Name: measuring spinal presynaptic inhibition in mice by dorsal root potential recording in vivo
Uploaded: 2014-03-29T14:00:00
Description: Watch this Scientific Journal Video about Measuring Spinal Presynaptic Inhibition in Mice By Dorsal Root Potential Recording In Vivo at JoVE.com

我们感谢曼弗雷德HECKMANN的有益讨论建立方法的过程中。此外，我们感谢克劳迪娅·索默的技术援助和弗兰克·舒伯特支持制作视频。 01EO1002和耶拿大学医院的多学科临床研究中心（IZKF）：这项工作是由德国联邦教育与研究部（BMBF），德国，FKZ支持。

在下面的协议中提到的所有实验程序批准了图林根州当局（图林根森Landesamt献给Verbraucherschutz，Reg.-Nr. 02-044/12）。 1。准备实验<ol><li>吸电极的制作<ol><li>使用标准的高硼硅玻璃毛细管用一个微量拉马， 比如一个标准的贴片电极拔微量。 </li><li>制动用金刚石文件到尖端的0.5-1毫米（比背根的直径稍大）直径的电极。 </li><li>热打磨尖，所以它不会损害后根当它被吸入。?...

<ol>
	<li>Eccles, J. C., Eccles, R. M., Magni, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Central+inhibitory+action+attributable+to+presynaptic+depolarization+produced+by+muscle+afferent+volleys.">Central inhibitory action attributable to presynaptic depolarization produced by muscle afferent volleys.</a> J. Physiol. 159, 147-166 (1961).</li><li>Levy, R. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+role+of+gaba+in+primary+afferent+depolarization.">The role of gaba in primary afferent depolarization.</a> Prog. Neurobiol. 9, 211-267 (1977).</li><li>Eccles, J. C., Magni, F., Willis, W. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Depolarization+of+central+terminals+of+Group+I+afferent+fibres+from+muscle.">Depolarization of central terminals of Group I afferent fibres from muscle.</a> J. Physiol. 160, 62-93 (1962).</li><li>Eccles, J. C., Schmidt, R., Willis, W. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pharmacological+Studies+on+Presynaptic+Inhibition.">Pharmacological Studies on Presynaptic Inhibition.</a> J. Physiol. 168, 500-530 (1963).</li><li>Maxwell, D. J., Bannatyne, B. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ultrastructure+of+muscle+spindle+afferent+terminations+in+lamina+VI+of+the+cat+spinal+cord.">Ultrastructure of muscle spindle afferent terminations in lamina VI of the cat spinal cord.</a> Brain Res. 288, 297-301 (1983).</li><li>Barber, R. P., Vaughn, J. E., Saito, K., McLaughlin, B. J., Roberts, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=GABAergic+terminals+are+presynaptic+to+primary+afferent+terminals+in+the+substantia+gelatinosa+of+the+rat+spinal+cord.">GABAergic terminals are presynaptic to primary afferent terminals in the substantia gelatinosa of the rat spinal cord.</a> Brain Res. 141, 35-55 (1978).</li><li>Wall, P. D., Lidierth, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Five+sources+of+a+dorsal+root+potential:+their+interactions+and+origins+in+the+superficial+dorsal+horn.">Five sources of a dorsal root potential: their interactions and origins in the superficial dorsal horn.</a> J. Neurophysiol. 78, 860-871 (1997).</li><li>Rudomin, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+search+of+lost+presynaptic+inhibition.">In search of lost presynaptic inhibition.</a> Exp. Brain Res. 196, 139-151 (2009).</li><li>Rudomin, P., Schmidt, R. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Presynaptic+inhibition+in+the+vertebrate+spinal+cord+revisited.">Presynaptic inhibition in the vertebrate spinal cord revisited.</a> Exp. Brain Res. 129, 1-37 (1999).</li><li>Kullmann, D. 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=Presynaptic,+extrasynaptic+and+axonal+GABAA+receptors+in+the+CNS:+where+and+why?.">Presynaptic, extrasynaptic and axonal GABAA receptors in the CNS: where and why?.</a> Prog. Biophys. Mol. Biol. 87, 33-46 (2005).</li><li>Hochman, S., Shreckengost, J., Kimura, H., Quevedo, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Presynaptic+inhibition+of+primary+afferents+by+depolarization:+observations+supporting+nontraditional+mechanisms.">Presynaptic inhibition of primary afferents by depolarization: observations supporting nontraditional mechanisms.</a> Ann. N.Y. Acad. Sci. 1198, 140-152 (2010).</li><li>Thompson, S. W., Wall, P. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+effect+of+GABA+and+5-HT+receptor+antagonists+on+rat+dorsal+root+potentials.">The effect of GABA and 5-HT receptor antagonists on rat dorsal root potentials.</a> Neurosci. Lett. 217, 153-156 (1996).</li><li>Enriquez-Denton, M., Manjarrez, E., Rudomin, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Persistence+of+PAD+and+presynaptic+inhibition+of+muscle+spindle+afferents+after+peripheral+nerve+crush.">Persistence of PAD and presynaptic inhibition of muscle spindle afferents after peripheral nerve crush.</a> Brain Res. 1027, 179-187 (2004).</li><li>Wall, P. D., Devor, 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+effect+of+peripheral+nerve+injury+on+dorsal+root+potentials+and+on+transmission+of+afferent+signals+into+the+spinal+cord.">The effect of peripheral nerve injury on dorsal root potentials and on transmission of afferent signals into the spinal cord.</a> Brain Res. 209, 95-111 (1981).</li><li>Witschi, R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Presynaptic+&#945;2-GABAA+Receptors+in+Primary+Afferent+Depolarization+and+Spinal+Pain+Control.">Presynaptic &#945;2-GABAA Receptors in Primary Afferent Depolarization and Spinal Pain Control.</a> J. Neurosci. 31, 8134-8142 (2011).</li><li>Calancie, 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=Evidence+that+alterations+in+presynaptic+inhibition+contribute+to+segmental+hypo-+and+hyperexcitability+after+spinal+cord+injury+in.">Evidence that alterations in presynaptic inhibition contribute to segmental hypo- and hyperexcitability after spinal cord injury in.</a> 89, 177-186 (1993).</li><li>Geis, C., 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=Stiff+person+syndrome-associated+autoantibodies+to+amphiphysin+mediate+reduced+GABAergic+inhibition.">Stiff person syndrome-associated autoantibodies to amphiphysin mediate reduced GABAergic inhibition.</a> Brain. 133, 3166-3180 (2010).</li><li>Geis, C., 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=Human+IgG+directed+against+amphiphysin+induces+anxiety+behavior+in+a+rat+model+after+intrathecal+passive+transfer.">Human IgG directed against amphiphysin induces anxiety behavior in a rat model after intrathecal passive transfer.</a> J. Neural Transm. 119 (8), 981-985 (2012).</li><li>Barron, D. H., Matthews, B. 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+interpretation+of+potential+changes+in+the+spinal+cord.">The interpretation of potential changes in the spinal cord.</a> J. Physiol. 92, 276-321 (1938).</li><li>Schmidt, R. F., Trautwein, W., Zimmermann, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dorsal+root+potentials+evoked+by+natural+stimulation+of+cutaneous+afferents.">Dorsal root potentials evoked by natural stimulation of cutaneous afferents.</a> Nature. 212, 522-523 (1966).</li><li>Eccles, J. C., Schmidt, R. F., Willis, W. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Presynaptic+inhibition+of+the+spinal+monosynaptic+reflex+pathway.">Presynaptic inhibition of the spinal monosynaptic reflex pathway.</a> J. Physiol. 161, 282-297 (1962).</li><li>Manjarrez, E., Rojas-Piloni, J. G., Jimenez, I., Rudomin, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Modulation+of+synaptic+transmission+from+segmental+afferents+by+spontaneous+activity+of+dorsal+horn+spinal+neurones+in+the+cat.">Modulation of synaptic transmission from segmental afferents by spontaneous activity of dorsal horn spinal neurones in the cat.</a> J. Physiol. 529 Pt 2, 445-460 (2000).</li><li>Geis, C., 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=Human+Stiff-Person+Syndrome+IgG+Induces+Anxious+Behavior+in+Rats.">Human Stiff-Person Syndrome IgG Induces Anxious Behavior in Rats.</a> PLoS One. 6, e16775 (2011).</li><li>Martinez-Gomez, J., Lopez-Garcia, J. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Electrophysiological+and+pharmacological+characterisation+of+ascending+anterolateral+axons+in+the+in+vitro+mouse+spinal+cord.">Electrophysiological and pharmacological characterisation of ascending anterolateral axons in the in vitro mouse spinal cord.</a> J. Neurosci. Methods. 146, 84-90 (2005).</li></ol>

体内的神经元活动和突触电位外和细胞内电生理记录是在调查中枢神经系统神经元功能和病理生理学的艺术手法状态。脊柱融合是运动功能， 如肢体运动和多式联运感官知觉的关键。突触前抑制是在这个计算过程，确保感官输入适当的反应的一个关键机制。在IA传入纤维的GABA能突触抑制运动神经元通过PAD的激励。背根电位记录在体内揭开了完整的动物直接测量垫的可能性。?...

突触前抑制是在脊髓中的最有力的抑制机制之一。它抑制兴奋性突触后电位（EPSPS）的单突触兴奋运动神经元而不改变突触后膜电位和运动神经元1-3的兴奋性。通过GABA能轴突轴突突触上诱导突触前感觉纤维初级传入去极化（PAD）是底层机制4-7（参见Figure1a）。这些突触含有GABA A和 GABA B-受体（GABA A R和γ-氨基丁酸乙 R）。 GABA A R活动导致的增加，氯电导而引发的PAD由于当地离子分布。这种去极化块的动作电位传播到轴突终末并降低其强度，导致降低的Ca 2 +内流和减少递质释放。 γ-氨基丁酸乙受体的激活并没吨有助于垫，但导致减少的Ca 2 +内流，从而提高突触前抑制的。而GABA A R的激活似乎涉及短期的抑制作用，γ-氨基丁酸乙 R的参与长期调制8-10。除了​​γ-氨基丁酸，占PAD和突触前抑制的主要部分，其它发射机系统也可能调节和促进这一机制11,12。 在突触前抑制病理变化似乎是至关重要的几种疾病状态，例如外周炎症和神经性疼痛13,14，以及异常的中枢性疼痛的处理15，脊髓损伤16，和中枢神经系统疾病与兴奋马达由有缺陷的GABA能传输17介导的18。因此，推定的突触前抑制是值得研究的实验病理条件下在体内对脊髓水平</em&gt;。垫产生了进行音量电位提供了一个直接测量突触前抑制脊髓。这些潜能被称为背根电位（DRP），可以从脊髓背根相邻背根7的刺激后进行测量。 DRP的第一测量已经报道在猫和青蛙19，并深入研究了猫被埃克尔斯，施密特等人在20世纪70年代初3,4,20,21。而DRP猫22和大鼠23 在体内的记录已被广泛使用，在小鼠中测量已经几乎完全执行在体外分离的脊髓制剂15,24。在这里，我们描述了一种方法， 在体内 ，允许直接测量突触前抑制在完整有机体的记录DRP麻醉小鼠。

<table border="0" cellpadding="0" cellspacing="0">
	<tbody>
		<tr>
			<td>Glass tubing (inner diameter 1.16 mm)</td>
			<td>Science Products (Hofheim, Germany)</td>
			<td>GB200F-10</td>
			<td>Other glass tubing might also be suitable</td>
		</tr>
		<tr>
			<td>Superfusion solution (sterile, 0,9% NaCl)</td>
			<td>Braun Melsungen AG&#160;</td>
			<td>3570350</td>
			<td></td>
		</tr>
		<tr>
			<td>(Melsungen, Germany)</td>
		</tr>
		<tr>
			<td>Rompun 2% (Xylazine)</td>
			<td>Bayer Animal Health GmbH (Leverkusen, Germany)</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Ketamin 10%</td>
			<td>Medistar GmbH (Ascheberg, Germany)</td>
			<td>KETAMIN 10%</td>
			<td></td>
		</tr>
		<tr>
			<td>30G micro needle/ Sterican</td>
			<td>Braun Melsungen AG&#160;</td>
			<td>4656300</td>
			<td></td>
		</tr>
		<tr>
			<td>(Melsungen, Geramny)</td>
		</tr>
		<tr>
			<td>Salts for aCSF</td>
			<td>Sigma-Aldrich&#160;</td>
			<td>Diverse</td>
			<td></td>
		</tr>
		<tr>
			<td>S88 Dual Output Square Pulse</td>
			<td>Grass Technologies (Warwick, USA)</td>
			<td>S88X</td>
			<td></td>
		</tr>
		<tr>
			<td>Stimulator</td>
		</tr>
		<tr>
			<td>SIU5 RF Transformer Isolation Unit</td>
			<td>Grass Technologies (Warwick, USA)</td>
			<td>SIU-V</td>
			<td></td>
		</tr>
		<tr>
			<td>InstruTECH LIH 8+8</td>
			<td>HEKA (Lambrecht, Deutschland)</td>
			<td>LIH 8+8 + Patchmaster software</td>
			<td></td>
		</tr>
		<tr>
			<td>Data acquisition&#160;</td>
		</tr>
		<tr>
			<td>Universal amplifier</td>
			<td>npi (Tamm, Deutschland)</td>
			<td>ELC-03X</td>
			<td></td>
		</tr>
		<tr>
			<td>Micropipette puller</td>
			<td>Sutter Instruments (Novato, USA)</td>
			<td>P-1000</td>
			<td></td>
		</tr>
		<tr>
			<td>Dissecting microscope</td>
			<td>Olympus (Tokyo, Japan)</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Micromanipulator</td>
			<td>Sutter Instruments (Novato, USA)</td>
			<td>MPC-200/MPC-325</td>
			<td>Mechanical micromanipulators also possible</td>
		</tr>
		<tr>
			<td>Homeothermic Blanket System</td>
			<td>Stoelting (Wood Dale, USA)</td>
			<td>50300V</td>
			<td></td>
		</tr>
		<tr>
			<td>Intra-/extracellular recording electrode holder</td>
			<td>Harvard Apparatus (Holliston, USA)</td>
			<td>641227</td>
			<td></td>
		</tr>
	</tbody>
</table>

measuring spinal presynaptic inhibition in mice by dorsal root potential recording in vivo

突触前抑制是在脊髓中的最有力的抑制机制之一。底层的生理机制是通过GABA能轴突 - 轴突突触（初级传入去极化）介导的初级传入纤维的去极化。初级传入去极化强度可以通过录音的音量进行电位在背根（背根电位，DRP）进行测量。突触前抑制的病理改变是至关重要的某些疼痛疾病的中央异常处理和电机兴奋的有些失常。在这里，我们描述了记录的DRP的方法体内对小鼠。在麻醉动物，并用吸电极记录过程脊髓背根的编制进行了说明。这种方法允许测量GABA能DRP和由此估计在活小鼠脊髓突触前抑制。与转基因小鼠模型的组合，DRP记录可能本身相比于体外分离脊髓的准备工作， 如同时记录或操纵棘上网络和感应DRP由外周神经刺激的可能性RVE作为一种强大的工具，调查疾病相关的脊髓病理生理学。 在体内录音有几个优点。

典型的DRP迹线示于图3。突出刺激伪迹通常后跟一个短的向下偏转。此后缓慢，持久向上偏转，占本DRP是清晰可辨。在录音的一个子集，背根反射是因为在DRP的顶部小幅冲高可见。在正常野生型小鼠，背根反射最常出现时，刺激电压过高。作为背根反射，不能以高的重现性在此制剂引起，它们不用于分析和注意采取减少刺激电压至最低，但仍然超最大强度。为DRP的峰值振幅分析，平均2...

Watch this Scientific Journal Video about Measuring Spinal Presynaptic Inhibition in Mice By Dorsal Root Potential Recording In Vivo at JoVE.com

Measuring Spinal Presynaptic Inhibition in Mice By Dorsal Root Potential Recording In Vivo

测量脊髓突触前抑制小鼠背根电位记录<em&gt;在体内</em

GABA能突触前抑制是在脊髓电机和脊髓网络感官信号集成的重要功能强大的抑制机制。底层的初级传入去极化可以通过记录背根电位（DRP）来测量。在这里，我们证明在体内录音DRP小鼠的方法。

Presynaptic inhibition is one of the most powerful inhibitory mechanisms in the spinal cord. The underlying physiological mechanism is a depolarization of ...

Research

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Neuroscience

Measuring Spinal Presynaptic Inhibition in Mice By Dorsal Root Potential Recording In Vivo

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科研

神经科学

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哺乳动物轴突再生: 成年小鼠背根神经节体内电穿孔的研究

Electroporation is an essential non-viral gene transfection approach to introduce DNA plasmids or small RNA molecules into cells. A sensory neuron in the ...

Use of In Vivo Single-fiber Recording and Intact Dorsal Root Ganglion with Attached Sciatic Nerve to Examine the Mechanism of Conduction Failure

使用 Vivo 单纤维记录和带附骨神经的固定背道根结条，以检查传导失败机制

Single-fiber recording has been a classical and effective electrophysiological technique over the last few decades because of its specific application for ...

Rapid Isolation of Dorsal Root Ganglion Macrophages

多尾根结鱼巨噬细胞的快速隔离

There are growing interests to study the molecular and cellular interactions among immune cells and sensory neurons in the dorsal root ganglia after ...