Name: large volume, behaviorally-relevant illumination for optogenetics in non-human primates
Uploaded: 2017-10-03T15:00:00
Description: Watch this Scientific Journal Video about Large Volume, Behaviorally-relevant Illumination for Optogenetics in Non-human Primates at JoVE.com

LCA は、NDSEG ・ フェローシップ、NSF GRFP とマクガバン研究所の友人からの資金を認めています。EP は、ハリーとユーニス野原支給基金、1995 支給基金の MIT クラスと MIT 支給基金の資金を認めています。ESB は、NIH 2R44NS070453 03A1、IET ハーヴェイ賞とニューヨーク幹細胞財団・ ロバートソン賞からの資金を認めています。RD は、NIH の EY017292 からの資金提供を認めています。マイケル ・ ウィリアムズ整理、撮影前に消耗品を収集してチームに貢献しました。

注: 動物のすべてのプロシージャは NIH のガイドラインに従ってされ、マサチューセッツ研究所の技術委員会によって動物のケアに関する承認された。
 1 照明作製 <ol><li>250 μ m 径プラスチック光ファイバー 10 cm 以上必要な総照明の長さよりも長いであるのセクションをカットする鋭いはさみのペアを使用します。</li>。
	<li>22 ゲージのワイヤーストリッパーを使用して?...

<ol>
	<li>Herculano-Houzel, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+human+brain+in+numbers:+a+linearly+scaled-up+primate+brain.">The human brain in numbers: a linearly scaled-up primate brain.</a> Front Hum Neurosci. 3, 31 (2009).</li><li>Tamura, K., 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=A+glass-coated+tungsten+microelectrode+enclosing+optical+fibers+for+optogenetic+exploration+in+primate+deep+brain+structures.">A glass-coated tungsten microelectrode enclosing optical fibers for optogenetic exploration in primate deep brain structures.</a> J Neurosci Meth. 211 (1), 49-57 (2012).</li><li>Diester, 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=An+optogenetic+toolbox+designed+for+primates.">An optogenetic toolbox designed for primates.</a> Nat Neurosci. 14 (3), 387-397 (2011).</li><li>Dai, J., Brooks, D. I., Sheinberg, D. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Optogenetic+and+Electrical+Microstimulation+Systematically+Bias+Visuospatial+Choice+in+Primates.">Optogenetic and Electrical Microstimulation Systematically Bias Visuospatial Choice in Primates.</a> Curr biol. 24 (1), 63-69 (2014).</li><li>Ozden, 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=A+coaxial+optrode+as+multifunction+write-read+probe+for+optogenetic+studies+in+non-human+primates.">A coaxial optrode as multifunction write-read probe for optogenetic studies in non-human primates.</a> J Neurosci Meth. 219 (1), 142-154 (2013).</li><li>Ruiz, O., 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=Optogenetics+through+windows+on+the+brain+in+the+nonhuman+primate.">Optogenetics through windows on the brain in the nonhuman primate.</a> J Neurophysiol. 110 (6), 1455-1467 (2013).</li><li>Chuong, A. S., 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=Noninvasive+optical+inhibition+with+a+red-shifted+microbial+rhodopsin.">Noninvasive optical inhibition with a red-shifted microbial rhodopsin.</a> Nat Neurosci. 17 (8), 1123-1129 (2014).</li><li>Acker, L., Pino, E. N., Boyden, E. S., Desimone, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=FEF+inactivation+with+improved+optogenetic+methods.">FEF inactivation with improved optogenetic methods.</a> Proc Natl Acad Sci U S A. 113 (46), 7297-7306 (2016).</li><li>Jazayeri, M., Lindbloom-Brown, Z., Horwitz, G. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Saccadic+eye+movements+evoked+by+optogenetic+activation+of+primate+V1.">Saccadic eye movements evoked by optogenetic activation of primate V1.</a> Nat Neurosci. 15 (10), 1368-1370 (2012).</li><li>Gerits, A., 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=Optogenetically+Induced+Behavioral+and+Functional+Network+Changes+in+Primates.">Optogenetically Induced Behavioral and Functional Network Changes in Primates.</a> Curr Biol. 22 (18), 1722-1726 (2012).</li><li>Ohayon, S., Grimaldi, P., Schweers, N., Tsao, D. Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Saccade+modulation+by+optical+and+electrical+stimulation+in+the+macaque+frontal+eye+field.">Saccade modulation by optical and electrical stimulation in the macaque frontal eye field.</a> J Neurosci. 33 (42), 16684-16697 (2013).</li><li>Cavanaugh, J., 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=Optogenetic+inactivation+modifies+monkey+visuomotor+behavior.">Optogenetic inactivation modifies monkey visuomotor behavior.</a> Neuron. 76 (5), 901-907 (2012).</li></ol>

病気および齧歯動物の生理学を研究する光遺伝学的ツールが広く使用されますが、照明の大脳ボリュームの技術的な課題は非ひと霊長類におけるオプトジェネティクスの使用を制限されています。おそらく小さなボリュームを照らす大きな光パワー密度 (20 W/mm2~ 100 mW/mm2 ) を使用サルの研究の先駆的な &#60; 1 mm3、および皮質4、興奮のオプシンと報...

生理機能や齧歯動物および無脊椎動物の行動を研究するミリ秒精度、光駆動型神経制御を可能にする光遺伝学的ツールが活躍しています。ただし、技術的な課題には、いる齧歯動物の脳1を超える x 数量 100 ~ 非ひと霊長類脳研究の使用が限られています。
2 つの競合する目標に対処するため設計された蛍光灯器具非ひと霊長類における光遺伝学の研究を促進する: 大量の照明と最小限の貫通ダメージ。これらの懸念の 1 つのアドレスに以前の試みは、他の高価で来ています。バンドル ファイバー照らす大きなボリュームが増加の直径と、したがって、ダメージ2,3。テーパー ガラス繊維を減らす狭くフォーカス ライトを発光表面積しますが、侵入被害 &#60; 100 μ m2 4,5。外部脳硬膜にある窓の照明が侵入被害の課題を回避大量照明を許可する可能性があります、いくつかの表面的な脳エリア6のみ使用できます。
大ボリューム ・小径照明 (図 1 a) を作成するには、プラスチック光学繊維は熱のテーパーし、コアとクラッドの先端エッチング (図 1 b, c)。狭いポイントにライトを集中する他のテーパ繊維とは異なり、エッチングは均等に大面積 (図 1 d, e) で光を広く配布するためが、先端側から脱出して光をことができます。侵入被害は浸透径に比例して、この照明は従来繊維よりもないより多くの浸透ダメージをまだそれはので &#62; 1 とより広く表面エリアと提供光発光 x 100/100 光パワー脳ファントム (1.75% の agarose) の密度 (図 1e)。彼らは彼らの発光面として等しい光パワー密度を持っているとき、モンテカルロ モデル (図 1 階) は従来ファイバーと大量の照明器具の光の広がりで違いを示しています。各照明を個別に (図 2 ab) ヒント (図 2 c) も光分布を確保するための積分球を使用して校正します。
この大量の照明器具は、動作と非ひと霊長類におけるニューロンの両方の光遺伝学的操作と検証されています。脳の任意の領域に、それぞれの動物の個々 の受容野マップに、繊維の先端の長さをカスタマイズできます。照明は、照明の長さにわたる神経録音用貫通電極と組み合わせられるかもしれない。さらに、繊維は、可視光の任意の色を運ぶことができます、ために、利用できる利用可能な光分子のいずれかと組み合わせることが。

<table border="0" cellpadding="0" cellspacing="0" width="853">
	<tbody>
		<tr height="63">
			<td height="63" style="height:63px;width:403px;">Plastic optical fiber</td>
			<td style="width:323px;">Industrial fiber optics</td>
			<td style="width:64px;">SK-10</td>
			<td style="width:64px;">250 micron diameter, Super Eska line</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:403px;">Wire stripper</td>
			<td style="width:323px;">Klein Tools</td>
			<td align="right" style="width:64px;">11047</td>
			<td>22 gauge</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:403px;">Vise Clamp</td>
			<td style="width:323px;">Wilton</td>
			<td align="right" style="width:64px;">11104</td>
			<td>Generic table mount vice clamp</td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:403px;">Dual temperature heat gun</td>
			<td style="width:323px;">Milwaukee</td>
			<td style="width:64px;">8975-6</td>
			<td>570 / 1000&#176;F</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:403px;">Lab marker</td>
			<td style="width:323px;">VWR</td>
			<td align="right" style="width:64px;">52877</td>
			<td></td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:403px;">Dissection microscope</td>
			<td style="width:323px;">VistaVision</td>
			<td style="width:64px;">82027-156</td>
			<td>Stereo microscope w/ dual incandescent light, 2x/4x magnification, available from VWR</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:403px;">Lab tape</td>
			<td style="width:323px;">VWR</td>
			<td style="width:64px;">89097-972</td>
			<td>4 pack of violet color; however, tape color does not matter</td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:403px;">Silicon carbide lapping sheet</td>
			<td style="width:323px;">ThorLabs</td>
			<td style="width:64px;">LF5P</td>
			<td>5 micron grit, 10 pack</td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:403px;">Aluminum oxide lapping sheet</td>
			<td style="width:323px;">ThorLabs</td>
			<td style="width:64px;">LF3P</td>
			<td>3 micron grit, 10 pack</td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:403px;">Aluminum oxide lapping&#160; sheet</td>
			<td style="width:323px;">ThorLabs</td>
			<td style="width:64px;">LF1P</td>
			<td>1 micron grit, 10 pack</td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:403px;">Calcined alumina lapping sheet</td>
			<td style="width:323px;">ThorLabs</td>
			<td style="width:64px;">LF03P</td>
			<td>0.3 micron grit, 10 pack</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:403px;">Hot knife</td>
			<td style="width:323px;">Industrial fiber optics</td>
			<td style="width:64px;">IF370012</td>
			<td>60 Watt, heavy duty</td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:403px;">Fiber inspection scope</td>
			<td style="width:323px;">ThorLabs</td>
			<td style="width:64px;">FS201</td>
			<td>optional</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:403px;">Stainless Steel Ferrule</td>
			<td style="width:323px;">Precision fiber optics</td>
			<td style="width:64px;">MM-FER2003SS-265</td>
			<td>265 micron inner diameter</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:403px;">1 mL syringe</td>
			<td style="width:323px;">BD</td>
			<td style="width:64px;">14-823-30</td>
			<td>Luer-lok tip is preferable to reduce risk of leakage, but not strictly needed</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:403px;">Plastic epoxy</td>
			<td style="width:323px;">Industrial fiber optics</td>
			<td>40 0005</td>
			<td></td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:403px;">18 gauge blunt needle</td>
			<td style="width:323px;">BD</td>
			<td align="right" style="width:64px;">305180</td>
			<td>1.5 inch length</td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:403px;">Lint-free wipe (KimWipe)</td>
			<td style="width:323px;">ThorLabs</td>
			<td>KW32</td>
			<td>available from many vendors</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:403px;">Light absorbing foil</td>
			<td style="width:323px;">ThorLabs</td>
			<td style="width:64px;">BKF12</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:403px;">Electrical tape</td>
			<td style="width:323px;">3M</td>
			<td style="width:64px;">Temflex 1700</td>
			<td>Optional, may substitute other brands / models</td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:403px;">26 gauge sharp needle&#160;</td>
			<td style="width:323px;">BD</td>
			<td align="right" style="width:64px;">305111</td>
			<td>0.5 inch length</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:403px;">Micromanipulator</td>
			<td style="width:323px;">Siskiyou</td>
			<td style="width:64px;">70750000E</td>
			<td>may substitute other brands/models</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:403px;">Steretactic arm</td>
			<td style="width:323px;">Kopf</td>
			<td align="right" style="width:64px;">1460</td>
			<td>may substitute other brands/models</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:403px;">Laser safety goggles</td>
			<td style="width:323px;">KenTeK</td>
			<td>KCM-6012</td>
			<td>must be selected based on the color of laser used, example given here</td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:403px;">Laser or other light source</td>
			<td style="width:323px;">vortran</td>
			<td style="width:64px;">Stradus 473-50</td>
			<td>example of blue laser</td>
		</tr>
		<tr height="64">
			<td height="64" style="height:64px;width:403px;">Integrating sphere</td>
			<td style="width:323px;">ThorLabs</td>
			<td>S142C</td>
			<td>Attached power meter, also available from ThorLabs, item #PM100D</td>
		</tr>
		<tr height="105">
			<td height="105" style="height:105px;width:403px;">Ultem recording chamber</td>
			<td style="width:323px;">Crist instrument company</td>
			<td style="width:64px;">6-ICO-J0</td>
			<td>Customized with alignment notch</td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:403px;">Tower microdrive with clamps</td>
			<td style="width:323px;">NAN</td>
			<td style="width:64px;">DRTBL-CMS</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:403px;">Guide tube</td>
			<td style="width:323px;">Custom</td>
			<td style="width:64px;">N/A</td>
			<td>Made from 25 gauge spinal needle (BD) or blunt tubing</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:403px;">NAN driver system</td>
			<td style="width:323px;">NAN</td>
			<td style="width:64px;">NANDrive</td>
			<td></td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:403px;">Custom grid design</td>
			<td style="width:323px;">custom</td>
			<td style="width:64px;">custom</td>
			<td>plans available upon request</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:403px;">Blunt forceps</td>
			<td style="width:323px;">FischerScientific</td>
			<td style="width:64px;">08-875-8A</td>
			<td>generic stainless steel blunt forceps</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:403px;">Digital calipers</td>
			<td style="width:323px;">Neiko</td>
			<td style="width:64px;">01407A</td>
			<td>available on amazon.com. May select a finer resolution caliper for more precise measurements.</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:403px;">Patch cable</td>
			<td style="width:323px;">ThorLabs</td>
			<td style="width:64px;">FG200LCC-custom</td>
			<td>This is one example of many possible patch cables. As long as the fiber diameter is less than or equal to the fiber diameter of the large volume illuminator and as long as the connectors interface, any patch cable (glass or plastic, vendor purchased or made in the lab) is fine for this application.</td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:403px;">Clear plastic dust caps</td>
			<td style="width:323px;">ThorLabs</td>
			<td style="width:64px;">CAPF</td>
			<td>Package of 25</td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:403px;">ceramic split mating sleeve</td>
			<td style="width:323px;">Precision Fiber Products, Inc.</td>
			<td style="width:64px;">SM-CS1140S</td>
			<td></td>
		</tr>
	</tbody>
</table>

large volume, behaviorally-relevant illumination for optogenetics in non-human primates

このプロトコルでは、ヒト以外の霊長類の脳の光遺伝学的操作用に開発された大容量の照明について説明します。照明器具はエッチングされたヒントを変更したプラスチック光ファイバー発光面積は &#62; 100 倍、従来の繊維。大量の照明器具の構造を記述する、に加えてこのプロトコルも光の分布を確認するために使用する品質管理校正を詳細します。さらに、このプロトコルでは、挿入し、大量の照明器具を削除するためのテクニックについて説明します。浅と深い構造を点灯可能性があります。この大量の照明が、電極に物理的に結合する必要はありません、照明器具、ガラスではなくプラスチックで作られているのでそれが単に曲がる状況で伝統的な光ファイバーを破るととき。この照明は、行動関連組織ボリューム (≈ 10 mm3) 従来の光ファイバーより大きい浸透損傷なしに光を提供するため、非ひと霊長類におけるオプトジェネティクスを用いた行動の研究を容易にします。

ヒト以外の霊長類の大脳ボリュームの照明は行動に関連した光遺伝学的操作のためことができます。Acker さんら(2016) ハロロドプシンの赤いシフト、2 アカゲザルにおける記憶誘導性サッカード発生する前頭眼野 (FEF) の一時的な貢献を研究する顎7でこの大量の照明器具を使用します。具体的には、FEF ニューロンを顎を含むウイルスのベクトルを注入され、適用先のプ?...

Watch this Scientific Journal Video about Large Volume, Behaviorally-relevant Illumination for Optogenetics in Non-human Primates at JoVE.com

Large Volume, Behaviorally-relevant Illumination for Optogenetics in Non-human Primates

最小直径と大量の光を提供するための組織貫通照明を構築するためのプロトコルが表示されます。

大ボリューム、非ひと霊長類におけるオプトジェネティクスの行動に関連した照明

This protocol describes a large-volume illuminator, which was developed for optogenetic manipulations in the non-human primate brain. The illuminator is a ...

The overall goal of this protocol is to create a large volume tissue illuminator that is well suited to optogenetic manipulations in non human primates. This method can help answer key questions in the field of neuroscience, such as how do populations of neurons in the different brain regions contribute to behavior, and how can temporally specific neuronal firing patterns in one region impact another region? The main advantage of this technique is that it uses light, and thus allows optogenetic manipulation of large behaviorally relevant brain volumes with minimal tissue damage.The implications of this technique extend toward translational medicine because of the similarities between the human and non-human primate brains. Generally, individuals new to this method will struggle because it takes some practice to apply the proper amount of pressure when pulling these fibers. To begin this procedure, use a pair of sharp shears to cut a section of 250 micrometer diameter plastic optical fiber that is at least 10 centimeters longer than the desired total illuminator length.Next, remove 15 to 20 centimeter of polyethylene jacket from one end of the plastic optical fiber using a 22-gauge wire stripper. Then, secure the distal-most three to five centimeters of the stripped end of the fiber in a table vice clamp. Hold the jacketed end of the fiber taut in one hand, with a constant steady pull.The fiber should be parallel to the floor, pulled away from the vice. Maintain this constant tension on the fiber throughout heating and cooling, so that the fiber remains straight and taut as it thins. Using the lowest setting of a dual temperature heat gun, heat the stripped section of the fiber until it is thinned to a diameter of 60 to 100 micrometers, or about the diameter of a human hair.Maintain the constant tension on the fiber while allowing it to cool. Failure to maintain tension or overheating the fiber may cause the fiber to curl and or break. To help maintain constant tension on the fiber as you heat it, use your non-dominant hand so you don't have to let go when you're using the heat gun.Also, you can gently wrap the fiber around your palm to get an even better grip. Once the fiber has fully cooled, and the desired diameter has been achieved, pinch the fiber three centimeters from the narrowest point on both sides. With a quick sharp pull along the axis of the fiber, separate the sides to create a tapered tip.Then, examine the tapered tip in low power under a dissection microscope. If the tip is forked or curled, discard the fiber. Even with a lot of practice, about a third of fibers will have forked or curled tips.We just discard those fibers rather than trying to repair them, since it's so quick and inexpensive to pull another one. Prepare to etch the tapered tip by placing a piece of lab tape near the end of the tip, leaving the desired tip length exposed. Here the last five millimeters are exposed.The tape protects the fiber from etching above the desired length of light emission. After that, fold a small square of five micrometer silicon carbide lapping sheet over between the thumb and forefinger. Place the tip of the fiber between the two sides of the lapping sheet, and gently etch the fiber using small circular motions.Frequently rotate the fiber so that all sides are etched evenly. Next, remove about five millimeters of the polyethylene jacket from the opposite end of the illuminator using a 22-gauge wire stripper. Then, cut the opposite end flat with a hot knife to smooth the surface.Affix a connector on the end of the illuminator opposite to the etched tip. This allows the illuminator to connect to an optical cable or laser. Afterward, polish the opposite end flat with successively finer lapping sheets.Subsequently, insert the flat opposite end into a 260 micrometer inner diameter stainless steel ferrule until it is flush with the end of the ferrule. Visually confirm that the opposite end is smoothly and evenly polished with a fiber microscope. Tape the ferrule vertically on the edge of the table with the smaller opening of the ferrule pointing down.Then, fill a one milliliter syringe with plastic epoxy and attach a blunt 18-gauge needle to the syringe. Following this, use the needle to apply epoxy down into the ferrule. Fill the ferrule completely with epoxy.Insert fiber into the ferrule so that it is flush with the bottom of the ferrule. If needed, wipe any excess epoxy from the polished surface of the fiber with a wet lint-free wipe prior to drying. In this step, prepare a tower microdrive prior to each testing session.Then affix a 25-gauge guide tube with a beveled tip to the lower clamp on the drive. Secure the large volume illuminator in the upper clamp on the same drive. This clamp is attached to the drive motor.Afterward, thread the large volume illuminator through the guide tube fully, and confirm that the tip of the illuminator extends past the tip of the guide tube. Alternately, lab tape may be applied to the fiber so that the tape, rather than the fiber itself, is clamped, to protect the fiber. Next, soak the guide tube and illuminator in an antiseptic solution for sterilization.Place a custom sterilized grid with one millimeter hole spacing inside the clean chamber, and secure it at a prespecified orientation with the screw on the side of the chamber. Retract the sterilized illuminator from the guide tube by pulling the illuminator up using sterile blunt forceps. The tip of the illuminator should be five to 10 millimeters above the tip of the guide tube.Affix the microdrive to the holder and place the guide tube into the target grid hole. Then, lower the microdrive stage until the guide tube is just through the level of the dura. At the same time, lower the illuminator manually with sterile forceps.If there is any resistance, retract the illuminator five to 10 millimeters, wait 10 minutes, and try again. If there is still resistance on the second attempt, retract the illuminator tip into the guide tube, lower the guide tube 0.25 millimeters, and try again. Connect the ferrule on the other end of the illuminator to the larger optic setup or laser.This figure shows that the error rates increased significantly with illumination. And the raster plots here show the inhibition of neurons spanning the 2.5 millimeter thickness of cortex. And the local field potential showing the light artifact spanning 4.5 millimeters.Once mastered, a large volume illuminator can be constructed using this technique in 10 to 15 minutes, plus epoxy drying time. Calibration adds another half hour or so, so to maximize efficiency, we usually make several illuminators at once. While attending this procedure, it's important to remember that many of the illuminators won't meet quality control standards.That's okay. If they fail tip inspection, we discard them. But if they fail calibration, we usually repolish them and repeat the calibration.After its development, this technique helped pave the way for researchers in the field of neuroscience to use optogenetics to suppress neuronal firing and modulate behavior in non-human primates. After watching this video, you should have a good understanding of how to construct a large volume tissue illuminator.

Research

JoVE Journal

Behavior

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

禁欲ラットで探しているヘロインに長期の食事制限の影響を研究するための手順

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

行動学

Barnes Maze Testing Strategies with Small and Large Rodent Models

小型·大齧歯類モデルとバーンズ迷路テスト戦略

Spatial learning and memory of laboratory rodents is often assessed via navigational ability in mazes, most popular of which are the water and dry-land ...

Methods to Explore the Influence of Top-down Visual Processes on Motor Behavior

運動行動上のトップダウン視覚プロセスの影響を探索する方法

Kinesthetic awareness is important to successfully navigate the environment. When we interact with our daily surroundings, some aspects of movement are ...

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

ビート難聴を暴く：同期指タッピングと知覚タイミングタスクを検出リズム障害

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

A Procedure to Observe Context-induced Renewal of Pavlovian-conditioned Alcohol-seeking Behavior in Rats

ラットにおけるパブロフ付きアルコール探索行動の状況誘発性のリニューアルを観察する手順

Environmental contexts in which drugs of abuse are consumed can trigger craving, a subjective Pavlovian-conditioned response that can facilitate ...

Assessment of Social Cognition in Non-human Primates Using a Network of Computerized Automated Learning Device (ALDM) Test Systems

Assessment of Social Cognition in Non-human Primates Using a Network of Computerized Automated Learning Device ALDM Test Systems

コンピューターの自動学習装置（ALDM）テストシステムのネットワークを用いた非ヒト霊長類における社会的認知の評価

Fagot &#38; Paleressompoulle1 and Fagot &#38; Bonte2 have published an automated learning device (ALDM) for the study of cognitive abilities of monkeys ...

The Social Dimension of Stress: Experimental Manipulations of Social Support and Social Identity in the Trier Social Stress Test

トリーアの社会ストレステストにおけるソーシャルサポートと社会的アイデンティティの実験操作：ストレスの社会的側面

In many situations humans are influenced by the behavior of other people and their relationships with them. For example, in stressful situations ...

Using Facial Electromyography to Assess Facial Muscle Reactions to Experienced and Observed Affective Touch in Humans

人間の経験と観測された情緒的なタッチに顔面筋の反応を評価するために顔面筋電図

"Affective" touch is believed to be processed in a manner distinct from discriminatory touch and to involve activation of C-tactile (CT) afferent fibers. ...

In Vivo Protocol of Controlled Subconcussive Head Impacts for the Validation of Field Study Data

フィールド研究データの検証コントロールの Subconcussive ヘッド影響の Vivo プロトコルで

Subconcussive hits pose a threat to neuronal health as they have shown to induce neuronal structural damage and functional impairment without causing ...

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

5分間の音声サンプル手順を用いた子どもに関する親の短い物語の一貫性の評価

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