Name: a novel method to model chronic traumatic encephalopathy in drosophila
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Description: Watch this Scientific Journal Video about A Novel Method to Model Chronic Traumatic Encephalopathy in Drosophila at JoVE.com

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1. Montaggio del Dispositivo Strike ( Figura 1 ) <ol><li> Rimuovere lo stantuffo da una siringa tubercolina da 1 ml. Tagliare la canna al punto di 1 ml. </li><li> Rimuovere una barriera aerosol (3 mm di altezza x 4 mm di diametro) da una punta di pipetta da 200 μl e utilizzarla come impattatore. Posizionare l&#39;impatto nell&#39;interno della barra della siringa. Toccare delicatamente il cilindro per spostare l&#39;impattatore all&#39;estremità della punta, con il lato piatto che copre...

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C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chronic+traumatic+encephalopathy:+where+are+we+and+where+are+we+going?.">Chronic traumatic encephalopathy: where are we and where are we going?.</a> Curr Neurol Neurosci Rep. 13 (12), 407 (2013).</li><li>McKee, A. 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=The+spectrum+of+disease+in+chronic+traumatic+encephalopathy.">The spectrum of disease in chronic traumatic encephalopathy.</a> Brain. 136 (Pt 1), 43-64 (2013).</li><li>Petraglia, A. L., 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=The+spectrum+of+neurobehavioral+sequelae+after+repetitive+mild+traumatic+brain+injury:+a+novel+mouse+model+of+chronic+traumatic+encephalopathy.">The spectrum of neurobehavioral sequelae after repetitive mild traumatic brain injury: a novel mouse model of chronic traumatic encephalopathy.</a> J Neurotrauma. 31 (13), 1211-1224 (2014).</li><li>Hirth, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Drosophila+melanogaster+in+the+study+of+human+neurodegeneration.">Drosophila melanogaster in the study of human neurodegeneration.</a> CNS Neurol Disord Drug Targets. 9 (4), 504-523 (2010).</li><li>Littleton, J. T., Ganetzky, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ion+channels+and+synaptic+organization:+analysis+of+the+Drosophila+genome.">Ion channels and synaptic organization: analysis of the Drosophila genome.</a> Neuron. 26 (1), 35-43 (2000).</li><li>Appel, L. F., 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=The+Drosophila+Stubble-stubbloid+gene+encodes+an+apparent+transmembrane+serine+protease+required+for+epithelial+morphogenesis.">The Drosophila Stubble-stubbloid gene encodes an apparent transmembrane serine protease required for epithelial morphogenesis.</a> Proc Natl Acad Sci USA. 90 (11), 4937-4941 (1993).</li><li>Piyankarage, S. C., Featherstone, D. E., Shippy, S. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nanoliter+hemolymph+sampling+and+analysis+of+individual+adult+Drosophila+melanogaster.">Nanoliter hemolymph sampling and analysis of individual adult Drosophila melanogaster.</a> Anal Chem. 84 (10), 4460-4466 (2012).</li><li>Katzenberger, R. 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=A+Drosophila+model+of+closed+head+traumatic+brain+injury.">A Drosophila model of closed head traumatic brain injury.</a> Proc Natl Acad Sci USA. 110 (44), E4152-E4159 (2013).</li><li>Barekat, 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=Using+Drosophila+as+an+integrated+model+to+study+mild+repetitive+traumatic+brain+injury.">Using Drosophila as an integrated model to study mild repetitive traumatic brain injury.</a> Sci Rep. 6, 25252 (2016).</li><li>Katzenberger, R. 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=A+Method+to+Inflict+Closed+Head+Traumatic+Brain+Injury+in+Drosophila.">A Method to Inflict Closed Head Traumatic Brain Injury in Drosophila.</a> J Vis Exp. (e52905), (2015).</li><li>Branson, K., Robie, A. A., Bender, J., Perona, P., Dickinson, M. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High-throughput+ethomics+in+large+groups+of+Drosophila.">High-throughput ethomics in large groups of Drosophila.</a> Nat Methods. 6 (6), 451-457 (2009).</li><li>Straw, A. D., Dickinson, M. H., 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=Motmot,+an+open-source+toolkit+for+realtime+video+acquisition+and+analysis.">Motmot, an open-source toolkit for realtime video acquisition and analysis.</a> Source Code Biol Med. 4 (5), 1-10 (2009).</li><li>Talavage, T. 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=Functionally-detected+cognitive+impairment+in+high+school+football+players+without+clinically-diagnosed+concussion.">Functionally-detected cognitive impairment in high school football players without clinically-diagnosed concussion.</a> J Neurotrauma. 31 (4), 327-338 (2014).</li><li>Theadom, 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=Frequency+and+impact+of+recurrent+traumatic+brain+injury+in+a+population-based+sample.">Frequency and impact of recurrent traumatic brain injury in a population-based sample.</a> J Neurotrauma. 32 (10), 674-681 (2015).</li><li>Drobysheva, D., 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+optimized+method+for+histological+detection+of+dopaminergic+neurons+in+Drosophila+melanogaster.">An optimized method for histological detection of dopaminergic neurons in Drosophila melanogaster.</a> J Histochem Cytochem. 56 (12), 1049-1063 (2008).</li></ol>

I modelli animali che modellano fedelmente le caratteristiche del CTE, comprese le alterazioni neurofisiologiche, i segni neuropatologici e i disordini neurobehaviorali, sono essenziali per scoprire i meccanismi della malattia e per sviluppare obiettivi diagnostici e terapeutici. È comprensibile che nessun modello animale di una malattia umana sia perfetto a imitare tutti gli endpoint clinicamente rilevanti. Tuttavia, riteniamo che un robusto modello CTE dovrebbe soddisfare i seguenti tre requisiti: (1) l&#39;impatto d...

L&#39;encefalopatia traumatica cronica (CTE) è stata recentemente riconosciuta come un disturbo neurodegenerativo distinto, separato da altre tauopatie come la malattia di Alzheimer 1 . A differenza della malattia di Alzheimer e di altre tauopatie comuni, i cui fattori di rischio più importanti stanno avanzando l&#39;età e una storia familiare di demenza, la CTE, come indicato dal suo nome, implica una stretta collaborazione con una storia di traumi cerebrali, probabilmente visto in atleti sportivi di contatto, Come pugili e calciatori, così come nei veterani militari 2 , 3 , 4 , 5 . Si pensa di essere iniziato da ripetuti colpi concussivi e subconcutici alla testa. I pazienti possono presentare sintomi e segni quali disordini cognitivi, cambiamenti di umore e comportamento e disfunzione di movimento, che si sovrappongono in modo significativo con la malattia di Alzheimer, frontotemporalDemenza, demenza del corpo di Lewy e malattia di Parkinson 6 . Al contrario, gli esami post-mortem del tessuto cerebrale rivelano un modello distinto di accumulo di tau per iperfosforilati che circonda i piccoli vasi sanguigni nelle profondità dei sulci corticali, caratteristica patognomonica non osservata nelle altre condizioni degenerative 7 . Tuttavia, finora poco si sa sulla patogenesi che conduce alla manifestazione delle malattie. Questo è in gran parte dovuto alla mancanza di un modello animale fedele - solo di recente sono stati generati modelli roditori 5 , 8 . Questi organismi di modello hanno gli svantaggi di una terapia intensiva e una durata di vita relativamente lunga, che non sono adatti per studi di malattia neurodegenerativa. Rispetto alle omologhe di mammiferi, gli animali invertebrati come Drosophila sono un&#39;ottima alternativa, con la loro manutenzione economica,Strumenti estensivi per la dissezione di determinanti genetici e durata relativamente breve 9 . Notevolmente, il cervello volante e umano condividono le vie molecolari e cellulari evolutamente conservate, nonché le somiglianze anatomiche 10 , 11 , 12 . Due geniali modelli Drosophila per studiare lesioni cerebrali traumatiche sono state riportate in precedenza 13 , 14 . Il primo dispositivo &quot;High Impact Trauma&quot; (HIT) progettato da Katzenberger e colleghi conteneva mosche in movimento in una flaconcina di plastica legata all&#39;estremità libera di una molla metallica 13 , 15 . Quando il flaconcino di plastica è stato inclinato in verticale e rilasciato, ha colpito un tampone di poliuretano e impartito il trauma alle mosche quando si sono rimbalzati alla parete del flacone e rimbalzato. Al contrario, Barekat e colleghi hanno progettato un diverso metodo di consegnaLa piattaforma omogeneizzatrice Omni Bead Ruptor-24 14 . Le mosche sono state inabilitate con CO 2 e poste in un tubo a vite da 2 mL che è stato fissato all&#39;omogenizzatore e sottoposto a condizioni di scossa preprogrammate. Un vantaggio di utilizzare il sistema di omogeneizzazione dei tessuti è che lo sperimentatore potrebbe modulare l&#39;intensità della lesione, la durata della lesione e il numero di attacchi di lesioni. Tuttavia, entrambi i regimi subiscono lo stesso inconveniente: lesioni primarie alla testa vengono inflitte casualmente in termini di posizione e forza di impatto. Inoltre, entrambi i metodi hanno determinato una notevole mortalità, causata da inevitabili danneggiamenti collaterali ad altre parti del corpo e degli organi interni. Qui descriviamo un nuovo metodo per indurre mTBI nelle mosche di frutta. Il nostro apparecchio è costituito da un impatto balistico a propulsione a gas. Rispetto ai modelli Drosophila 14 , 15 , il nostro metodo ha il vantaggio unico di fornire measImpatto urra, diretto solo alla testa di mosca libera, consentendo così il controllo accurato di vari fattori, come la gravità dell&#39;impatto, l&#39;intervallo di tempo tra gli impatti e il numero totale di impatti sostenuti.

<table border="0" cellpadding="0" cellspacing="0" width="692">
	<tbody>
		<tr height="20">
			<td height="20" style="height:20px;width:216px;">Aerosol Barrier</td>
			<td style="width:151px;">USA Scientific</td>
			<td style="width:119px;">1120-8810</td>
			<td style="width:207px;">Used as an impactor</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:216px;">200 ul Pipette Tip</td>
			<td style="width:151px;">USA Scientific</td>
			<td style="width:119px;">1111-0706</td>
			<td>Used as a fly head holder</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:216px;">1000 ul Pipette Tip</td>
			<td style="width:151px;">USA Scientific</td>
			<td style="width:119px;">1122-1830</td>
			<td>Used as a connector</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:216px;">1 ml Tuberculin Syringe</td>
			<td style="width:151px;">Becton Dickinson</td>
			<td style="width:119px;">309625</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:216px;">60 mm Petri Dishes</td>
			<td style="width:151px;">Fisher Scientific</td>
			<td>FB0875713A</td>
			<td>Used as a tracking arenas</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:216px;">Flow Regulator</td>
			<td>Genesee Scientific</td>
			<td style="width:119px;">59-122WC</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:216px;">Standard Clamp Holder/stand</td>
			<td style="width:151px;">EISCO Scientific</td>
			<td style="width:119px;">CH0688</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:216px;">Fine Brush</td>
			<td style="width:151px;">Genesee Scientific</td>
			<td style="width:119px;">59-204</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:216px;">Flypad</td>
			<td style="width:151px;">Genesee Scientific</td>
			<td style="width:119px;">59-114</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:216px;">Sylgard Silicone Elastomer</td>
			<td style="width:151px;">Dow Corning</td>
			<td>4019862</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">CCD Camera</td>
			<td style="width:151px;">Microsoft&#160;</td>
			<td style="width:119px;">HD-5000</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Ctrax Walking Fly Tracker</td>
			<td style="width:151px;">Caltech</td>
			<td style="width:119px;">Ctrax 0.2.11</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:216px;">MATLAB Image Processing Toolbox</td>
			<td style="width:151px;">MATLAB</td>
			<td style="width:119px;">R2015b</td>
			<td></td>
		</tr>
	</tbody>
</table>

a novel method to model chronic traumatic encephalopathy in drosophila

L&#39;encefalopatia traumatica cronica (CTE) è una malattia neurodegenerativa consolidata che è strettamente associata all&#39;esposizione a un mite traumatico (TMT) ripetitivo. I meccanismi responsabili dei suoi complessi cambiamenti patologici rimangono in gran parte sfuggenti, nonostante un recente consenso per definire i criteri neuropatologici. Qui descriviamo un nuovo metodo per sviluppare un modello di CTE in Drosophila melanogaster ( Drosophila ) nel tentativo di identificare i geni chiave e le vie che portano al caratteristico accumulo tau iperfosforilato e alla morte neuronale nel cervello. Impatti di forza di regolazione per infliggere lesioni chiuse lievi vengono consegnati direttamente alla testa di mosca sottoponendo la testa ad accelerazione e decelerazione rapida. Il nostro metodo elimina i problemi potenziali inerenti ad altri modelli moto Drosophila ( ad esempio, la morte animale potrebbe essere indotta da danni aAltre parti del corpo o agli organi interni). La mancanza di manodopera a basso costo e a basso costo, una breve durata di vita e estesi strumenti genetici rendono la frutta ideale per studiare la patogenesi della CTE e rendere possibile la realizzazione di schermi genetici e farmacologici avanzati su vasta scala genomica. Prevediamo che la continua caratterizzazione del modello genera importanti conoscenze meccaniche sulla prevenzione delle malattie e sugli approcci terapeutici.

Per stabilire un modello CTE con l&#39;adulto Drosophila abbiamo determinato l&#39;efficacia del nostro dispositivo per infliggere una singola lesione alla testa chiusa. Per eliminare le variazioni relative al genotipo, al sesso o all&#39;età, abbiamo usato le mosche femminili Canton-S WT di due giorni nell&#39;esperimento. Potremmo facilmente controllare la resistenza dell&#39;impulso di impatto regolando la portata di CO 2 ad una pressione costante di 100 kPa. Le m...

Watch this Scientific Journal Video about A Novel Method to Model Chronic Traumatic Encephalopathy in Drosophila at JoVE.com

A Novel Method to Model Chronic Traumatic Encephalopathy in Drosophila

Qui descriviamo un nuovo approccio per infliggere danni traumatici al cervello chiuso a Drosophila melanogaster . Il nostro metodo ha il vantaggio di trasmettere direttamente impatti ripetitivi con forza regolabile solo alla testa. Un&#39;ulteriore esplorazione del sistema invertebrato contribuirà ad illuminare la patogenesi di encefalopatia traumatica cronica.

Un nuovo metodo per modellare l&#39;encefalopatia traumatica cronica in<em&gt; Drosophila</em

Chronic Traumatic Encephalopathy (CTE) is an established neurodegenerative disease that is closely associated with exposure to repetitive mild Traumatic ...

The overall goal of this novel procedure is to inflict closed traumatic brain injuries in the fruit fly, Drosophila, to establish a new chronic traumatic encephalopathy model. This method can help answer key questions in the chronic traumatic encephalopathy field, such as how the repetitive trauma induces neuron degeneration. The main advantage of this technique is that it delivers the repetitive impacts of adjustable strengths to the fly head directly without causing damage to other parts of the animal body.Real world demonstration of this method is critical as assembly of the device and fly loading steps are difficult to learn, because the Drosophila head is so small and it is important to apply the impact only to the head. To make the apparatus used to concuss or strike the fruit flies, first, make the impactor portion of the apparatus. Remove the plunger from a one milliliter polycarbonate tuberculin syringe and then cut the barrel at the one milliliter mark.For an impactor, use the aerosol barrier found on a 200 microliter pipette tip. This impactor should be three millimeters long and four millimeters in diameter. Then, pack the impactor into the syringe barrel with the flat side towards the nozzle.To provide force, attach the syringe barrel to a carbon dioxide flow regulator. The regulator must have an On Off toggle switch and good control of the gas flow rate. Then, clamp the barrel vertically to an iron stand, such that the impactor will fall to the bottom of the barrel.Next, make the fly holder. Start with a 200 microliter pipette tip. Cut the pipette's tip five millimeters from the small end to make a 8 millimeter diameter opening.A fly head will fit through this hole, but the thorax will not. Lastly, make a for the fly holder and impactor device. First, cut a one milliliter pipette tip 44 millimeters from the small opening.Second, cut a six millimeter length from the needle cover for a one milliliter syringe. Then, push the two together. Anesthetize a single, two-day old adult female fly using carbon dioxide.From the carbon dioxide pad, use a fine brush to gently transfer the fly into the holder. Then, gently tap the holder until the head protrudes from the tip. Of the proboscis is exposed, use a blunted, one milliliter syringe needle to gently tuck it back inside the tip.It is critical to keep the fly body, especially the mouth part, inside the holder. Otherwise, the fly may die from damage to the internal organs or from not being able to ingest food. Next, set the gas pressure to 100 kilopascals and adjust the flow as needed.Once the fly is loaded, tighten the holder to the syringe barrel using the connector, such that the fly head faces downward. Then, using the toggle switch, send a burst of gas that moves the impactor to strike the fly once and only once. Now, detach the fly holder and dump the fly back onto the carbon dioxide pad.Brush the fly into an empty vial until it recovers. Keep one fly per vial. The recovery takes only a few minutes.Repeat the process to test four flies per experimental group. Processing two groups, a test and a sham, should only take 20 to 30 minutes. Treat shams just like test animals, except do not include an aerosol barrier in the impactor tube.To track fly movements in a Petri dish, first, fill a six centimeter dish with transparent silicon elastomer to make the tracking arena. Leave a three millimeter gap between the silicon and the lid so that flies can walk freely in the gap, but not take flight. Beneath the tracking arena, have a white background.Next, anesthetize four flies from the same treatment group and place them in the arena. Let the flies acclimate for an hour to the arena, which should be fully lit and at about 22 degrees Celsius. Not hotter than room temperature.After an hour, use a charge-coupled device camera positioned above the arena to record the fly's activity for five minutes. After the five minute recording, anesthetize the flies in the arena and return them to a new home vial. Then, dispose of the arena.Later, analyze the recordings using the freely available Ctrax software. This software generates tracking data, which can be exported in a programmable language compatible format, such as the MATLAB format. From the data, calculate the distance traveled per frame, the mean walking distance for each fly, and the average distance traveled per fly.For statistical significance, expect to test about 100 flies per treatment group. To establish a chronic traumatic encephalopathy model, the effectiveness of a single, closed-head injury made by the impacting apparatus at different speeds was analyzed using two-day old Canton-S females. The gas pressure was held at 100 kilopascals.Flies exposed to a single strike at the highest tested flow rate, exhibited minimal external defects. Despite no clear evidence for external damage, injuries at a flow rate of 15 liters per minute were acutely lethal, resulting in less than a 10%survival within 24 hours. Survival increased with lower flow rates.One hundred percent survival was achieved at five liter per minute or less, so five liters per minute was selected as the standard for the model. Just after being struck, flies gradually recovered their mobility within four minutes. Recovery for the shams was about a minute quicker.Post-impact, video-assisted movement tracking was used to measure locomotive function. During the two days after the impact, locomotion slowly returned to normal. This evidence for gradual recovery is in line with CTE recovery in humans.To evaluate long term effects, a repetitive traumatic brain injury protocol of five strikes over five days was applied. Twenty days post-treatment, the locomotion of surviving members in each group was compared and the treated group clearly walked less. The lifespan of the treated group was also substantially reduced, with a median at 26 days compared to 37 days for controls.After its development, this technique paves the way for researchers in the field of mild repetitive traumatic brain injury to explore pathogenic mechanisms in the classic animal model Drosophila melanogaster. While attempting this procedure, it is important to remember to shield the proboscis, thorax and abdomen from direct impact by the aerosol barrier.

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Neuroscience

A Novel Method to Model Chronic Traumatic Encephalopathy in Drosophila

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A Method to Inflict Closed Head Traumatic Brain Injury in Drosophila

A Method to Inflict Closed Head Traumatic Brain Injury in Drosophila

Un metodo per infliggere Chiuso Testa trauma cranico in<em&gt; Drosophila</em

Traumatic brain injury (TBI) affects millions of people each year, causing impairment of physical, cognitive, and behavioral functions and death. Studies ...

Recording EEG in Freely Moving Neonatal Rats Using a Novel Method

Registrazione di EEG nei ratti neonatali liberamente in movimento utilizzando un metodo novello

EEG is a useful method to detect electrical activity in the brain. Moreover, it is a widely used diagnostic tool for various neurological conditions, such ...

Novel Assay for Cold Nociception in Drosophila Larvae

Novel Assay for Cold Nociception in Drosophila Larvae

Nuovo metodo di analisi per Cold nocicezione in<em&gt; Drosophila</em&gt; Le larve

How organisms sense and respond to noxious temperatures is still poorly understood. Further, the mechanisms underlying sensitization of the sensory ...

A Novel In Vitro Model of Blast Traumatic Brain Injury

A Novel In Vitro Model of Blast Traumatic Brain Injury

Un modello di romanzo In Vitro di ferita di cervello traumatica di scoppio

Traumatic brain injury is a leading cause of death and disability in military and civilian populations. Blast traumatic brain injury results from the ...