Name: two-photon imaging of microglial processes' attraction toward atp or serotonin in acute brain slices
Uploaded: 2019-01-31T13:30:00
Description: Watch this Scientific Journal Video about Two-photon Imaging of Microglial Processes' Attraction Toward ATP or Serotonin in Acute Brain Slices at JoVE.com

我们感谢穆林研究所的细胞和组织成像设施, 在那里进行了所有的图像采集和分析。这项工作得到了国家科学研究中心、国家研究和研究研究所、索邦大学科学研究所以及索邦大学-皮埃尔·玛丽·居里大学赠款的部分支持 (emargemce-upmc 方案, Recherche sur le cerveau 基金会, 法国基金会, "equipe frm deq2010" 基金会, 法国研究部 (国家研究机构, Recherche)anr-17-ce16-008 和 "bio-psy labex" anr-11-idex-000-02) 和计算神经科学方案合作研究, 国家科学基金会/法国国家研究机构 (编号: 1515686)。所有作者都附属于巴黎神经科学学院 (enp) 和 bio-psy labex 的研究小组。f. e. 是法国巴黎 f-75005 索邦大学附属博士生, 由 bio-psy labex 资助。v. m. 是由法国国家科学基金会/法国国家研究机构计算神经科学合作研究项目资助的博士后研究员 (编号: 1515686)。提交人感谢参与启动该项目的 marta kolodziejczak。

所有实验都得到了地方道德委员会 (达尔文委员会、协议 #1170 和 #10921) 的批准。
1. 用于局部应用的玻璃微管的制备
<ol><li>用电极拉拔器从硼硅酸盐薄壁玻璃毛细管中准备移液器。调整参数以获得在其末端直径为4-5 微米的移液器。图 2d显示了一个低放大倍率的明亮场移液器。</li>
</ol>2. 解决方案
<ol><li>确保只使用通过高压灭菌器循环?...

<ol>
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急性脑片通过保持一种结构完整性和有限的网络调整, 与离解或有机切片培养不同, 使研究人员能够在其生理环境中研究微胶质细胞。然而, 其中一个主要的限制是, 切片程序造成的伤害, 可以迅速损害神经元的生存能力, 特别是在成人大脑。由于微胶质细胞对细胞损伤的反应特别严重, 因此尽可能地限制神经元细胞死亡以保持微胶质细胞接近其生理状态是很重要的。这反过来又通过良性循环有助于<e...

微胶质细胞是大脑的巨噬细胞, 在生理和病理条件 1,2中都起着一定的作用.它们具有高度分枝的形态, 并不断扩展和收缩其过程3,4。这种 "扫描" 行为被认为与对周围环境的调查有关, 也是必要的。微胶质细胞的形态可塑性以三种模式表示。首先, 一些化合物迅速调节微胶质细胞的形态: 在中浴急性脑片中加入 atp5、6或 nmda5, 7 增加了微胶质细胞影响的复杂性,而去甲肾上腺素会降低它 6。这些效应要么是由微胶质受体 (atp 和去甲肾上腺素) 直接介导的, 要么是需要神经元的 atp 释放 (对于 nmda)。第二, 微胶质过程的生长和收缩速度, 称为 "动力或监视", 可以受到细胞外因子8, 稳态中断9,10,或突变9, 10,11。第三, 除了这些各向同性的形态和动力的变化, 微胶质细胞有能力将其过程定向扩展到提供 atp3,5,12的移液器,13,14、在培养中、在急性脑片或体内, 或在急性脑片15中提供5-ht。这种微胶质过程的定向生长, 也被称为定向运动, 首先被描述为对局部激光病变3,4的反应。因此, 从生理上讲, 它可能与受伤的反应有关, 也可能与针对微胶质过程的目标, 以突触或大脑区域为目标, 在发育15、16或生理上17需要修剪 ,18,19或病理情况9,18,19,20在成年。三种类型的形态变化依赖于不同的细胞内机制11,13, 20 和一个给定的化合物不一定调节所有这些机制 (例如, nmda, 间接作用于微胶质细胞, 对形态有影响, 但不会引起方向动力5,7)。因此, 当目标是表征化合物、突变或病理对微胶质细胞的影响时, 重要的是要表征其形态可塑性的三个组成部分。在这里, 我们描述了一种方法来研究微胶质过程的定向生长向一个局部来源的化合物, 这是, 在这里, atp 或5-ht。
研究微胶质细胞过程吸引力的模型有几个: 三维环境中的初级培养 6、18、19、急性大脑切片 6、13、15和体内成像3,13。体内入路是保存微胶质细胞生理状态的最佳方法。然而, 深部区域的宫内成像需要复杂的外科手术, 因此, 它往往被限制在浅层皮质层。使用微胶质原代培养是用数量有限的动物测试大量条件的最简单的技术。然而, 它是不可能获得相同的细胞形态在体内, 细胞失去其与神经元和星形胶质细胞的生理相互作用。急性脑片代表了这两种方法之间的妥协。该模型允许研究人员研究在体内难以到达的大脑结构和高分辨率的图像, 并从新生儿阶段的切片进行研究, 而经颅显微镜主要是在成年时进行的。最后, 可以实时观察局部药物应用的效果, 并在使用数量有限的动物的同时重复实验。然而, 急性脑片的一个问题是细胞存活的时间有限 (几个小时), 特别是两周以上小鼠的切片, 以及随着时间的推移, 微胶质细胞形态可能发生变化.
在这里, 我们描述了一个方案, 以准备急性脑片的年轻和成人 cx3cr1gfp/c + 小鼠长达两个月大, 保持微胶质的形态和运动几个小时。然后, 我们描述如何使用这些切片来研究微胶质过程对 atp 或5-ht 等化合物的吸引力。

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			<td>for pipettes preparation</td>
			<td></td>
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			<td>Clark Borosilicate Thin Wall Capillaries</td>
			<td>Harvard Apparatus</td>
			<td>30-0065</td>
			<td>Borosilicate Thin Wall without Filament, 1.5 mm OD, 1.17 mm ID, 75 mm L , Pkg. of 225</td>
		</tr>
		<tr>
			<td>DMZ Universal Puller</td>
			<td>Zeitz Instrumente</td>
			<td></td>
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			<td>Name</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td>Comments</td>
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			<td>for solutions</td>
			<td></td>
			<td></td>
			<td></td>
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			<td>Calcium Chloride dihydrate (CaCl2)</td>
			<td>Sigma</td>
			<td>C5080</td>
			<td></td>
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			<td>Choline Chloride</td>
			<td>Sigma</td>
			<td>C7527</td>
			<td></td>
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			<td>D-(+)-Glucose</td>
			<td>Sigma</td>
			<td>G8270</td>
			<td></td>
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			<td>L-Ascorbic acid</td>
			<td>Sigma</td>
			<td>A5960</td>
			<td></td>
		</tr>
		<tr>
			<td>Magnesium Chloride solution 1M (MgCl2)</td>
			<td>Sigma</td>
			<td>63020</td>
			<td></td>
		</tr>
		<tr>
			<td>Potassium chloride SigmaUltra &#62;99,0% (KCl)</td>
			<td>Sigma</td>
			<td>P9333</td>
			<td></td>
		</tr>
		<tr>
			<td>Sodium bicarbonate (NaHCO3)</td>
			<td>Sigma</td>
			<td>S5761</td>
			<td></td>
		</tr>
		<tr>
			<td>Sodium Chloride (NaCl)</td>
			<td>Sigma</td>
			<td>S5886</td>
			<td></td>
		</tr>
		<tr>
			<td>Sodium phosphate monobasic</td>
			<td>Sigma</td>
			<td>S5011</td>
			<td></td>
		</tr>
		<tr>
			<td>Sodium pyruvate</td>
			<td>Sigma</td>
			<td>P2256</td>
			<td></td>
		</tr>
		<tr>
			<td>Ultrapure water</td>
			<td>MilliQ</td>
			<td></td>
			<td>for all the solutions</td>
		</tr>
		<tr>
			<td>Name</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td>Comments</td>
		</tr>
		<tr>
			<td>for slice preparation</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>2x 200 mL crystalizing dishes</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>80 mL Pyrex beaker</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Antlia-3C Digital Peristaltic pump</td>
			<td>DD Biolab</td>
			<td>178961</td>
			<td>For mice perfusion and 2-photon chamber perfusion (aCSF)</td>
		</tr>
		<tr>
			<td>Carbogen 5% CO2/95% O2</td>
			<td>Air Liquide France Industrie</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Dolethal</td>
			<td>Vetoquinol</td>
			<td>Dolethal 50 mg/mL</td>
			<td></td>
		</tr>
		<tr>
			<td>Filter papers (Whatman)</td>
			<td>Sigma</td>
			<td>WHA1001042</td>
			<td>Whatman qualitative filter paper, Grade 1 (Pore size: 11&#181;M)</td>
		</tr>
		<tr>
			<td>Fine Scissors - Sharp</td>
			<td>Fine Science Tools</td>
			<td>14060-60</td>
			<td></td>
		</tr>
		<tr>
			<td>Food box 10 cm diameter, 8 cm Height</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>glue (ethyl cyanoacrylate)</td>
			<td>Loctite</td>
			<td>super glue 3 power flex</td>
			<td></td>
		</tr>
		<tr>
			<td>Hippocampal Tool (spatula)</td>
			<td>Fine Science Tools</td>
			<td>10099-15</td>
			<td>The largest extremity has to be angled at 90 &#176;</td>
		</tr>
		<tr>
			<td>Ice</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Iris Forceps (curved)</td>
			<td>Moria</td>
			<td>MC31</td>
			<td></td>
		</tr>
		<tr>
			<td>Lens cleaning tissue</td>
			<td>THOR LABS</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Nylon mesh strainer</td>
			<td></td>
			<td></td>
			<td>diameter 7 cm</td>
		</tr>
		<tr>
			<td>Razor blades</td>
			<td>Electron Microscopy Sciences</td>
			<td>72000</td>
			<td>For the slicer</td>
		</tr>
		<tr>
			<td>scalpel blade</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Slice interface holder</td>
			<td></td>
			<td></td>
			<td>home-made, the file for 3D printing is provided in Supplemental Material</td>
		</tr>
		<tr>
			<td>Surgical Scissors - Sharp</td>
			<td>Fine Science Tools</td>
			<td>14002-14</td>
			<td></td>
		</tr>
		<tr>
			<td>Vibrating slicer</td>
			<td>Thermo Scientific</td>
			<td>720-2709</td>
			<td>Model: HM 650V (Vibrating blade microtome)</td>
		</tr>
		<tr>
			<td>Water bath</td>
			<td></td>
			<td></td>
			<td>Set at 32&#176;C (first recovery step)</td>
		</tr>
		<tr>
			<td>Name</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td>Comments</td>
		</tr>
		<tr>
			<td>for slice imaging</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>&#215; 25 0.95 NA water-immersion objective</td>
			<td>Leica Microsystems (Germany)</td>
			<td>HCX Irapo</td>
			<td></td>
		</tr>
		<tr>
			<td>2-photon MP5 upright microscope with resonant scanners (8 kHz) and two HyD Hybrid detectors</td>
			<td>Leica Microsystems (Germany)</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Antlia-3C Digital Peristaltic pump</td>
			<td>DD Biolab</td>
			<td>178961</td>
			<td>For 2-photon chamber perfusion with aCSF</td>
		</tr>
		<tr>
			<td>Carbogen 5% CO2/95% O2</td>
			<td>Air Liquide France Industrie</td>
			<td>I1501L50R2A001</td>
			<td></td>
		</tr>
		<tr>
			<td>Chameleon Ultra2 Ti:sapphire laser</td>
			<td>Coherent (Germany)</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>disposable transfer pipettes , wide mouth</td>
			<td>ThermoFischer scientific</td>
			<td>for example : 232-11</td>
			<td>5.8ml with fin tip, but we cut it (approx 7cm) to have a 4 mm diameter mouth</td>
		</tr>
		<tr>
			<td>emission filter SP680</td>
			<td>Leica Microsystems (Germany)</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>fluorescent cube containing a 525/50 emission filter and a 560 dichroic filter (for fluorescence collection)</td>
			<td>Leica Microsystems (Germany)</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>glass beaker with 50 mL of ACSF to maintain constant perfusion of the slice</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Heating system</td>
			<td>Warner Instrument Corporation</td>
			<td>Automatic Heater Controller TC-324B</td>
			<td>to maintain perfusion solution at 32&#176;C</td>
		</tr>
		<tr>
			<td>perfusion chamber</td>
			<td></td>
			<td></td>
			<td>home-made, the file for 3D printing is provided in Supplemental Material</td>
		</tr>
		<tr>
			<td>slice holder (&#34;harp&#34;)</td>
			<td></td>
			<td></td>
			<td>home made : hairpin made of platinum with the two branches joined by parallel nylon threads</td>
		</tr>
		<tr>
			<td>Name</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td>Comments</td>
		</tr>
		<tr>
			<td>for slice stimulation </td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Adenosine 5&#8242;-triphosphate disodium salt hydrate (ATP)</td>
			<td>Sigma</td>
			<td>A-26209</td>
			<td>to be prepared ex-temporaneously : 1mg/ml (3mM) stock solution prepared the day of the experiment, kept at 4&#176;C (a few hours) and diluted just before use</td>
		</tr>
		<tr>
			<td>Fluorescein (optional)</td>
			<td>Sigma</td>
			<td>F-6377</td>
			<td>use at 1 &#181;M final</td>
		</tr>
		<tr>
			<td>Micromanipulator</td>
			<td>Luigs and Neumann</td>
			<td>SM7</td>
			<td>connected to the micropipette holde</td>
		</tr>
		<tr>
			<td>Micropipette holder</td>
			<td></td>
			<td></td>
			<td>same as for eletrophysiology</td>
		</tr>
		<tr>
			<td>Serotonin hydrochloride</td>
			<td>Sigma</td>
			<td>H-9523</td>
			<td>aliquots of 50mM stock solution in H20 kept at -20&#176;C. 500&#181;M solution prepared the day of the experiment.</td>
		</tr>
		<tr>
			<td>Syringe 5mL (without needle)</td>
			<td>Terumo medical products</td>
			<td>SS+05S1</td>
			<td></td>
		</tr>
		<tr>
			<td>Transparent tubing</td>
			<td>Fischer Scientific</td>
			<td>11750105</td>
			<td>Saint Gobain Performance Plastics&#8482; Tygon&#8482; E-3603 Non-DEHP Tubing</td>
		</tr>
		<tr>
			<td>Name</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td>Comments</td>
		</tr>
		<tr>
			<td>for image analysis</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Fiji</td>
			<td></td>
			<td>https://fiji.sc</td>
			<td>Schindelin, J. et al Nat. Methods (2012) doi 10.1038</td>
		</tr>
		<tr>
			<td>Icy</td>
			<td>Institut Pasteur</td>
			<td>http://icy.bioimageanalysis.org</td>
			<td>de Chaumont, F. et al. Nat. Methods (2012)</td>
		</tr>
		<tr>
			<td>Name</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td>Comments</td>
		</tr>
		<tr>
			<td>mice</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>CX3CR1-GFP mice</td>
			<td>Jung et al, 2000</td>
			<td></td>
			<td>male or females, P3 to 2 months-old ; we have backcrossed these mice on 129sv background.</td>
		</tr>
		<tr>
			<td>CX3CR1creER-YFP mice</td>
			<td>Parkhurst et al 2013</td>
			<td></td>
			<td>male or females, P3 to 2 months-old ; we have backcrossed these mice on 129sv background.</td>
		</tr>
	</tbody>
</table>

two-photon imaging of microglial processes' attraction toward atp or serotonin in acute brain slices

微胶质细胞是大脑中的固有免疫细胞, 它们在长时间的过程中不断地扫描它们的环境, 并在破坏稳态时经历快速的形态变化。例如, 激光病变在几分钟内诱导一个定向的微胶质过程的生长, 也被称为 "定向动力", 向受伤部位。通过局部 atp 或血清素 (5-羟基色胺 [5-ht]), 也可以获得类似的效果。在本文中, 我们描述了一种方案, 以诱导微胶质过程的定向生长, 向局部应用 atp 或5-ht 在急性脑片的急性脑片的年轻和成年小鼠, 并通过多光子显微镜图像这种吸引力随着时间的推移。提出了一种利用自由和开源图像分析软件进行量化的简单方法。急性脑片的一个特点仍然是时间有限, 随着年龄的推移而减少, 在此期间, 细胞保持生理状态。因此, 该协议强调了一些技术改进 (介质、气液界面室、双灌注成像室), 旨在在几个小时内优化微胶质细胞的活力, 特别是在成年小鼠的切片中。

该协议描述了一种方法, 以诱导, 观察和量化定向生长的微胶质过程向局部应用的化合物, 例如 atp 或 5-ht, 急性脑切片从年轻或成人 (至少两个月) 小鼠。在有助于将成年动物的大脑切片保持在健康状态数小时的因素中, 使用了两个工具, 旨在在协议的两个步骤中优化细胞存活率。首先, 接口腔中的界面片支架 (图 1) 改善了切割后切片的保存。其次, 记录?...

Watch this Scientific Journal Video about Two-photon Imaging of Microglial Processes' Attraction Toward ATP or Serotonin in Acute Brain Slices at JoVE.com

Two-photon Imaging of Microglial Processes' Attraction Toward ATP or Serotonin in Acute Brain Slices

小胶质细胞是大脑中的常驻免疫细胞, 随着环境变化的形态变化而迅速反应。该协议描述了如何使用双光子显微镜研究微胶质过程对血清素或 atp 的吸引力, 在急性脑切片的小鼠。

微胶质过程对急性脑片 atp 或血清素的双光子成像

Microglial cells are resident innate immune cells of the brain that constantly scan their environment with their long processes and, upon disruption of ...

要充分描述微胶质表型，必须同时解决基础和定向运动问题。该协议可用于利用定制的3D打印界面和灌注室，在小鼠脑片中通过双光子成像来测试微胶质的方向性。展示这个程序的将是范妮·艾蒂安，一个博士生，和文森佐·马斯特里奥利亚，一个博士后研究员，都来自我们的实验室。解剖前至少30分钟，开始在冰上冒泡70毫升胆碱人工脑脊液，或胆碱ACSF。在32摄氏度下加入150毫升胆碱ACSF，加入碳原。将一个200毫升的结晶盘与棒磁铁放入密封的食物盒，并在菜中加入200毫升的ACSF。将 3D 打印界面切片架放在盘上，从结晶盘中去除多余的液体，直到仅保留覆盖界面滑动架网格的薄膜溶液。在食品盒底部加入几毫米的 aCSF，然后开始用碳原冒泡液体。然后，关闭密封盒，同时保持恒定的碳化物，形成一个加湿的、95%的氧气、5%富含二氧化碳的界面室。收获后，将大脑放在一块CSF浸泡的滤纸上，根据切片的首选角度解剖出感兴趣的区域。对于日冕切片，将大脑的冠状面固定到连接到振动切片器切割块上的 10 厘米 Petri 盘上，并在充满冰的大腔室中将块放在振动切片器的储层室中。用冰冷的胆管 aCSF 填充盘子，并使用切片器获得 300 微米厚的脑片，使用直径四毫米直径的一次性转移移液器在刀片的每一次通过后收集每片。让每片在32摄氏度的胆碱ACSF中恢复约10分钟后，它得到，然后转移片片到片片清洁纸，并加一滴胆碱ACSF。然后吸出多余的胆碱ACSF，并使用铲将切片转移到接口室的网格上，使切片在此环境中恢复至少30分钟。开始录音前 30 分钟，将活体泵连接到具有顶部和底部灌注的定制记录室，以优化组织切片的氧合和生存能力，并用 50 毫升超纯水清洁整个灌注系统。在清洁周期结束时，在不断的碳化下，在玻璃烧杯中用 50 毫升 aCSF 开始灌注记录室，并使用一次性宽口移液器将第一片片转移到烧杯上以取出透镜纸。当截面沉入烧杯底部时，将截面转移到记录室，并将切片架放在切片上，以最大限度地减少灌注流量引起的运动。使用亮场照明以 5 到 10 倍的放大倍率定位感兴趣的大脑区域。然后，使用 25 倍目标与 0.35 倍水浸透镜调整观察场的位置。使用荧光照明定位荧光微胶质细胞，并在移液器上回填10微升的ACSF，其中含有最终浓度时感兴趣的化合物。用柔和的摇动向下点尖，以去除夹在尖端中的任何气泡，将填充的移液器安装到连接到五毫升注射器的移液器支架上，柱塞位于五毫升位置，并安装在三轴微操纵器上。轻轻地向切片下部移液器，同时控制和调整目标，直到移液器尖端轻轻接触切片表面。现在调谐激光并切换到多光子模式。确保室从任何光源中筛选出来，然后打开非扫描式探测器。设置增益并使用具有颜色编码上限的查找表，以避免使图像中的像素饱和。然后，开始记录，总持续时间至少30分钟，慢慢地将注射器柱塞从5毫升位置压到1毫升，5分钟后，在5秒内，将化合物应用到该部分。对于图像分析，首先在感兴趣的文件上使用 ImageJ 执行 z 投影和漂移校正。然后，打开"冰"中修改的文件，绘制一个圆形的、直径为 35 微米的感兴趣区域，以注射部位为中心。使用 ROI 再次运行影片，以确保影片处于有利位置。然后，使用兴趣强度演化插件区域测量感兴趣区域中一段时间的均度强度，并保存结果为 xls 文件。该协议允许测量不同化合物（如ATP或血清素）引起的反应。虽然 ATP 的注射会导致荧光增加，但大多数 ATP 注射后，由于几分钟后恢复的组织变形，荧光会立即略有减少。感兴趣区域的大小也会影响定量，因为增加直径会降低实验之间的变异性，但会降低检测到的响应的准确性和幅度。评估特定时间点的微胶质反应可用于对不同化合物的统计比较，或测试添加到灌注溶液中的特定拮抗剂的效果。要在三维中量化活动性，请了解您可能需要更改 z 步长间隔和采集采样率，以符合分析要求。

Research

JoVE Journal

Neuroscience

In vivo Imaging of the Mouse Spinal Cord Using Two-photon Microscopy

In vivo Imaging of the Mouse Spinal Cord Using Two-photon Microscopy

在体内脊髓的鼠标线使用双光子显微镜成像

In vivo imaging using two-photon microscopy 1 in mice that have been genetically engineered to express fluorescent proteins in specific cell types 2-3 has ...

科研

神经科学

Preparation of Acute Subventricular Zone Slices for Calcium Imaging

钙成像的急性下区片的制备

The subventricular zone (SVZ) is one of the two neurogenic zones in the postnatal brain. The SVZ contains densely packed cells, including neural ...

In Vivo Two-photon Imaging Of Experience-dependent Molecular Changes In Cortical Neurons

In Vivo Two-photon Imaging Of Experience-dependent Molecular Changes In Cortical Neurons

在体内双光子成像的经验依赖的分子在大脑皮质神经元的变化

The brain's ability to change in response to experience is essential for healthy brain function, and abnormalities in this process contribute to a variety ...

In vivo Postnatal Electroporation and Time-lapse Imaging of Neuroblast Migration in Mouse Acute Brain Slices

In vivo Postnatal Electroporation and Time-lapse Imaging of Neuroblast Migration in Mouse Acute Brain Slices

成神经细胞迁移的小鼠急性脑片在体内电穿孔产后和时间推移成像

The subventricular zone (SVZ) is one of the main neurogenic niches in the postnatal brain. Here, neural progenitors proliferate and give rise to ...

Two-photon Imaging of Cellular Dynamics in the Mouse Spinal Cord

细胞动力学在小鼠脊髓双光子成像

Two-photon (2P) microscopy is utilized to reveal cellular dynamics and interactions deep within living, intact tissues. Here, we present a method for ...

Recording Synaptic Plasticity in Acute Hippocampal Slices Maintained in a Small-volume Recycling-, Perfusion-, and Submersion-type Chamber System

小容积回收-灌注-浸没式室系统中维持的急性海马切片的突触可塑性记录

Even though experiments on brain slices have been in use since 1951, problems remain that reduce the probability of achieving a stable and successful ...

Two-photon Calcium Imaging in Neuronal Dendrites in Brain Slices

脑切片神经元树突的双光子钙成像

Calcium (Ca2+) imaging is a powerful tool to investigate the spatiotemporal dynamics of intracellular Ca2+ signals in neuronal dendrites. Ca2+ ...

In Vivo Two-photon Imaging of Cortical Neurons in Neonatal Mice

In Vivo Two-photon Imaging of Cortical Neurons in Neonatal Mice

体内新生小鼠皮层神经元的双光子成像

Two-photon imaging is a powerful tool for the in vivo analysis of neuronal circuits in the mammalian brain. However, a limited number of in vivo imaging ...

Imaging of Intracellular ATP in Organotypic Tissue Slices of the Mouse Brain using the FRET-based Sensor ATeam1.03YEMK

Imaging of Intracellular ATP in Organotypic Tissue Slices of the Mouse Brain using the FRET-based Sensor ATeam1.03YEMK

使用基于 FRET 的传感器 ATeam1.03YEMK在小鼠大脑的组织切片中进行细胞内 ATP 成像

Neuronal activity in the central nervous system (CNS) evokes a high demand on cellular energy provided by the breakdown of adenosine triphosphate (ATP). A ...

Transpupillary Two-Photon In Vivo Imaging of the Mouse Retina

小鼠视网膜的经瞳双光子体内成像

The retina transforms light signals from the environment into electrical signals that are propagated to the brain. Diseases of the retina are prevalent ...