这项工作得到了中国国家自然科学基金 （82204088， 82273669） 和中国山东省自然科学基金 （ZR2021QH209） 的财政支持。

我们使用雄性 C57BL/6J 小鼠 （年龄：6-8 周;体重：20-22 g） 进行所有行为测试。小鼠经受稳定的条件（即温度，23 ± 1 °C;湿度，55% ± 5%，以及 7：00 开灯的 12/12 小时明暗循环）。所有行为测试均在 10：00 至 17：00 之间进行。所有动物实验均经青岛大学动物实验专业委员会伦理委员会批准。1 周的驯化期后，所有小鼠均暴露于可吸入环境污染物。
1. 接触污染物
<ol><li>滴注鼻内给药<ol><li>将可吸入的环境污染物溶解在 0.9% 盐水溶液中。使用 5% 硫酸锌溶液一次，已被证明会导致嗅觉功能障碍17。对于对照小鼠，施用 0.9% 生理盐水溶液。</li>
		<li>通过腹膜内注射 1% 戊巴比妥钠 18 麻醉小鼠，并使用脚趾捏反射评估麻醉深度。将兽医药膏涂抹在老鼠的眼睛上以防止干燥。将鼠标仰卧在倾斜的表面上，头部朝下。</li>
		<li>使用移液枪，在小鼠的一个鼻孔中注入 10 μL 溶液，让溶液自然吸入鼻腔。</li>
		<li>为确保鼠标的健康并防止任何潜在的不适，请在 10 分钟间隔后对另一个鼻孔重复吸气。</li>
		<li>在鼻内给药后 3 天，通过滴注在小鼠中，进行埋藏食物试验。</li>
	</ol></li>
</ol>2. 埋藏食物测试
<ol><li>在测试前 18-24 小时通过从家笼的食物料斗中取出所有食物颗粒来剥夺食物。更换老鼠的床上用品。不要取下水瓶。</li>
	<li>如下所述布置手术台。<ol><li>在测试开始前 1 小时，将装有小鼠的笼子带到手术室休息。</li>
		<li>在此期间安排手术室。使用透明 PVC 标准鼠标笼并将其标记为 A，因为这些笼子可以营造熟悉的环境。使用并将测试笼标记为 B，这是一种透明的 PVC 标准鼠笼。将实验后用于放置小鼠的公共笼子标记为笼子 C。</li>
		<li>用 3 厘米的垫料覆盖笼子 A 和 B，并用尺子测量它们。并排排列笼子，每个笼子之间的距离为 0.5 m（图 1）。</li>
		<li>将实验区域定义为半径为 2 m 的区域，中心是笼子的中心。将 2 m 范围之外的区域定义为观察区域。</li>
		<li>将笼子 C 和装有未经测试的小鼠的笼子尽可能远离实验区域。</li>
	</ol></li>
	<li>记录寻找食物的时间，如下所述。<ol><li>在笼子 B 中随机选择一个位置，将食物埋在被褥表面以下 1 厘米处，并将被褥表面抹平。</li>
		<li>将鼠标放入笼 A 中 4 分钟。定时结束时将小鼠转移到笼 B，打开视频设备，返回观察区域。</li>
		<li>当老鼠用前爪捡起食物块时，停止视频录制。在某些情况下，可以看到老鼠低头吃东西。这种行为也表明试验成功，即使鼠标没有用前爪夹住食物。</li>
		<li>记录数据。记录从接触笼 B 底部的垫子到为每只老鼠找到食物的时间。如果小鼠在 4 分钟后没有找到食物，则将找到时间记录为 240 秒的延迟时间。</li>
		<li>测试后将鼠标放入笼 C 中。</li>
		<li>从笼子 B 中取出饲料并将其放入密封袋中。更换笼子 B 中的食物，更换床上用品后测试下一只鼠标，如下所述。<ol><li>对于饲养在同一笼子中的小鼠，在笼子 B 中使用同一组垫料进行测试。对于不同笼子里的老鼠，用酒精清洗笼子，更换垫料。</li>
		</ol></li>
		<li>实验后向小鼠添加饲料和水。1 天后进行社会气味辨别实验。 注意：整个过程需要戴口罩，每只鼠标完成实验后需要更换透明手套，以尽可能避免气味交叉。保持运动的幅度尽可能小，以避免鼠标压力。</li>
	</ol></li>
</ol>3. 社会气味辨别实验
<ol><li>尿液采集<ol><li>分别收集性成熟雄性小鼠和雌性小鼠的尿液，并分装到等体积的试管中19。装在 2 mL 微量离心管中，每管含 300 μL 尿液。</li>
		<li>将样品储存在 -80°C 直至使用。解冻后摇动试管以均匀分布样品。不要反复解冻和冷冻样品。</li>
	</ol></li>
	<li>如下所述布置手术台。<ol><li>在测试开始前 1 小时，将装有小鼠的笼子带到手术室休息。</li>
		<li>在此期间，按照步骤 2.2.2-2.2.5 中的说明安排手术室。</li>
	</ol></li>
	<li>记录查找尿液的时间，如下所述。<ol><li>在笼 B 的两个宽边周围用胶带制作一个凹槽，其大小足以容纳含有 300 μL 男性和女性尿液的微量离心管。放置试管并暂时关闭两个试管盖。</li>
		<li>将鼠标放入笼 A 中并设置 4 分钟的倒计时。在计时结束时打开笼 B 的两管。</li>
		<li>将鼠标转移到笼 B 的中间位置，与两根管保持相同的距离，然后打开录像设备，轻轻缓慢地撤退到观察区域。</li>
		<li>当鼠标嗅到管壁/嘴巴甚至管内时，管子被成功闻到。此时按下秒表，将时间记录为闻到管子的时间（M s），然后继续记录停留时间（X s），直到鼠标离开管子。</li>
		<li>继续计时，嗅探另一根管子。当鼠标嗅到管壁/嘴巴甚至管内时，管子被成功闻到。按下秒表并记录闻到另一根管子所需的时间 （N s），然后继续记录停留时间 （Y s），直到鼠标离开管子。</li>
		<li>实验结束时将鼠标转移到笼 C。</li>
		<li>每次小鼠测试后更换笼 B 的尿管和床上用品。根据性别将用过的大鼠尿液放入干净的袋子中。</li>
		<li>如上所述更换手套并测试下一只鼠标。</li>
	</ol></li>
	<li>在无明显气味的实验室进行实验。避免使用散发强烈气味的个人产品。在整个手术过程中戴上手套和口罩，以尽可能避免异味传播。保持运动的幅度尽可能小，以避免鼠标压力。</li>
</ol>

评估吸入环境污染物导致小鼠嗅觉损害

<ol>
	<li>Sch&#228;fer, L., Schriever, V. A., Croy, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Human+olfactory+dysfunction:+causes+and+consequences.">Human olfactory dysfunction: causes and consequences.</a> Cell Tissue Res. 383 (1), 569-579 (2021).</li><li>Keller, A., Malaspina, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hidden+consequences+of+olfactory+dysfunction:+a+patient+report+series.">Hidden consequences of olfactory dysfunction: a patient report series.</a> BMC Ear Nose Throat Disord. 13 (1), 8 (2013).</li><li>Schroeter, J. D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Application+of+physiological+computational+fluid+dynamics+models+to+predict+interspecies+nasal+dosimetry+of+inhaled+acrolein.">Application of physiological computational fluid dynamics models to predict interspecies nasal dosimetry of inhaled acrolein.</a> Inhal Toxicol. 20 (3), 227-243 (2008).</li><li>Schroeter, J. D., Garcia, G. J., Kimbell, J. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+computational+fluid+dynamics+approach+to+assess+interhuman+variability+in+hydrogen+sulfide+nasal+dosimetry.">A computational fluid dynamics approach to assess interhuman variability in hydrogen sulfide nasal dosimetry.</a> Inhal Toxicol. 22 (4), 277-286 (2010).</li><li>Keyhani, K., Scherer, P. W., Mozell, M. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Numerical+simulation+of+airflow+in+the+human+nasal+cavity.">Numerical simulation of airflow in the human nasal cavity.</a> J Biomech Eng. 117 (4), 429-441 (1995).</li><li>Hahn, I., Scherer, P. W., Mozell, M. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Velocity+profiles+measured+for+airflow+through+a+large-scale+model+of+the+human+nasal+cavity.">Velocity profiles measured for airflow through a large-scale model of the human nasal cavity.</a> J Appl Physiol. 75 (5), 2273-2287 (1993).</li><li>Zhang, Z., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Exposure+to+particulate+matter+air+pollution+and+Anosmia.">Exposure to particulate matter air pollution and Anosmia.</a> JAMA Netw Open. 4 (5), e2111606 (2021).</li><li>Ekstr&#246;m, I. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Environmental+air+pollution+and+olfactory+decline+in+aging.">Environmental air pollution and olfactory decline in aging.</a> Environ Health Perspect. 130 (2), 27005 (2022).</li><li>Adams, D. R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nitrogen+dioxide+pollution+exposure+is+associated+with+olfactory+dysfunction+in+older+U.S.+adults.">Nitrogen dioxide pollution exposure is associated with olfactory dysfunction in older U.S. adults.</a> Int Forum Allergy Rhinol. 6 (12), 1245-1252 (2016).</li><li>Prah, J. D., Benignus, V. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Decrements+in+olfactory+sensitivity+due+to+ozone+exposure.">Decrements in olfactory sensitivity due to ozone exposure.</a> Percept Mot Skills. 48 (1), 317-318 (1979).</li><li>Shi, Z., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chronic+exposure+to+environmental+pollutant+ammonia+causes+damage+to+the+olfactory+system+and+behavioral+abnormalities+in+mice.">Chronic exposure to environmental pollutant ammonia causes damage to the olfactory system and behavioral abnormalities in mice.</a> Environ Sci Technol. 57 (41), 15412-15421 (2023).</li><li>Hern&#225;ndez-Soto, R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chronic+intermittent+hypoxia+alters+main+olfactory+bulb+activity+and+olfaction.">Chronic intermittent hypoxia alters main olfactory bulb activity and olfaction.</a> Exp Neurol. 340, 113653 (2021).</li><li>Islam, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Odor+preference+and+olfactory+memory+are+impaired+in+Olfaxin-deficient+mice.">Odor preference and olfactory memory are impaired in Olfaxin-deficient mice.</a> Brain Res. 1688, 81-90 (2018).</li><li>Wang, H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Inducible+and+conditional+activation+of+ERK5+MAP+kinase+rescues+mice+from+cadmium-induced+olfactory+memory+deficits.">Inducible and conditional activation of ERK5 MAP kinase rescues mice from cadmium-induced olfactory memory deficits.</a> Neurotoxicology. 81, 127-136 (2020).</li><li>Chamero, P., Leinders-Zufall, T., Zufall, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=From+genes+to+social+communication:+molecular+sensing+by+the+vomeronasal+organ.">From genes to social communication: molecular sensing by the vomeronasal organ.</a> Trends Neurosci. 35 (10), 597-606 (2012).</li><li>Mohrhardt, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Signal+detection+and+coding+in+the+accessory+olfactory+system.">Signal detection and coding in the accessory olfactory system.</a> Chem Senses. 43 (9), 667-695 (2018).</li><li>Liu, X., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Type+3+adenylyl+cyclase+in+the+MOE+is+involved+in+learning+and+memory+in+mice.">Type 3 adenylyl cyclase in the MOE is involved in learning and memory in mice.</a> Behav Brain Res. 383, 112533 (2020).</li><li>Li, X., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Polyhexamethylene+guanidine+aerosol+triggers+pulmonary+fibrosis+concomitant+with+elevated+surface+tension+via+inhibiting+pulmonary+surfactant.">Polyhexamethylene guanidine aerosol triggers pulmonary fibrosis concomitant with elevated surface tension via inhibiting pulmonary surfactant.</a> J Hazard Mater. 420, 126642 (2021).</li><li>Yang, M., Crawley, J. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Simple+behavioral+assessment+of+mouse+olfaction.">Simple behavioral assessment of mouse olfaction.</a> Curr Protoc Neurosci. , (2009).</li><li>Huang, K. L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Zinc+oxide+nanoparticles+induce+eosinophilic+airway+inflammation+in+mice.">Zinc oxide nanoparticles induce eosinophilic airway inflammation in mice.</a> J Hazard Mater. 297, 304-312 (2015).</li><li>Burd, G. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Morphological+study+of+the+effects+of+intranasal+zinc+sulfate+irrigation+on+the+mouse+olfactory+epithelium+and+olfactory+bulb.">Morphological study of the effects of intranasal zinc sulfate irrigation on the mouse olfactory epithelium and olfactory bulb.</a> Microsc Res Tech. 24 (3), 195-213 (1993).</li><li>Zou, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Methods+to+measure+olfactory+behavior+in+mice.">Methods to measure olfactory behavior in mice.</a> Curr Protoc Toxicol. 63, 1-21 (2015).</li><li>Ryalls, J. M. W., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Anthropogenic+air+pollutants+reduce+insect-mediated+pollination+services.">Anthropogenic air pollutants reduce insect-mediated pollination services.</a> Environ Pollut. 297, 118847 (2022).</li><li>Schiffman, S. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Livestock+odors:+implications+for+human+health+and+well-being.">Livestock odors: implications for human health and well-being.</a> J Anim Sci. 76 (5), 1343-1355 (1998).</li><li>Albrechet-Souza, L., Gilpin, N. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+predator+odor+avoidance+model+of+post-traumatic+stress+disorder+in+rats.">The predator odor avoidance model of post-traumatic stress disorder in rats.</a> Behav Pharmacol. 30, 105-114 (2019).</li><li>Davies, D. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Inactivation+of+medial+prefrontal+cortex+or+acute+stress+impairs+odor+span+in+rats.">Inactivation of medial prefrontal cortex or acute stress impairs odor span in rats.</a> Learn Mem. 20 (12), 665-669 (2013).</li><li>Landers, M. S., Sullivan, R. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+development+and+neurobiology+of+infant+attachment+and+fear.">The development and neurobiology of infant attachment and fear.</a> Dev Neurosci. 34 (2-3), 101-114 (2012).</li><li>Drobyshevsky, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Antenatal+insults+modify+newborn+olfactory+function+by+nitric+oxide+produced+from+neuronal+nitric+oxide+synthase.">Antenatal insults modify newborn olfactory function by nitric oxide produced from neuronal nitric oxide synthase.</a> Exp Neurol. 237 (2), 427-434 (2012).</li><li>Arbuckle, E. P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Testing+for+odor+discrimination+and+habituation+in+mice.">Testing for odor discrimination and habituation in mice.</a> J Vis Exp. (99), e52615 (2015).</li></ol>

作者声明没有利益冲突。

本文介绍了两种旨在快速评估小鼠嗅觉障碍的基本方案。各种可吸入环境污染物导致小鼠不同程度的嗅觉功能障碍。埋藏食物测试用于评估检测挥发性气味的能力，而社会气味辨别实验用于评估动物辨别和区分各种社会气味的能力。这里的协议用于评估环境污染物对嗅觉功能的毒性影响。
吸入的环境污染物，包括臭氧、氨和柴油机尾气，具有可能影响小鼠在样品嗅探过程中?...

嗅觉障碍已成为一个值得注意的公共卫生问题，并且与神经退行性疾病的风险增加独立相关。这种情况会对整体健康状况产生不利影响，导致抑郁症状的发展，并导致生活质量下降。它的影响在对食物的感知改变、社交沟通的阻碍和负面情绪的加剧中得到了突出的体现1。包括鼻窦疾病、上呼吸道感染和创伤性脑损伤在内的各种因素被认为是导致人类嗅觉受损的原因2。值得注意的是，PM2.5 等可吸入环境污染物（估计在 2% 到 16% 之间）通过吸入的空气进入人体，穿过鼻腔，并到达特定的嗅觉专用区域，在那里它们被沉积 3,4,5,6,7。最近的研究结果表明，可吸入的环境污染物，包括 PM2.5 和氨，确实会伤害嗅觉感觉神经元 8,9,10。然而，需要进一步验证以确定这种损伤是否直接导致嗅觉功能障碍。因此，仔细评估可吸入环境污染物对嗅觉功能的影响尤为重要。
目前，许多研究实验室使用小鼠作为替代脊椎动物模型进行行为实验，旨在理解嗅觉功能的变化 11,12,13,14。小鼠被选为研究脊椎动物化学交流的首选模型系统，它表现出对觅食和社会交流至关重要的非凡嗅觉敏感性15。此外，用于观察和影响小鼠行为的一系列不断发展的工具使该物种对嗅觉功能研究极具吸引力16。
在这项研究中，我们采用埋藏食物测试和社会气味鉴别实验来评估暴露于可吸入环境污染物的小鼠模型的嗅觉损害。为了提高评估的精度，我们选择了最具代表性的方法来评估嗅觉功能。我们系统地改进了这种方法，以确保简单和清晰，使我们能够有效地衡量可吸入环境污染物引起的嗅觉功能障碍的程度。

<table><tbody><tr><td>0.5-10 &amp;mu;L&amp;nbsp; adjustable micropipette</td><td>Eppendorf, Germany</td><td>3123000225</td><td>Intranasal instillation</td></tr><tr><td>0.9% saline solution</td><td>Solarbio</td><td>7647-14-5</td><td>Dissolve pollutants</td></tr><tr><td>Anhydrous zinc sulfate</td><td>Macklin</td><td>7733-02-0</td><td>Expose mice</td></tr><tr><td>Centrifuge tube (2 mL)</td><td>Biosharp Incorporated</td><td>BS-20-M</td><td>Place urine</td></tr><tr><td>Electronic balance</td><td>Changzhou Ohaus Co.</td><td>EX125DZH</td><td>Weight anesthetics and pollutants</td></tr><tr><td>GraphPad Prism</td><td>GraphPad Software</td><td>8.0.1</td><td>statistic analysis</td></tr><tr><td>Handheld Dust detector</td><td>TSI&amp;nbsp;Incorporated</td><td>DuatTrak &lt;img alt=&quot;Equation 1&quot; src=&quot;/files/ftp_upload/66818/66818eq01.jpg&quot; style=&quot;margin: auto;&quot;/&gt;8532</td><td>Inhalation-exposed mice</td></tr><tr><td>Video recording equipment</td><td>Apple Inc.</td><td>iPhone 6s Plus</td><td>The activity time of mice was recorded</td></tr><tr><td>Vortex mixer</td><td>Haimen Kylin-Bell Lab Instruments Co.</td><td>Vortex-5&amp;nbsp;</td><td>Mix solution</td></tr></tbody></table>

studying the effects of inhaled environmental pollutants on olfactory function in mice

嗅觉障碍是一个重大的公共卫生问题，可独立预测神经退行性疾病的风险。吸入环境污染物暴露可能会损害嗅觉;因此，迫切需要评估吸入环境污染物暴露对嗅觉影响的方法。小鼠是嗅觉实验的理想模型，因为它们的高度发达的嗅觉系统和行为特征。为评估吸入环境污染物暴露对小鼠嗅觉功能的影响，提供了详细的埋藏食物检测和社会气味鉴别实验，包括实验准备、实验设施的选择和建设、检测过程和时间指标。同时，讨论了计时设备、操作细节和实验环境，以确保检测的成功。以硫酸锌为处理方法，证明了实验方法的可行性。该协议提供了一个简单明了的操作过程，用于评估吸入环境污染物对小鼠嗅觉功能的影响。

可吸入的环境污染物会损害小鼠的嗅觉功能。焚化炉和机动车辆排放的大气锌已被证明是一种吸入污染物，可导致过敏性肺部炎症20。硫酸锌被认为是导致嗅觉功能障碍的典型化合物之一21。因此，我们以硫酸锌为处理，通过鼻内滴注暴露小鼠，并使用埋藏食物测试和社会气味鉴别实验进行测试。在食物埋葬试验中，与对照组相比，暴露于硫酸锌的小鼠找到食物...

作者聚焦：评估空气污染物的嗅觉影响 - 埋藏的食物和社会气味测试

本文对埋藏食物试验和社会气味鉴别实验进行了详细说明，以评估吸入环境污染物暴露对小鼠嗅觉功能的影响。

研究吸入环境污染物对小鼠嗅觉功能的影响

Olfactory impairment is a significant public health problem and independently predicts the risk of neurodegenerative diseases. Inhaled environmental ...

studying-effects-inhaled-environmental-pollutants-on-olfactory

Research

JoVE Journal

Environment

Studying the Effects of Inhaled Environmental Pollutants on Olfactory Function in Mice

665 次观看.  Qingdao University. 我们的研究方向是环境毒理学。它要求提供两种行为测试方法，用于启动嗅觉功能的影响或可遗传的环境污染物。我们通过某些食物测试和社会秩序判别实验提供了两种简单有效的方法来确定可遗传环境污染物对嗅觉功能的毒性。我们可以通过说服变量测试和社会秩序区分实验来综合评估可遗传环境污染物对嗅觉功能的影响。我们的协议使用视...

视频: 作者聚焦：评估空气污染物的嗅觉影响 - 埋藏的食物和社会气味测试

Testing for Odor Discrimination and Habituation in Mice

This video demonstrates a technique to establish the presence of a normally functioning olfactory system in a mouse. The test helps determine whether the mouse can discriminate between non-social odors and social odors, whether the mouse habituates to a repeatedly presented odor, and whether the mouse demonstrates dishabituation when presented with a novel odor. Since many social behavior tests measure the experimental animal&#8217;s response to a familiar or novel mouse, false positives can be avoided by establishing that the animals can detect and discriminate between social odors. There are similar considerations in learning tests such as fear conditioning that use odor to create a novel environment or olfactory cues as an associative stimulus. Deficits in the olfactory system would impair the ability to distinguish between contexts and to form an association with an olfactory cue during fear conditioning. 
In the odor habitation/dishabituation test, the mouse is repeatedly presented with several odors. Each odor is presented three times for two minutes. The investigator records the sniffing time directed towards the odor as the measurement of olfactory responsiveness. A typical mouse shows a decrease in response to the odor over repeated presentations (habituation). The experimenter then presents a novel odor that elicits increased sniffing towards the new odor (dishabituation). After repeated presentation of the novel odor the animal again shows habituation. This protocol involves the presentation of water, two or more non-social odors, and two social odors. In addition to reducing experimental confounds, this test can provide information on the function of the olfactory systems of new knockout, knock-in, and conditional knockout mouse lines.

This manuscript describes a protocol to examine the olfactory system of rodents. The olfactory habituation/dishabituation test will allow investigators to determine whether a mouse habituates to a repeatedly presented odor and whether the mouse demonstrates dishabituation when presented a novel odor.

在小鼠试验的气味辨别和习惯化

This video demonstrates a technique to establish the presence of a normally functioning olfactory system in a mouse. The test helps determine whether the ...

科研

行为学

Simple and Computer-assisted Olfactory Testing for Mice

Olfaction is highly conserved among species and is required for reproduction and survival.
In humans, olfaction is also one of the senses that is affected with aging and is a strong predictor of neurodegenerative diseases. Thus, olfaction testing is used as a non-invasive diagnostic method to detect neurological deficits early on. In order to understand the mechanisms underlying olfactory network susceptibility, olfactory research in rodents has gained momentum in the past decade.
Here, we present a very simple, time efficient and reproducible olfactory testing method of innate odor perception and sensitivity in mice without the need of any prior food or water restriction. The tests are performed in a familiar environment to the mice, require only the scents and a 2 min session of odorant exposure. The analysis is performed, post-hoc, using computer-assisted commands on ImageJ and can be, therefore, carried out from start to end by one researcher.
This protocol does not require any special hardware or setup and is indicated for any laboratory interested in testing olfactory perception and sensitivity.

We present a simple and unbiased olfactory test in mice. With this protocol olfactory discrimination, preference, avoidance and sensitivity to a novel odor as compared to water can be assessed in single behavioral sessions. This method is indicated for a single experimenter and analysis is based on computer-assisted video processing.

简单和计算机辅助测试嗅觉的小鼠

Olfaction is highly conserved among species and is required for reproduction and survival.
In humans, olfaction is also one of the senses that is affected ...

Olfactory Behavioral Testing in the Adult Mouse

The rodent olfactory system is of increasing interest to scientists, studied, in part, in systems biology because of its stereotyped, yet accessible circuitry. In addition, this area's unique ability to generate new neurons throughout an organism's lifetime makes it an attractive system for developmental and regenerative biologists alike. Such interest necessitates a means for a quick, yet reliable assessment of olfactory function. Many tests of olfactory ability are complex, variable or not specifically designed for mice. Also, some tests are sensitive to memory deficits as well as defects in olfactory abilities, confounding obtained results.
Here, we describe a simple battery of tests designed to identify defects in olfactory sensitivity and preference. First, an initial general health assessment allows for the identification of animals suitable for further testing. Second, mice are exposed to various dilutions of scents to ascertain whether there is a threshold difference. Third, mice are presented with various scents, both attractive and aversive, that allow for the assessment of olfactory preference. These simple studies should make the initial characterization of olfactory behavior accessible for labs of varied resources and expertise.

Fundamental, yet unique properties of the rodent olfactory system have led to its increasing study among biologists. A relatively simple assessment of its function is then also needed. Here we describe sensitive tests for the characterization of mouse olfactory sensitivity and preference.

在成年小鼠的嗅觉行为测试

The rodent olfactory system is of increasing interest to scientists, studied, in part, in systems biology because of its stereotyped, yet accessible ...

生物学

Olfactory Assays for Mouse Models of Neurodegenerative Disease

In many neurodegenerative diseases and particularly in Parkinson&#8217;s disease, deficits in olfaction are reported to occur early in the disease process and may be a useful behavioral marker for early detection. Earlier detection in neurodegenerative disease is a major goal in the field because this is when neuroprotective therapies have the best potential to be effective. Therefore, in preclinical studies testing novel neuroprotective strategies in rodent models of neurodegenerative disease, olfactory assessment could be highly useful in determining therapeutic potential of compounds and translation to the clinic. In the present study we describe a battery of olfactory assays that are useful in measuring olfactory function in mice. The tests presented in this study were chosen because they measure olfaction abilities in mice related to food odors, social odors, and non-social odors. These tests have proven useful in characterizing novel genetic mouse models of Parkinson&#8217;s disease as well as in testing potential disease-modifying therapies.

Impairment in olfactory function is a common feature in many neurodegenerative disorders including Parkinson, Alzheimer, and Huntington diseases. In the present article, we describe a set of tests for assessing olfaction discrimination and detection in mice that can be used to measure olfactory abilities in mouse models of neurodegenerative diseases.

嗅觉测定的神经退行性疾病的小鼠模型

In many neurodegenerative diseases and particularly in Parkinson&#8217;s disease, deficits in olfaction are reported to occur early in the disease process ...

神经科学

Automated Measurement of Pulmonary Emphysema and Small Airway Remodeling in Cigarette Smoke-exposed Mice

COPD is projected to be the third most common cause of mortality world-wide by 2020(1). Animal models of COPD are used to identify molecules that contribute to the disease process and to test the efficacy of novel therapies for COPD. Researchers use a number of models of COPD employing different species including rodents, guinea-pigs, rabbits, and dogs(2). However, the most widely-used model is that in which mice are exposed to cigarette smoke. Mice are an especially useful species in which to model COPD because their genome can readily be manipulated to generate animals that are either deficient in, or over-express individual proteins. Studies of gene-targeted mice that have been exposed to cigarette smoke have provided valuable information about the contributions of individual molecules to different lung pathologies in COPD(3-5). Most studies have focused on pathways involved in emphysema development which contributes to the airflow obstruction that is characteristic of COPD. However, small airway fibrosis also contributes significantly to airflow obstruction in human COPD patients(6), but much less is known about the pathogenesis of this lesion in smoke-exposed animals. To address this knowledge gap, this protocol quantifies both emphysema development and small airway fibrosis in smoke-exposed mice. This protocol exposes mice to CS using a whole-body exposure technique, then measures respiratory mechanics in the mice, inflates the lungs of mice to a standard pressure, and fixes the lungs in formalin. The researcher then stains the lung sections with either Gill&#8217;s stain to measure the mean alveolar chord length (as a readout of emphysema severity) or Masson&#8217;s trichrome stain to measure deposition of extracellular matrix (ECM) proteins around small airways (as a readout of small airway fibrosis). Studies of the effects of molecular pathways on both of these lung pathologies will lead to a better understanding of the pathogenesis of COPD.

The goal of this protocol is to provide automated methods to quantify chronic lung pathologies in a murine model of COPD. The protocol includes exposing mice to cigarette smoke (CS), measuring pulmonary function, inflating the lungs, and using morphometry methods to measure emphysema and small airway remodeling in mice.

肺气肿，小气道重塑的香烟烟雾暴露小鼠自动测量

COPD is projected to be the third most common cause of mortality world-wide by 2020(1).  Animal models of COPD are used to identify molecules that ...

医学

Whole Mount Labeling of Cilia in the Main Olfactory System of Mice

The mouse olfactory system comprises 6-10 million olfactory sensory neurons in the epithelium lining the nasal cavity. Olfactory neurons extend a single dendrite to the surface of the epithelium, ending in a structure called dendritic knob. Cilia emanate from this knob into the mucus covering the epithelial surface. The proteins of the olfactory signal transduction cascade are mainly localized in the ciliary membrane, being in direct contact with volatile substances in the environment. For a detailed understanding of olfactory signal transduction, one important aspect is the exact morphological analysis of signaling protein distribution. Using light microscopical approaches in conventional cryosections, protein localization in olfactory cilia is difficult to determine due to the density of ciliary structures. To overcome this problem, we optimized an approach for whole mount labeling of cilia, leading to improved visualization of their morphology and the distribution of signaling proteins. We demonstrate the power of this approach by comparing whole mount and conventional cryosection labeling of Kirrel2. This axon-guidance adhesion molecule is known to localize in a subset of sensory neurons and their axons in an activity-dependent manner. Whole mount cilia labeling revealed an additional and novel picture of the localization of this protein.

Cilia of olfactory sensory neurons contain proteins of the signal transduction cascade, but a detailed spatial analysis of their distribution is difficult in cryosections. We describe here an optimized approach for whole mount labeling and en face visualization of ciliary proteins.

纤毛对小鼠的主要嗅觉系统整装标签

The mouse olfactory system comprises 6-10 million olfactory sensory neurons in the epithelium lining the nasal cavity. Olfactory neurons extend a single ...

An Objective and Reproducible Test of Olfactory Learning and Discrimination in Mice

Olfaction is the predominant sensory modality in mice and influences many important behaviors, including foraging, predator detection, mating, and parenting. Importantly, mice can be trained to associate novel odors with specific behavioral responses to provide insight into olfactory circuit function. This protocol details the procedure for training mice on a Go/No-Go operant learning task. In this approach, mice are trained on hundreds of automated trials daily for 2&#8211;4 weeks and can then be tested on novel Go/No-Go odor pairs to assess olfactory discrimination, or be used for studies on how odor learning alters the structure or function of the olfactory circuit. Additionally, the mouse olfactory bulb (OB) features ongoing integration of adult-born neurons. Interestingly, olfactory learning increases both the survival and synaptic connections of these adult-born neurons. Therefore, this protocol can be combined with other biochemical, electrophysiological, and imaging techniques to study learning and activity-dependent factors that mediate neuronal survival and plasticity.

Here, we train mice on an associative learning task to test odor discrimination. This protocol also allows for studies on learning-induced structural changes in the brain.

小鼠嗅觉学习和鉴别的客观和可重复性试验

Olfaction is the predominant sensory modality in mice and influences many important behaviors, including foraging, predator detection, mating, and ...

Generation of a Chronic Obstructive Pulmonary Disease Model in Mice by Repeated Ozone Exposure

Chronic obstructive pulmonary disease (COPD) is characterized by persistent airflow limitation and lung parenchymal destruction. It has a very high incidence in aging populations. The current conventional therapies for COPD focus mainly on symptom-modifying drugs; thus, the development of new therapies is urgently needed. Qualified animal models of COPD could help to characterize the underlying mechanisms and can be used for new drug screening. Current COPD models, such as lipopolysaccharide (LPS) or the porcine pancreatic elastase (PPE)-induced emphysema model, generate COPD-like lesions in the lungs and airways but do not otherwise resemble the pathogenesis of human COPD. A cigarette smoke (CS)-induced model remains one of the most popular because it not only simulates COPD-like lesions in the respiratory system, but it is also based on one of the main hazardous materials that causes COPD in humans. However, the time-consuming and labor-intensive aspects of the CS-induced model dramatically limit its application in new drug screening. In this study, we successfully generated a new COPD model by exposing mice to high levels of ozone. This model demonstrated the following: 1) decreased forced expiratory volume 25, 50, and 75/forced vital capacity (FEV25/FVC, FEV50/FVC, and FEV75/FVC), indicating the deterioration of lung function; 2) enlarged lung alveoli, with lung parenchymal destruction; 3) reduced fatigue time and distance; and 4) increased inflammation. Taken together, these data demonstrate that the ozone exposure (OE) model is a reliable animal model that is similar to humans because ozone overexposure is one of the etiological factors of COPD. Additionally, it only took 6 - 8 weeks, based on our previous work, to create an OE model, whereas it requires 3 - 12 months to induce the cigarette smoke model, indicating that the OE model might be a good choice for COPD research.

This study describes the successful generation of a new chronic obstructive pulmonary disease (COPD) animal model by repeatedly exposing mice to high concentrations of ozone.

反复臭氧暴露对小鼠慢性阻塞性肺疾病模型的产生

Chronic obstructive pulmonary disease (COPD) is characterized by persistent airflow limitation and lung parenchymal destruction. It has a very high ...

Evaluation of Respiratory System Mechanics in Mice using the Forced Oscillation Technique

The forced oscillation technique (FOT) is a powerful, integrative and translational tool permitting the experimental assessment of lung function in mice in a comprehensive, detailed, precise and reproducible manner. It provides measurements of respiratory system mechanics through the analysis of pressure and volume signals acquired in reaction to predefined, small amplitude, oscillatory airflow waveforms, which are typically applied at the subject's airway opening. The present protocol details the steps required to adequately execute forced oscillation measurements in mice using a computer-controlled piston ventilator (flexiVent; SCIREQ Inc, Montreal, Qc, Canada). The description is divided into four parts: preparatory steps, mechanical ventilation, lung function measurements, and data analysis. It also includes details of how to assess airway responsiveness to inhaled methacholine in anesthetized mice, a common application of this technique which also extends to other outcomes and various lung pathologies. Measurements obtained in na&iuml;ve mice as well as from an oxidative-stress driven model of airway damage are presented to illustrate how this tool can contribute to a better characterization and understanding of studied physiological changes or disease models as well as to applications in new research areas.

The present protocol provides a detailed step-by-step description of the procedures required to execute measurements of respiratory system mechanics as well as the assessment of airway responsiveness to inhaled methacholine in mice using the forced oscillation technique (flexiVent; SCIREQ Inc, Montreal, Qc, Canada).

使用强迫振动技术在小鼠呼吸系统力学评价

The  forced oscillation technique (FOT) is a powerful, integrative and translational  tool permitting the experimental assessment of lung function in mice ...

Assessment of Respiratory Function in Conscious Mice by Double-chamber Plethysmography

Air volume changes created by a conscious subject breathing spontaneously within a body box are at the basis of plethysmography, a technique used to non-invasively assess some features of the respiratory function in humans as well as in laboratory animals. The present article focuses on the application of the double-chamber plethysmography (DCP) in small animals. It provides background information on the methodology as well as a detailed step-by-step procedure to successfully assess respiratory function in conscious, spontaneously breathing animals in a non-invasive manner. The DCP can be used to monitor the respiratory function of multiple animals in parallel, as well as to identify changes induced by aerosolized substances over a chosen time period and in a repeated manner. Experiments on control and allergic mice are used herein to demonstrate the utility of the technique, explain the associated outcome parameters, as well as to discuss the related advantages and shortcomings. Overall, the DCP provides valid and theoretically sound readouts that can be trusted to evaluate the respiratory function of conscious small animals both at baseline and after challenges with aerosolized substances.

The objective of the present article is to provide a detailed description of the recommended procedures to evaluate respiratory function in conscious mice by double-chamber plethysmography.

双室 Plethysmography 对清醒小鼠呼吸功能的评价

Air volume changes created by a conscious subject breathing spontaneously within a body box are at the basis of plethysmography, a technique used to ...