我们衷心感谢加拿大卫生研究院 (NSERC) 和加拿大科学与工程研究所的支持。波斯特得到了亚历山大·格雷厄姆加拿大研究生奖学金 (NSERC) 的支持。

所有的程序都是按照国家研究委员会的《动物保护和使用实验指南》 (第八版; 2011) 和加拿大动物保护委员会关于试验动物的护理和使用指南 (1993) 进行的, 并由西安大略大学动物学委员会动物保育委员会批准。
下面的过程可以应用于实验研究皮层贡献的运动控制在步行猫。
1. 仪器
<ol><li>构造用于评估障碍物记忆的仪器。 注: 该仪器由2.43 米长, 29 厘米宽的走道包围, 18 厘米高清晰丙烯酸墙 (图 1)。一个狭窄的插槽沿设备的一半, 允许25.8 厘米宽 x 3 毫米厚的障碍, 以提高或从人行道上使用杠杆安装在行走表面。</li>
	<li>为了确保动物的注意力保持在进食上, 避免用手抬高或降低障碍物。相反, 障碍可以提高或降低使用实验者的腿移动的杠杆下的走道, 使实验者继续喂养的动物。</li>
	<li>适当地维护杠杆系统, 以确保障碍可以提高或降低无声。</li>
	<li>使用一个小的高架平台 (23 厘米长的 x 23 厘米宽 x 16 厘米高), 在软食物放置, 以指导动物的运动。</li>
	<li>记录所有使用以太网摄像头 (54 帧/秒) 安装在三脚架1.85 米远离走道中线的试验。</li>
</ol>2. 培训程序
注意: 为了获得成功的数据, 在行为测试之前的训练期间, 确保每只动物都能适当地适应测试室和仪器。反复接触到一个新的环境将有助于减少令人吃惊或其他压力的行为。 驯化可能因动物而异, 可能需要1-2 月的训练。根据动物进食的重点和动机, 最初的驯化疗程可达5分钟。随后的会议应旨在增加动物的工作时间 (通常约20-25 分钟)。
<ol><li>获得成熟 (> 6 月的年龄) 国内短毛猫从任何重量或性别的商业实验室饲养员。 注意: 在考虑哪些动物应该包括在研究中时, 食物的工作动机和合作性的性格构成了选择标准。</li>
	<li>使每只动物都能佩戴1米长的皮带。将皮带拴在走道中点上方的架子上。 注意: 这使得动物可以沿着器械的中心部分不带任何张力, 从而鼓励动物留在仪器的这一部分。建立这样的边界有助于使用移动测试对象。</li>
	<li>把动物放在人行道上, 让它从平台上吃软食物。 注意: 这项初步训练的目的之一是确保动物在向前移动时很容易跟随食物平台, 并能舒适地与安全带和皮带行走。使用软性食物作为积极的强化, 鼓励动物在每次训练或测试期间保持专注, 并促进舒适的工作环境。</li>
	<li>确保动物对搬运感到舒适, 包括必须将动物移到人行道起始区域的情况。</li>
</ol>3. 行为训练和测试协议
注意: 障碍记忆被评估在两个范例: 一个视觉依赖障碍记忆任务, 和一个触觉相关的障碍记忆任务。这两种范式都应在最初的培训和随后的测试中使用。
<ol><li>
视觉障碍记忆
	<ol>
<li>要评估视觉障碍存储器, 请将障碍物提升到人行道上 (图 2A)。将平台放置在障碍物的远处。将动物放在人行道的开始区域。</li>
		<li>允许动物接近食物, 只用前腿踩在障碍物上, 以便从站台上进食。</li>
		<li>当动物继续进食时, 降低障碍物, 使其变得与人行道齐平, 以防止任何进一步的视觉或触觉输入。</li>
		<li>在一个可变的延迟期间, 移动食物批转再鼓励动物恢复步行;这种延迟可以小于 1 s 到2分钟以上。</li>
		<li>重要的是, 在没有障碍的情况下进行试验, 以防止习惯性的障碍和学习回避反应的发展。在这种视觉障碍缺席试验中, 在将动物放在人行道的起始区之前, 确保障碍物不会被抬到人行道上。</li>
		<li>观察 hindleg 步在障碍目前和障碍缺席试验, 以验证典型的运动行为和完整的视觉障碍记忆前冷却。确保动物能在无接触的情况下清除障碍, 而在障碍性试验中, 四条腿的跨步明显升高。 注意: 观看训练试验的视频可能有助于验证。</li>
	</ol>
</li>
	<li>
触觉障碍记忆
	<ol>
<li>要评估触觉障碍记忆, 确保在将动物放在人行道的起始区域之前, 不会将障碍物抬到人行道上 (图 2B)。</li>
		<li>允许动物走向位于障碍物的远侧的食物平台。</li>
		<li>当动物进食时, 将障碍物抬高到食物盘下的人行道上, 防止障碍物的任何视觉输入。</li>
		<li>当食物被向前移动时, 请注意, 动物在踏过前应先用前腿接触障碍物。</li>
		<li>允许动物继续进食, 同时跨越他们的前后腿的障碍。在此期间, 降低障碍, 使其成为冲洗与人行道, 以防止任何进一步的视觉或触觉输入。</li>
		<li>在一个可变的延迟期间, 移动食物再次批转鼓励动物恢复步行。</li>
		<li>重要的是, 在不存在障碍的情况下进行试验, 避免前腿接触, 防止习惯性障碍和学习回避反应的发展。<ol>
<li>在这些触觉障碍缺席试验, 有动物的方法和吃的食物平台, 如步骤3.2.1 所述。然而, 在把食物向前推进的步骤3.2.3 前, 提高和降低障碍 (步骤 3.2.2)。确保允许动物继续进食的类似的延迟期 (步骤 3.2.4) 在运动的最后的继续之前 (步 3.2.5)。</li>
		</ol>
</li>
		<li>观察 hindleg 步在障碍目前和障碍缺席试验, 以验证正常的运动行为和完整的视觉障碍记忆前冷却。</li>
	</ol>
</li>
</ol>4. 视频分析
注: 为了评估障碍记忆, 在初始训练期间进行的分析和随后的测试后, 冷却环植入涉及量化的峰值步高度, 步隙, 以及脚趾和障碍物之间的水平距离在每个步骤的高峰期视觉和触觉范例 (图 2C)。
<ol><li>使用自定义手写脚本分析视频。</li>
	<li>每一次试验, 每步通过标记脚趾最靠近相机的位置来跟踪每个脚印。</li>
	<li>在每个步进轨迹的最高点 (图 2C) 中, 测量峰值台阶高度作为脚趾和人行道表面之间的垂直距离。</li>
	<li>在障碍物的试验中, 将台阶间隙测量为在障碍物的高度上减去的台阶高度。</li>
	<li>此外, 测量障碍-目前试验中每一步的峰值处的脚趾与障碍物之间的水平距离。</li>
	<li>确认在冷却回路植入之前, 障碍记忆能力是完整的, 通过验证在障碍试验中的峰值台阶高度是否提升, 与在无障碍试验中的步进相比。</li>
</ol>5. 冷却回路 (Cryoloop) 植入
<ol><li>根据先前报告的外科手术程序8 (图 3), 在5和7区域 cryoloops 种植体。</li>
	<li>简而言之, 对于每个半球, 执行一个开颅和 durotomy 从斯科特·霍斯利-克拉克坐标20 A15 到 A25, 以揭露 ansate 和侧面沟的接合点。</li>
	<li>从23口径不锈钢皮下注射导管中定位单个冷却回路, 与顶叶区5或7的皮质表面直接接触。</li>
	<li>安全的基础, 每个 cryoloop 的头骨与牙科丙烯酸锚定不锈钢螺丝。</li>
	<li>关闭开颅与额外的牙科丙烯酸;绘制到丙烯酸边缘的皮肤边缘和缝合在一起。</li>
</ol>6. 皮质冷却协议
<ol><li>
实验设置 注意: 在将动物带入测试室之前, 要准备并测试冷却回路。该冷却电路由一甲醇储气罐 (3.2 毫米外径, 1.6 毫米身份证), 往复式活塞泵, 和干冰浴连接通过聚四氟乙烯管 (1.6 毫米外径, 0.5 毫米身份证;图 4)。此外, 还需要一个数字温度计。<ol>
<li>在冰浴中加入500毫升干冰至 200 mL 甲醇。配合油管两端紧贴在入口和出口的一个虚拟 cryoloop, 以完成冷却电路。</li>
		<li>将热电偶插头连接到数字温度计上, 使用由两个热电偶连接器和热电偶线组成的电缆进行连续温度监测。当一端插入温度计时, 确保此电缆的长度足以到达动物的头部。</li>
		<li>使用开关打开活塞泵。 注: 甲醇应从油层中抽出, 通过泵到干冰浴, 在那里流动的甲醇在油管中将冷却到-75 ° c。冷却后的甲醇将退出冰浴, 并通过附着的 cryoloop, 然后返回甲醇库。</li>
		<li>确保泵的设置, 在冰浴内的油管长度, 和从冰浴到假循环的油管长度是最佳的, 使假 cryoloop 温度可以达到一个稳定的状态在-5.0 ° c 左右。 注: 此初始设置期间所达到的温度通常足以达到3.0 ±1.0 ° c 时的测试温度, 而同一系统用于冷却植入的 cryoloop。通过调整泵的转速、增加在冰浴中浸没的油管长度和/或最小化从冰浴到 cryoloop 的油管长度, 可以解决实现足够冷却的困难。</li>
		<li>如有必要, 通过管端管接头和法兰端部的管子端部用翻边工具来延长油管的一端。使用连接器以类似的法兰端连接所需长度的油管。</li>
		<li>验证所有连接是否舒适, 并且不存在泄漏。一旦满意的初始设置, 开关泵关闭, 并删除假 cryoloop;这个电路现在已经为一个试验动物准备好了。</li>
	</ol>
</li>
	<li>
行为测试
	<ol>
<li>将动物放在测试仪器上。滑动线束在头上, 并确保表带舒服周围的动物。系上皮带</li>
		<li>卸下植入的 cryoloop 的保护盖, 露出入口和出口管。适合油管两端紧贴在 cryoloop 的入口和出口管。将热电偶插头连接到数字温度计。</li>
		<li>开始测试阶段与视觉 (步骤 3.1) 或触觉 (步骤 3.2) 障碍记忆试验。跟随对所有四类型的另外的试验 (视觉障碍-当前, 视觉障碍-缺席, 触觉障碍-存在, 触觉障碍-缺席) 以一个随机的方式。 注: 一个典型的测试环节由一个 "温暖" 的试验块组成, 在没有冷却的情况下观察记忆引导的避障以建立基线测量。</li>
		<li>打开活塞泵, 等待 cryoloop 达到3.0 ±1.0 ° c (1-2 分钟) 的温度。然后, 运行一个 ' 凉爽 ' 的试验块后, 活塞泵已接通。在这段试验中, 如果需要, 评估冷却区的贡献, 以记忆为导向。确保 cryoloop 的温度保持在3.0 ±1.0 ° c 整块。 注: 所有四种试验类型应在整个区块中随机穿插。</li>
		<li>在活塞泵关闭后, 运行最后的 "复" 测试块, cryoloop 已恢复到原来的温度。 注意: 在此块中重新建立基线步进行为。同样, 所有四种试验类型都应在整个区块中随机穿插。</li>
	</ol>
</li>
	<li>
清理
	<ol>
<li>当行为测试结束时, 从入口和出口管中取出油管。注意剩余的甲醇可能从油管末端滴下, 可能会刺激动物。</li>
		<li>确保更换防护帽。在把动物送回蚁群之前, 去掉皮带和安全带。修剪油管两端 (3-4 毫米) 使用油管切割器, 以防止在下一个测试日漏连接。</li>
	</ol>
</li>
</ol>7. 验证冷却范围
<ol><li>在行为测试结束时, 请确认使用先前报告的技术8, 将停用的范围限制在每个 cryoloop 下方的皮层区域。 注: 这可以通过跃层映射12或与热成像相机13、14、19进行验证。</li>
</ol>

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Cortex. 25, 84-96 (2015).</li><li>Coomber, 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=Cortical+inactivation+by+cooling+in+small+animals.">Cortical inactivation by cooling in small animals.</a> Front. Syst. Neurosci. 5, 53 (2011).</li><li>Malmierca, M. S., Anderson, L. A., Antunes, F. 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+cortical+modulation+of+stimulus-specific+adaptation+in+the+auditory+midbrain+and+thalamus:+a+potential+neuronal+correlate+for+predictive+coding.">The cortical modulation of stimulus-specific adaptation in the auditory midbrain and thalamus: a potential neuronal correlate for predictive coding.</a> Front. Syst. Neurosci. 9, 19 (2015).</li><li>Antunes, F. M., Malmierca, M. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+auditory+cortex+deactivation+on+stimulus-specific+adaptation+in+the+medial+geniculate+body.">Effect of auditory cortex deactivation on stimulus-specific adaptation in the medial geniculate body.</a> J. Neurosci. 31, 17306-17316 (2011).</li><li>Peel, T. R., Johnston, K., Lomber, S. G., Corneil, B. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bilateral+saccadic+deficits+following+large+and+reversible+inactivation+of+unilateral+frontal+eye+field.">Bilateral saccadic deficits following large and reversible inactivation of unilateral frontal eye field.</a> J. Neurophysiol. 111, 415-433 (2014).</li><li>Wong, C., Wong, G., Pearson, K. G., Lomber, S. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Memory-guided+stumbling+correction+in+the+hindlimb+of+quadrupeds+relies+on+parietal+area+5.">Memory-guided stumbling correction in the hindlimb of quadrupeds relies on parietal area 5.</a> Cereb. Cortex. , (2016).</li><li>Horsley, V., Clarke, R. 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+structure+and+function+of+the+cerebellum+examined+by+a+new+method.">The structure and function of the cerebellum examined by a new method.</a> Brain Behav Evol. 31, 45-124 (1908).</li><li>Lomber, S. G., Malhotra, S., Hall, 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=Functional+specialization+in+non-primary+auditory+cortex+of+the+cat:+areal+and+laminar+contributions+to+sound+localization.">Functional specialization in non-primary auditory cortex of the cat: areal and laminar contributions to sound localization.</a> Hear. Res. 229, 31-45 (2007).</li><li>Johnston, K., Koval, M. J., Lomber, S. G., Everling, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Macaque+dorsolateral+prefrontal+cortex+does+not+suppress+saccade-related+activity+in+the+superior+colliculus.">Macaque dorsolateral prefrontal cortex does not suppress saccade-related activity in the superior colliculus.</a> Cereb. Cortex. 24, 1373-1388 (2014).</li></ol>

作者没有什么可透露的。

所描述的范例采用冷却诱导的离散皮层区 deactivations 的 cryoloop, 以研究记忆制导障碍的运动在猫。视觉和触觉障碍记忆范例是相当简单的动物执行, 因为他们利用自然主义的运动行为, 发生在最小的努力, 当一个动物的动机是跟随一个移动的食物来源。因此, 大部分的培训阶段是专门适应动物的测试室和冷却设备。大多数动物都需要反复接触, 佩戴安全带, 并通过皮带拴住, 然后才能舒适地和自然地在...

在自然的、不平坦的地形上, 关于障碍物的感官信息可以通过视觉或触觉获得, 可以迅速地改变运动以躲避。步进运动的细致协调涉及多个皮层区域1,2。例如, 电机皮层3,4和顶叶皮层5,6,7在复杂的运动任务中被牵连, 如避障。在足动物中, 避免障碍所需的阶跃调制必须伸展至前腿和后腿。如果向前运动被延迟在前腿和 hindleg 障碍间隙之间 (可能出现作为动物踏入小心地通过一个复杂, 自然主义的环境跟踪猎物), 关于在记忆中维护的障碍的信息被用来指导一旦走了, hindleg 就会越过障碍物。
实验技术旨在解除离散皮层区域可以用来研究皮层贡献的记忆引导障碍运动。冷却诱发的皮质失活提供了一种可逆的, 可靠的, 可重现的方法来评估皮质对显性行为的贡献8。Cryoloops 由不锈钢管制成, 是特定于皮层的兴趣, 确保高度选择性和离散的基因座的失活。一旦植入, 冷冻甲醇泵通过管腔的 cryoloop 冷却的皮层区域直接下方的循环到 < 20 ° c。在这个临界温度下, 突触的传递在大脑皮层的区域被抑制。这种钝化可以通过停止甲醇的流动来逆转。该方法已用于研究皮层对感官处理和行为的贡献9,10,11,12,13,14,15,16,17, 以及跳眼移动的马达控制18和内存引导障碍运动19。
该协议的目的是使用可逆冷却诱导 deactivations 评估的参与, 顶叶皮层区的运动协调在猫。特别地, 记忆引导障碍运动被检查与或没有活跃顶叶皮层。这些方法已经被用来成功地证明了顶骨面积5在记忆制导避障中的作用, 在行走 cat19。

<table>
	<tbody>
		<tr>
			<td>Camera</td>
			<td>IDS Imaging Development Systems GmbH</td>
			<td>Model: UI-5240CP-C-HQ</td>
			<td></td>
		</tr>
		<tr>
			<td>Intake tubing</td>
			<td>Restek</td>
			<td>25306</td>
			<td>Unflanged end is submerged in the methanol reservoir while the flanged end is connected to the pump</td>
		</tr>
		<tr>
			<td>Pump</td>
			<td>Fluid Metering, Inc.</td>
			<td>Model: QG 150</td>
			<td></td>
		</tr>
		<tr>
			<td>Nalgene Dewar vacuum flask</td>
			<td>Sigma-Aldrich</td>
			<td>F9401</td>
			<td></td>
		</tr>
		<tr>
			<td>Teflon tubing</td>
			<td>Ezkem</td>
			<td>A051754</td>
			<td></td>
		</tr>
		<tr>
			<td>Microprobe thermometer</td>
			<td>Physitemp</td>
			<td>Model: BAT-12</td>
			<td></td>
		</tr>
		<tr>
			<td>Flanged tube end fittings</td>
			<td>Valco Instruments Co. Inc.</td>
			<td>CF-1BK</td>
			<td>Assorted colours available for colour coding. Packages include the same number of washers as fittings</td>
		</tr>
		<tr>
			<td>Washers</td>
			<td>Valco Instruments Co. Inc.</td>
			<td>CF-W1</td>
			<td>Extra washers</td>
		</tr>
		<tr>
			<td>Flanging kit</td>
			<td>Pro Liquid GmbH</td>
			<td>201553</td>
			<td></td>
		</tr>
		<tr>
			<td>Tubing connector</td>
			<td>Restek</td>
			<td>25323</td>
			<td></td>
		</tr>
		<tr>
			<td>Tubing cutter</td>
			<td>Restek</td>
			<td>25069</td>
			<td></td>
		</tr>
		<tr>
			<td>Male thermocouple connector</td>
			<td>Omega</td>
			<td>SMPW-T-M</td>
			<td>Used to make cable connection to thermometer</td>
		</tr>
		<tr>
			<td>Thermocouple wire</td>
			<td>Omega</td>
			<td>PP-T-24S</td>
			<td>Used to make cable connection to thermometer</td>
		</tr>
		<tr>
			<td>MATLAB</td>
			<td>MathWorks</td>
			<td>n/a</td>
			<td></td>
		</tr>
	</tbody>
</table>

reversible cooling-induced deactivations to study cortical contributions to obstacle memory in the walking cat

在复杂的、自然主义的地形上, 关于环境障碍的感官信息可以用来快速调整运动动作以躲避。例如, 在猫, 关于一个即将到来的障碍的视觉信息可以调节步避。运动适应也可以发生独立的视觉, 因为突然的触觉输入到腿部由一个预期的障碍可以修改的步的所有四条腿, 以避免。这种复杂的运动协调涉及上结构, 如顶叶皮质。该协议描述了使用可逆的, 冷却诱导的皮层失活, 以评估顶叶皮层贡献的记忆引导障碍运动在猫。小的冷却回路, 被称为 cryoloops, 是特殊的形状, 以关闭分散的兴趣区域, 以评估其贡献的公开行为。这种方法已被用来阐明的作用, 5 在记忆引导的障碍, 避免在猫的顶叶区。

该协议已成功地用于检查顶叶皮层贡献的障碍记忆在步行猫19。在这项研究中, cryoloops 在5和7在三成人 (> 6 月龄) 雌性猫 (图 5A) 上被植入了双侧壁区。在没有冷却 (温暖、控制条件) 的情况下, 或者当5或7区被双边钝化时, 在触觉障碍记忆模式中对动物进行了评估。
该研究的结果表明, 当5区...

Watch this Scientific Journal Video about Reversible Cooling-induced Deactivations to Study Cortical Contributions to Obstacle Memory in the Walking Cat at JoVE.com

Reversible Cooling-induced Deactivations to Study Cortical Contributions to Obstacle Memory in the Walking Cat

复杂的运动在自然主义的环境需要仔细的肢体协调涉及顶叶皮层的区域。下面的协议描述了可逆冷却诱导失活的使用, 以证明顶骨面积5的作用, 在记忆制导避障的步行猫。

可逆冷却诱导 Deactivations 对行走猫的障碍记忆的皮层贡献研究

On complex, naturalistic terrain, sensory information about an environmental obstacle can be used to rapidly adjust locomotor movements for avoidance. For ...

reversible-cooling-induced-deactivations-to-study-cortical

Universidad de Cartagena UDEC

Research

JoVE Journal

Behavior

6.9K 次观看.  University of Western Ontario. 复杂的运动在自然主义的环境需要仔细的肢体协调涉及顶叶皮层的区域。下面的协议描述了可逆冷却诱导失活的使用, 以证明顶骨面积5的作用, 在记忆制导避障的步行猫。

视频: Reversible Cooling-induced Deactivations to Study Cortical Contributions to Obstacle Memory in the Walking Cat

Stereotaxic Microinjection of Viral Vectors Expressing Cre Recombinase to Study the Role of Target Genes in Cocaine Conditioned Place Preference

Microinjecting recombinant adenoassociated viral (rAAV) vectors expressing Cre recombinase into distinct mouse brain regions to selectively knockout genes of interest allows for enhanced temporally- and regionally-specific control of gene deletion, compared to existing methods. While conditional deletion can also be achieved by mating mice that express Cre recombinase under the control of specific gene promoters with mice carrying a floxed gene, stereotaxic microinjection allows for targeting of discrete brain areas at experimenter-determined time points of interest. In the context of cocaine conditioned place preference, and other cocaine behavioral paradigms such as self-administration or psychomotor sensitization that can involve withdrawal, extinction and/or reinstatement phases, this technique is particularly useful in exploring the unique contribution of target genes to these distinct phases of behavioral models of cocaine-induced plasticity. Specifically, this technique allows for selective ablation of target genes during discrete phases of a behavior to test their contribution to the behavior across time. Ultimately, this understanding allows for more targeted therapeutics that are best able to address the most potent risk factors that present themselves during each phase of addictive behavior.

This article describes how to microinject viral vectors into mouse brain and then test in a conditioned place preference paradigm that includes an acquisition, extinction and reinstatement phase.

立体定向显微注射病毒载体表达Cre重组酶研究的靶基因的作用在可卡因条件性位置偏爱

Microinjecting recombinant adenoassociated viral (rAAV) vectors expressing Cre recombinase into distinct mouse brain regions to selectively knockout genes ...

科研

行为学

A Procedure to Study the Effect of Prolonged Food Restriction on Heroin Seeking in Abstinent Rats

In human drug addicts, exposure to drug-associated cues or environments that were previously associated with drug taking can trigger relapse during abstinence. Moreover, various environmental challenges can exacerbate this effect, as well as increase ongoing drug intake.
The procedure we describe here highlights the impact of a common environmental challenge, food restriction, on drug craving that is expressed as an augmentation of drug seeking in abstinent rats.
Rats are implanted with chronic intravenous i.v. catheters, and then trained to press a lever for i.v. heroin over a period of 10-12 days. Following the heroin self-administration phase the rats are removed from the operant conditioning chambers and housed in the animal care facility for a period of at least 14 days. While one group is maintained under unrestricted access to food (sated group), a second group (FDR group) is exposed to a mild food restriction regimen that results in their body weights maintained at 90% of their nonrestricted body weight. On day 14 of food restriction the rats are transferred back to the drug-training environment, and a drug-seeking test is run under extinction conditions (i.e.&#160;lever presses do not result in heroin delivery).
The procedure presented here results in a highly robust augmentation of heroin seeking on test day in the food restricted rats. In addition, compared to the acute food deprivation manipulations we have used before, the current procedure is a more clinically relevant model for the impact of caloric restriction on drug seeking. Moreover, it might be closer to the human condition as the rats are not required to go through an extinction-training phase before the drug-seeking test, which is an integral component of the popular reinstatement procedure.

A procedure that allows a demonstration of robust augmentation of drug seeking in food-restricted rats is described. Following heroin self-administration training, rats go through an abstinence period, in a drug-free environment, during which they are mildly food restricted. Drug seeking is then tested in the drug-associated environment.

一个过程来研究延长食品的限制对海洛因寻求戒断大鼠的影响

In human drug addicts, exposure to drug-associated cues or environments that were previously associated with drug taking can trigger relapse during ...

Uncovering Beat Deafness: Detecting Rhythm Disorders with Synchronized Finger Tapping and Perceptual Timing Tasks

A set of behavioral tasks for assessing perceptual and sensorimotor timing abilities in the general population (i.e., non-musicians) is presented here with the goal of uncovering rhythm disorders, such as beat deafness. Beat deafness is characterized by poor performance in perceiving durations in auditory rhythmic patterns or poor synchronization of movement with auditory rhythms (e.g., with musical beats). These tasks include the synchronization of finger tapping to the beat of simple and complex auditory stimuli and the detection of rhythmic irregularities (anisochrony detection task) embedded in the same stimuli. These tests, which are easy to administer, include an assessment of both perceptual and sensorimotor timing abilities under different conditions (e.g., beat rates and types of auditory material) and are based on the same auditory stimuli, ranging from a simple metronome to a complex musical excerpt. The analysis of synchronized tapping data is performed with circular statistics, which provide reliable measures of synchronization accuracy (e.g., the difference between the timing of the taps and the timing of the pacing stimuli) and consistency. Circular statistics on tapping data are particularly well-suited for detecting individual differences in the general population. Synchronized tapping and anisochrony detection are sensitive measures for identifying profiles of rhythm disorders and have been used with success to uncover cases of poor synchronization with spared perceptual timing. This systematic assessment of perceptual and sensorimotor timing can be extended to populations of patients with brain damage, neurodegenerative diseases (e.g., Parkinson&#8217;s disease), and developmental disorders (e.g., Attention Deficit Hyperactivity Disorder).

Behavioral tasks that allow for the assessment of perceptual and sensorimotor timing abilities in the general population (i.e., non-musicians) are presented. Synchronization of finger tapping to the beat of an auditory stimuli and detecting rhythmic irregularities provides a means of uncovering rhythm disorders.

揭开节拍耳聋:检测节律紊乱与同步手指敲击和感知定时任务

A set of behavioral tasks for assessing perceptual and sensorimotor timing abilities in the general population (i.e., non-musicians) is presented here ...

Applying Stereotactic Injection Technique to Study Genetic Effects on Animal Behaviors

Stereotactic injection is a useful technique to deliver high titer lentiviruses to targeted brain areas in mice. Lentiviruses can either overexpress or knockdown gene expression in a relatively focused region without significant damage to the brain tissue. After recovery, the injected mouse can be tested on various behavioral tasks such as the Open Field Test (OFT) and the Forced Swim Test (FST). The OFT is designed to assess locomotion and the anxious phenotype in mice by measuring the amount of time that a mouse spends in the center of a novel open field. A more anxious mouse will spend significantly less time in the center of the novel field compared to controls. The FST assesses the anti-depressive phenotype by quantifying the amount of time that mice spend immobile when placed into a bucket of water. A mouse with an anti-depressive phenotype will spend significantly less time immobile compared to control animals. The goal of this protocol is to use the stereotactic injection of a lentivirus in conjunction with behavioral tests to assess how genetic factors modulate animal behaviors.

Stereotactic injection of lentiviruses expressing cDNAs or shRNAs can modulate gene expression in specific brain areas of mice. Here, we present a protocol to combine stereotactic injections with behavioral tasks, such as the Open Field Test (OFT) and the Forced Swim Test (FST).

应用立体定向注射技术在研究动物行为的遗传效应

Stereotactic injection is a useful technique to deliver high titer lentiviruses to targeted brain areas in mice. Lentiviruses can either overexpress or ...

Utilizing the Modified T-Maze to Assess Functional Memory Outcomes After Cardiac Arrest

Background: Evaluating mild to moderate cognitive impairment in a global cerebral ischemia (i.e. cardiac arrest) model can be difficult due to poor locomotion after surgery. For example, rats who undergo surgical procedures and are subjected to the Morris water maze may not be able to swim, thus voiding the experiment.
New Method: We established a modified behavioral spontaneous alternation T-maze test. The major advantage of the modified T-maze protocol is its relatively simple design that is powerful enough to assess functional learning/memory after ischemia. Additionally, the data analysis is simple and straightforward. We used the T-maze to determine the rats&#39; learning/memory deficits both in the presence or absence of mild to moderate (6 min) asphyxial cardiac arrest (ACA). Rats have a natural tendency for exploration and will explore the alternate arms in the T-maze, whereas hippocampal-lesioned rats tend to adopt a side-preference resulting in decreased spontaneous alternation ratios, revealing the hippocampal-related functional learning/memory in the presence or absence of ACA.
Results: ACA groups have higher side-preference ratios and lower alternations as compared to control.
Comparison with Existing Method(s): The Morris water and Barnes maze are more prominent for assessing learning/memory function. However, the Morris water maze is more stressful than other mazes. The Barnes maze is widely used to measure reference (long-term) memory, while ACA-induced neurocognitive deficits are more closely related to working (short-term) memory.
Conclusions: We have developed a simple, yet effective strategy to delineate working (short-term) memory via the T-maze in our global cerebral ischemia model (ACA).

This protocol describes the use of a modified T-maze to evaluate functional learning/memory in asphyxia cardiac arrest-induced cerebral ischemia.

应用改良 T 迷宫评估心脏骤停后的功能记忆结果

Background: Evaluating mild to moderate cognitive impairment in a global cerebral ischemia (i.e. cardiac arrest) model can be difficult due to poor ...

Modified Blood Collection from Tail Veins of Non-anesthetized Mice with a Vacuum Blood Collection System and Eyeglass Magnifier

Blood sample collection is the basis of experimental animal research. It is of importance to obtain adequate blood samples for various research purposes. The tail veins of mice are small, and it is sometimes difficult to obtain the required blood volume by conventional puncture methods. This study investigates the superiority of repeated blood sample collection from tail veins of mice through use of a vacuum blood collection system and eyeglass magnifier (experimental group) compared to conventional blood sampling methods (conventional group), performed by beginners and experts, respectively. With the help of an eyeglass magnifier, a butterfly needle tip is inserted into the tail vein of each mouse in the experimental group. When the vein is penetrated successfully, a blood sample is collected in the vacuum collection tube by insertion of the rubber end of a butterfly needle into the vacuum blood collection tube. The plunger is then used to collect blood without the help of the eyeglass magnifier in the conventional group. Success rates of blood sample collection by the beginners and experts were shown to be 70% vs. 100% (p &#60; 0.01) in the experimental group and 35% vs. 85% (p &#60; 0.01) in the conventional group. For both beginners and experts, puncture times required for obtaining required blood sample were significantly lower in the experimental group compared to the conventional group (2.40 &#177; 0.75 vs. 2.90 &#177; 0.31, p &#60; 0.05; 1.15 &#177; 0.37 vs. 1.55 &#177; 0.76, p &#60; 0.05). In conclusion, the presented blood sampling technique is feasible and easy to practice and enables frequent sampling of adequate blood volumes from non-anesthetized mice.&#160;

This study reports blood sampling from tail vein in mice using a vacuum extraction tube system with eyeglass magnifier. Our method is easy to practice and could be used for repeat blood sampling in mice.

真空采血系统和眼镜放大镜从非麻醉小鼠尾静脉中修饰采血

Blood sample collection is the basis of experimental animal research. It is of importance to obtain adequate blood samples for various research purposes. ...

Assessing the Coherence of Parents' Short Narratives Regarding their Child Using the Five-Minute Speech Sample Procedure

Valid and efficient measures for assessing the quality of parent-child relationships are needed to facilitate research and evidence-based practice with parents. This paper focuses on such a method, namely Five-Minute Speech Sample-Coherence (FMSS-Coherence). In this method, a parent is asked to speak for five uninterrupted minutes about her/his child and their relationship. The resulted narrative is coded for coherence, namely the extent to which the parent provides&#160;in the narrative&#160;a clear, consistent, multidimensional and well-supported portrayal of the child. FMSS-Coherence is based on attachment research that shows that the coherence of parents&#8217; narratives is indicative of the quality of the parent-child relationship and child adjustment. It overcomes the limitations of attachment narrative measures which are typically labor intensive. FMSS-Coherence is less nuanced than extant attachment narrative measures. Yet, studies of families from different cultural backgrounds, across different child ages and in the context of typically developing children as well as children with special needs suggest that coherence can be reliably evaluated using the FMSS procedure. Furthermore, parents&#8217; FMSS-Coherence is associated with parenting quality and children&#8217;s socio-emotional adjustment. Thus, it holds promise for researchers and practitioners who seek a relatively time- and cost-effective method for assessing the coherence of parents&#8217; narratives regarding their child.

This paper introduces a method for assessing the coherence of parents' short narratives regarding their child and their relationship using the five-minute speech sample procedure.

使用五分钟言语样本程序评估家长对孩子的简短叙述的一致性

Valid and efficient measures for assessing the quality of parent-child relationships are needed to facilitate research and evidence-based practice with ...

Measuring the Switch Cost of Smartphone Use While Walking

This paper presents a study protocol to measure the task-switching cost of using a smartphone while walking. This method involves having participants walk on a treadmill under two experimental conditions: a control condition (i.e., simply walking) and a multitasking condition (i.e., texting while walking). During these conditions, the participants must switch between the tasks related to the experimental condition and a direction determining task. This direction task is done with a point-light walker figure, seemingly walking towards the left or the right of the participant. Performance on the direction task represents the participant&#8217;s task-switching costs. There were two performance measures: 1) correct identification of the direction and 2) response time. EEG data are recorded in order to measure the alpha oscillations and cognitive engagement occurring during the task switch. This method is limited in its ecological validity: pedestrian environments have many stimuli occurring simultaneously and competing for attention. Nonetheless, this method is appropriate for pinpointing task-switching costs. The EEG data allow the study of the underlying mechanisms in the brain that are related to differing task-switching costs. This design allows the comparison between task switching when doing one task at a time, as compared to task switching when multitasking, prior to the stimulus presentation. This allows understanding and pinpointing both the behavioral and neurophysiological impact of these two different task-switching conditions. Furthermore, by correlating the task-switching costs with the brain activity, we can learn more about what causes these behavioral effects. This protocol is an appropriate base for studying the switching cost of different smartphone uses. Different tasks, questionnaires, and other measures can be added to it in order to understand the different factors involved in the task-switching cost of smartphone use while walking.

This study design measures the task-switching cost of using a smartphone while walking. Participants undergo two experimental conditions: a control condition (walking) and a multitasking condition (texting while walking). Participants switch between these tasks and a direction determining task. EEG data as well as behavioral measures are recorded.

测量边走边使用智能手机的开关成本

This paper presents a study protocol to measure the task-switching cost of using a smartphone while walking. This method involves having participants walk ...

Assessment of Memory Function in Pilocarpine-induced Epileptic Mice

Cognitive impairment is one of the most common comorbidities in temporal lobe epilepsy. To recapitulate epilepsy-associated cognitive decline in an animal model of epilepsy, we generated pilocarpine-treated chronic epileptic mice. We present a protocol for three different behavioral tests using these epileptic mice: novel object location (NL), novel object recognition (NO), and pattern separation (PS) tests to evaluate learning and memory for places, objects, and contexts, respectively. We explain how to set the behavioral apparatus and provide experimental procedures for the NL, NO, and PS tests following an open field test that measures the animals&#8217; basal locomotor activities. We also describe the technical advantages of the NL, NO, and PS tests with respect to other behavioral tests for assessing memory function in epileptic mice. Finally, we discuss possible causes and solutions for epileptic mice failing to make 30 s of good contact with the objects during the familiarization sessions, which is a critical step for successful memory tests. Thus, this protocol provides detailed information about how to assess epilepsy-associated memory impairments using mice. The NL, NO, and PS tests are simple, efficient assays that are appropriate for the evaluation of different kinds of memory in epileptic mice.

This article presents experimental procedures for assessing memory impairments in pilocarpine-induced epileptic mice. This protocol can be used to study the pathophysiologic mechanisms of epilepsy-associated cognitive decline, which is one of the most common comorbidities in epilepsy.

皮洛卡平诱导癫痫小鼠记忆功能评估

Cognitive impairment is one of the most common comorbidities in temporal lobe epilepsy. To recapitulate epilepsy-associated cognitive decline in an animal ...

Using a Virtual Reality Walking Simulator to Investigate Pedestrian Behavior

To cross a road successfully, individuals must coordinate their movements with moving vehicles. This paper describes use of a walking simulator in which people walk on a treadmill to intercept gaps between two moving vehicles in an immersive virtual environment. Virtual reality allows for a safe and ecologically varied investigation of gap crossing behavior. Manipulating the initial starting distance can further the understanding of a participant&#8217;s speed regulation while approaching a gap. The speed profile may be assessed for various gap crossing variables, such as initial distance, vehicle size, and gap size. Each walking simulation results in a position/time series that can inform how velocity is adjusted differently depending on the gap characteristics. This methodology can be used by researchers investigating pedestrian behavior and behavioral dynamics while employing human participants in a safe and realistic setting.

This protocol describes use of a walking simulator that serves as a safe and ecologically valid method to study pedestrian behavior in the presence of moving traffic.

使用虚拟现实步行模拟器调查行人行为

To cross a road successfully, individuals must coordinate their movements with moving vehicles. This paper describes use of a walking simulator in which ...