作者希望感谢肯塔基大学微电极技术中心 (CenMeT) 和俄亥俄州立大学实验动物资源中心 (乌拉尔) 的贡献。

仔猪 (Sus 鲷) 是通过一个当地农场接收的, 由俄亥俄州立大学机构动物保育和使用委员会 (IACUC) 预先批准。经过该议定书的批准, 动物实验是按照 IACUC 政策进行的。
1. 仔猪和仔猪的搬运
<ol><li>以系统和随机的方式利用雄性和雌性仔猪, 以消除任何潜在的基于性别的混杂根据到达指南19。 注: 由于最大脑发育期在仔猪出生后3-5 天内, 实验仅以仔猪3-5 天的年龄进行。</li>
	<li>确保仔猪在试验前至少24小时到达 vivarium, 以便对环境进行驯化。 注: 训练有素的兽医人员提供日常动物护理。仔猪被保存在单独的温度维持, 不断监测的网箱, 并获得营养完整的牛奶乳粉ad 随意, 以防止低血糖。仔猪还保持不加奶 (零每 os), 至少3小时麻醉前, 并提供毛毯和玩具, 以确保正常水平的刺激。如果可能的话, 把多只小猪放在同一个笼子里, 允许社会化。</li>
</ol>2. 仔猪模型研究中的环境协定的开发和定制
注: 该技术采用酶促生物协定, 预涂酶和电镀的 m 苯二胺盐酸盐 (mPD)。电极是定制的设计与40毫米刚性轴和生产用于仔猪 (图 1)。
<ol><li>为避免冗余, 请参阅前面描述的15 (图 2) 的详细说明和校准。</li>
</ol>3. 用于仔猪的自定义立体定向装置的麻醉和使用
<ol><li>麻醉动物使用一个麻醉工作站与适当的呼吸机和监测设备, 并监测生理参数, 如脉搏氧, 无创血压, 心电图和温度贯穿整个以前描述的19的整个实验。对仔猪进行插管和通气, 并对七氟醚进行1最小肺泡浓度 (MAC) (约 2.5-3%) 的麻醉, 以达到3.5 小时的水平. 确保这些实验的训练有素的实验室工作人员在场。在准备皮肤之前, 用电动快刀将手术部位上的皮毛去掉。 注: 使用麻醉的浓度和持续时间允许实验模拟手术过程中实际麻醉暴露的时间长度。此外, 七氟醚是儿科人群中最常用的全麻, 使调查对其安全性具有极其重要的意义。</li>
	<li>在放置到立体定向的框架, 开始一个罗库溴铵负荷剂量2.5 毫克/千克和输液1.5 毫克/公斤/小时, 以防止动物运动, 而在安全的框架。<ol>
<li>一旦适当的麻醉深度被脚趾捏确认, 把小猪放在小猪特定的立体定位框架内。</li>
		<li>在立体定向的框架内提供足够的填料, 将小猪放在强制空气变暖垫上, 附加填料 (例如, 流化变位机) 以防止压力溃疡。</li>
		<li>将上颌骨的牙齿放在齿条上 (图 3)。 注: 牙棒应在足够的水平, 以保持头骨非常牢固到位。</li>
		<li>固定和收紧耳朵内的穿透耳棒, 注意确保小猪处于中线位置。将侧面的尖尖尖端放置在耳道内, 并牢固地插入耳棒, 以听到与鼓膜穿透有关的 "pop" 声音。将耳条牢固地附着在头骨上, 并在每一侧插入到相等的深度, 以防止头骨在实验期间的移动 (图 4, 面板 A)。 注意: 保持仔猪的体温 (37.8-38.6 °c) 并持续监测整个过程中的温度对常温的维护至关重要。这可以通过通过毯子和/或热灯完成。一定要在适当的距离放置散热灯, 以避免烧伤动物的皮肤。</li>
	</ol>
</li>
	<li>用小心的方法在头骨上创建一个4-6 厘米的中线切口, 以避免用手术刀把头骨打进。一旦切口, 使用温和的缩回和钝的解剖, 以提高头皮从头骨。用纱布垫轻轻擦洗头骨, 清除任何结缔组织并暴露缝合线 (图 4, 面板 B)。 注意: 没有必要在非生存实验中保持不孕。然而, 在生存实验中必须保持不孕。</li>
	<li>如果需要, 进一步反映头皮, 以暴露感兴趣的区域并确定开颅手术的预定位置 (图 5, 面板 A)。使用外科演练创建大约0.25 厘米2的开颅手术窗口 (根据实验目标可能会更大或更小) 覆盖感兴趣的结构, 使用谨慎不伤害硬脑膜或底层大脑 (图5 , 面板 B)。使用精细的外科剪刀来切除脑组织覆盖的硬脑膜 (图 5, 面板 C)。</li>
	<li>通过确保 headstage 的金属臂与小猪 stereotax 的机器人, 将电极尽可能垂直地放置在 bregma 上。尽量降低电极, 不接触头骨表面。记录 bregma 的坐标 (图 6, 面板 A)。<ol>
<li>确定前、后外侧坐标以及与 bregma 相关的兴趣结构的深度。使用一个物种和年龄合适的立体定向图谱确定立体定向坐标。在这种情况下, 使用专门为小猪20开发的立体定向图谱。</li>
		<li>重新定位电极, 使其具有适当的前后和内侧侧位置, 确保微电极和仪器都垂直于表面。将伪参考电极置于头皮下, 确保与动物接触 (图 6, 面板 B)。</li>
		<li>慢慢地, 轻轻地, 用立体定向的手臂将电极降低到大脑的适当深度。在最后2毫米, 使用液压 microdrive 进一步驱动电极到确切的位置 (图 6, 面板 C)。 注: 电极位置应位于开颅窗。电极深度会因感兴趣的大脑结构而异。在非生存实验数据收集完成后, 没有必要关闭切口。</li>
	</ol>
</li>
</ol>4. 七氟醚麻醉下胞外谷氨酸的测定
<ol><li>确保仔猪在整个过程中持续进行生理监测。仔猪麻醉 inhalationally (通过面锥) 为程序准备。</li>
	<li>在该多边环境协定的植入后, 等待30分钟, 允许电极达到基线, 以确保为 3 h (图 7) 获得正确的测量结果。</li>
	<li>0.25% 布比卡因 (1 毫升/千克) 在术后疼痛管理的手术现场被管理。此外, 持续释放丁丙诺啡是根据需要皮下 q72h。</li>
</ol>5. 灌注和牺牲
<ol><li>执行以前描述的21的灌注和脑组织收集过程。对于非生存手术, 动物在试验后立即被安乐死, 但仍在全身麻醉下。</li>
	<li>采取固定的小猪大脑的大体横断面, 并使用光显微镜可视化电极的轨道, 如前所述的22 , 以允许验证多边环境协定的放置, 确保适当的位置在感兴趣的领域。</li>
</ol>

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D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Extracellular+hippocampal+glutamate+and+spontaneous+seizure+in+the+conscious+human+brain.">Extracellular hippocampal glutamate and spontaneous seizure in the conscious human brain.</a> Lancet. 341 (8861), 1607-1610 (1993).</li><li>Reinstrup, 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=Intracerebral+microdialysis+in+clinical+practice:+baseline+values+for+chemical+markers+during+wakefulness,+anesthesia,+and+neurosurgery.">Intracerebral microdialysis in clinical practice: baseline values for chemical markers during wakefulness, anesthesia, and neurosurgery.</a> Neurosurgery. 47 (3), 701-710 (2000).</li><li>Cavus, I., 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=Extracellular+metabolites+in+the+cortex+and+hippocampus+of+epileptic+patients.">Extracellular metabolites in the cortex and hippocampus of epileptic patients.</a> Ann Neurol. 57 (2), 226-235 (2005).</li></ol>

格雷格葛哈德是 Quanteon 有限责任公司的主要所有者。乔治. 金特罗·库维德斯和杰森 Burmeister 曾担任 Quanteon 有限责任公司的顾问。

从实验开始, 小猪的生理动态平衡必须按照本实验室先前发布的21中的描述进行维护。最小的监测应包括脉冲氧, 心电图, capnography, 无创血压和温度。需要训练有素的调查人员, 以便能够适当纠正生理扰动 (例如, 如、低体温/热疗、缺氧、低血压、心律失常)。
在归纳之前,体外在已知条件下进行校准, 以建立多边环境协定的功能和选择性。多边环境协定的校准和电镀是有效利用该技术的关键。在校准过程中可能会出现许多可能的错误。校准可以识别这些问题以及不正确的电镀, 导致错误的干扰响应。编写了一个更详细的表格, 说明了在多边环境协定响应中可能发生的错误, 以及值得注意的原因和建议的解决方案, 这些都应该成为可能的疑难解答的有用工具 (表 1)。必须注意的是, 在校准和电镀之前, 应检查玻璃参考电极是否存在气泡或白色变色, 因为两者都将消极地影响多边环境协定的功能和记录的准确性。
<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always"><tbody>
<tr>
<td>症状</td>
			<td>原因</td>
			<td>纠正措施</td>
		</tr>
<tr>
<td rowspan="2">无信号</td>
			<td>电极未连接</td>
			<td>正确地将电极连接到 headstage 和 headstage, 以快速探头系统。</td>
		</tr>
<tr>
<td>对快速探头系统没有动力</td>
			<td>打开快速系统背面的电源开关</td>
		</tr>
<tr>
<td rowspan="5">信号噪声</td>
			<td rowspan="2">被血液污染的电极</td>
			<td>在电极插入过程中持续冲洗脑表面</td>
		</tr>
<tr>
<td>立即在 dH2O 中漂洗电极</td>
		</tr>
<tr>
<td>酶膜松动</td>
			<td>清洁和 recoat 电极</td>
		</tr>
<tr>
<td>未插入或涂覆参考电极</td>
			<td>外套和放置参考电极更远的头皮下</td>
		</tr>
<tr>
<td>电极检测脑表面运动</td>
			<td>通常发生在浅层结构中。如果可能, 将电极深度插入 (一次1毫米)</td>
		</tr>
<tr>
<td rowspan="2">动物运动</td>
			<td>动物没有得到充分的保护</td>
			<td>移动后方向的动物, 以更好地安全 earbars 的头骨。如有必要, 提升躯干, 使身体更好地对准。</td>
		</tr>
<tr>
<td>动物麻醉不充分</td>
			<td>验证麻醉设备的完整性。滴定麻醉的有效剂量和管理肌内的罗库溴铵剂量 (5 毫克/千克)</td>
		</tr>
<tr>
<td rowspan="3">电极放置不准确</td>
			<td>电极未正确对齐</td>
			<td>调整电极, 同时保持与 headstage 的正确连接。</td>
		</tr>
<tr>
<td rowspan="2">立体定向坐标不准确</td>
			<td>确保所引用的小猪图谱不使用另一个参照点或对齐平面。</td>
		</tr>
<tr>
<td>小心不要通过评分头骨来掩盖缝合痕迹。</td>
		</tr>
</tbody></table>表 1: 在仔猪中使用的疑难解答说明.可能的原因和纠正措施, 以协助优化和故障排除。
一个用于小猪的立体定向图谱用于确定感兴趣区域的立体定位坐标, 例如 bregma18。应妥善保护耳棒, 以确保头骨水平和完全固定。应注意在头皮中线切口, 以避免评分的头骨, 因为这可能会影响可视化缝合线。开颅窗应大到足以容纳多边环境协定。
该议定书提出了一些技术挑战, 需要有一个库存充足的操作套件和一个熟练掌握该议定书的外科和麻醉方面的调查员/小组。该模型还提出了财政限制, 因为小猪模型比啮齿动物模型更昂贵;然而, 它的成本大大低于使用非人类灵长类动物, 这可以花费上千美元。使用多边环境协定技术提出了自己的挑战, 因为手工涂敷和电镀电极的过程需要熟练的调查员或助理, 以确保足够的选择性和可靠的功能。电极本身是脆弱的, 因为它们是由陶瓷制成的, 因此如果没有观察到适当的谨慎, 就容易损坏。电极受到来自其他电气设备的干扰, 这些装置可以在录音中制造噪音, 并可在操作现场的血液中产生噪声, 咬合记录地点。需要专用设备带来额外的负担, 因为立体定向手术框架必须是定制的, 以固定的小猪头骨在植入过程中。立体定向的框架, 谷氨酸氧化酶, 和电极本身都是昂贵的。此外, 在过去十年中缺乏一只小猪立体定位图谱, 造成技术上的限制, 需要特定的专门知识来确定小猪大脑深层结构的具体位置。开发一个新的立体定向图谱, 也许使用磁共振成像, 将大大提高在仔猪中使用这种技术的能力。
仔猪是一种临床上相关的模型, 主要是由于这一物种和人类新生儿之间存在的相似性, 因为两者都具有相似的大脑结构和发展。与更常用的模型, 如老鼠或老鼠, 小猪有一个更大的中枢神经系统的相似性与人类, 这借给了模型的结果可译性。小猪模型是另外便宜和涉及较少复杂处理比非人类灵长类动物模型。小猪模型旨在研究麻醉可能诱发发育性神经毒性的过程, 测量其对神经损伤的贡献, 并与混淆变量引起的损害问题作斗争。例如, 缺氧可能被误解为麻醉剂造成的损害, 因为它对大脑有全球性的影响。仔猪的使用与人类医学中使用的外科和麻醉条件相同, 以确保结果的逼真度。
用陶瓷为基础的多边环境技术消除了与现代微透析技术相关的一些缺点。微透析与诸如多边环境协定等安培方法相比, 具有有限的时间和空间分辨率, 可以在多达10赫兹23的多个微观区域连续记录谷氨酸事件。这种快速采样率消除了局部神经递质扩散的混杂因子, 这是慢采样方法 (如微透析24) 所固有的。此外, 多边环境协定是一种比微透析探针更少的侵入性方法, 在插入的过程中可能导致显著的胶质, 并且可以改变插入点22的神经递质活动。
以前的研究使用了一系列哺乳动物模型, 测量技术和大脑区域, 表明了基本的谷氨酸水平, 可与使用这种技术发现。这表明, 在适应仔猪模型时, 多边环境协定技术提供了对体内谷氨酸浓度 (表 2) 的有效记录。
<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always"><tbody>
<tr>
<td>作者 (年)</td>
			<td>录音技术</td>
			<td>动物模型</td>
			<td>年龄</td>
			<td>脑区 (s)</td>
			<td>平均基底谷氨酸浓度 (µM)</td>
		</tr>
<tr>
<td>Hascup 等 (2008)23
</td>
			<td>(以酶为基础)</td>
			<td>啮齿动物</td>
			<td>20-24 周</td>
			<td>前额叶皮质, 纹状体</td>
			<td>3.3 ~ 1.0;5.0 @ 1。2</td>
		</tr>
<tr>
<td>Hascup 等 (2010)25
</td>
			<td>(以酶为基础)</td>
			<td>啮齿动物</td>
			<td>3-6 月</td>
			<td>海马</td>
			<td>4.7-10。4</td>
		</tr>
<tr>
<td>卢瑟福等 (2007)9
</td>
			<td>(以酶为基础)</td>
			<td>啮齿动物</td>
			<td>3-6 月</td>
			<td>前额叶皮质, 纹状体</td>
			<td>44.9 ~ 4.7;7.3 @ 0。9</td>
		</tr>
<tr>
<td>Miele 等 (1996)26
</td>
			<td>微透析 (酶基)</td>
			<td>啮齿动物</td>
			<td>-</td>
			<td>纹 状 体</td>
			<td>3.6 @ 0。5</td>
		</tr>
<tr>
<td>日 et 等 (2006)27
</td>
			<td>(以酶为基础)</td>
			<td>啮齿动物</td>
			<td>3-6 月</td>
			<td>额叶皮层, 纹状体</td>
			<td>1.6 @ 0.3; 1.4 @ 0。2</td>
		</tr>
<tr>
<td>金特罗·库维德斯等 (2007)28
</td>
			<td>(以酶为基础)</td>
			<td>非人类灵长类动物</td>
			<td>5.3-5.5 年</td>
			<td>前运动神经皮质, 运动皮层</td>
			<td>3.8 ~ 1.7;3.7 @ 0。9</td>
		</tr>
<tr>
<td>斯蒂芬斯等。 (2010)29
</td>
			<td>协定 [Spencer-Gerhardt-2 (SG-2)]</td>
			<td>非人类灵长类动物</td>
			<td>11-21 年</td>
			<td>壳</td>
			<td>8.53</td>
		</tr>
<tr>
<td>Kodama 等 (2002)30
</td>
			<td>微透析 (酶基)</td>
			<td>非人类灵长类动物</td>
			<td>-</td>
			<td>前额叶皮质</td>
			<td>1.29-2.21</td>
		</tr>
<tr>
<td>盖文等 (2003)31
</td>
			<td>微透析 (酶基)</td>
			<td>非人类灵长类动物</td>
			<td>青少年</td>
			<td>纹 状 体</td>
			<td>28.74 @ 2.73</td>
		</tr>
<tr>
<td>期间和斯宾塞 (1993)32
</td>
			<td>微透析 (酶基)</td>
			<td>人类</td>
			<td>18-35 年</td>
			<td>海马</td>
			<td>20.3 @ 6。6</td>
		</tr>
<tr>
<td>Reinstrup 等 (2000)33
</td>
			<td>微透析 (酶基)</td>
			<td>人类</td>
			<td>-</td>
			<td>额叶皮质</td>
			<td>16 @ 16</td>
		</tr>
<tr>
<td>Cavus 等 (2005)34
</td>
			<td>微透析 (酶基)</td>
			<td>人类</td>
			<td>15-52 年</td>
			<td>大脑皮层</td>
			<td>2.6 @ 0。3</td>
		</tr>
</tbody></table>表2。不同动物模型基底胞外谷氨酸 levelsacross 的比较。对在健康清醒和麻醉动物中使用微透析或电极建立正常胞外谷氨酸水平的研究进行的选择性审查。
使用多边环境协定技术来监测仔猪模型中的体内谷氨酸浓度, 可以使今后对仔猪神经功能的评估成为麻醉后的结果。已经计划了生存实验, 这将进一步了解麻醉对人类新生儿认知幸福的长期影响。生存实验将允许在麻醉暴露后长时间进行行为测试和谷氨酸变化监测。儿童在有可能以手术干预的形式经历生理压力的情况下接受麻醉也很常见。未来的研究, 以解决的影响, 手术的神经损伤和增加神经毒性将允许更准确的模拟一个共同的临床设置为儿童。使用替代动物模型也是可行的, 这也是通过慢性植入对这些模型进行研究, 使我们能够追踪与神经毒性相关的行为变化。技术本身是多才多艺的, 所以未来的研究不需要局限于谷氨酸水平的分析 (例如, GABA, 胆碱, 赖氨酸,等).可以分析)。

每年, 数以百万计的儿童在美国的1个侵入性和非侵入性程序中都接受麻醉。多年来, 麻醉提供者已经向父母们保证了麻醉药的安全性, 即使是在小孩和新生儿中也是如此。然而, 在1999年发现, 在早期的生命中, n-甲基-d-天门冬氨酸受体亚型的瞬态阻断可能会导致大鼠的神经细胞凋亡2。最近, FDA 发布了一项药物安全通讯, 要求麻醉药品的标签包括对一般麻醉药的警告, 以及它们对3岁以下儿童发育大脑的潜在负面影响 (美国食品和药物管理局, 2017)。然而, 仍然需要阐明可能的机制和潜在的保护措施。
谷氨酸和伽玛氨基丁酸 (GABA) 等神经递质的正常活动对正常神经发育的发生至关重要。虽然大部分涉及的路径仍然是难以捉摸的, 神经递质系统很可能会参与, 因为麻醉剂是已知的调节这些途径, 以产生无意识。特别是, 兴奋性神经递质谷氨酸导致兴奋时, 其活动失调。这种神经递质通常涉及神经发生、神经系统可塑性、突触和神经生长, 以及其他一些重要的脑功能。然而, 长期活化谷氨酸受体可能导致兴奋和神经细胞凋亡, 特别是在压力条件下, 如手术, 缺氧, 减少能源可用性3。谷氨酸与 NMDA 受体的结合已被证明导致 Na+和 Cl−流入。随后的退极化被认为导致 ca2 +通道打开和释放胞内 Ca2 +存储4。这种功能障碍可能导致一连串的代谢紊乱, 最终减少神经细胞的增殖, 增加炎症, 并导致神经元死亡。尽管有这些假设, 但实际机制仍不清楚5。谷氨酸失调由于其在细胞凋亡中的作用, 它代表了一种新的途径, 它可能有助于以前记录的神经元凋亡机制, 这是一种重要的特征。
神经过程研究的障碍之一是它们的高度复杂性, 特别是在神经元发育的设置中。最初几个月的生命是最容易受伤的时期, 在这期间, 神经发育的重要步骤, 如生理凋亡 (神经修剪), synaptogenesis, gliogenesis 和髓鞘形成发生6.鉴于神经元通信的复杂性以及在不破坏正常中枢神经功能的情况下研究这些过程的困难, 已经开发出了针对体内检测和量化重要元素的新技术。神经元通讯。
本研究采用酶联的多边环境技术, 作为研究仔猪模型中的作用机理的一种新方法。该技术可用于研究复杂的体内电化学脑过程, 包括谷氨酸失调。纳入多边环境协定的4通道白金记录站点 (2 个谷氨酸敏感的站点和2个定点站点) 允许自我引用, 这有助于检测的准确性。此外, 将排除层应用于每个电极, 通过防止其他干扰分子检测到7来赋予选择性。此外, 与早期技术相比, 该多边环境协定的低调设计使组织创伤最小。这一特性赋予了更高的空间分辨率, 有利于研究大脑中的微观区域。例如, 海马体 (齿状回、CA1、CA2) 的离散区域可以专门研究8。关于多边环境协定的功能的具体细节以前已被描述为9。
与电化学技术相比, 微透析结合了在兴趣溶液和类似成分溶液之间放置的膜, 允许检测胞外流体变化10。虽然微透析是神经系统的重要支柱, 长期以来一直用于神经递质的检测, 但它具有低时间分辨、谷氨酸变化延迟检测和重大组织损伤的缺点11。
多边环境协定可以间接检测神经递质, 如谷氨酸, 乙酰胆碱, 胆碱, 通过使用适当的氧化酶, 产生活性记者分子, 如 H2O2或 O212,13.
多边环境协定技术已被广泛应用于大鼠和非人类灵长类动物, 用于研究病理生理学过程中的神经毒性, 而不是7,14。在其中一些病理生理学过程中, 多边环境协定技术已被用于研究阿尔茨海默病、癫痫、外伤性脑损伤, 以及药理作用对突触通讯的影响8,15,16,17. 尽管多边环境协定已被用于研究大鼠和非人类灵长类动物的这些病态, 但人类与仔猪大脑的高度发展相似性使得对仔猪的气候变化技术的适应成为一种非常适宜的研究方法。(s) 机制18。

<table><tbody><tr><td>Advance Liqui-Wean Pig Milk Replacer</td><td>PBS Animal Health</td><td>292-13</td><td></td></tr><tr><td>Piglet Anesthesia Face-Cone Mask</td><td>VetEquip</td><td>921428</td><td></td></tr><tr><td>Integra SL Anesthesia Workstation</td><td>DRE Veterinary</td><td>2350</td><td>This anesthesia workstation is chosen to best mimic the clinical monitoring experienced by pediatric patients in the operating room. Any anesthesia machine can be used as long as it allows for sufficient physiologic monitoring and intervention.</td></tr><tr><td>Sevoflurane</td><td>Ultane</td><td>0074-4456-04</td><td></td></tr><tr><td>Rocuronium Bromide Injection</td><td>Hospira</td><td>0409-9558-05</td><td></td></tr><tr><td>Medfusion 4000 IV Infusion&amp;nbsp;</td><td>Smiths Medical</td><td></td><td></td></tr><tr><td>Model 1530 Heavy-Duty Research Model Stereotax</td><td>Kopf</td><td></td><td>custom made</td></tr><tr><td>Model 1541 Piglet Adaptor</td><td>Kopf</td><td></td><td>custom made</td></tr><tr><td>Infrared Spot Lamp</td><td>Amazon</td><td>B000HHQ94C</td><td></td></tr><tr><td>Bair Hugger Torso Blanket</td><td>3M</td><td>540</td><td></td></tr><tr><td>Bair Hugger</td><td>3M</td><td>750</td><td></td></tr><tr><td>Sterile Alcohol Prep Pad</td><td>Fisherbrand</td><td>22-363-750</td><td></td></tr><tr><td>Carbon Steel Rib-Back Surgical Blade</td><td>Bard-Parker</td><td>#10</td><td></td></tr><tr><td>Scalpel Handel</td><td>Havel&#039;s</td><td>HAN-G4</td><td></td></tr><tr><td>Surgical Scissors</td><td>World Precision Instruments</td><td>504615</td><td></td></tr><tr><td>Mosquito Forceps</td><td>Sklar Surgical Instruments</td><td>17-1225</td><td></td></tr><tr><td>Gauze Pads</td><td>Fisherbrand</td><td>22-246-069</td><td></td></tr><tr><td>Adson Tissue Forceps</td><td>Teleflex</td><td>181223</td><td></td></tr><tr><td>Dremel 111 Engraving Cutter</td><td>Amazon</td><td>Dremel 111</td><td></td></tr><tr><td>Microelectrode Array</td><td>Center for Microelectrdoe Technology, University of Kentucky</td><td></td><td>S2 4Ch MEA; custom made</td></tr><tr><td>Headstage</td><td>Quanteon</td><td></td><td>2pA/mV</td></tr><tr><td>Wire, silver, PFA, .008&quot; Bare, .0110&quot; coated</td><td>A-M Systems</td><td>786500</td><td></td></tr><tr><td>Fine Micromanipulator</td><td>Narishige Scientific Instrument Lab</td><td>MO-8</td><td></td></tr></tbody></table>

adaptation of microelectrode array technology for the study of anesthesia-induced neurotoxicity in the intact piglet brain

每年都有数以百万计的儿童接受麻醉以进行多种手术。然而, 在动物和人类的研究中, 对儿童麻醉的安全性提出质疑, 将麻醉药作为发展中大脑的潜在毒性。迄今为止, 没有研究成功地阐明了麻醉可能是神经毒性的机制。动物研究允许对这种机制进行调查, 而新生仔猪则是研究这些效应的优良模型, 因为它们与人脑的发育相似性显著。
该协议采用酶基微电极阵列技术作为研究麻醉诱发神经毒性机制的新方法。多边环境协定允许对体内神经递质活动进行实时监测, 并提供异常的时间和空间分辨率。假设麻醉神经毒性是由谷氨酸失调部分引起的, 而多边环境协定则提供一种测定谷氨酸的方法。该技术在仔猪模型中的新实现, 为研究这一问题提供了一个独特的机会。

这一概念验证的研究与酶为基础的陶瓷多边环境技术在小猪模型可以提供特殊的洞察力的谷氨酸动力学基础。这项研究进一步表明, 酶为基础的多边环境协定技术可以成功地适应于仔猪模型, 以测量生理和麻醉相关的变化, 神经递质活动, 高灵敏度, 高的空间和时间分辨率。在我们的实验中, 通过临床相关的方法和标准维持生理动态平衡, 没有小猪表现出生理扰动的迹象。
所获得的数据表明, 多边环境协定能够精确和空间地解决神经递质测量的皮质和亚皮层脑结构。使用立体定向设备可以明确识别参考表面结构 (bregma), 以便始终如一地定位感兴趣的区域, 而不管小猪的大小和解剖的个体差异。清晰的缝线可视化有助于在微米范围内一致地在区域内放置多边环境协定的准确性 (图 4)。获得对大脑皮质表面的接触是微小的创伤, 微小的出血, 确保任何体内谷氨酸动力学不是由于无意的系统性或局部侮辱 (图 5)。自定义, 刚性的多边环境, 然后与小猪的额平面垂直对齐 (图 6)。在插入之前不能正确地对齐多边环境协定, 可能会阻止目标区域的精确空间记录, 尤其是皮层下区域。
实时体内谷氨酸测量是在3-4 天的老仔猪 (n = 4) 下, 在 2.5-3% 七氟醚麻醉 (大约 1 MAC) 的海马中采取的。探头测量被记录在4赫兹和转换成浓度使用基于校准参数的线性回归 (图 8, 面板 a)。对于每一个时间点, 两个谷氨酸敏感点的信号平均之前, 减去平均哨兵信号, 以产生一个纠正谷氨酸信号。通过应用移动平均来更好地可视化总趋势 (图 8, 面板 B), 这些连续测量得到平滑。平均基底谷氨酸浓度为 4.61, 0.02 µM, 在麻醉暴露过程中保持相对稳定。通过分析信号中与哨兵信号不相关的峰值 (R2 < 0.5), 并超过了信噪比为 3 (图 9, 面板 a), 在一个动物中发现了瞬态谷氨酸活动。在 3.5 h (图 10, 面板 a) 期间检测到总共116个瞬态峰值。在1µM 范围内 (图 10, 面板 B) 中通常观察到产生的瞬态峰值的振幅。为了量化每个瞬态的持续时间, 获得了每个最大峰值值到衰变80% 所需的时间 (t80) (图 9, 面板 B)。3.5 小时记录期间所有谷氨酸瞬态的平均 t80为 4.68 @ 0.82 s (图 10, 面板 C)。这些数据表明, 有可能准确地测量长期和瞬态神经递质的活动, 在皮层下区域的麻醉仔猪大脑。
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/57391/57391fig1.jpg"> 图 1: SG-2 微电极阵列类型的可视比较.SG-2 阵列包含两个谷氨酸敏感的站点和两个谷氨酸不敏感的定点站点 (每个站点150µm x 20 µm)。(A) 左侧显示挠性轴微电极阵列。刚性轴微电极阵列是为仔猪使用而设计的, 允许在大型动物中进行更深的植入。<a href="https://cloudflare.jove.com/files/ftp_upload/57391/57391fig1large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/57391/57391fig2.jpg"> 图 2: 微电极阵列的制备和校准过程概述.总的多边环境协定的准备和校准持续约一周。涂层酶、排斥层和校准分析物是特定于感兴趣的神经递质的。<a href="https://cloudflare.jove.com/files/ftp_upload/57391/57391fig2large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<img alt="Figure 3" class="xfigimg" src="/files/ftp_upload/57391/57391fig3.jpg"> 图 3: 在立体定向设备中放置小猪.小猪的嘴巴直接放在嘴上, 在犬齿后面。穿透耳棒插入耳道, 以确保头骨的后端。<a href="https://cloudflare.jove.com/files/ftp_upload/57391/57391fig3large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<img alt="Figure 4" class="xfigimg" src="/files/ftp_upload/57391/57391fig4.jpg"> 图 4: 将小猪放置在用于开颅手术的立体定向装置中.(A) 小猪的头部在自定义立体定向框架内紧紧地固定在一起, 确保了多边环境协定的一致放置。穿透耳棒的等距位置是可见的。(B) 沿头皮中线前后切口。避免颅骨的评分, 以可视化冠状和矢状缝线和优化 bregma 的可视性。显示的刻度条表明切口的相对大小和开颅窗的位置。<a href="https://cloudflare.jove.com/files/ftp_upload/57391/57391fig4large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<img alt="Figure 5" class="xfigimg" src="/files/ftp_upload/57391/57391fig5.jpg"> 图 5: 用于访问海马体的开颅手术.(A) 头皮进一步反映出根据立体定向坐标来暴露多边环境协定插入的近似位置。带圆圈区域被标记 (黑点) 以指导开颅手术。(B) 将颅骨瓣移除的开颅窗 (0.25 厘米2), 以暴露底层硬脑膜。(C) 将脑膜仔细移除, 以暴露未组织外伤的表面皮层。<a href="https://cloudflare.jove.com/files/ftp_upload/57391/57391fig5large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<img alt="Figure 6" class="xfigimg" src="/files/ftp_upload/57391/57391fig6.jpg"> 图 6: 将定位和插入到海马中.(A) 在 bregma 上放置多边环境协定, 以确定海马体的相对立体定位位置。(B) 在脑表面立体定向放置多边环境协定, 以确定海马插入深度。银伪参考电极安全地放置在头皮下 (用箭头表示)。(C) 在适当深度插入的多边环境协定, 以获得实时的体内胞外谷氨酸测量海马。<a href="https://cloudflare.jove.com/files/ftp_upload/57391/57391fig6large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<img alt="Figure 7" class="xfigimg" src="/files/ftp_upload/57391/57391fig7.jpg"> 图 7: 在30分钟基线期间的协定行为.最初的快速增加对应于使用机器人的环境协定的下降到海马体。一旦多边环境协定达到适当的深度 (虚线), 基线周期就开始了。胞外谷氨酸的测量将减少30分钟的时间, 不应该被解释为生理学读数。<a href="https://cloudflare.jove.com/files/ftp_upload/57391/57391fig7large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<img alt="Figure 8" class="xfigimg" src="/files/ftp_upload/57391/57391fig8.jpg"> 图 8: 在七氟醚麻醉下的一只新生猪海马体内的实时胞外谷氨酸测量.(A) 在七氟醚麻醉下, 一只新生猪海马谷氨酸浓度的移动平均值 (使用10数据点)。测量是在4Hz 的3小时后, 一个简短的30分钟基线期。(B) 用移动平均每15分钟平滑谷氨酸测量, 以更好地可视化趋势。<a href="https://cloudflare.jove.com/files/ftp_upload/57391/57391fig8large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<img alt="Figure 9" class="xfigimg" src="/files/ftp_upload/57391/57391fig9.jpg"> 图 9: 七氟醚麻醉下新生儿猪海马中瞬态谷氨酸活性的鉴定.(A) 在实时谷氨酸跟踪中指示瞬时谷氨酸峰值 (以红色表示)。当信噪比超过 3, 其信号与哨兵信号 (R2 < 0.5) 不相关时, 峰值被认为是重要的。(B) 在图 9(面板 A) 中标识的具有代表性的瞬态峰值。虚线表示峰值衰减80% 所需的总工期。<a href="https://cloudflare.jove.com/files/ftp_upload/57391/57391fig9large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<img alt="Figure 10" class="xfigimg" src="/files/ftp_upload/57391/57391fig10.jpg"> 图 10: 七氟醚麻醉下新生猪海马谷氨酸瞬态的特征.(A) 在15分钟箱中有许多瞬时谷氨酸峰值。当信噪比超过 3, 其信号与哨兵信号 (R2 < 0.5) 不相关时, 峰值被认为是重要的。(B) 瞬时谷氨酸峰值的振幅。误差线表示平均值的标准错误。(C) 表示瞬时谷氨酸峰值的 T80 。误差线表示平均值的标准错误。<a href="https://cloudflare.jove.com/files/ftp_upload/57391/57391fig10large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>

Watch this Scientific Journal Video about Adaptation of Microelectrode Array Technology for the Study of Anesthesia-induced Neurotoxicity in the Intact Piglet Brain at JoVE.com

Adaptation of Microelectrode Array Technology for the Study of Anesthesia-induced Neurotoxicity in the Intact Piglet Brain

微电极阵列技术在完整仔猪脑麻醉诱导神经毒性研究中的应用

本研究探讨了用酶基微电极阵列技术监测仔猪体内的活体神经递质活性的新方法.假设谷氨酸失调有助于麻醉神经毒性的机制。在此, 我们提出了一种适应多边环境技术的协议, 研究麻醉诱发神经毒性的机制。

Every year, millions of children undergo anesthesia for a multitude of procedures. However, studies in both animals and humans have called into question ...

adaptation-microelectrode-array-technology-for-study-anesthesia

Research

JoVE Journal

Neuroscience

9.4K 次观看.  Ohio State University College of Medicine. 本研究探讨了用酶基微电极阵列技术监测仔猪体内的活体神经递质活性的新方法.假设谷氨酸失调有助于麻醉神经毒性的机制。在此, 我们提出了一种适应多边环境技术的协议, 研究麻醉诱发神经毒性的机制。

视频: Adaptation of Microelectrode Array Technology for the Study of Anesthesia-induced Neurotoxicity in the Intact Piglet Brain

A Piglet Model of Neonatal Hypoxic-Ischemic Encephalopathy

Birth asphyxia, which causes hypoxic-ischemic encephalopathy (HIE), accounts for 0.66 million deaths worldwide each year, about a quarter of the world&#8217;s 2.9 million neonatal deaths. Animal models of HIE have contributed to the understanding of the pathophysiology in HIE, and have highlighted the dynamic process that occur in brain injury due to perinatal asphyxia. Thus, animal studies have suggested a time-window for post-insult treatment strategies. Hypothermia has been tested as a treatment for HIE in pdiglet models and subsequently proven effective in clinical trials. Variations of the model have been applied in the study of adjunctive neuroprotective methods and piglet studies of xenon and melatonin have led to clinical phase I and II trials1,2. The piglet HIE model is further used for neonatal resuscitation- and hemodynamic studies as well as in investigations of cerebral hypoxia on a cellular level. However, it is a technically challenging model and variations in the protocol may result in either too mild or too severe brain injury. In this article, we demonstrate the technical procedures necessary for establishing a stable piglet model of neonatal HIE. First, the newborn piglet (&#60; 24 hr old, median weight 1500 g) is anesthetized, intubated, and monitored in a setup comparable to that found in a neonatal intensive care unit. Global hypoxia-ischemia is induced by lowering the inspiratory oxygen fraction to achieve global hypoxia, ischemia through hypotension and a flat trace amplitude integrated EEG (aEEG) indicative of cerebral hypoxia. Survival is promoted by adjusting oxygenation according to the aEEG response and blood pressure. Brain injury is quantified by histopathology and magnetic resonance imaging after 72 hr.

Hypoxic-ischemic encephalopathy following perinatal asphyxia can be studied using animal models. We demonstrate the procedures necessary for establishing a piglet model of neonatal hypoxic-ischemic encephalopathy.

新生儿缺氧缺血性脑病的小猪模型

Birth asphyxia, which causes hypoxic-ischemic encephalopathy (HIE), accounts for 0.66 million deaths worldwide each year, about a quarter of the ...

科研

医学

Extracellular Recording of Neuronal Activity Combined with Microiontophoretic Application of Neuroactive Substances in Awake Mice

Differences in the activity of neurotransmitters and neuromodulators, and consequently different neural responses, can be found between anesthetized and awake animals. Therefore, methods allowing the manipulation of synaptic systems in awake animals are required in order to determine the contribution of synaptic inputs to neuronal processing unaffected by anesthetics. Here, we present methodology for the construction of electrodes to simultaneously record extracellular neural activity and release multiple neuroactive substances at the vicinity of the recording sites in awake mice. By combining these procedures, we performed microiontophoretic injections of gabazine to selectively block GABAA receptors in neurons of the inferior colliculus of head-restrained mice. Gabazine successfully modified neural response properties such as the frequency response area and stimulus-specific adaptation. Thus, we demonstrate that our methods are suitable for recording single-unit activity and for dissecting the role of specific neurotransmitter receptors in auditory processing.
The main limitation of the described procedure is the relatively short recording time (~3 hr), which is determined by the level of habituation of the animal to the recording sessions. On the other hand, multiple recording sessions can be performed in the same animal. The advantage of this technique over other experimental procedures used to manipulate the level of neurotransmission or neuromodulation (such as systemic injections or the use of optogenetic models), is that the drug effect is confined to the local synaptic inputs to the target neuron. In addition, the custom-manufacture of electrodes allows adjustment of specific parameters according to the neural structure and type of neuron of interest (such as the tip resistance for improving the signal-to-noise ratio of the recordings).

We present methods for the construction of electrodes to simultaneously record extracellular neural activity and release multiple neuroactive substances at the vicinity of the recording sites in awake mice. This technique allows the detailed analysis of putative local synaptic inputs to the neuron of interest.

神经元电活动外记录与神经活性物质在清醒小鼠微电泳应用相结合

Differences in the activity of neurotransmitters and neuromodulators, and consequently different neural responses, can be found between anesthetized and ...

神经科学

Exposure of the Pig CNS for Histological Analysis: A Manual for Decapitation, Skull Opening, and Brain Removal

Pigs have become increasingly popular in large-animal translational neuroscience research as an economically and ethically feasible substitute to non-human primates. The large brain size of the pig allows the use of conventional clinical brain imagers and the direct use and testing of neurosurgical procedures and equipment from the human clinic. Further macroscopic and histological analysis, however, requires postmortem exposure of the pig central nervous system (CNS) and subsequent brain removal. This is not an easy task, as the pig CNS is encapsulated by a thick, bony skull and spinal column. The goal of this paper and instructional video is to describe how to expose and remove the postmortem pig brain and the pituitary gland in an intact state, suitable for subsequent macroscopic and histological analysis.

The goal of this paper and instructional video is to describe how to expose and remove the postmortem pig brain and pituitary gland in an intact state, suitable for subsequent macroscopic and histological analysis.

猪中枢神经系统的组织学分析的曝光:一个手册斩首，颅骨打开，并去除脑

Pigs have become increasingly popular in large-animal translational neuroscience research as an economically and ethically feasible substitute to ...

Use of a Piglet Model for the Study of Anesthetic-induced Developmental Neurotoxicity (AIDN): A Translational Neuroscience Approach

Anesthesia cannot be avoided in many cases when surgery is required, particularly in children. Recent investigations in animals have raised concerns that anesthesia exposure may lead to neuronal apoptosis, known as anesthesia-induced developmental neurotoxicity (AIDN). Furthermore, some clinical studies in children have suggested that anesthesia exposure may lead to neurodevelopmental deficits later in life. Nonetheless, an ideal animal model for preclinical study has yet to be developed. The neonatal piglet represents a valuable model for preclinical study, as they share a striking number of developmental similarities with humans.
The anatomy and physiology of piglets allow for implementation of rigorous human perioperative conditions in both survival and non-survival procedures. Femoral artery catheterization allows for close monitoring, thus enabling prompt correction of any deviation of the piglet's vital signs and chemistries. In addition, there are multiple developmental similarities between piglets and human neonates. The techniques required to use piglets for experimentation will require experience to master. A pediatric anesthesiologist is a critical member of the investigative team. We describe, in a general sense, the appropriate use of a piglet model for neurodevelopmental study.

Use of a Piglet Model for the Study of Anesthetic-induced Developmental Neurotoxicity AIDN: A Translational Neuroscience Approach

Anesthesia-induced developmental neurotoxicity (AIDN) research has focused on rodents, which are not broadly applicable to humans. Non-human primate models are more relevant, but are cost-prohibitive and difficult to use for experimentation. The piglet, in contrast, is a clinically relevant, practical animal model ideal for the study of anesthetic neurotoxicity.

使用仔猪模型研究麻醉诱发的发育神经毒性（AIDN）:一种转化神经科学方法

Anesthesia cannot be avoided in many cases when surgery is required, particularly in children.  Recent investigations in animals have raised concerns that ...

Noninvasive EEG Recordings from Freely Moving Piglets

The method allows the recording of high-quality electroencephalograms (EEGs) from freely moving piglets directly in the pigpen. We use a one-channel telemetric electroencephalography system in combination with standard self-adhesive hydrogel electrodes. The piglets are calmed down without the use of sedatives. After their release into the pigpen, the piglets behave normally&#8212;they drink and sleep in the same cycle as their siblings. Their sleep phases are used for the EEG recordings.

Here, we present a protocol to record telemetric electroencephalograms (EEGs) from freely moving piglets directly in the pigpen without the use of a sedative, making it possible to record typical EEG patterns during non-REM sleep, like spindle bursts.

自由移动仔猪的无创脑电图记录

The method allows the recording of high-quality electroencephalograms (EEGs) from freely moving piglets directly in the pigpen. We use a one-channel ...

Subdural Soft Electrocorticography (ECoG) Array Implantation and Long-Term Cortical Recording in Minipigs

Neurological impairments and diseases can be diagnosed or treated using electrocorticography (ECoG) arrays. In drug-resistant epilepsy, these help delineate the epileptic region to resect. In long-term applications such as brain-computer interfaces, these epicortical electrodes are used to record the movement intention of the brain, to control the robotic limbs of paralyzed patients. However, current stiff electrode grids do not answer the need for high-resolution brain recordings and long-term biointegration. Recently, conformable electrode arrays have been proposed to achieve long-term implant stability with high performance. However, preclinical studies for these new implant technologies are needed to validate their long-term functionality and safety profile for their translation to human patients. In this context, porcine models are routinely employed in developing medical devices due to their large organ sizes and easy animal handling. However, only a few brain applications are described in the literature, mostly due to surgery limitations and integration of the implant system on a living animal.
Here, we report the method for long-term implantation (6 months) and evaluation of soft ECoG arrays in the minipig model. The study first presents the implant system, consisting of a soft microfabricated electrode array integrated with a magnetic resonance imaging (MRI)-compatible polymeric transdermal port that houses instrumentation connectors for electrophysiology recordings. Then, the study describes the surgical procedure, from subdural implantation to animal recovery. We focus on the auditory cortex as an example target area where evoked potentials are induced by acoustic stimulation. We finally describe a data acquisition sequence that includes MRI of the whole brain, implant electrochemical characterization, intraoperative and freely moving electrophysiology, and immunohistochemistry staining of the extracted brains.
This model can be used to investigate the safety and function of novel design of cortical prostheses; mandatory preclinical study to envision translation to human patients.

Subdural Soft Electrocorticography ECoG Array Implantation and Long-Term Cortical Recording in Minipigs

Here, we present a method for the long-term performance and safety assessment of soft subdural electrode arrays in a minipig model, describing surgical method and tools, postoperative magnetic resonance imaging, electrophysiology of the auditory cortex, electrochemical properties of the implant, and postmortem immunochemistry.

硬膜下软皮质电图 （ECoG） 阵列植入和小型猪的长期皮质记录

Neurological impairments and diseases can be diagnosed or treated using electrocorticography (ECoG) arrays. In drug-resistant epilepsy, these help ...

生物工程

<meta charset="utf8"></head><body>小鼠深部脑干结构的倾斜硅多电极记录

Acute Single-Unit Multi-Electrode Recordings from the Brainstem of Head-Fixed Mice

Silicon multielectrode-based recordings are increasingly popular for studying neuronal activity at the temporal resolution of action potentials in many brain regions. However, recording neuronal activity from deep caudal structures like the brainstem using multi-channel probes remains challenging. A significant concern is finding a trajectory for probe insertion that avoids large blood vessels, such as the superior sagittal venous sinus and the transverse venous sinus. Injuring these large veins can cause extensive bleeding, damage to the underlying brain tissue, and potentially death. This approach describes targeting brainstem structures by coupling anterior coordinates with an angled approach, allowing the recording probe to penetrate the brain below high-risk vascular structures. Compared to a strictly vertical approach, the angled approach maximizes the number of brain regions that can be targeted. Using this strategy, the ventrolateral periaqueductal gray (vlPAG), a brainstem region associated with REM sleep, can be reproducibly and reliably accessed to obtain single-unit, multi-electrode recordings in head-fixed mice before and during sevoflurane anesthesia. The ability to record neuronal activity in the vlPAG and surrounding nuclei with high temporal resolution is a step forward in advancing the understanding of the relationship between REM sleep and anesthesia.

<meta charset="utf8"></head><body>作者聚焦：研究小鼠麻醉诱导的睡眠通路和神经元兴奋性

This protocol describes a method to obtain in vivo, high-density single-neuron recordings from the brainstem of head-fixed mice. This approach is deployed to measure the action potential firing of neurons in the ventrolateral periaqueductal gray - a brainstem region inactive during Rapid Eye Movement (REM) sleep - before and during general anesthesia.

来自头部固定小鼠脑干的急性单单位多电极记录

Silicon multielectrode-based recordings are increasingly popular for studying neuronal activity at the temporal resolution of action potentials in many ...

Measuring Brain Glutamate Levels Under Anesthesia in Neonatal Piglets Using a Microelectrode Array

Source: Geyer, E. D., et al. <a href="https://app.jove.com/v/57391">Adaptation of Microelectrode Array Technology for the Study of Anesthesia-induced Neurotoxicity in the Intact Piglet Brain.</a> J. Vis. Exp. (2018).
This video demonstrates the measurement of glutamate levels in the brain of an anesthetized neonatal piglet using a microelectrode array (MEA). The procedure involves creating a craniotomy window in the pig skull and inserting the MEA into the brain region of interest. The glutamate-oxidase-coated recording sites of the MEA convert extracellular brain glutamate to hydrogen peroxide, which generates a measurable current. This current is measured and normalized against background noise to determine brain glutamate levels under anesthesia.

使用微电极阵列测量新生仔猪麻醉下脑谷氨酸水平

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.
1. ...

微电极阵列技术在完整仔猪脑麻醉诱导神经毒性研究中的应用

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