突直接支配前庭毛细胞不必要的紧张。 </li><li> CN八横断后取出大脑全盘 。 </li><li>观察含耳蜗和中颅窝周围前庭器官的骨迷路。让两个小切口，每个骨迷路旁边，握住前半规管和后拉切除整个结构盟友。 </li><li>立即浸入切除迷宫中含有冰冷ACSF溶液（如在步骤3.1中所述），同时连续地卡波金灌注解剖盘中。 </li><li>在立体显微镜下，按住迷宫由耳蜗并将其固定钳盘的底部。 <ol><li>直接使用细镊子划伤小开在上面的前半规管（SSC）壶腹骨。 </li><li>轻轻达到立即骨下方，向外弹离壶腹放大此开幕。谨慎行事这里没有推入钳开放和破坏脆弱的膜迷路下方。继续以这种方式，直到椭圆囊，前部和侧壶腹都暴露出来。如果可能的话删除后壶腹也。 </li></ol></li><li>用细镊子，轻轻抬起囊，并从骨迷路壶腹距离，直到他们完全不沾边。如果可能的话，他们持有由它们相关联的半规管，以避免损坏感觉上皮。在一些情况下，半圆形膜质导管的近端部分可能需要与虹膜剪切以从骨释放壶腹。 </li><li>牢固地抓住镊子的顶端和地点之间的前庭器官到先前在步骤3.2中制备的裂解缓冲液。轻轻绕摇动镊子在缓冲，以确保前庭器官已经从钳子脱离。检查，这是通过使钳子的立体显微镜下的情况。 <ol><li>拧上微管盖，并立即冷冻在液氮中的样品。 </li></ol></li></ol> 4. RNA提取和RT-PCR <ol><li>根据制造商的说明或优选的实验方案遵循信使RNA（mRNA）的提取的标准方法。 注:利用载体分子RNA，以帮助隔离mRNA的小产量商品化试剂盒在此使用协议。低洗脱体积可用于增加最终浓度的mRNA。 注:负"无酶"的控制（NEC的）和"无DNA模板"对照（NTC却）也必须被完成，以确保观察到的效果的有效性。 </li><li>适用的mRNA逆转录，以互补的DNA的靶基因的21-23（cDNA中）和扩增的标准方法。 </li></ol>

<ol><li>Agrawal, Y., Carey, J. P., Della Santina,, C, C., Schubert, M. C., Minor, L. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Disorders+of+balance+and+vestibular+function+in+US+adults%3A+data+from+the+National+Health+and+Nutrition+Examination+Survey%2C+2001-2004%5BTitle%5D)+AND+%22Archives+of+internal+medicine%22%5BJournal%5D)">Disorders of balance and vestibular function in US adults: data from the National Health and Nutrition Examination Survey, 2001-2004.</a> Archives of internal medicine. 169, 938-944 (2009).</li>
<li>Bessho, K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Effect+of+subthreshold+infrared+laser+treatment+for+drusen+regression+on+macular+autofluorescence+in+patients+with+age-related+macular+degeneration%5BTitle%5D)+AND+%22Retina%22%5BJournal%5D)">Effect of subthreshold infrared laser treatment for drusen regression on macular autofluorescence in patients with age-related macular degeneration.</a> Retina. 25, 981-988 (2005).</li>
<li>Olk, R. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Therapeutic+benefits+of+infrared+%28810-nm%29+diode+laser+macular+grid+photocoagulation+in+prophylactic+treatment+of+nonexudative+age-related+macular+degeneration%3A+two-year+results+of+a+randomized+pilot+study%5BTitle%5D)+AND+%22Ophthalmology%22%5BJournal%5D)">Therapeutic benefits of infrared (810-nm) diode laser macular grid photocoagulation in prophylactic treatment of nonexudative age-related macular degeneration: two-year results of a randomized pilot study.</a> Ophthalmology. 106, 2082-2090 (1999).</li>
<li>Rodanant, N., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Predictors+of+drusen+reduction+after+subthreshold+infrared+%28810+nm%29+diode+laser+macular+grid+photocoagulation+for+nonexudative+age-related+macular+degeneration%5BTitle%5D)+AND+%22American+journal+of+ophthalmology%22%5BJournal%5D)">Predictors of drusen reduction after subthreshold infrared (810 nm) diode laser macular grid photocoagulation for nonexudative age-related macular degeneration.</a> American journal of ophthalmology. 134, 577-585 (2002).</li>
<li>De Taboada, L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Transcranial+laser+therapy+attenuates+amyloid-beta+peptide+neuropathology+in+amyloid-beta+protein+precursor+transgenic+mice%5BTitle%5D)+AND+%22Journal+of+Alzheimer%27s+disease+%3A+JAD%22%5BJournal%5D)">Transcranial laser therapy attenuates amyloid-beta peptide neuropathology in amyloid-beta protein precursor transgenic mice.</a> Journal of Alzheimer's disease : JAD. 23, 521-535 (2011).</li>
<li>Grillo, S. L., Duggett, N. A., Ennaceur, A., Chazot, P. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Non-invasive+infra-red+therapy+%281072+nm%29+reduces+beta-amyloid+protein+levels+in+the+brain+of+an+Alzheimer%27s+disease+mouse+model%5BTitle%5D)+AND+%22TASTPM.+Journal+of+photochemistry+and+photobiology.+B%2C+Biology%22%5BJournal%5D)">Non-invasive infra-red therapy (1072 nm) reduces beta-amyloid protein levels in the brain of an Alzheimer's disease mouse model.</a> TASTPM. Journal of photochemistry and photobiology. B, Biology. 123, 13-22 (2013).</li>
<li>Purushothuman, S., Johnstone, D. M., Nandasena, C., Mitrofanis, J., Stone, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Photobiomodulation+with+near+infrared+light+mitigates+Alzheimer%27s+disease-related+pathology+in+cerebral+cortex+-+evidence+from+two+transgenic+mouse+models%5BTitle%5D)+AND+%22Alzheimer%27s+researc.%2C+%26+therapy%22%5BJournal%5D)">Photobiomodulation with near infrared light mitigates Alzheimer's disease-related pathology in cerebral cortex - evidence from two transgenic mouse models.</a> Alzheimer's researc., & therapy. 6, 2(2014).</li>
<li>Sommer, A. P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((670+nm+laser+light+and+EGCG+complementarily+reduce+amyloid-beta+aggregates+in+human+neuroblastoma+cells%3A+basis+for+treatment+of+Alzheimer%27s+disease%5BTitle%5D)+AND+%22Photomedicine+and+laser+surgery%22%5BJournal%5D)">670 nm laser light and EGCG complementarily reduce amyloid-beta aggregates in human neuroblastoma cells: basis for treatment of Alzheimer's disease.</a> Photomedicine and laser surgery. 30, 54-60 (2012).</li>
<li>Moro, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Photobiomodulation+preserves+behaviour+and+midbrain+dopaminergic+cells+from+MPTP+toxicity%3A+evidence+from+two+mouse+strains%5BTitle%5D)+AND+%22BMC+neuroscience%22%5BJournal%5D)">Photobiomodulation preserves behaviour and midbrain dopaminergic cells from MPTP toxicity: evidence from two mouse strains.</a> BMC neuroscience. 14, 40(2013).</li>
<li>Peoples, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Photobiomodulation+enhances+nigral+dopaminergic+cell+survival+in+a+chronic+MPTP+mouse+model+of+Parkinson%27s+disease%5BTitle%5D)+AND+%22Parkinsonis.%2C+%26+related%22%5BJournal%5D)">Photobiomodulation enhances nigral dopaminergic cell survival in a chronic MPTP mouse model of Parkinson's disease.</a> Parkinsonis., & related. 18, 469-476 (2012).</li>
<li>Shaw, V. E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Neuroprotection+of+midbrain+dopaminergic+cells+in+MPTP-treated+mice+after+near-infrared+light+treatment%5BTitle%5D)+AND+%22The+Journal+of+comparative+neurology%22%5BJournal%5D)">Neuroprotection of midbrain dopaminergic cells in MPTP-treated mice after near-infrared light treatment.</a> The Journal of comparative neurology. 518, 25-40 (2010).</li>
<li>Ying, R., Liang, H. L., Whelan, H. T., Eells, J. T., Wong-Riley, M. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Pretreatment+with+near-infrared+light+via+light-emitting+diode+provides+added+benefit+against+rotenone-+and+MPP%2B-induced+neurotoxicity%5BTitle%5D)+AND+%22Brain+research%22%5BJournal%5D)">Pretreatment with near-infrared light via light-emitting diode provides added benefit against rotenone- and MPP+-induced neurotoxicity.</a> Brain research. 1243, 167-173 (2008).</li>
<li>Rajguru, S. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Infrared+photostimulation+of+the+crista+ampullaris%5BTitle%5D)+AND+%22The+Journal+of+physiology%22%5BJournal%5D)">Infrared photostimulation of the crista ampullaris.</a> The Journal of physiology. 589, 1283-1294 (2011).</li>
<li>Chung, H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((The+nuts+and+bolts+of+low-level+laser+%28light%29+therapy%5BTitle%5D)+AND+%22Annals+of+biomedical+engineering%22%5BJournal%5D)">The nuts and bolts of low-level laser (light) therapy.</a> Annals of biomedical engineering. 40, 516-533 (2012).</li>
<li>Desmet, K. D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Clinical+and+experimental+applications+of+NIR-LED+photobiomodulation%5BTitle%5D)+AND+%22Photomedicine+and+laser+surgery%22%5BJournal%5D)">Clinical and experimental applications of NIR-LED photobiomodulation.</a> Photomedicine and laser surgery. 24, 121-128 (2006).</li>
<li>Huang, Y. Y., Chen, A. C., Carroll, J. D., Hamblin, M. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Biphasic+dose+response+in+low+level+light+therapy%5BTitle%5D)+AND+%22Dose-response+%3A+a+publication+of+International+Hormesis+Society%22%5BJournal%5D)">Biphasic dose response in low level light therapy.</a> Dose-response : a publication of International Hormesis Society. 7, 358-383 (2009).</li>
<li>Rojas, J. C., Gonzalez-Lima, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Low-level+light+therapy+of+the+eye+and+brain%5BTitle%5D)+AND+%22Eye+and+brain%22%5BJournal%5D)">Low-level light therapy of the eye and brain.</a> Eye and brain. 3, 49-67 (2011).</li>
<li>Vranceanu, F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Striated+organelle%2C+a+cytoskeletal+structure+positioned+to+modulate+hair-cell+transduction%5BTitle%5D)+AND+%22Proceedings+of+the+National+Academy+of+Sciences+of+the+United+States+of+America%22%5BJournal%5D)">Striated organelle, a cytoskeletal structure positioned to modulate hair-cell transduction.</a> Proceedings of the National Academy of Sciences of the United States of America. 109, 4473-4478 (2012).</li>
<li>Johnson, K. R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Separate+and+combined+effects+of+Sod1+and+Cdh23+mutations+on+age-related+hearing+loss+and+cochlear+pathology+in+C57BL%2F6J+mice%5BTitle%5D)+AND+%22Hearing+research%22%5BJournal%5D)">Separate and combined effects of Sod1 and Cdh23 mutations on age-related hearing loss and cochlear pathology in C57BL/6J mice.</a> Hearing research. 268, 85-92 (2010).</li>
<li>Tung, V. W., Di Marco, S., Lim, R., Brichta, A. M., Camp, A. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((An+isolated+semi-intact+preparation+of+the+mouse+vestibular+sensory+epithelium+for+electrophysiology+and+high-resolution+two-photon+microscopy%5BTitle%5D)+AND+%22Journal+of+visualized+experiments+%3A+JoVE%22%5BJournal%5D)">An isolated semi-intact preparation of the mouse vestibular sensory epithelium for electrophysiology and high-resolution two-photon microscopy.</a> Journal of visualized experiments : JoVE. , e50471(2013).</li>
<li>Kirby, J., Menzies, F. M., Cookson, M. R., Bushby, K., Shaw, P. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Differential+gene+expression+in+a+cell+culture+model+of+SOD1-related+familial+motor+neurone+disease%5BTitle%5D)+AND+%22Human+molecular+genetics%22%5BJournal%5D)">Differential gene expression in a cell culture model of SOD1-related familial motor neurone disease.</a> Human molecular genetics. 11, 2061-2075 (2002).</li>
<li>Parry, S. N., Ellis, N., Li, Z., Maitz, P., Witting, P. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Myoglobin+induces+oxidative+stress+and+decreases+endocytosis+and+monolayer+permissiveness+in+cultured+kidney+epithelial+cells+without+affecting+viability%5BTitle%5D)+AND+%22Kidney+and+Blood+Pressure+Research%22%5BJournal%5D)">Myoglobin induces oxidative stress and decreases endocytosis and monolayer permissiveness in cultured kidney epithelial cells without affecting viability.</a> Kidney and Blood Pressure Research. 31, 16-28 (2008).</li>
<li>Saee-Rad, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Analysis+of+superoxide+dismutase+1%2C+dual-specificity+phosphatase+1%2C+and+transforming+growth+factor%2C+beta+1+genes+expression+in+keratoconic+and+non-keratoconic+corneas%5BTitle%5D)+AND+%22Molecular+vision%22%5BJournal%5D)">Analysis of superoxide dismutase 1, dual-specificity phosphatase 1, and transforming growth factor, beta 1 genes expression in keratoconic and non-keratoconic corneas.</a> Molecular vision. 19, 2501-2507 (2013).</li>
<li>Albert, E. S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((TRPV4+channels+mediate+the+infrared+laser-evoked+response+in+sensory+neurons%5BTitle%5D)+AND+%22Journal+of+neurophysiology%22%5BJournal%5D)">TRPV4 channels mediate the infrared laser-evoked response in sensory neurons.</a> Journal of neurophysiology. 107, 3227-3234 (2012).</li>
<li>Moro, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Photobiomodulation+inside+the+brain%3A+a+novel+method+of+applying+near-infrared+light+intracranially+and+its+impact+on+dopaminergic+cell+survival+in+MPTP-treated+mice%5BTitle%5D)+AND+%22Journal+of+neuroscience%22%5BJournal%5D)">Photobiomodulation inside the brain: a novel method of applying near-infrared light intracranially and its impact on dopaminergic cell survival in MPTP-treated mice.</a> Journal of neuroscience. 120, 670-683 (2014).</li>
<li>Moreno, L. E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Infrared+neural+stimulation%3A+beam+path+in+the+guinea+pig+cochlea%5BTitle%5D)+AND+%22Hearing+research%22%5BJournal%5D)">Infrared neural stimulation: beam path in the guinea pig cochlea.</a> Hearing research. 282, 289-302 (2011).</li>
<li>Curthoys, I. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((A+red+thread+as+a+guide+in+the+vestibular+labyrinth%5BTitle%5D)+AND+%22The+Journal+of+physiology%22%5BJournal%5D)">A red thread as a guide in the vestibular labyrinth.</a> The Journal of physiology. 589, 1241-1241 (2011).</li>
<li>Chakrabarti, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Mitochondrial+Dysfunction+during+Brain+Aging%3A+Role+of+Oxidative+Stress+and+Modulation+by+Antioxidant+Supplementation%5BTitle%5D)+AND+%22Aging+and+disease%22%5BJournal%5D)">Mitochondrial Dysfunction during Brain Aging: Role of Oxidative Stress and Modulation by Antioxidant Supplementation.</a> Aging and disease. 2, 242-256 (2011).</li>
<li>Petrosillo, G., De Benedictis, V., Ruggiero, F. M., Paradies, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Decline+in+cytochrome+c+oxidase+activity+in+rat-brain+mitochondria+with+aging.+Role+of+peroxidized+cardiolipin+and+beneficial+effect+of+melatonin%5BTitle%5D)+AND+%22Journal+of+bioenergetics+and+biomembranes%22%5BJournal%5D)">Decline in cytochrome c oxidase activity in rat-brain mitochondria with aging. Role of peroxidized cardiolipin and beneficial effect of melatonin.</a> Journal of bioenergetics and biomembranes. 45, 431-440 (2013).</li>
<li>Zhu, H., Sun, A., Zou, Y., Ge, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Inducible+metabolic+adaptation+promotes+mesenchymal+stem+cell+therapy+for+ischemia%3A+a+hypoxia-induced+and+glycogen-based+energy+prestorage+strategy%5BTitle%5D)+AND+%22Arteriosclerosis%2C+thrombosis%2C+and+vascular+biology%22%5BJournal%5D)">Inducible metabolic adaptation promotes mesenchymal stem cell therapy for ischemia: a hypoxia-induced and glycogen-based energy prestorage strategy.</a> Arteriosclerosis, thrombosis, and vascular biology. 34, 870-876 (2014).</li>
</ol>

作者宣称，他们没有竞争的财务权益。

这里所描述的代表性的结果表明，当与假治疗的小鼠相比较简短颅递送NIR光的（90秒/天5天）足以提高抗氧化剂的表达在老年小鼠的水平。而发射的热可以代表线粒体和/或神经元激活的一个来源，据报道为大鼠前庭传入神经​​24 -我们的热量由NIR发射测量LED器件是&lt;0.2℃，90秒，因此不太可能导致这里描述的变化。此外，与报告所强调以上，这里使用的NIR治疗不直接向感觉上皮或前庭传?...

余额递减性能和后续的瀑布是常见的，而自然老化1的不幸往往定义功能。这种下降的影响，既可以是自然和社会，和显著降低生活质量的老年人。作为回应，物理治疗和康复一直是研究的重点为瀑布，但尚未有一致的减少重复跌倒的发生率有关。与此同时，工作调查在周边或中央前庭系统（负责保持平衡的系统）是稀少，并且针对这些系统的潜在的治疗策略的变化和不平衡有限的根本原因。 在年龄相关的神经变性病症，包括年龄相关性黄斑变性2-4，阿耳茨海默氏病模型5-8，和帕金森氏病9-12最近的工作已经表明SI的神经保护作用mple非侵入性的应用的近红外（NIR）光。另外，在前庭系统，NIR已被用于提高在体外 13前庭初级传入神经 ​​元的活性。而没有很好理解NIR光的机构，利用NIR大多数的研究表明，NIR刺激线粒体复合物IV（细胞色素C氧化酶）14-17，以促进细胞代谢。在前庭感觉上皮I型毛细胞的表皮下板是致密的线粒体18，因此可以代表用于治疗NIR治疗作用的部位。 这里，经颅的简要的，非侵入性的治疗方案应用于NIR可用于测量细胞代谢（和暗示线粒体功能）在小鼠前庭感觉上皮进行说明。还讨论了为制备的前庭感觉上皮和它表明NIR增加一个ubiquito的表达我们的抗氧化剂（超氧化物歧化酶1）中的感觉上皮-先前证明是对耳蜗毛细胞存活19重要。

<table><tbody><tr><td>Quantum WARP 10</td><td>Quantum Devices</td><td>2070N030-A</td><td></td></tr><tr><td>Screw top microtubules</td><td>Quality Scientific Plastics</td><td>520-GRD-Q</td><td></td></tr><tr><td>Ketamine</td><td>Parnell, Alexandria Australia</td><td></td><td></td></tr><tr><td>Standard pattern scissors</td><td>FST</td><td>14001-12</td><td></td></tr><tr><td>Carbon steel surgical blades #22</td><td>Livingstone</td><td>SBLDCL 22</td><td></td></tr><tr><td>Friedman-Pearson rongeurs</td><td>FST</td><td>16221-14</td><td></td></tr><tr><td>Stereo microscope</td><td>Leica Microsystems</td><td>A60S</td><td></td></tr><tr><td>Dumont #5 SF forceps</td><td>FST</td><td>11252-00</td><td></td></tr><tr><td>Isolate II RNA Micro Kit</td><td>Bioline</td><td>BIO-52075</td><td></td></tr></tbody></table>

near infrared (nir) light increases expression of a marker of mitochondrial function in the mouse vestibular sensory epithelium

策略随着年龄的增加衰减下降平衡功能主要是专注于物理疗法，包括平衡工作和锻炼。然而，这些方法没有解决平衡下降的根本原因。使用小鼠，近红外光（NIR）的细胞上，在前庭感觉上皮的代谢的影响进行了评估。收集的数据表明，这种简单和安全的干预可能保护这些脆弱的细胞从自然老化的有害影响。的mRNA从分离的前庭外周感觉上皮（壶腹嵴和椭圆囊斑）萃取，随后转录成cDNA文库。这个库然后探查无处不抗氧化剂（SOD-1）的表达。抗氧化剂的基因表达，然后用于量化细胞代谢。用年轻的经颅交付NIR（4周）及以上（8 - 9个月）小鼠，并简要治疗方案（90秒/天5天AYS），这项工作表明NIR单独可足以改善线粒体功能的前庭感觉上皮。由于目前没有可用的，负担得起的，非侵入性的治疗方法，提高前庭毛细胞功能，外部NIR辐射的应用程序提供，以抵消对细胞在矿井前庭感觉上皮代谢老化的影响，潜在的战略。

比较近红外治疗的年轻年长的影响（4周）和（8 - 9个月）小鼠，我们测得的抗氧化剂超氧化物歧化酶1（SOD-1）的表达年轻的（N = 16）和老年人（N = 20）小鼠是NIR治疗，假治疗，或NIR封; 图2示出了相比于年轻假处理的动物一个显著增加β肌动蛋白标准化的SOD-1的表达超过2倍于年轻NIR处理的动物（P &lt;0.01），年轻的NIR阻断动物（P &lt;0.01）。较旧的NIR处理的动物也显示出超过一个时与旧NI...

Watch this Scientific Journal Video about Near Infrared NIr Light Increases Expression of a Marker of Mitochondrial Function in the Mouse Vestibular Sensory Epithelium at JoVE.com

Near Infrared NIr Light Increases Expression of a Marker of Mitochondrial Function in the Mouse Vestibular Sensory Epithelium

线粒体功能障碍是细胞衰老的标志。本文使用的非侵入性的近红外（NIR）处理，以改善在老化小鼠前庭感觉上皮线粒体功能。

近红外（NIR）光增加线粒体功能的标志物的表达，在鼠标前庭上皮细胞

Strategies for attenuating decline in balance function with increasing age are predominantly focused on physical therapies including balance tasks and ...

near-infrared-nir-light-increases-expression-marker-mitochondrial

Research

JoVE Journal

Biology

Near Infrared (NIr) Light Increases Expression of a Marker of Mitochondrial Function in the Mouse Vestibular Sensory Epithelium

9.9K 次观看.  University of Sydney. 线粒体功能障碍是细胞衰老的标志。本文使用的非侵入性的近红外（NIR）处理，以改善在老化小鼠前庭感觉上皮线粒体功能。 

视频: Near Infrared NIr Light Increases Expression of a Marker of Mitochondrial Function in the Mouse Vestibular Sensory Epithelium

An Isolated Semi-intact Preparation of the Mouse Vestibular Sensory Epithelium for Electrophysiology and High-resolution Two-photon Microscopy

Understanding vestibular hair cells function under normal conditions, or how trauma, disease, and aging disrupt this function is a vital step in the development of preventative approaches and/or novel therapeutic strategies. However, the majority of studies looking at abnormal vestibular function have not been at the cellular level but focused primarily on behavioral assays of vestibular dysfunction such as gait analyses and vestibulo-ocular reflex performance. While this work has yielded valuable data about what happens when things go wrong, little information is gleaned regarding the underlying causes of dysfunction. Of the studies that focus on the cellular and subcellular processes that underlie vestibular function, most have relied on acutely isolated hair cells, devoid of their synaptic connections and supporting cell environment. Therefore, a major technical challenge has been access to the exquisitely sensitive vestibular hair cells in a preparation that is least disrupted, physiologically. Here we demonstrate a semi-intact preparation of the mouse vestibular sensory epithelium that retains the local micro-environment including hair cell/primary afferent complexes.

Analysis of vestibular hair cell function is complicated by their location deep within the hardest part of the skull, the petrous temporal bone. Most functional hair cell studies have used acutely isolated hair cells. Here we describe a semi-intact preparation of mouse vestibular epithelium for electrophysiological and two-photon microscopy studies.

一个孤立的半完整的上下游鼠标前庭上皮细胞电生理和高分辨率的双光子显微镜

Understanding vestibular hair cells function  under normal conditions, or how trauma, disease, and aging disrupt this  function is a vital step in the ...

科研

神经科学

Labelling and Visualization of Mitochondrial Genome Expression Products in Baker's Yeast Saccharomyces cerevisiae

Mitochondria are essential organelles of eukaryotic cells capable of aerobic respiration. They contain circular genome and gene expression apparatus. A mitochondrial genome of baker&#8217;s yeast encodes eight proteins: three subunits of the cytochrome c oxidase (Cox1p, Cox2p, and Cox3p), three subunits of the ATP synthase (Atp6p, Atp8p, and Atp9p), a subunit of the ubiquinol-cytochrome c oxidoreductase enzyme, cytochrome b (Cytb), and mitochondrial ribosomal protein Var1p. The purpose of the method described here is to specifically label these proteins with 35S methionine, separate them by electrophoresis and visualize the signals as discrete bands on the screen. The procedure involves several steps. First, yeast cells are cultured in a galactose-containing medium until they reach the late logarithmic growth stage. Next, cycloheximide treatment blocks cytoplasmic translation and allows 35S methionine incorporation only in mitochondrial translation products. Then, all proteins are extracted from yeast cells and separated by polyacrylamide gel electrophoresis. Finally, the gel is dried and incubated with the storage phosphor screen. The screen is scanned on a phosphorimager revealing the bands. The method can be applied to compare the biosynthesis rate of a single polypeptide in the mitochondria of a mutant yeast strain versus the wild type, which is useful for studying mitochondrial gene expression defects. This protocol gives valuable information about the translation rate of all yeast mitochondrial mRNAs. However, it requires several controls and additional experiments to make proper conclusions.

Labelling and Visualization of Mitochondrial Genome Expression Products in Baker's Yeast Saccharomyces cerevisiae

Baker&#8217;s yeast mitochondrial genome encodes eight polypeptides. The goal of the current protocol is to label all of them and subsequently visualize them as separate bands.

贝克酵母糖精中线粒体基因组表达产品的标记和可视化

Mitochondria are essential organelles of eukaryotic cells capable of aerobic respiration. They contain circular genome and gene expression apparatus. A ...

生物化学

Three-dimensional Imaging and Analysis of Mitochondria within Human Intraepidermal Nerve Fibers

The goal of this protocol is to study mitochondria within intraepidermal nerve fibers. Therefore, 3D imaging and analysis techniques were developed to isolate nerve-specific mitochondria and evaluate disease-induced alterations of mitochondria in the distal tip of sensory nerves. The protocol combines fluorescence immunohistochemistry, confocal microscopy and 3D image analysis techniques to visualize and quantify nerve-specific mitochondria. Detailed parameters are defined throughout the procedures in order to provide a concrete example of how to use these techniques to isolate nerve-specific mitochondria. Antibodies were used to label nerve and mitochondrial signals within tissue sections of skin punch biopsies, which was followed by indirect immunofluorescence to visualize nerves and mitochondria with a green and red fluorescent signal respectively. Z-series images were acquired with confocal microscopy and 3D analysis software was used to process and analyze the signals. It is not necessary to follow the exact parameters described within, but it is important to be consistent with the ones chosen throughout the staining, acquisition and analysis steps. The strength of this protocol is that it is applicable to a wide variety of circumstances where one fluorescent signal is used to isolate other signals that would otherwise be impossible to study alone.

This protocol uses three-dimensional (3D) imaging and analysis techniques to visualize and quantify nerve-specific mitochondria. The techniques are applicable to other situations where one fluorescent signal is used to isolate a subset of data from another fluorescent signal.

人 Intraepidermal 神经纤维内线粒体三维成像及分析

The goal of this protocol is to study mitochondria within intraepidermal nerve fibers. Therefore, 3D imaging and analysis techniques were developed to ...

In Vivo Imaging of Cx3cr1gfp/gfp Reporter Mice with Spectral-domain Optical Coherence Tomography and Scanning Laser Ophthalmoscopy

Spectral domain optical coherence tomography (SD-OCT) and scanning laser ophthalmoscopy (SLO) are extensively used in experimental ophthalmology. In the present protocol, mice expressing green fluorescent protein (gfp) under the promoter of Cx3cr1 (BALB/c-Cx3cr1gfp/gfp) were used to image microglia cells in vivo in the retina. Microglia are resident macrophages of the retina and have been implicated in several retinal diseases1,2,3,4,5,6. This protocol provides a detailed approach for generation of retinal B-scans, with SD-OCT, and imaging of microglia cell distribution in Cx3cr1gfp/gfp mice with SLO in vivo, using an ophthalmic imaging platform system. The protocol can be used in several reporter mouse lines. However, there are some limitations to the protocol presented here. First, both SLO and SD-OCT, when used in the high-resolution mode, collect data with high axial resolution but the lateral resolution is lower (3.5 &#181;m and 6 &#181;m, respectively). Moreover, the focus and saturation level in SLO is highly dependent on parameter selection and correct alignment of the eye. Additionally, using devices designed for human patients in mice is challenging due to the higher total optical power of the mouse eye compared to the human eye; this can lead to lateral magnification inaccuracies7, which are also dependent on the magnification by the mouse lens among others. However, despite that the axial scan position is dependent upon lateral magnification, the axial SD-OCT measurements are accurate8.

In Vivo Imaging of Cx3cr1gfp/gfp Reporter Mice with Spectral-domain Optical Coherence Tomography and Scanning Laser Ophthalmoscopy

This protocol describes how high-resolution imaging techniques such as spectral domain optical coherence tomography and scanning laser ophthalmoscopy can be utilized in small rodents, using an ophthalmic imaging platform system, to obtain information on retinal thickness and microglial cell distribution, respectively.

在体内图像的Cx3cr1gfp/gfp 记者光谱域光学相干层析成像和扫描激光眼底

Spectral domain optical coherence tomography (SD-OCT) and scanning laser ophthalmoscopy (SLO) are extensively used in experimental ophthalmology. In the ...

医学

In Vitro and In Vivo Detection of Mitophagy in Human Cells, C. Elegans, and Mice

Mitochondria are the powerhouses of cells and produce cellular energy in the form of ATP. Mitochondrial dysfunction contributes to biological aging and a wide variety of disorders including metabolic diseases, premature aging syndromes, and neurodegenerative diseases such as Alzheimer's disease (AD) and Parkinson's disease (PD). Maintenance of mitochondrial health depends on mitochondrial biogenesis and the efficient clearance of dysfunctional mitochondria through mitophagy. Experimental methods to accurately detect autophagy/mitophagy, especially in animal models, have been challenging to develop. Recent progress towards the understanding of the molecular mechanisms of mitophagy has enabled the development of novel mitophagy detection techniques. Here, we introduce several versatile techniques to monitor mitophagy in human cells, Caenorhabditis elegans (e.g., Rosella and DCT-1/ LGG-1 strains), and mice (mt-Keima). A combination of these mitophagy detection techniques, including cross-species evaluation, will improve the accuracy of mitophagy measurements and lead to a better understanding of the role of mitophagy in health and disease.

In Vitro and In Vivo Detection of Mitophagy in Human Cells, C. Elegans, and Mice

Mitophagy, the process of clearing damaged mitochondria, is necessary for mitochondrial homeostasis and health maintenance. This article presents some of the latest mitophagy detection methods in human cells, Caenorhabditis elegans, and mice.

体外和体内检测人类细胞中的 Mitophagy,线虫和小鼠

Mitochondria are the powerhouses of cells and produce cellular energy in the form of ATP. Mitochondrial dysfunction contributes to biological aging and a ...

A Screenable In Vivo Assay for Mitochondrial Modulators Using Transgenic Bioluminescent Caenorhabditis elegans

The multicellular model organism Caenorhabditis elegans is a small nematode of approximately 1 mm in size in adulthood that is genetically and experimentally tractable. It is economical and easy to culture and dispense in liquid medium which makes it well suited for medium-throughput screening. We have previously validated the use of transgenic luciferase expressing C. elegans strains to provide rapid in vivo assessment of the nematode&#8217;s ATP levels.1-3 Here we present the required materials and procedure to carry out bioassays with the bioluminescent C. elegans strains PE254 or PE255 (or any of their derivative strains). The protocol allows for in vivo detection of sublethal effects of drugs that may identify mitochondrial toxicity, as well as for in vivo detection of potential beneficial drug effects. Representative results are provided for the chemicals paraquat, rotenone, oxaloacetate and for four firefly luciferase inhibitory compounds. The methodology can be scaled up to provide a platform for screening drug libraries for compounds capable of modulating mitochondrial function. Pre-clinical evaluation of drug toxicity is often carried out on immortalized cancerous human cell lines which derive ATP mostly from glycolysis and are often tolerant of mitochondrial toxicants.4,5 In contrast, C. elegans depends on oxidative phosphorylation to sustain development into adulthood, drawing a parallel with humans and providing a unique opportunity for compound evaluation in the physiological context of a whole live multicellular organism.

A Screenable In Vivo Assay for Mitochondrial Modulators Using Transgenic Bioluminescent Caenorhabditis elegans

A protocol is described for in vivo detection of effects of mitochondrial inhibitors in the model organism Caenorhabditis elegans and for identification of potential enhancing compounds. This protocol can be used to screen drug libraries for compounds modulating mitochondrial function.

可筛选的<em&gt;在体内</em&gt;测定线粒体调节剂使用转基因生物发光<em&gt;秀丽隐杆线虫</em

The multicellular model organism Caenorhabditis elegans is a small nematode of approximately 1 mm in size in adulthood that is genetically and ...

生物学

Measurement of Mitochondrial Membrane Potential In Vivo using a Genetically Encoded Voltage Indicator

Mitochondrial membrane potential (MMP, &#916;&#936;m) is critical for mitochondrial functions, including ATP synthesis, ion transport, reactive oxygen species (ROS) generation, and the import of proteins encoded by the nucleus. Existing methods for measuring &#916;&#936;m typically use lipophilic cation dyes, such as Rhodamine 800 and tetramethylrhodamine methyl ester (TMRM), but these are limited by low specificity and are not well-suited for in vivo applications. To address these limitations, we have developed a novel protocol utilizing genetically encoded voltage indicators (GEVIs). Genetically encoded voltage indicators (GEVIs), which generate fluorescent signals in response to membrane potential changes, have demonstrated significant potential for monitoring plasma membrane and neuronal potentials. However, their application to mitochondrial membranes remains unexplored. Here, we developed protein-based mitochondrial-targeted GEVIs capable of detecting &#916;&#936;m fluctuations in cells and the motor cortex of living animals. The mitochondrial potential indicator (MPI)offers a non-invasive approach to study &#916;&#936;m dynamics in real-time, providing a method to investigate mitochondrial function under both normal and pathological conditions.

This protocol describes the application of mitochondria-targeted genetically encoded voltage indicators (GEVIs). These GEVIs offer a significant advantage over traditional mitochondrial membrane potential dyes by enabling specific, in vivo, and real-time monitoring of mitochondrial membrane potential.

使用基因编码的电压指示器测量 体内 线粒体膜电位

Mitochondrial membrane potential (MMP, &#916;&#936;m) is critical for mitochondrial functions, including ATP synthesis, ion transport, reactive oxygen ...

定量 秀丽隐杆线 虫和肝癌细胞系中的线粒体自噬

Detection of Mitophagy in Caenorhabditis elegans and Mammalian Cells Using Organelle-Specific Dyes

Mitochondria are essential for various biological functions, including energy production, lipid metabolism, calcium homeostasis, heme biosynthesis, regulated cell death, and the generation of reactive oxygen species (ROS). ROS are vital for key biological processes. However, when uncontrolled, they can lead to oxidative injury, including mitochondrial damage. Damaged mitochondria release more ROS, thereby intensifying cellular injury and the disease state. A homeostatic process named mitochondrial autophagy (mitophagy) selectively removes damaged mitochondria, which are then replaced by new ones. There are multiple mitophagy pathways, with the common endpoint being the breakdown of the damaged mitochondria in lysosomes. 
Several methodologies, including genetic sensors, antibody immunofluorescence, and electron microscopy, use this endpoint to quantify mitophagy. Each method for examining mitophagy has its advantages, such as specific tissue/cell targeting (with genetic sensors) and great detail (with electron microscopy). However, these methods often require expensive resources, trained personnel, and a lengthy preparation time before the actual experiment, such as for creating transgenic animals. Here, we present a cost-effective alternative for measuring mitophagy using commercially available fluorescent dyes targeting mitochondria and lysosomes. This method effectively measures mitophagy in the nematode Caenorhabditis elegans and human liver cells, which indicates its potential efficiency in other model systems.

作者聚焦：使用细胞器特异性染料检测秀丽隐杆线虫和哺乳动物细胞中的线粒体自噬  

Exploring mitophagy through electron microscopy, genetic sensors, and immunofluorescence requires costly equipment, skilled personnel, and a significant time investment. Here, we demonstrate the efficacy of a commercial fluorescence dye kit in quantifying the mitophagy process in both Caenorhabditis elegans and a liver cancer cell line.

使用细胞器特异性染料检测 秀丽隐杆 线虫和哺乳动物细胞中的线粒体自噬

Mitochondria are essential for various biological functions, including energy production, lipid metabolism, calcium homeostasis, heme biosynthesis, ...

近红外（NIR）光增加线粒体功能的标志物的表达，在鼠标前庭上皮细胞

摘要

探索更多视频

一个孤立的半完整的上下游鼠标前庭上皮细胞电生理和高分辨率的双光子显微镜

贝克酵母糖精中线粒体基因组表达产品的标记和可视化

人 Intraepidermal 神经纤维内线粒体三维成像及分析

在体内图像的Cx3cr1^gfp/gfp 记者光谱域光学相干层析成像和扫描激光眼底

体外和体内检测人类细胞中的 Mitophagy,线虫和小鼠

可筛选的

使用基因编码的电压指示器测量体内线粒体膜电位

使用细胞器特异性染料检测秀丽隐杆线虫和哺乳动物细胞中的线粒体自噬

使用细胞器特异性染料检测秀丽隐杆线虫和哺乳动物细胞中的线粒体自噬

使用细胞器特异性染料检测秀丽隐杆线虫和哺乳动物细胞中的线粒体自噬

近红外（NIR）光增加线粒体功能的标志物的表达，在鼠标前庭上皮细胞

摘要

探索更多视频

一个孤立的半完整的上下游鼠标前庭上皮细胞电生理和高分辨率的双光子显微镜

贝克酵母糖精中线粒体基因组表达产品的标记和可视化

人 Intraepidermal 神经纤维内线粒体三维成像及分析

在体内图像的Cx3cr1gfp/gfp 记者光谱域光学相干层析成像和扫描激光眼底

体外和体内检测人类细胞中的 Mitophagy,线虫和小鼠

可筛选的

使用基因编码的电压指示器测量 体内 线粒体膜电位

使用细胞器特异性染料检测 秀丽隐杆 线虫和哺乳动物细胞中的线粒体自噬

使用细胞器特异性染料检测 秀丽隐杆 线虫和哺乳动物细胞中的线粒体自噬

使用细胞器特异性染料检测 秀丽隐杆 线虫和哺乳动物细胞中的线粒体自噬

在体内图像的Cx3cr1^gfp/gfp 记者光谱域光学相干层析成像和扫描激光眼底

使用基因编码的电压指示器测量体内线粒体膜电位

使用细胞器特异性染料检测秀丽隐杆线虫和哺乳动物细胞中的线粒体自噬

使用细胞器特异性染料检测秀丽隐杆线虫和哺乳动物细胞中的线粒体自噬

使用细胞器特异性染料检测秀丽隐杆线虫和哺乳动物细胞中的线粒体自噬