Name: alternate immersion in glucose to produce prolonged hyperglycemia in zebrafish
Uploaded: 2021-05-05T15:00:00
Description: Watch this Scientific Journal Video about Alternate Immersion in Glucose to Produce Prolonged Hyperglycemia in Zebrafish at JoVE.com

我们感谢 VPC、CJR 和 MCP 为本协议的制定。EMM 获得美国大学文理学院研究生支持的资助，以开展此项研究。这项工作还得到了美国大学梅隆学院奖的支持，并通过美国大学艺术与科学学院（VPC）提供资金。

所有程序都得到了美国大学动物护理和使用机构委员会的批准。
1. 准备解决方案罐
<ol><li>获得六个水箱，每个实验组两个（葡萄糖、曼尼托和水）。给两个储罐中的一个贴上"储罐"（它将容纳鱼）的标签，并给另一个"溶液罐"贴上标签（它将容纳溶液）。 注：曼尼托治疗组为渗透控制，水处理组为处理/应力控制。重要的是要保持坦克，航空公司/航空石，盖子和清洁用品?...

<ol>
	<li>Rubinstein, A. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Zebrafish:+from+disease+modeling+to+drug+discovery.">Zebrafish: from disease modeling to drug discovery.</a> Current Opinion in Drug Discovery and Development. 6 (2), 218-223 (2003).</li><li>Gerlai, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Associative+learning+in+zebrafish+(Danio+rerio).">Associative learning in zebrafish (Danio rerio).</a> Methods in Cell Biology. 101, 249-270 (2011).</li><li>Goldsmith, J. R., Jobin, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Think+small:+zebrafish+as+a+model+system+of+human+pathology.">Think small: zebrafish as a model system of human pathology.</a> BioMed Research International. , (2012).</li><li>Moss, J. B., 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=Regeneration+of+the+pancreas+in+adult+zebrafish.">Regeneration of the pancreas in adult zebrafish.</a> Diabetes. 58 (8), 1844-1851 (2009).</li><li>Connaughton, V. P., Baker, C., Fonde, L., Gerardi, E., Slack, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Alternate+immersion+in+an+external+glucose+solution+differentially+affects+blood+sugar+values+in+older+versus+younger+zebrafish+adults.">Alternate immersion in an external glucose solution differentially affects blood sugar values in older versus younger zebrafish adults.</a> Zebrafish. 13 (2), 87-94 (2016).</li><li>Etuk, E. U. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Animal+models+for+studying+diabetes+mellitus.">Animal models for studying diabetes mellitus.</a> Agriculture and Biology Journal of North America. 1 (2), 130-134 (2010).</li><li>Agardh, E., Bruun, A., Agardh, C. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Retinal+glial+cell+immunoreactivity+and+neuronal+cell+changes+in+rats+with+STZ-induced+diabetes.">Retinal glial cell immunoreactivity and neuronal cell changes in rats with STZ-induced diabetes.</a> Current Eye Research. 23 (4), 276-284 (2001).</li><li>Carmo, A., Cunha-Vaz, J. G., Carvalho, A. P., Lopes, M. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nitric+oxide+synthase+activity+in+retinas+from+non-insulin-dependent+diabetic+Goto-Kakizaki+rats:+correlation+with+blood-retinal+barrier+permeability.">Nitric oxide synthase activity in retinas from non-insulin-dependent diabetic Goto-Kakizaki rats: correlation with blood-retinal barrier permeability.</a> Nitric Oxide. 4 (6), 590-596 (2000).</li><li>Ramsey, D. J., Ripps, H., Qian, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+electrophysiological+study+of+retinal+function+in+the+diabetic+female+rat.">An electrophysiological study of retinal function in the diabetic female rat.</a> Investigative Ophthalmology &amp; Visual Science. 47 (11), 5116-5124 (2006).</li><li>Biessels, G. J., Gispen, W. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+impact+of+diabetes+on+cognition:+what+can+be+learned+from+rodent+models.">The impact of diabetes on cognition: what can be learned from rodent models.</a> Neurobiology of Aging. 26 (1), 36-41 (2005).</li><li>Intine, R. V., Olsen, A. S., Sarras, M. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+zebrafish+model+of+diabetes+mellitus+and+metabolic+memory.">A zebrafish model of diabetes mellitus and metabolic memory.</a> Journal of Visualized Experiments. (72), e50232 (2013).</li><li>Sarras, M. P., Intine, R. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Use+of+zebrafish+as+a+disease+model+provides+a+unique+window+for+understanding+the+molecular+basis+of+diabetic+metabolic+memory.">Use of zebrafish as a disease model provides a unique window for understanding the molecular basis of diabetic metabolic memory.</a> Research on Diabetes. , (2013).</li><li>Gleeson, M., Connaughton, V., Arneson, L. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Induction+of+hyperglycaemia+in+zebrafish+(Danio+rerio)+leads+to+morphological+changes+in+the+retina.">Induction of hyperglycaemia in zebrafish (Danio rerio) leads to morphological changes in the retina.</a> Acta Diabetologica. 44 (3), 157-163 (2007).</li><li>Capiotti, K. M., 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=Hyperglycemia+induces+memory+impairment+linked+to+increased+acetylcholinesterase+activity+in+zebrafish+(Danio+rerio).">Hyperglycemia induces memory impairment linked to increased acetylcholinesterase activity in zebrafish (Danio rerio).</a> Behavioural Brain Research. 274, 319-325 (2014).</li><li>Capiotti, K. M., 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=Persistent+impaired+glucose+metabolism+in+a+zebrafish+hyperglycemia+model.">Persistent impaired glucose metabolism in a zebrafish hyperglycemia model.</a> Comparative Biochemistry and Physiology Part B: Biochemistry and Molecular Biology. 171, 58-65 (2014).</li><li>Capiotti, K. M., 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=Hyperglycemia+alters+E-NTPDases,+ecto-5'-nucleotidase,+and+ectosolic+and+cytosolic+adenosine+deaminase+activities+and+expression+from+encephala+of+adult+zebrafish+(Danio+rerio).">Hyperglycemia alters E-NTPDases, ecto-5'-nucleotidase, and ectosolic and cytosolic adenosine deaminase activities and expression from encephala of adult zebrafish (Danio rerio).</a> Purinergic Signaling. 12 (2), 211-220 (2016).</li><li>Ali, 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=Photoreceptor+Degeneration+Accompanies+Vascular+Changes+in+a+Zebrafish+Model+of+Diabetic+Retinopathy.">Photoreceptor Degeneration Accompanies Vascular Changes in a Zebrafish Model of Diabetic Retinopathy.</a> Investigative Ophthalmology &amp; Visual Science. 61 (2), 43 (2020).</li><li>Wiggenhauser, L. M., 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=Activation+of+Retinal+Angiogenesis+in+Hyperglycemic+pdx1-/-+Mutants.">Activation of Retinal Angiogenesis in Hyperglycemic pdx1-/- Mutants.</a> Diabetes. 69 (5), 1020-1031 (2020).</li><li>Chen, X. 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=Involvement+of+HMGB1+mediated+signalling+pathway+in+diabetic+retinopathy:+evidence+from+type+2+diabetic+rats+and+ARPE-19+cells+under+diabetic+condition.">Involvement of HMGB1 mediated signalling pathway in diabetic retinopathy: evidence from type 2 diabetic rats and ARPE-19 cells under diabetic condition.</a> Journal of Ophthalmology. 97, 1598-1603 (2013).</li><li>Costa, E., 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=Effects+of+light+exposure,+pH,+osmolarity,+and+solvent+on+the+retinal+pigment+epithelial+toxicity+of+vital+dyes.">Effects of light exposure, pH, osmolarity, and solvent on the retinal pigment epithelial toxicity of vital dyes.</a> American Journal of Ophthalmology. 155, 705-712 (2013).</li><li>Alvarez, Y., 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=Predominant+cone+photoreceptor+dysfunction+in+a+hyperglycemic+model+of+non-proliferative+diabetic+retinopathy.">Predominant cone photoreceptor dysfunction in a hyperglycemic model of non-proliferative diabetic retinopathy.</a> Disease Models and Mechanisms. 3, 236-245 (2010).</li><li>Fletcher, E. L., Phipps, J. A., Wilkinson-Berka, J. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dysfunction+of+retinal+neurons+and+glia+during+diabetes.">Dysfunction of retinal neurons and glia during diabetes.</a> Clinical and Experimental Optometry. 88, 132-145 (2005).</li><li>Fletcher, E. L., Phipps, J. A., Ward, M. M., Puthussery, T., Wilkinson-Berka, J. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neuronal+and+glial+abnormality+as+predictors+of+progression+of+diabetic+retinopathy.">Neuronal and glial abnormality as predictors of progression of diabetic retinopathy.</a> Current Pharmaceutical Design. 13, 2699-2712 (2007).</li><li>Agardh, E., Bruun, A., Agardh, C. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Retinal+glial+cell+immunoreactivity+and+neuronal+cell+changes+in+rats+with+STZ-+induced+diabetes.">Retinal glial cell immunoreactivity and neuronal cell changes in rats with STZ- induced diabetes.</a> Current Eye Research. 23, 276-284 (2001).</li><li>Barber, A. J., Antonetti, D. A., Gardner, T. W., Group, T. P. S. R. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Altered+expression+of+retinal+occludin+and+glial+fibrillary+acidic+protein+in+experimental+diabetes.">Altered expression of retinal occludin and glial fibrillary acidic protein in experimental diabetes.</a> Investigative Ophthalmology &amp; Visual Science. 41, 3561-3568 (2000).</li><li>Lieth, E., 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=Glial+reactivity+and+impaired+glutamate+metabolism+in+short-term+experimental+diabetic+retinopathy.">Glial reactivity and impaired glutamate metabolism in short-term experimental diabetic retinopathy.</a> Diabetes. 47, 815-820 (1998).</li><li>Rungger-Brandle, E., Dosso, A. A., Leuenberger, P. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Glial+reactivity,+an+early+feature+of+diabetic+retinopathy.">Glial reactivity, an early feature of diabetic retinopathy.</a> Investigative Ophthalmology &amp; Visual Science. 41, 1971-1980 (2000).</li><li>Zeng, X. X., Ng, Y. K., Ling, E. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neuronal+and+microglial+response+in+the+retina+of+streptozotocin-induced+diabetic+rats.">Neuronal and microglial response in the retina of streptozotocin-induced diabetic rats.</a> Visual Neuroscience. 17, 463-471 (2000).</li><li>Mizutani, M., Gerhardinger, C., Lorenzi, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Muller+cell+changes+in+human+diabetic+retinopathy.">Muller cell changes in human diabetic retinopathy.</a> Diabetes. 47, 445-449 (1998).</li><li>Tanvir, Z., Nelson, R., DeCicco-Skinner, K., Connaughton, V. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=One+month+of+hyperglycemia+alters+spectral+responses+of+the+zebrafish+photopic+electroretinogram.">One month of hyperglycemia alters spectral responses of the zebrafish photopic electroretinogram.</a> Disease Models and Mechanisms. 11, (2018).</li><li>Hancock, H. A., Kraft, T. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Oscillatory+potential+analysis+and+ERGs+of+normal+and+diabetic+rats.">Oscillatory potential analysis and ERGs of normal and diabetic rats.</a> Investigative Ophthalmology &amp; Visual Science. 45, 1002-1008 (2004).</li><li>Layton, C. J., Safa, R., Osborne, N. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Oscillatory+potentials+and+the+b-wave:+partial+masking+and+interdependence+in+dark+adaptation+and+diabetes+in+the+rat.">Oscillatory potentials and the b-wave: partial masking and interdependence in dark adaptation and diabetes in the rat.</a> Graefe's Archives for Clinical and Experimental Ophthalmology. 245, 1335-1345 (2007).</li><li>Li, Q., Zemel, E., Miller, B., Perlman, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Early+retinal+damage+in+experimental+diabetes:+electroretinographical+and+morphological+observations.">Early retinal damage in experimental diabetes: electroretinographical and morphological observations.</a> Experimental Eye Research. 74, 615-625 (2002).</li><li>Kohzaki, K., Vingrys, A. J., Bui, B. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Early+inner+retinal+dysfunction+in+streptozotocin-induced+diabetic+rats.">Early inner retinal dysfunction in streptozotocin-induced diabetic rats.</a> Investigative Ophthalmology &amp; Visual Science. 49, 3595-3604 (2008).</li><li>Phipps, J. A., Yee, P., Fletcher, E. L., Vingrys, A. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rod+photoreceptor+dysfunction+in+diabetes:+activation,+deactivation,+and+dark+adaptation.">Rod photoreceptor dysfunction in diabetes: activation, deactivation, and dark adaptation.</a> Investigative Ophthalmology &amp; Visual Science. 47, 3187-3194 (2006).</li></ol>

糖尿病是一个全国性的问题。研究表明，到2030年，估计有4亿人将患有某种形式的糖尿病。在啮齿动物模型中，使用基因操作研究2型DM。在大鼠中，扎克糖尿病脂肪大鼠（ZDF）和大须卡长埃文斯德岛脂肪大鼠（OLETF）提供了更多关于2型DM10的影响的信息。此外，高脂肪饮食已用于啮齿动物诱发高血糖。这反映了本文中提出的非侵入性程序。使用我们的非侵入性协议，我们可以诱发?...

斑马鱼（丹尼奥·雷里奥）正在迅速成为一个广泛使用的动物模型，以研究疾病和认知1。基因操纵的易受干扰和胚胎透明性在早期发育阶段，使他们成为研究人类疾病的主要候选者与已知的遗传基础。例如，斑马鱼已经被用来研究霍尔特-奥拉姆综合征，心肌病，球状细胞性肾脏疾病，肌肉萎缩症和糖尿病（DM）等疾病1。此外，斑马鱼模型是理想的，因为该物种的体积小，维护方便，和高粪便2，3。
斑马鱼胰腺在解剖学上和功能上都与哺乳动物胰腺4相似。因此，斑马鱼具有体型大、肥大和内分泌结构相似的独特特征，因此斑马鱼成为研究DM相关并发症的合适人选。在斑马鱼中，有两种实验方法用于诱导DM特有的长时间高血糖：葡萄糖的流入（建模2型）和胰岛素分泌的停止（建模1型）5、6。在实验中，为了阻止胰岛素分泌，胰腺β细胞可以通过链球菌素（STZ）或Aloxan注射进行化学破坏。STZ已成功地用于啮齿动物和斑马鱼，导致并发症与视网膜病变7，8，9，认知障碍10，和肢体再生11。然而，在斑马鱼中，β细胞在治疗后再生，导致STZ的"助推器注射"成为维持糖尿病状况所必需的。或者，斑马鱼的胰腺可以去除6。这些都是高度侵入性的程序，由于多次注射，和广泛的恢复时间。
相反，高血糖可以通过接触外源性葡萄糖来非侵入性地诱发。在此协议中，鱼被淹没在高度浓缩的葡萄糖溶液中24小时5，13或连续2周14，15，16。外源性葡萄糖通过摄入和/或穿过刺，导致血糖水平升高而被跨皮吸走。由于这种非侵入性技术不直接操纵胰岛素水平，因此不能声称诱导2型DM。然而，它可以用来检查高血糖引起的并发症，这是2型DM的主要症状之一。
最近，斑马鱼突变pdx1-/-通过操纵胰腺和十二指肠家谱1基因，一种与人类2型DM的遗传原因相关的基因而开发的。利用这种突变体，研究人员已经能够复制胰腺发育中断，高血糖，并研究高血糖引起的糖尿病视网膜病变17，18。
本文描述了一种使用交替浸入协议的非侵入性高血糖诱导方法。此协议维持高血糖条件长达 8 周，并观察到随后的并发症。简言之，成年斑马鱼被放置在糖溶液中24小时，然后放入水溶液中24小时。与持续浸入外部葡萄糖溶液相反，糖和水之间的交替天模拟糖尿病中血糖的上升和下降。此外，交替葡萄糖协议还允许高血糖在较长时间内诱发，因为斑马鱼无法补偿高外部葡萄糖条件。作为原则的证明，我们提供的数据表明，使用此协议诱导的血糖过高会改变视网膜化学和生理学。

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	<tbody>
		<tr>
			<td>Airline Tubing</td>
			<td>petsmart</td>
			<td>5291863</td>
			<td>This can be used in the tank to circulate air</td>
		</tr>
		<tr>
			<td>Airpump</td>
			<td>petsmart</td>
			<td>5094984</td>
			<td>This can be used in the tank to circulate air</td>
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		<tr>
			<td>Airstones</td>
			<td>petsmart</td>
			<td>5149683</td>
			<td>This can be used in the tank to circulate air</td>
		</tr>
		<tr>
			<td>D-glucose</td>
			<td>Sigma</td>
			<td>G8270-5KG</td>
			<td></td>
		</tr>
		<tr>
			<td>D-mannitol</td>
			<td>Acros Organics</td>
			<td>AC125340050</td>
			<td></td>
		</tr>
		<tr>
			<td>Freestyle Lite Meter</td>
			<td>Amazon</td>
			<td>B01LMOMLTU</td>
			<td></td>
		</tr>
		<tr>
			<td>Freestyle Lite Strips</td>
			<td>Amazon</td>
			<td>B074ZN3H2Z</td>
			<td></td>
		</tr>
		<tr>
			<td>Net</td>
			<td>petsmart</td>
			<td>5175115</td>
			<td></td>
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		<tr>
			<td>Tanks</td>
			<td>Amazon</td>
			<td>B0002APZO4</td>
			<td></td>
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</table>

alternate immersion in glucose to produce prolonged hyperglycemia in zebrafish

斑马鱼（Danio rerio）是研究慢性高血糖影响的优秀模型，这是 II 型糖尿病 （T2DM） 的标志。这种替代浸入协议是一种非侵入性的、循步骤的方法，可诱导高血糖长达八周。成年斑马鱼交替接触糖（葡萄糖）和水，每次24小时。斑马鱼开始用1%的葡萄糖溶液治疗2周，然后用2%的溶液治疗2周，最后在剩下的4周内进行3%的溶液治疗。与水处理（压力）和曼尼托处理（渗透）对照相比，葡萄糖处理斑马鱼的血糖水平显著较高。经过葡萄糖处理的斑马鱼的血糖水平是对照组的3倍，这表明在4周和8周后可以达到高血糖。持续性高血糖与胶质纤维酸性蛋白 （GFAP） 增加、心网膜核因子 Kappa B （NF-kB） 水平增加、生理反应下降以及认知缺陷有关，表明此协议可用于模拟疾病并发症。

使用此协议（图 1），经过 4 周和 8 周的治疗（图 2A），血糖值显著升高，高血糖的定义是水处理组和曼尼托治疗组的控制平均值的 3 倍。水处理控制每天进出水，提供压力/处理控制。曼尼托在体外葡萄糖研究19，20中充当渗透控制，因为它是一种6碳糖，如葡萄糖，但不被细胞占用。为了与这些研究和斑马鱼...

Watch this Scientific Journal Video about Alternate Immersion in Glucose to Produce Prolonged Hyperglycemia in Zebrafish at JoVE.com

Alternate Immersion in Glucose to Produce Prolonged Hyperglycemia in Zebrafish

此协议非侵入性地诱发斑马鱼高血糖长达 8 周。利用这一协议，可以深入研究高血糖的不利影响。

交替浸入葡萄糖中，在斑马鱼中产生长期血糖超血症

Zebrafish (Danio rerio) are an excellent model to investigate the effects of chronic hyperglycemia, a hallmark of Type II Diabetes Mellitus (T2DM). This ...

此协议在斑马鱼中诱发高血糖，其上升和下降模式模仿二型糖尿病中出现的高血糖。这项技术的主要优点是，它是一种非侵入性技术，可以确保高血糖是高血糖鱼类中观察到的任何改变的原因。该协议可能用于探索治疗途径或药物，针对高血糖并发症。此协议有很多步骤，但当步骤正确和谨慎地遵循时，它应迅速成为第二性质。但是，请记住，在整个协议中，始终温和、人道地对待动物。首先建立六个储罐，每个实验组两个。使用两升鱼缸，如果鱼总数小于20升，如果鱼缸总数超过20升。标记两个储罐之一，外壳罐和其他解决方案罐。将水箱保存在28至29摄氏度的水浴中，以保持水温。第一天，将鱼放入各自的治疗解决方案中24小时。第二天，将鱼从处理液转移到水中24小时。第三天，将鱼从水中转移到处理解决方案。在实验的剩余部分继续这种交替曝光。每天将经过水处理的控制鱼从水转移到水中。确保在整个实验期间，每天在同一两个小时的窗口内喂养和转移鱼。使用标准鱼网将每个处理组中的鱼从外壳罐转移到相应的溶液罐中。将装有鱼的鱼缸放回水浴池，更换空气石和油箱盖。这个坦克现在是外壳罐，以前装鱼的鱼缸现在是溶液罐。丢弃旧溶液，与油箱盖、气管、空气结石和网一起清洁罐体，以防止葡萄糖和曼尼托的积聚。使用水和专用海绵为每个处理条件，以适当清洁水箱。用纸巾擦干新清洁的溶液罐，并使用此油箱为第二天的解决方案做好准备。确保其他物品干燥，由适当的治疗组分离。要准备糖溶液，请向每个溶液罐中加满两到四升系统水。使用顶部装载秤测量葡萄糖和曼尼托的正确量，并为每种化学品分别测量称重船。将称重葡萄糖或曼尼托阿利库特添加到适当的清洁溶液罐中。用单独的玻璃搅拌棒搅拌溶液，直到糖完全溶解。然后将溶液罐返回水浴池，并盖上相应的盖子。要准备水溶液，请向实验罐灌注系统水，将这些溶液罐返回水浴池，并盖上相应的盖子。经过四周和八周的葡萄糖治疗后，血糖值显著升高。高血糖的定义是水处理组和曼尼托治疗组的控制平均值的三倍。在高血糖四周后收集的视网膜组织有胶质纤维酸性蛋白质或GFAP水平增加。GFAP表达在视网膜中的穆勒胶质细胞中观察到，这些细胞在糖尿病视网膜病变中改变。GFAP的增加与核因素卡帕布水平的增加有关。暗示诱发的血糖过高会引发炎症反应和反应性胶质。经过四周治疗后，ERG记录发现，与曼尼托治疗的对照组相比，葡萄糖治疗视网膜的反应有所降低。高血糖鱼类中光感受器和双极细胞成分的振幅均有所下降。这个程序可以通过其他高血糖测试来补充。例如，记忆检测可用于观察认知缺陷或记录基于视觉的反应，如光运动反应以评估基于视觉的并发症。一旦这项技术建立起来，我们的实验室就可以用它来研究斑马鱼模型中的高血糖诱发并发症。经过四周的治疗，我们观察到这些并发症的速度相对较快。

Research

JoVE Journal

Neuroscience

Human Fear Conditioning Conducted in Full Immersion 3-Dimensional Virtual Reality

人类的恐惧充分浸泡三维虚拟现实中进行的空调

Fear conditioning is a widely used paradigm in non-human animal research to investigate the neural mechanisms underlying fear and anxiety. A major ...

科研

神经科学

Time-lapse Live Imaging of Clonally Related Neural Progenitor Cells in the Developing Zebrafish Forebrain

在发展中国家斑马鱼的前脑的神经祖细胞克隆相关的定时实时成像

Precise patterns of division, migration and differentiation of neural progenitor cells are crucial for proper brain development and function1,2. To ...

Retrograde Fluorescent Labeling Allows for Targeted Extracellular Single-unit Recording from Identified Neurons In vivo

Retrograde Fluorescent Labeling Allows for Targeted Extracellular Single-unit Recording from Identified Neurons In vivo

逆行荧光染料标记允许有针对性的外单细胞记录技术鉴定神经元<em&gt;在体内</em

The overall goal of this method is to record single-unit responses from an identified population of neurons. In vivo electrophysiological recordings from ...

Dual Electrophysiological Recordings of Synaptically-evoked Astroglial and Neuronal Responses in Acute Hippocampal Slices

双突触诱发的星形胶质细胞和神经元的反应，在急性海马脑片电生理记录

Astrocytes form together with neurons tripartite synapses, where they integrate and modulate neuronal activity. Indeed, astrocytes sense neuronal inputs ...

Multi-unit Recording Methods to Characterize Neural Activity in the Locust (Schistocerca Americana) Olfactory Circuits

Multi-unit Recording Methods to Characterize Neural Activity in the Locust Schistocerca Americana Olfactory Circuits

多单位记录方法来描述蝗虫的神经活动（<em&gt; Schistocerca美洲</em&gt;）嗅觉电路

Detection and interpretation of olfactory cues are critical for the survival  of many organisms. Remarkably, species across phyla have strikingly similar ...

Direct Intraventricular Delivery of Drugs to the Rodent Central Nervous System

啮齿动物的中枢神经系统药物直接脑室交货

Due to an inability to cross the blood brain barrier, certain drugs need to be directly delivered into the central nervous system (CNS). Our lab focuses ...

A Simple and Inexpensive Method for Determining Cold Sensitivity and Adaptation in Mice

测定冷灵敏度和适应小鼠一种简单廉价的方法

Cold hypersensitivity is a serious clinical problem, affecting a broad subset of patients and causing significant decreases in quality of life.  The cold ...

Analyzing Synaptic Modulation of Drosophila melanogaster Photoreceptors after Exposure to Prolonged Light

Analyzing Synaptic Modulation of Drosophila melanogaster Photoreceptors after Exposure to Prolonged Light

分析突触调制<em&gt;果蝇</em&gt;曝光时间延长后，光光感受器

The nervous system has the remarkable ability to adapt and respond to various stimuli. This neural adjustment is largely achieved through plasticity at ...

A Revised Surgical Approach to Induce Endolymphatic Hydrops in the Guinea Pig

几内亚猪诱导内部水解的修正手术方法

Endolymphatic hydrops is an enlargement of scala media that is most often associated with Meniere&#39;s disease, though the pathophysiologic mechanism(s) ...

Combining Behavior and EEG to Study the Effects of Mindfulness Meditation on Episodic Memory

结合行为和脑电图研究正念冥想对情景记忆的影响

Although there has been recent interest in how mindfulness meditation can affect episodic memory as well as brain structure and function, no study has ...