a technique to inflict closed-head traumatic brain injury in drosophila flies

A Technique to Inflict Closed-Head Traumatic Brain Injury in Drosophila Flies

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.
If the proboscis is exposed, use a blunted 1 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.

a-technique-to-inflict-closed-head-traumatic-brain-injury-drosophila

Research

JoVE Encyclopedia of Experiments

Neuroscience

Neuropathology

Trauma

EoE Sub Articles

eoe sub articles

1. Assembly of the Strike Device (Figure 1)
<ol>
	<li>Remove the plunger from a 1 mL tuberculin syringe. Cut the barrel at the 1 mL mark.</li>
	<li>Remove an aerosol barrier (3 mm height x 4 mm diameter) from a 200 &#181;L pipette tip and use it as an impactor. Place the impactor inside the syringe barrel. Gently tap the barrel to move the impactor to the tip end, with the flat side covering the nozzle opening.</li>
	<li>Attach the barrel tip end to plastic tubing that is connected to the carbon dioxide (CO2) flow regulator of a Drosophila anesthesia station.</li>
	<li>Hold the barrel vertically and clamp it to a standard clamp holder stand so that the impactor stays at the bottom of the barrel.</li>
	<li>Modify a 200 &#181;L pipette tip to make the fly holder.
	<ol>
		<li>Cut 4 mm from the tip to make a 0.8 mm-diameter opening, allowing only the fly head to be exposed. 
		&#8203;NOTE: The thorax and all the other parts of the fly body will stay inside the pipette tip.</li>
	</ol>
	</li>
	<li>Modify a 1,000 &#181;L pipette tip and a 1 mL syringe needle cap to make the connector.
	<ol>
		<li>Cut off 44 mm from the opening of the tip. Take a 6 mm length of a 1 mL syringe needle cap and push it tightly into the remaining segment of the 1,000 &#181;L pipette tip.</li>
	</ol>
	</li>
</ol>
2. Operation of the Strike Device
<ol>
	<li>Anesthetize a single 2-day-old adult female fly using CO2 on a fly pad.</li>
	<li>Gently transfer it to the fly holder using a fine brush. Tap the holder gently so that the fly head is seen outside of the tip end. If the fly proboscis is exposed outside the tip, gently tuck it back inside the tip with a blunted 1 mL syringe needle. 
	NOTE: Make sure to keep the fly proboscis inside the holder. Otherwise, the fly may die from a sucking proboscis injury.</li>
	<li>Tighten the fly holder to the syringe barrel with the connector so that the fly head is facing downwards.</li>
	<li>Set the gas pressure to 100 kPa. Adjust the flow rate according to the experiment design.</li>
	<li>Quickly turn the flow regulator toggle switch on and off so that the impactor strikes the fly head once.</li>
	<li>Lift the fly holder and move it over a fly pad. Reverse the fly holder and gently tap the side to let the fly out. Leave the fly in an empty vial to recover.</li>
</ol>

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Aerosol Barrier</td>
			<td>USA Scientific</td>
			<td>1120-8810</td>
			<td>Used as an impactor</td>
		</tr>
		<tr>
			<td>200 ul Pipette Tip</td>
			<td>USA Scientific</td>
			<td>1111-0706</td>
			<td>Used as a fly head holder</td>
		</tr>
		<tr>
			<td>1000 ul Pipette Tip</td>
			<td>USA Scientific</td>
			<td>1122-1830</td>
			<td>Used as a connector</td>
		</tr>
		<tr>
			<td>1 ml Tuberculin Syringe</td>
			<td>Becton Dickinson</td>
			<td>309625</td>
			<td></td>
		</tr>
		<tr>
			<td>60 mm Petri Dishes</td>
			<td>Fisher Scientific</td>
			<td>FB0875713A</td>
			<td>Used as a tracking arenas</td>
		</tr>
		<tr>
			<td>Flow Regulator</td>
			<td>Genesee Scientific</td>
			<td>59-122WC</td>
			<td></td>
		</tr>
		<tr>
			<td>Standard Clamp Holder/stand</td>
			<td>EISCO Scientific</td>
			<td>CH0688</td>
			<td></td>
		</tr>
		<tr>
			<td>Fine Brush</td>
			<td>Genesee Scientific</td>
			<td>59-204</td>
			<td></td>
		</tr>
		<tr>
			<td>Flypad</td>
			<td>Genesee Scientific</td>
			<td>59-114</td>
			<td></td>
		</tr>
	</tbody>
</table>

Source: Sun, M., et al. <a href="https://app.jove.com/v/55602">A Novel Method to Model Chronic Traumatic Encephalopathy in Drosophila.</a> J. Vis. Exp. (2017).
This video demonstrates a procedure to inflict a closed-head traumatic injury in anesthetized Drosophila flies using a strike device with a gas-propelled impactor. The injury affects neuronal function and impairs walking ability.

<img alt="Figure 1" src="/files/ftp_upload/23073/23073_Figure1.jpg" /> 
Figure 1. Diagram of the Strike Device. The device used materials that are readily available in a Drosophila lab: 1) 200 &#181;L pipette tip, 2) 1 mL tuberculin syringe, 3) impactor, 4) connecting tube hooked to a Drosophila anesthesia station, and 5) connector to tighten the tip onto the syringe.

1. Assembly of the Strike Device (Figure 1)

	Remove the plunger from a 1 mL tuberculin syringe. Cut the barrel at the 1 mL mark.
	Remove an aerosol ...

Take control and test adult Drosophila flies under anesthesia.
Place the test fly in a holder and gently tap it to position the fly&#39;s head outside the tip. Ensure the fly&#39;s mouthparts and body remain inside the holder to avoid non-target injuries.
Attach the holder to a strike device with the fly&#39;s head facing downward.
The device is connected to a controlled gas supply and contains a gas-propelled impactor to inflict injury.
Deliver a gas burst to propel the impactor and strike the fly&#39;s head, inducing a closed-head traumatic brain injury without external damage.
The resulting mechanical stress damages neurons, impairing their function.&#160;&#160;
Detach the holder to release the fly, then transfer it to a vial for recovery.
For the control fly, apply a gas burst without striking it with the impactor.
The control and test flies are now ready to assess the impact of the injury.&#160;&#160;&#160;&#160;&#160;

17 Views. טכניקה לגרימת פגיעה מוחית טראומטית בראש סגור בזבובי דרוזופילה

Video: טכניקה לגרימת פגיעה מוחית טראומטית בראש סגור בזבובי דרוזופילה - Experiment

Investigating Alterations in Caecum Microbiota After Traumatic Brain Injury in Mice

Increasing evidence shows that the microbiota-gut-brain axis plays an important role in the pathogenesis of brain diseases. Several studies also demonstrate that traumatic brain injuries cause changes to the gut microbiota. However, mechanisms underlying the bidirectional regulation of the brain-gut axis remain unknown. Currently, few models exist for studying the changes in gut microbiota after traumatic brain injury. Therefore, the presented study combines protocols for inducing traumatic brain injury using a lateral fluid percussion device and analysis of caecum samples following injury for investigating alterations in the gut microbiome. Alterations of the gut microbiota composition after traumatic brain injury are determined using 16S-rDNA sequencing. This protocol provides an effective method for studying the relationships between enteric microorganisms and traumatic brain injury.

Presented here is a protocol to induce diffuse traumatic brain injury using a lateral fluid percussion device followed by the collection of the caecum content for gut microbiome analysis.

חקירת שינויים Caecum Microbiota לאחר פציעה טראומטית במוח עכברים

Increasing evidence shows that the microbiota-gut-brain axis plays an important role in the pathogenesis of brain diseases. Several studies also ...

JoVE Journal

Assessing Changes in Synaptic Plasticity Using an Awake Closed-Head Injury Model of Mild Traumatic Brain Injury

Mild traumatic brain injuries (mTBIs) are a prevalent health issue in North America. There is increasing pressure to utilize ecologically valid models of closed-head mTBI in the preclinical setting to increase translatability to the clinical population. The awake closed-headed injury (ACHI) model uses a modified controlled cortical impactor to deliver closed-headed injury, inducing clinically relevant behavioral deficits without the need for a craniotomy or the use of an anesthetic.
This technique does not normally induce fatalities, skull fractures, or brain bleeds, and is more consistent with being a mild injury. Indeed, the mild nature of the ACHI procedure makes it ideal for studies investigating repetitive mTBI (r-mTBI). Growing evidence indicates that r-mTBI can result in a cumulative injury that produces behavioral symptoms, neuropathological changes, and neurodegeneration. r-mTBI is common in youths playing sports, and these injuries occur during a period of robust synaptic reorganization and myelination, making the younger population particularly vulnerable to the long-term influences of r-mTBI.
Further, r-mTBI occurs in cases of intimate partner violence, a condition for which there are few objective screening measures. In these experiments, synaptic function was assessed in the hippocampus in juvenile rats that had experienced r-mTBI using the ACHI model. Following the injuries, a tissue slicer was utilized to make hippocampal slices to evaluate bidirectional synaptic plasticity in the hippocampus at either 1 or 7 days following the r-mTBI. Overall, the ACHI model provides researchers with an ecologically valid model to study changes in synaptic plasticity following mTBI and r-mTBI.

Here, it is demonstrated how an awake closed-head injury model can be used for examining the effects of repeated mild traumatic brain injury (r-mTBI) on synaptic plasticity in the hippocampus. The model replicates important features of r-mTBI in patients and is used in conjunction with in vitro electrophysiology.

הערכת שינויים בפלסטיות סינפטית באמצעות מודל פגיעת ראש סגורה ערה של פגיעה מוחית טראומטית קלה

Mild traumatic brain injuries (mTBIs) are a prevalent health issue in North America. There is increasing pressure to utilize ecologically valid models of ...

אימות תגובה לפציעה והערכות התנהגותיות במודל של חולדות mTBI

Rat Model of Closed-Head Mild Traumatic Injury and its Validation

Animal models are crucial for advancing our understanding of mild traumatic brain injury (mTBI) and guiding clinical research. To achieve meaningful insights, developing a stable and reproducible animal model is essential. In this study, we report a detailed description of a closed-head mTBI model and a representative validation method using Sprague-Dawley rats to verify the modeling effect. The model involves dropping a 550 g mass weight from a height of 100 cm directly onto the head of a rat on a destructible surface, followed by a 180-degree turn. To assess the injury, rats underwent a series of neurobehavioral assessments 10 min post-injury, including time of loss of consciousness, first seeking-behavior time, escape ability, and beam balance ability test. During the acute and subacute stages following the injury, behavioral tests were conducted to assess motor coordination ability (Beam task), anxiety (Open Field test), and learning and memory abilities (Morris Water Maze test). The closed-head mTBI model produced a consistent injury response with minimal mortality and replicated real-life situations. The validation method effectively verified the model development and ensured the stability and consistency of the model.

מחבר Spotlight: Advanceancing Traumatic Brain Injury Research - מודל ראש סגור לשכפול מדויק והערכה מהירה 

Here, we present a closed-head mild traumatic brain injury (mTBI) rat model and its validation exhibiting remarkable similarity to human mTBI concerning behavioral manifestations during the acute and subacute stages.

מודל חולדה של פגיעה טראומטית קלה בראש סגור ותיקוף שלה

Animal models are crucial for advancing our understanding of mild traumatic brain injury (mTBI) and guiding clinical research. To achieve meaningful ...

A Technique to Inflict Closed-Head Traumatic Brain Injury in Drosophila Flies

Transcript

A Technique to Inflict Closed-Head Traumatic Brain Injury in Drosophila Flies