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 anerkender støtte fra et NDSEG stipendium, NSF GRFP og venner af McGovern Institute. EP anerkender finansiering Harry og Eunice Nohara UROP fond, MIT klasse af 1995 UROP fond, og MIT UROP fond. ESB anerkender midler fra NIH 2R44NS070453-03A1, IET Harvey præmie og New York stamcelle Foundation-Robertson Award. RD anerkender midler fra NIH EY017292. Michael Williams hjalp holdet til at organisere og samle leverancer inden optagelserne.

Bemærk: alle dyr, der var i overensstemmelse med retningslinjerne for NIH og blev godkendt af Massachusetts Institut for teknologi Udvalget om Animal Care.
 1. illuminator fabrikation <ol><li>bruger et par skarpe Sakse til at skære en del af 250 µm diameter plast optisk fiber, der er mindst 10 cm længere end den ønskede samlede illuminator.</li>
	<li>Fjerne 15-20 cm af polyethylen jakke fra den ene ende af den plast optisk fiber ved hjælp af en 22 gauge wire stripper.</li>
	<li>Sikre ...

<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>

Mens optogenetic værktøjer er meget udbredt at studere sygdom og fysiologi i gnavere, har den tekniske udfordring af lysende store hjerne diskenheder begrænset anvendelse af optogenetics i ikke-menneskelige primater. Banebrydende undersøgelser i aber bruges store lys effekttætheder (~ 100 mW/mm2 til 20 W/mm2) til at belyse små mængder, måske &#60; 1 mm3og rapporterede beskedne adfærdsmæssige effekter med excitatoriske opsins i cortex4, <sup cl...

Optogenetic værktøjer, som gør det muligt for millisekund-præcis, lys-drevet neuronal kontrol er meget udbredt at studere funktionelle fysiologi og adfærd i gnavere og hvirvelløse dyr. Tekniske udfordringer har dog begrænset brug af optogenetics i primat hjerne, som har en volumen ~ 100 x større end gnaver hjernen 1.
For at lette optogenetics undersøgelser i ikke-menneskelige primater, en illuminator var designet til at løse to konkurrerende mål: stor mængde belysning og minimal penetration skader. Tidligere forsøg på at løse en af disse bekymringer er kommet på de dyre af den anden. Bundter af fibre belyse større mængder, men med øget diameter og dermed skade2,3. Koniske glas fibre reducere penetration skader, men snævert fokus lys til lysemitterende overflade områder &#60; 100 µm2 4,5. Eksterne hjernen belysning gennem et vindue i dura omgår udfordring af penetration skader og giver mulighed for stor mængde belysning, men det kan kun bruges til et par overfladiske hjernen områder6.
Hvis du vil oprette et stort volumen, lille diameter illuminator (figur 1a), ætset spidsen af en optisk fiber er varme tilspidset og core og beklædning plast (figur 1bc). I modsætning til andre tilspidset fibre, der fokuserer lyset til en smal punkt, giver ætsning lys til at undslippe jævnt ud af siderne af den aflæsse, således distribuere lys bredt over et stort område (figur 1 de). Fordi penetration skader er proportional med penetration diameter, denne illuminator har ingen mere penetration skade end en konventionel fiber, men det har &#62; 100 x lysemitterende overflade område og leverer lys mere bredt med 1/100 lys magt tæthed i en hjerne phantom (1,75% Agarosen) (figur 1e). Monte Carlo model (figur 1f) illustrerer forskellen i lys spredes mellem en konventionel fiber og den store mængde illuminator, når de har lige lys effekttætheder som deres lysemitterende flader. Hver illuminator kalibreres individuelt ved hjælp af en Integrationskuglens (figur 2a, b) at sikre selv lysfordeling langs spidsen (figur 2 c).
Denne store mængde illuminator er blevet valideret med optogenetic manipulation af både adfærd og neuronal fyring i ikke-menneskelige primater. Fiber tip længde kan tilpasses ethvert område, hjernen og enkelte dyrs individuelle modtagelig feltforbindelse. Af belysningen kan være parret med en gennemtrængende elektrode til neuronal optagelser, der strækker sig over længden af belysning. Yderligere, fordi fiber kan bære alle farver af synligt lys, kan det være parret med nogen af de tilgængelige optogenetic molekyler tilgængelige.

<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

Denne protokol beskriver et stort volumen illuminator, som blev udviklet for optogenetic manipulationer i primat hjerne. Af belysningen er en modificeret plast optisk fiber med ætset tip, således, at den lysemitterende overflade &#62; 100 x, at af en konventionel fiber. Ud over at beskrive opbygningen af store mængder illuminator, detaljer denne protokol kvalitetskontrol kalibreringen bruges til at sikre endnu lysfordeling. Yderligere, denne protokol beskriver teknikker til at indsætte og fjerne den store mængde illuminator. Både overfladisk og dyb strukturer kan blive belyst. Denne store mængde illuminator behøver ikke at være fysisk koblet til en elektrode, og fordi illuminator er lavet af plastic, ikke glas, det simpelthen vil bøje under omstændigheder, når traditionelle optiske fibre vil splintre. Fordi denne illuminator leverer lys over behaviorally relevante væv diskenheder (≈ 10 mm3) uden større penetration skader end en konventionel optisk fiber, letter det adfærdsmæssige undersøgelser ved hjælp af optogenetics i ikke-menneskelige primater.

Belysningen af store hjerne mængder i ikke-menneskelige primater tillader behaviorally relevante optogenetic manipulation. Acker et al. (2016) anvendes denne store mængde illuminator med rød-forskudt Halorhodopsin, Jaws 7 at studere den tidsmæssige bidrag af feltet frontal øje (FEF) til hukommelse-styrede saccades i to rhesus aber. Specifikt, blev FEF neuroner injiceret med en viral vektor indeholdende kæber og derefter belyst med rødt-lys ved hjælp af den store mængde illuminato...

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

En protokol til at opbygge en vævet gennemtrængende illuminator for at levere lys over store mængder med minimale diameter er præsenteret.

Stor volumen, Behaviorally relevante belysning til Optogenetics i ikke-menneskelige primater

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

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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

Den sociale dimension af stress: Eksperimentelle manipulationer af social støtte og social identitet i Trier Social stresstesten

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

Ved hjælp af Facial Elektromyografi at vurdere Facial muskel reaktioner til erfarne og observerede affektive Touch hos mennesker

"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

I Vivo protokol af kontrollerede Subconcussive hoved virkninger for validering af felt undersøgelse Data

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