We are grateful to Louis Chiocchio for animal care and maintenance, Morgane Regniez for assistance with the intracardiac perfusion procedure, and David Castonguay for assistance with the cryostat. This work was supported by the Caroline Durand Foundation Chair in acute traumatology of the Universite de Montreal awarded to LDB.

The animal protocol for this project obtained the approval from the Animal Care Committee of the Hopital du Sacre-C&#339;ur de Montreal in compliance with the guidelines of the Canadian Council on Animal Care.
NOTE: A schematic outline of the research protocol is presented in Figure 1.
1. Animal Preparation
<ol>
	<li>Order Sprague-Dawley rats from a standard laboratory animal supplier to be delivered between 43 and 50 days of age and at a weight between 151 and 200 g.</li>
	<li>House all rats individually in a cycle of 12:12 h light: darkness, at 24-26 &#176;C with ad libitum access to water and food.</li>
	<li>During the 2 weeks before starting the protocol, handle the rats for 5 min on a daily basis to facilitate their habituation in contact with researchers. Rats should be aged about 10 weeks old and their weight should be between 295 and 351 g at the time of concussion or sham injury induction.</li>
</ol>
2. Microdialysis Guide Cannula Implantation Surgery
<ol>
	<li>Perform the surgery under sterile conditions. Wear sterile gloves, a hair bonnet and a surgical mask throughout the procedure. Autoclave and sterilize all the materials and surgical instruments beforehand. Clean and disinfect the working area and stereotaxic apparatus thoroughly with a solution of ethanol (70%).</li>
	<li>Anesthetize the animals by injecting a cocktail of ketamine (70 mg/kg) and xylazine (10 mg/kg) intraperitoneally. Asses anesthetic depth by testing the reflex to a toe pinch.</li>
	<li>Remove fur from the head of the animal using electric clippers. Clean shaved head using a solution of 2% isopropyl alcohol and 2% chlorhexidine gluconate (3 times). Apply lubricating eye ointment during the anesthesia to prevent dryness.</li>
	<li>Drape-off the surgical field so that only the head of the animal is exposed. Place the head of the rat in a stereotaxic apparatus, insert the ear bars into the ear canals with great care then tighten the nose clamp. Fix a 26 G stainless-steel guide cannula to the holder arm on the stereotaxic apparatus.</li>
	<li>Locally inject an anesthetic cocktail of bupivacaine (1.5 mg/kg) and lidocaine (1.5 mg/kg) subcutaneously on the head, 10 min prior to incision.</li>
	<li>Maintain anesthesia during the whole procedure by delivering sodium isoflurane (2.5%) at 0.5 L/min oxygen flow with a nose cone.</li>
	<li>Make a midline incision (3 cm) along the scalp with a scalpel. Leave the skull clear by installing 4 clamps around the incision.</li>
	<li>Scrape firmly the periosteum from the skull with a surgical blade until the Bregma and Lambda sutures are visible. Maintain firm pressure on the skull with a gauze pad or cotton tipped applicator if there is bleeding.</li>
	<li>Confirm if the skull is correctly aligned on the stereotaxic apparatus by comparing the dorsoventral coordinates of the Bregma and Lambda sutures. Identify the anteroposterior, mediolateral and dorsoventral coordinates of the Bregma suture as the reference points for the coordinates of the guide cannula.</li>
	<li>Taking the Bregma suture coordinates as references, calculate the coordinates of the guide cannula implantation site in the hippocampus. 
	NOTE: The following coordinates were determined according to the rat brain atlas from Paxinos and Watson (anteroposterior: -0.60 cm; mediolateral: &#177;0.58 cm; dorsoventral: -0.16 cm, Figure 2A)32.</li>
	<li>Mark the precise implantation site using a marker.</li>
	<li>Drill a 0.5 mm hole through the cranium at the target site of the guide cannula. Drill 3 other holes approximately 5 mm around this point to thread 3 anchor screws into the skull that will solidify the cannula after acrylic dental cement is applied.</li>
	<li>Insert the cannula into the hippocampus and fix it with dental cement. This cannula will be used to insert the probe into the region of interest 7 days later during the microdialysis procedure. Be careful as to not spill excess dental cement around the site where the weight will be dropped.</li>
	<li>Leave the cement to dry for 2 min, then remove the holder arm from the cannula. Insert a stainless-steel removeable obturator into the cannula to avoid cerebrospinal fluid seepage and risks of infection.</li>
	<li>Remove the 4 clamps, pull back the retracted skin and stitch it with a surgical suture thread 4-0.</li>
	<li>Remove the rat from the apparatus and inject buprenorphine subcutaneously to treat pain (0.05 mg/kg, after surgery then once per day during the following 2 days). Place the rodent back in its cage with a heating pad under until it becomes conscious, then return it to the animal care facility for a 7 days recovery period under close monitoring.</li>
</ol>
3. Microdialysis Procedure
<ol>
	<li>While performing the microdialysis procedure, wear sterile gloves, a hair bonnet and a surgical mask. Seven days following the cannula implantation surgery, anesthetize the rat with sodium isoflurane (2.5%) at 0.5 L/min oxygen flow.</li>
	<li>Remove the obturator from the cannula and insert slowly a microdialysis probe, perfused with artificial cerebral spinal fluid (ACSF) (26&#8201;mmol/L NaHCO3, 3&#8201;mmol/L NaH2PO4, 1.3&#8201;mmol/L MgCl2, 2.3&#8201;mmol/L CaCl2, 3.0&#8201;mmol/L KCl, 126&#8201;mmol/L NaCl, 0.2&#8201;mmol/L L-ascorbic acid), through the cannula into the hippocampus or other region of interest. 
	NOTE: Rats need to be anesthetized only while removing the obturator and inserting the microdialysis probe, and during induction of concussion or sham injury. The probes used here are laboratory-constructed, I-shaped, and comprised of fused side-by-side silica inlet-outlet lines [internal diameter (ID): 50 &#956;m] encased in polyethylene tubing (ID: 0.58-0.38 mm). The end of the cannula is secured with a length of regenerated hollow cellulose membrane [molecular weight cut-off: 13 kDa, outer diameter (OD): 216 &#956;m; ID: 200 &#956;m] using cyanoacrylate adhesive and the tip sealed with epoxy. The active membrane measures 2.5 mm for implantation in the hippocampus but can be adjusted according to the depth of the region of interest. The connection of the indwelling cannula of the rat to the probe is secured with a fitted, threaded stainless-steel collar.</li>
	<li>Fix the probe assembly to a stainless-steel spring tethered to a liquid swivel and counter balance lever arm suspended above the cage with a ring stand and clamps so that the animal can move freely within its cage. Tethered rats spend the entire duration of the microdialysis procedure with ad libitum access to water and food.</li>
	<li>Use a microinfusion pump to send perfusate to the probes, and collect the dialysate from the fused silica outlet line (dead volume: 0.79&#8201;&#956;L).</li>
	<li>At least 1 h and 30 min before the procedure begins, turn up the probe to its working flow rate (1&#8201;&#956;L/min). Verify that the flow rate of the probe is consistent by measuring volume over time with a pipette. 
	NOTE: The flow rate can be more or less depending on the neurotransmitters sampled and the brain region of interest. Dialysis samples are taken before, during, and after concussion or sham injury induction. Sampling interval depends on the brain region of interest, neurotransmitters being analyzed, dialysate concentrations of the analyte, and sensitivity of the analytical chemistry equipment used. The collecting phases done here in the hippocampus for glutamate and GABA sampling are as follows: 
	1. Baseline: At beginning of experiment, collect dialysis samples at 10 min intervals for 60 min. 
	2. Post-concussion or sham injury: After concussion or sham injury, collect samples for an additional 90 min (9 samples).</li>
	<li>Collect each dialysate sample in a fraction vial preloaded with 1 &#956;L of 0.25&#8201;mol/L perchloric acid to prevent analyte degradation. Store the samples at 4 &#176;C for subsequent analysis.</li>
	<li>Following the collection of the last dialysate sample, re-anesthetize the rat with a nose cone delivering sodium isoflurane (2.5%) at 0.5&#8201;L/min oxygen flow.</li>
	<li>Remove the microdialysis probe from the cannula, re-insert the obturator and then return the rat to the animal care facility.</li>
</ol>
4. Concussion Apparatus Installation
<ol>
	<li>Prior to commencement of procedure, carve a weight to be used to inflict the concussion (19 mm in diameter) from solid brass to obtain a mass of 450 g. Insert a metal loop at the top of the brass weight. Drill holes preliminary at a distance of 1.0 m inside a vertical polyvinyl chloride (PVC) guide tube.</li>
	<li>Slit an aluminum sheet with a sharp razor blade. The slotted aluminum sheet should support the weight of the rat (295 to 351 g) without interfering with the acceleration of its body after head impact from the brass weight.</li>
	<li>Tape the slotted aluminum sheet tightly to a U-shaped Plexiglas frame (38 cm long x 27 cm wide x 30 cm deep, Figure 3A,B) that contains a foam cushion (37 cm long x 26 cm wide x 12 cm deep).</li>
	<li>Position the Plexiglas frame under a PVC guide tube (20 mm diameter x 1.5 m length).</li>
	<li>Hold the PVC guide tube in place with a clamp stand 3.5 cm above the slotted aluminum.</li>
	<li>Attach a nylon fly fishing line (capacity of 9.1 kg, 0.46 mm diameter) through the metal loop so that the bottom of the weight hangs 2.5 cm over the slotted aluminum as to prevent multiple hits when the rat is falling on the foam cushion following impact.</li>
	<li>Attach the nylon fly fishing line to the clamp stand.</li>
	<li>Pull up the weight through the PVC tube with the nylon fly fishing line then keep it in place by inserting a hex key through the preliminary drilled holes at 1.0 m.</li>
</ol>
5. Concussion Induction
<ol>
	<li>After the baseline phase of the dialysis samples collection, re-anesthetize the rat lightly by placing a nose cone delivering sodium isoflurane (2.5% isoflurane at 0.5&#8201;L/min oxygen flow) until it as no response to a toe pinch (as mentioned in section 3.1).</li>
	<li>Place the animal on its chest on the slotted aluminum sheet so that its head is positioned directly in the path of the brass weight (Figure 3C,D). Maintain anesthesia with the nose cone to make sure that the rat does not move or wake-up before the weight strikes it.</li>
	<li>Remove the nose cone and pull the hex key. The weight will fall vertically through the PVC tube and impact the head of the rat. The rat will undergo a rapid 180&#176; rotation and land on its back (Figure 3E).</li>
	<li>Remove the rat from the foam cushion and place it on its back in its cage.</li>
	<li>Use a digital timer to measure the righting reflex time as a sign of recovery and injury severity. The righting reflex time is the total time from the impact until the rodents wake up and spontaneously right themselves to the prone position from the supine position, or start walking. Note any signs of death, fracture, or bleeding. 
	NOTE: The procedure can be repeated on the same subject at different time points for repeated concussions.</li>
</ol>
6. Sham Induction
<ol>
	<li>After the baseline phase of the dialysis samples collection, re-anesthetize the rat lightly by placing a nose cone delivering sodium isoflurane (2.5%) at 0.5 L/min oxygen flow until it as no response to a toe pinch (as mentioned in section 3.1).</li>
	<li>Place the animal on its chest on the slotted aluminum sheet so that its head lays directly in the path of the brass weight. Maintain anesthesia with the nose cone to make sure the rat does not move or wake-up.</li>
	<li>Remove the nose cone and remove the animal from the aluminum sheet without pulling the hex key. The rat will not undergo a rapid 180&#176; rotation.</li>
	<li>Place the rat on its back in its cage.</li>
	<li>Use a digital timer to measure the righting time as an indicator of neurologic restoration.</li>
</ol>
7. High-performance Liquid Chromatography
<ol>
	<li>Determine neurotransmitter levels (i.e., glutamate and GABA) by precolumn derivatization using high-performance liquid chromatography with rapid separation fluorescence detection, and a system consisting of a rapid separation autosampler and a pump coupled to a 3.0 x 50&#8201;mm 5&#8201;&#956;m analytical column.</li>
	<li>Prepare a mobile phase with 100 mmol/L sodium phosphate dibasic (Na2HPO4), 3.5% acetonitrile and 20% methanol. Adjust the pH to 6.7 with phosphoric acid (85%) as needed.</li>
	<li>Set the flow rate to 0.5 mL/min.</li>
	<li>Prepare fresh daily derivatization reagents and working standards (100 ng/mL) from stock solutions. Load them into a refrigerated (10 &#176;C) rapid separation autosampler with samples.</li>
	<li>Mix each fraction sequentially into the analytical column with 20&#8201;&#956;L of 3-mercaptopropionic acid (0.071&#8201;mol/L) diluted with H2O and 20&#8201;&#956;L of o-phthaldehyde (0.0143&#8201;mol/L) diluted with 0.1&#8201;mol/L sodium tetraborate. Allow 10 min for the mix to react.</li>
	<li>To prevent contamination of next samples, flush the injection loop with methanol (20%), following each injection. 
	NOTE: The glutamate retention time would be of 1 min approximately in this protocol, for a total run time of 30 min for each sample.</li>
	<li>During analysis of chromatographic peaks, identify unknown peaks using samples matched according to time of retention from known standards. Express levels of analytes as &#956;g/mL.</li>
</ol>
8. Histology
<ol>
	<li>A month following the microdialysis procedure and concussion or sham injury induction, anesthetize the animals by injecting a cocktail of ketamine (70 mg/kg) and xylazine (10 mg/kg) intraperitoneally and euthanize them by paraformaldehyde (4%) and saline intracardiac perfusion.</li>
	<li>Decapitate the rodents then dissect the brains.</li>
	<li>Store the brains in paraformaldehyde (4%) and subsequently cryoprotect them in a solution of sucrose (30%).</li>
	<li>Slice the brains in coronal sections of 50 &#956;m with a cryostat.</li>
	<li>Stain the brain slices with cresyl violet for histological verification of injury and probe placement (Nissl staining).</li>
</ol>

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T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intensity-+and+interval-specific+repetitive+traumatic+brain+injury+can+evoke+both+axonal+and+microvascular+damage.">Intensity- and interval-specific repetitive traumatic brain injury can evoke both axonal and microvascular damage.</a> Journal of Neurotrauma. 29 (12), 2172-2180 (2012).</li><li>Paxinos, G., Watson, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> The Rat Brain in Stereotaxic Coordinates. , (1998).</li><li>Bales, J. W., Wagner, A. K., Kline, A. E., Dixon, C. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Persistent+cognitive+dysfunction+after+traumatic+brain+injury:+A+dopamine+hypothesis.">Persistent cognitive dysfunction after traumatic brain injury: A dopamine hypothesis.</a> Neuroscience &amp; Biobehavioral Reviews. 33 (7), 981-1003 (2009).</li><li>Sumbria, R. K., Klein, J., Bickel, U. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Acute+depression+of+energy+metabolism+after+microdialysis+probe+implantation+is+distinct+from+ischemia-induced+changes+in+mouse+brain.">Acute depression of energy metabolism after microdialysis probe implantation is distinct from ischemia-induced changes in mouse brain.</a> Neurochemical Research. 36 (1), 109-116 (2011).</li></ol>

No competing financial interests exist.

Critical steps in the protocol 
For the generation of reliable results, critical steps in this protocol require particular attention. During the cannula implantation surgery, avoid using more cement than necessary, especially when it is very liquid as to prevent spilling over the impact site. To avoid blocking the implantation site, use an obturator that is the same length as the cannula. During the microdialysis procedure, insert the probe slowly into the cannula and make sure that it is inserted completely for dialysate sampling. Before the concussion induction, make sure that the aluminum sheet is properly slotted with a sharp razor blade. Otherwise, the impact from the brass weight won&#8217;t be sufficient to rip the aluminum sheet and the rat will remain chest down instead of undergoing an 180&#176; rotation and landing on its back. If this is the case, injuries induced will result from the blunt impact, not unlike what is seen in the open-skull weight drop models and be significantly more severe. During the concussion induction, avoid impacting the cannula with the weight as this would generate critical damage to the skull of the rat. It is highly recommended to work in teams of 2 to restrict manipulation errors during the experiment.
Modifications and troubleshooting 
During the microdialysis procedure, flow should be constant and yield a volume appropriate to the rate of perfusion, once the probe is linked to the pump. Lower volumes can indicate the presence of clogging in the membrane of the probe or air bubbles in the lines. In the case of clogging, the probe should be discarded and replaced. However, air bubbles can be ejected by circulating ACSF in the lines. If there is no clogging or air bubbles noted and there is still no flow, a small part of the outflow tube nearest to the end can be cut.
Limitations of the method 
Other studies using the Wayne State University weight-drop have evaluated some fundamental structural and molecular changes18. However, a more extensive investigation would maintain the legitimacy of this procedure. Information regarding the biological and neuroanatomical changes that take place at epigenetic and cellular levels would further solidify the reliable and translational value of our method. Furthermore, evaluation of cognitive function is a reliable measure of outcome related to mTBI in rodent models33. While the time-to-right was measured in this protocol and was significantly delayed in injured cases compared to sham cases, studies in the future should concentrate on methodically measuring cognitive function following trauma induction in rodents.
Significance of the method with respect to existing/alternative methods. 
The main significance of the method is twofold: Firstly, it allows the successful induction of a concussion with the Wayne State University procedure, which allows rapid acceleration and deceleration of the head and torso. With this method, serious injury outcomes such as cardiorespiratory arrests, skull fracture, high mortality and signs of visible cerebral contusions at the impact site were avoided. Secondly, this microdialysis technique successfully replicated the previously demonstrated the acute and short-lived extracellular glutamate release taking place within the first 10 min following trauma induction14,16. Moreover, keeping the probe inserted throughout the whole procedure significantly reduces the likelihood of inducing damage to the mTBI-sensitive blood-brain barrier linked to repeated microdialysis probe insertion34.
Future applications or directions of the method. 
Given the easy-to-use aspects of the Wayne State University weight-drop procedure and the acute extracellular neurotransmitter level changes measured by microdialysis, our rat model combining microdialysis and concussion provides researchers with a reliable tool to faithfully reproduces the biomechanics of human craniocerebral trauma and longitudinally characterize the molecular effects of concussions. Our rat model could also be used in a wide variety of therapeutical studies as it offers a valuable opportunity to study the mechanism and efficacy of pharmacologic agents in vivo, continuously and without having to sacrifice the animal. Furthermore, the availability of a rat model such as the one presented here could greatly facilitate the better understanding of the relationship between the neurotransmitter imbalances and the behavioral consequences of concussions.

The purpose of this method is to provide researchers with a reliable tool that faithfully reproduces the biomechanics of human craniocerebral trauma while allowing longitudinal characterization of the molecular effects of concussions/mild traumatic brain injury (mTBIs). This method combines cerebral microdialysis with the Wayne State weight-drop model to document, in lightly anesthetized and unrestrained adult rats, the acute changes in extracellular neurotransmitter levels following concussion. With this minimally-invasive method, neurotransmitters such as glutamate, GABA, taurine, glycine and serine can be rapidly and continuously quantified following trauma, in vivo, while not having to sacrifice the animal.
Concussion/mTBI is a pathophysiological disruption that affect brain functioning caused by an external force mechanism. Concussion/mTBI is the most common form of traumatic brain injury, accounting for 70-90% of cases1. Most of the acute functional disruptions following a concussion can be attributed to a primary and a secondary brain injury2,3: (1) the primary brain injury is induced by the rapid acceleration and deceleration of the head and torso which damages brain tissues by compression followed by the stretching and shearing of axons during the backlash4,5,6 and (2) the secondary brain injury is the indirect cellular response to the trauma. It takes place hours and days after the primary brain injury and plays an important role in the motor and cognitive impairment observed over time. Many of the symptoms can be attributed to altered neurotransmission such as the previously demonstrated excessive extracellular glutamate release in the first 10 min following injury7,8,9. Given its high density of terminals and receptors, the hippocampus is a brain structure particularly vulnerable to this excitotoxic response following injury. Being heavily involved in cognitive function10,11, studies in rodents reported that hippocampal damage associated with concussion can lead to impairments in fear conditioning and learning spatial memory12,13. The primary objective of this methodology was to work out a rat model of concussion/mTBI, using the Wayne State closed head weight-drop procedure to faithfully reproduce the mechanisms of the primary brain injury, and incorporate cerebral microdialysis to study in vivo, the acute extracellular neurotransmitter changes due to the secondary brain injury following a concussion. Concentrations of extracellular glutamate and GABA were measured in the hippocampus to act as representative results of our method.
Previous rodent studies have combined microdialysis and other models of injury, such as the open-skull weight drop and controlled cortical impact, to demonstrate the acute changes in extracellular neurotransmitter levels following an injury of varying severity degrees14,15,16,17. However, in addition to the high degrees of variability, the translational value of models like the open-skull weight drop and controlled cortical impact is hampered by an inherent lack of ecological validity due to 2 factors: (1) these models induce injuries much more severe than sport-related concussions suffered in humans, involving direct brain loading and (2) these models necessitate a craniectomy or a craniotomy, the head of the rodent being completely restrained in a stereotaxic frame, impeding the rapid acceleration and deceleration of the head and torso, thus poorly reproducing the biomechanics of concussion.
Microdialysis is a minimally-invasive method which offers the clear advantage of sampling neurotransmitters such as glutamate, GABA, taurine, glycine and serine, in vivo and continuously following trauma, while not having to sacrifice the animal. In addition to the advantages offered by microdialysis, the Wayne State University developed a closed-skull weight-drop model (as opposed to open-skull from other models), which allows the induction of a mTBI on a lightly anesthetized and unrestrained rodent, thus allowing the rapid acceleration and deceleration of the head and torso18. As mentioned previously, the acceleration and deceleration of the head and torso is a core biomechanical feature of sport-related concussions seen in humans that previous rodent mTBI models have failed to address. The weight-drop procedure can be done very quickly and does not require any prior surgery or scalp incision. Following induction of the concussion, rodents recover the righting reflex almost spontaneously and do not experience paralysis, seizures or respiratory distress after a single impact. Intracranial bleedings and skull fractures are rare, and only minor deficits in motor coordination have been reported in rodents. This rat model is easy-to-use, inexpensive and facilitates the quantification of neurotransmitters released in the acute phase following a concussion without removing the microdialysis probe during the impact.
Our rat model combining microdialysis and concussion is appropriate for researchers seeking to characterize longitudinally the molecular effects of concussion and could be used in a wide variety of therapeutical studies. Indeed, despite several years of research and an overwhelming need, no drug to prevent the long-term effects of concussions has passed the clinical trial phase19. One of the potential reasons for these failures could be the use of animal models that do not faithfully reproduce the traumatic biomechanical forces of concussions as experienced by humans. The method presented here meets the definition of human concussions which specifies that the primary brain injury is induced by a blunt impact as well as rapid acceleration and deceleration of the head and torso2,3.
Furthermore, our combined model is appropriate for researchers studying the effects of repeated mild traumatic brain injury (rmTBI) since one of its key characteristics that sets it apart from other animal models of concussion is that it makes it possible to induce repeated, mild injuries to the same case18. In humans, rmTBI is associated with more severe post-traumatic symptoms, longer recovery times, and aggravated motor and cognitive impairments that tend to spread over time20,21. Other relevant animal models have also made it possible to better understand the post-traumatic pathophysiology of rmTBI22,23,24,25,26,27. Increased brain vulnerability has been demonstrated in rodents after a minimum of 5 mTBI at 24 h intervals. Neuroinflammation increases with the number of mTBI experienced and markers of neurodegeneration appear28. Repeated mTBI would prevent the transition of microglia from a proinflammatory mode to a normal mode of recovery, resulting in prolonged excitotoxic activity and activation of neurodegenerative mechanisms 29. With our model, rats could be exposed to 1 impact per day over the period of 1 week for a total of 5 exposures. Given the simplicity of this animal model, it could facilitate the characterization of the cumulative effects of the acute indiscriminate neurotransmitter release arising immediately after a mTBI.
This model also allows animals to be readily exposed to 2 impacts per day, making it possible to study even more severe conditions such as when an athlete receives another traumatic impact within a short time from the first blow30. As demonstrated in a previous study31, the timing of a second blow to the head can dramatically affect vascular and axonal damage. The closer the second blow is to the first blow, the more damaging the consequences. This model is appropriate for investigating how this particular condition affects extracellular neurotransmitter release.
In this method, the hippocampus was used as region of interest due to its relevancy in concussion research but microdialysis samples can be collected from other regions of interest as well. However, any other brain region has to be considered on account of the space left by the impact site from the guide cannula, including the dental cement surrounding it, can take up a considerable amount of space on the rat&#8217;s head. In addition to this, the microdialysis parameters presented in this method such as the membrane&#8217;s molecular weight cut-off and active length, the sampling time intervals and the flow rate can be adjusted according to the type of molecule studied. The efficient collection of pro-inflammatory cytokines involved in concussions, for example, would require a membrane with a much larger pore size.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Animal Preparation</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Sprague Dawley Rats</td>
			<td>Charles River Laboratories</td>
			<td>SAS SD 40</td>
			<td></td>
		</tr>
		<tr>
			<td>Name</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td>Comments</td>
		</tr>
		<tr>
			<td>Microdialysis Guide Cannula Implantation Surgery</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Ketamine Hydrochloride (100 mg/ml)</td>
			<td>Bioniche</td>
			<td>1989529</td>
			<td></td>
		</tr>
		<tr>
			<td>Xylazine Hydrochloride (100 mg/ml)</td>
			<td>Bimeda</td>
			<td>8XYL004C</td>
			<td></td>
		</tr>
		<tr>
			<td>Solution of Chlorhexidine Gluconate 2% and Isopropyl Alcohol 2%</td>
			<td>Carefusion</td>
			<td>260100C</td>
			<td></td>
		</tr>
		<tr>
			<td>Lidocaine Hydrochloride</td>
			<td>Alveda Pharma</td>
			<td>0122AG01</td>
			<td></td>
		</tr>
		<tr>
			<td>Bupivacaine Hydrochloride</td>
			<td>Hospira</td>
			<td>1559</td>
			<td></td>
		</tr>
		<tr>
			<td>Ophthalmic Ointment</td>
			<td>Baussh and Lomb inc.</td>
			<td>2125706</td>
			<td></td>
		</tr>
		<tr>
			<td>Stereotaxic Frame</td>
			<td>Stoelting</td>
			<td>51600</td>
			<td></td>
		</tr>
		<tr>
			<td>Stereotaxic Cannula Holder Arm</td>
			<td>Harvard Apparatus</td>
			<td>72-4837</td>
			<td></td>
		</tr>
		<tr>
			<td>Drill</td>
			<td>Dremel</td>
			<td>8050-N/18</td>
			<td></td>
		</tr>
		<tr>
			<td>Suture Thread Coated Vicryl Rapide 4-0</td>
			<td>Ethicon</td>
			<td>VR2297</td>
			<td></td>
		</tr>
		<tr>
			<td>Dental Acrylic Cement</td>
			<td>Harvard Apparatus</td>
			<td>72-6906</td>
			<td></td>
		</tr>
		<tr>
			<td>Screws</td>
			<td>JI Morris Company</td>
			<td>P0090CE125</td>
			<td></td>
		</tr>
		<tr>
			<td>Isoflurane</td>
			<td>Baxter</td>
			<td>CA2L9100</td>
			<td></td>
		</tr>
		<tr>
			<td>Cannula Gauge 20 10.55mm</td>
			<td>HRS Scientific</td>
			<td>C311G/SPC</td>
			<td></td>
		</tr>
		<tr>
			<td>Dummy-Cannula 10.55mm</td>
			<td>HRS Scientific</td>
			<td>C311DC/1/SPC</td>
			<td></td>
		</tr>
		<tr>
			<td>Name</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td>Comments</td>
		</tr>
		<tr>
			<td>Microdialysis Procedure</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>CMA 402 Syringe Pump</td>
			<td>Harvard Apparatus Canada</td>
			<td>CMA-8003110</td>
			<td></td>
		</tr>
		<tr>
			<td>Microsyringe 2.5ml Glass</td>
			<td>Harvard Apparatus Canada</td>
			<td>CMA-8309021</td>
			<td></td>
		</tr>
		<tr>
			<td>Syringe Clip Medium For 1-2.5ml</td>
			<td>Harvard Apparatus Canada</td>
			<td>CMA-3408310</td>
			<td></td>
		</tr>
		<tr>
			<td>Low-Torque Dual Channel Quartz-Lined Swivel</td>
			<td>Instech Laboratories Inc.</td>
			<td>375/D/22QM</td>
			<td></td>
		</tr>
		<tr>
			<td>GSC Cast Iron Support Ring Stand</td>
			<td>Fisher Scientifique</td>
			<td>S13748</td>
			<td></td>
		</tr>
		<tr>
			<td>Fisherbrand Castaloy Adjustable-Angle Clamps</td>
			<td>Fisher Scientifique</td>
			<td>05769Q</td>
			<td></td>
		</tr>
		<tr>
			<td>NaHCO3 Sodium Bicarbonate</td>
			<td>Sigma-Aldrich Canada</td>
			<td>S5761-500G</td>
			<td>For Artificial Cerebrospinal Fluid (aCSF)</td>
		</tr>
		<tr>
			<td>MgCl2 Magnesium Chloride</td>
			<td>Sigma-Aldrich Canada</td>
			<td>M8266-100G</td>
			<td>For Artificial Cerebrospinal Fluid (aCSF)</td>
		</tr>
		<tr>
			<td>NaCl Sodium Chloride</td>
			<td>Sigma-Aldrich Canada</td>
			<td>S7653-1KG</td>
			<td>For Artificial Cerebrospinal Fluid (aCSF)</td>
		</tr>
		<tr>
			<td>L-Ascorbic Acid</td>
			<td>Sigma-Aldrich Canada</td>
			<td>A5960-25G</td>
			<td>For Artificial Cerebrospinal Fluid (aCSF)</td>
		</tr>
		<tr>
			<td>KCl Potassium Chloride</td>
			<td>Sigma-Aldrich Canada</td>
			<td>P9333-500G</td>
			<td>For Artificial Cerebrospinal Fluid (aCSF)</td>
		</tr>
		<tr>
			<td>NaH2PO4 Sodium Phosphate Monobasic</td>
			<td>Sigma-Aldrich Canada</td>
			<td>S0751-1KG</td>
			<td>For Artificial Cerebrospinal Fluid (aCSF)</td>
		</tr>
		<tr>
			<td>CaCl2 Calcium Chloride</td>
			<td>Sigma-Aldrich Canada</td>
			<td>383147-100G</td>
			<td>For Artificial Cerebrospinal Fluid (aCSF)</td>
		</tr>
		<tr>
			<td>Lighter</td>
			<td>Canadian Tire</td>
			<td></td>
			<td>For Laboratory Constructed Probes / Available at most hardware stores</td>
		</tr>
		<tr>
			<td>Epoxy Glue</td>
			<td>Canadian Tire</td>
			<td></td>
			<td>For Laboratory Constructed Probes / Available at most hardware stores</td>
		</tr>
		<tr>
			<td>Super Glue Gel</td>
			<td>Canadian Tire</td>
			<td></td>
			<td>For Laboratory Constructed Probes / Available at most hardware stores</td>
		</tr>
		<tr>
			<td>Heat Shrink Tube 0.063&#34; Inner Diameter Gardner Bender</td>
			<td>Canadian Tire</td>
			<td></td>
			<td>For Laboratory Constructed Probes / Available at most hardware stores</td>
		</tr>
		<tr>
			<td>Cut-Off Wheels Dremel #409</td>
			<td>Canadian Tire</td>
			<td></td>
			<td>For Laboratory Constructed Probes / Available at most hardware stores</td>
		</tr>
		<tr>
			<td>BD Needle 26 Gauge 0.5 Inch PrecisionGlide Sterile 305111</td>
			<td>Fisher Scientifique</td>
			<td>14-826-15</td>
			<td>For Laboratory Constructed Probes</td>
		</tr>
		<tr>
			<td>BD Needle 21 Gauge 1.5 Inch PrecisionGlide Sterile 305167</td>
			<td>Fisher Scientifique</td>
			<td>14-826-5B</td>
			<td>For Laboratory Constructed Probes</td>
		</tr>
		<tr>
			<td>26G Stainless Steel Tubing One Foot</td>
			<td>HRS Scientific</td>
			<td>SST-26/FT</td>
			<td>For Laboratory Constructed Probes</td>
		</tr>
		<tr>
			<td>Polyethylene Tubing PE/20 .024&#34; OD X .015&#34; ID</td>
			<td>HRS Scientific</td>
			<td>C315CT</td>
			<td>For Laboratory Constructed Probes</td>
		</tr>
		<tr>
			<td>Polyethylene Tubing PE/10 .024&#34; OD X .011&#34; ID</td>
			<td>HRS Scientific</td>
			<td>C314CT</td>
			<td>For Laboratory Constructed Probes</td>
		</tr>
		<tr>
			<td>Polyethylene Tubing PE/50 .038&#34; OD X .023&#34; ID</td>
			<td>HRS Scientific</td>
			<td>C313CT</td>
			<td>For Laboratory Constructed Probes</td>
		</tr>
		<tr>
			<td>30S WIRE ST.ST 0.008X 1&#8217; Long</td>
			<td>HRS Scientific</td>
			<td>008BSH/30S</td>
			<td>For Laboratory Constructed Probes</td>
		</tr>
		<tr>
			<td>Polymicro Technologies Flexible Fused Silica Capillary Tubing Inner Diameter 50&#181;m, Outer Diameter 150&#181;m</td>
			<td>Molex LLC Polymicro Technologies</td>
			<td>106815-0015</td>
			<td>For Laboratory Constructed Probes</td>
		</tr>
		<tr>
			<td>Spectra Por 132294 Micro-Dialysis Hollow Fiber Membranes 13 kD MWCO</td>
			<td>Spectrum Labs</td>
			<td>FSSP9778671</td>
			<td>For Laboratory Constructed Probes</td>
		</tr>
		<tr>
			<td>Stainless Steel Collar</td>
			<td>Sirnay In.c</td>
			<td>304</td>
			<td>For Laboratory Constructed Probes / Custome made</td>
		</tr>
		<tr>
			<td>Name</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td>Comments</td>
		</tr>
		<tr>
			<td>Concussion Apparatus</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Brass Weight</td>
			<td>Rapido M&#233;tal Inc.</td>
			<td></td>
			<td>Attach metal loop to base</td>
		</tr>
		<tr>
			<td>Metal Loop</td>
			<td>Rona Inc.</td>
			<td></td>
			<td>Available at most hardware stores</td>
		</tr>
		<tr>
			<td>PVC Guide Tube</td>
			<td>Rona Inc.</td>
			<td></td>
			<td>Available at most hardware stores</td>
		</tr>
		<tr>
			<td>Alluminum Foil</td>
			<td>Alcan</td>
			<td></td>
			<td>Available at most grocery stores</td>
		</tr>
		<tr>
			<td>Tape</td>
			<td></td>
			<td></td>
			<td>Available commercially</td>
		</tr>
		<tr>
			<td>GSC Cast Iron Support Ring Stand</td>
			<td>Fisher Scientifique</td>
			<td>S13748</td>
			<td></td>
		</tr>
		<tr>
			<td>U-Shaped Plexiglas Frame</td>
			<td>Pr&#233;sentoirs PlexiPlus Inc.</td>
			<td></td>
			<td>Custom made</td>
		</tr>
		<tr>
			<td>Foam Cushion</td>
			<td>Mousse D&#38;R Foam Inc.</td>
			<td></td>
			<td>Custom made</td>
		</tr>
		<tr>
			<td>Razor Blades</td>
			<td>VWR International</td>
			<td>55411-055</td>
			<td></td>
		</tr>
		<tr>
			<td>Super Strong Trilene XT 20 lb. Berkley</td>
			<td>Canadian Tire</td>
			<td></td>
			<td>Available at most hardware stores</td>
		</tr>
		<tr>
			<td>Isoflurane</td>
			<td>Baxter</td>
			<td>CA2L9100</td>
			<td></td>
		</tr>
		<tr>
			<td>Stop Watch</td>
			<td></td>
			<td></td>
			<td>Available at most sporting goods retailer</td>
		</tr>
	</tbody>
</table>
<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Animal Preparation</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Sprague Dawley Rats</td>
			<td>Charles River Laboratories</td>
			<td>SAS SD 40</td>
			<td></td>
		</tr>
		<tr>
			<td>Name</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td>Comments</td>
		</tr>
		<tr>
			<td>Microdialysis Guide Cannula Implantation Surgery</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Ketamine Hydrochloride (100 mg/ml)</td>
			<td>Bioniche</td>
			<td>1989529</td>
			<td></td>
		</tr>
		<tr>
			<td>Xylazine Hydrochloride (100 mg/ml)</td>
			<td>Bimeda</td>
			<td>8XYL004C</td>
			<td></td>
		</tr>
		<tr>
			<td>Solution of Chlorhexidine Gluconate 2% and Isopropyl Alcohol 2%</td>
			<td>Carefusion</td>
			<td>260100C</td>
			<td></td>
		</tr>
		<tr>
			<td>Lidocaine Hydrochloride</td>
			<td>Alveda Pharma</td>
			<td>0122AG01</td>
			<td></td>
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			<td>Bupivacaine Hydrochloride</td>
			<td>Hospira</td>
			<td>1559</td>
			<td></td>
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			<td>Ophthalmic Ointment</td>
			<td>Baussh and Lomb inc.</td>
			<td>2125706</td>
			<td></td>
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			<td>Stereotaxic Frame</td>
			<td>Stoelting</td>
			<td>51600</td>
			<td></td>
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			<td>Stereotaxic Cannula Holder Arm</td>
			<td>Harvard Apparatus</td>
			<td>72-4837</td>
			<td></td>
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			<td>Drill</td>
			<td>Dremel</td>
			<td>8050-N/18</td>
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			<td>Suture Thread Coated Vicryl Rapide 4-0</td>
			<td>Ethicon</td>
			<td>VR2297</td>
			<td></td>
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		<tr>
			<td>Dental Acrylic Cement</td>
			<td>Harvard Apparatus</td>
			<td>72-6906</td>
			<td></td>
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		<tr>
			<td>Screws</td>
			<td>JI Morris Company</td>
			<td>P0090CE125</td>
			<td></td>
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		<tr>
			<td>Isoflurane</td>
			<td>Baxter</td>
			<td>CA2L9100</td>
			<td></td>
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		<tr>
			<td>Cannula Gauge 20 10.55mm</td>
			<td>HRS Scientific</td>
			<td>C311G/SPC</td>
			<td></td>
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			<td>Dummy-Cannula 10.55mm</td>
			<td>HRS Scientific</td>
			<td>C311DC/1/SPC</td>
			<td></td>
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			<td>Name</td>
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			<td>Catalog Number</td>
			<td>Comments</td>
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			<td>Microdialysis Procedure</td>
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			<td>CMA 402 Syringe Pump</td>
			<td>Harvard Apparatus Canada</td>
			<td>CMA-8003110</td>
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			<td>Microsyringe 2.5ml Glass</td>
			<td>Harvard Apparatus Canada</td>
			<td>CMA-8309021</td>
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			<td>Syringe Clip Medium For 1-2.5ml</td>
			<td>Harvard Apparatus Canada</td>
			<td>CMA-3408310</td>
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			<td>Low-Torque Dual Channel Quartz-Lined Swivel</td>
			<td>Instech Laboratories Inc.</td>
			<td>375/D/22QM</td>
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			<td>Fisher Scientifique</td>
			<td>S13748</td>
			<td></td>
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			<td>Fisherbrand Castaloy Adjustable-Angle Clamps</td>
			<td>Fisher Scientifique</td>
			<td>05769Q</td>
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			<td>NaHCO3 Sodium Bicarbonate</td>
			<td>Sigma-Aldrich Canada</td>
			<td>S5761-500G</td>
			<td>For Artificial Cerebrospinal Fluid (aCSF)</td>
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		<tr>
			<td>MgCl2 Magnesium Chloride</td>
			<td>Sigma-Aldrich Canada</td>
			<td>M8266-100G</td>
			<td>For Artificial Cerebrospinal Fluid (aCSF)</td>
		</tr>
		<tr>
			<td>NaCl Sodium Chloride</td>
			<td>Sigma-Aldrich Canada</td>
			<td>S7653-1KG</td>
			<td>For Artificial Cerebrospinal Fluid (aCSF)</td>
		</tr>
		<tr>
			<td>L-Ascorbic Acid</td>
			<td>Sigma-Aldrich Canada</td>
			<td>A5960-25G</td>
			<td>For Artificial Cerebrospinal Fluid (aCSF)</td>
		</tr>
		<tr>
			<td>KCl Potassium Chloride</td>
			<td>Sigma-Aldrich Canada</td>
			<td>P9333-500G</td>
			<td>For Artificial Cerebrospinal Fluid (aCSF)</td>
		</tr>
		<tr>
			<td>NaH2PO4 Sodium Phosphate Monobasic</td>
			<td>Sigma-Aldrich Canada</td>
			<td>S0751-1KG</td>
			<td>For Artificial Cerebrospinal Fluid (aCSF)</td>
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		<tr>
			<td>CaCl2 Calcium Chloride</td>
			<td>Sigma-Aldrich Canada</td>
			<td>383147-100G</td>
			<td>For Artificial Cerebrospinal Fluid (aCSF)</td>
		</tr>
		<tr>
			<td>Lighter</td>
			<td>Canadian Tire</td>
			<td></td>
			<td>For Laboratory Constructed Probes / Available at most hardware stores</td>
		</tr>
		<tr>
			<td>Epoxy Glue</td>
			<td>Canadian Tire</td>
			<td></td>
			<td>For Laboratory Constructed Probes / Available at most hardware stores</td>
		</tr>
		<tr>
			<td>Super Glue Gel</td>
			<td>Canadian Tire</td>
			<td></td>
			<td>For Laboratory Constructed Probes / Available at most hardware stores</td>
		</tr>
		<tr>
			<td>Heat Shrink Tube 0.063&#34; Inner Diameter Gardner Bender</td>
			<td>Canadian Tire</td>
			<td></td>
			<td>For Laboratory Constructed Probes / Available at most hardware stores</td>
		</tr>
		<tr>
			<td>Cut-Off Wheels Dremel #409</td>
			<td>Canadian Tire</td>
			<td></td>
			<td>For Laboratory Constructed Probes / Available at most hardware stores</td>
		</tr>
		<tr>
			<td>BD Needle 26 Gauge 0.5 Inch PrecisionGlide Sterile 305111</td>
			<td>Fisher Scientifique</td>
			<td>14-826-15</td>
			<td>For Laboratory Constructed Probes</td>
		</tr>
		<tr>
			<td>BD Needle 21 Gauge 1.5 Inch PrecisionGlide Sterile 305167</td>
			<td>Fisher Scientifique</td>
			<td>14-826-5B</td>
			<td>For Laboratory Constructed Probes</td>
		</tr>
		<tr>
			<td>26G Stainless Steel Tubing One Foot</td>
			<td>HRS Scientific</td>
			<td>SST-26/FT</td>
			<td>For Laboratory Constructed Probes</td>
		</tr>
		<tr>
			<td>Polyethylene Tubing PE/20 .024&#34; OD X .015&#34; ID</td>
			<td>HRS Scientific</td>
			<td>C315CT</td>
			<td>For Laboratory Constructed Probes</td>
		</tr>
		<tr>
			<td>Polyethylene Tubing PE/10 .024&#34; OD X .011&#34; ID</td>
			<td>HRS Scientific</td>
			<td>C314CT</td>
			<td>For Laboratory Constructed Probes</td>
		</tr>
		<tr>
			<td>Polyethylene Tubing PE/50 .038&#34; OD X .023&#34; ID</td>
			<td>HRS Scientific</td>
			<td>C313CT</td>
			<td>For Laboratory Constructed Probes</td>
		</tr>
		<tr>
			<td>30S WIRE ST.ST 0.008X 1&#8217; Long</td>
			<td>HRS Scientific</td>
			<td>008BSH/30S</td>
			<td>For Laboratory Constructed Probes</td>
		</tr>
		<tr>
			<td>Polymicro Technologies Flexible Fused Silica Capillary Tubing Inner Diameter 50&#181;m, Outer Diameter 150&#181;m</td>
			<td>Molex LLC Polymicro Technologies</td>
			<td>106815-0015</td>
			<td>For Laboratory Constructed Probes</td>
		</tr>
		<tr>
			<td>Spectra Por 132294 Micro-Dialysis Hollow Fiber Membranes 13 kD MWCO</td>
			<td>Spectrum Labs</td>
			<td>FSSP9778671</td>
			<td>For Laboratory Constructed Probes</td>
		</tr>
		<tr>
			<td>Stainless Steel Collar</td>
			<td>Sirnay In.c</td>
			<td>304</td>
			<td>For Laboratory Constructed Probes / Custome made</td>
		</tr>
		<tr>
			<td>Name</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td>Comments</td>
		</tr>
		<tr>
			<td>Concussion Apparatus</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Brass Weight</td>
			<td>Rapido M&#233;tal Inc.</td>
			<td></td>
			<td>Attach metal loop to base</td>
		</tr>
		<tr>
			<td>Metal Loop</td>
			<td>Rona Inc.</td>
			<td></td>
			<td>Available at most hardware stores</td>
		</tr>
		<tr>
			<td>PVC Guide Tube</td>
			<td>Rona Inc.</td>
			<td></td>
			<td>Available at most hardware stores</td>
		</tr>
		<tr>
			<td>Alluminum Foil</td>
			<td>Alcan</td>
			<td></td>
			<td>Available at most grocery stores</td>
		</tr>
		<tr>
			<td>Tape</td>
			<td></td>
			<td></td>
			<td>Available commercially</td>
		</tr>
		<tr>
			<td>GSC Cast Iron Support Ring Stand</td>
			<td>Fisher Scientifique</td>
			<td>S13748</td>
			<td></td>
		</tr>
		<tr>
			<td>U-Shaped Plexiglas Frame</td>
			<td>Pr&#233;sentoirs PlexiPlus Inc.</td>
			<td></td>
			<td>Custom made</td>
		</tr>
		<tr>
			<td>Foam Cushion</td>
			<td>Mousse D&#38;R Foam Inc.</td>
			<td></td>
			<td>Custom made</td>
		</tr>
		<tr>
			<td>Razor Blades</td>
			<td>VWR International</td>
			<td>55411-055</td>
			<td></td>
		</tr>
		<tr>
			<td>Super Strong Trilene XT 20 lb. Berkley</td>
			<td>Canadian Tire</td>
			<td></td>
			<td>Available at most hardware stores</td>
		</tr>
		<tr>
			<td>Isoflurane</td>
			<td>Baxter</td>
			<td>CA2L9100</td>
			<td></td>
		</tr>
		<tr>
			<td>Stop Watch</td>
			<td></td>
			<td></td>
			<td>Available at most sporting goods retailer</td>
		</tr>
	</tbody>
</table>

a novel and translational rat model of concussion combining force and rotation with in vivo cerebral microdialysis

Persistent cognitive and motor symptoms are known consequences of concussions/mild traumatic brain injury (mTBIs) that can be partly attributable to altered neurotransmission. Indeed, cerebral microdialysis studies in rodents have demonstrated an excessive extracellular glutamate release in the hippocampus within the first 10 min following trauma. Microdialysis offers the clear advantage of in vivo neurotransmitter continuous sampling while not having to sacrifice the animal. In addition to the aforementioned technique, a closed head injury model that exerts rapid acceleration and deceleration of the head and torso is needed, as such a factor is not available in many other animal models. The Wayne State weight-drop model mimics this essential component of human craniocerebral trauma, allowing the induction of an impact on the head of an unrestrained rodent with a falling weight. Our novel and translational rat model combines cerebral microdialysis with the Wayne State weight-drop model to study, in lightly anesthetized and unrestrained adult rats, the acute changes in extracellular neurotransmitter levels following concussion. In this protocol, the microdialysis probe was inserted inside the hippocampus as region of interest, and was left inserted in the brain at impact. There is a high density of terminals and receptors in the hippocampus, making it a relevant region to document altered neurotransmission following concussion. When applied to adult Sprague-Dawley rats, our combined model induced increases in hippocampal extracellular glutamate concentrations within the first 10 min, consistent with the previously reported post-concussion symptomology. This combined weight-drop model provides a reliable tool for researchers to study early therapeutic responses to concussions in addition to repetitive brain injury, since this protocol induces a closed-head mild trauma.

Using our model of concussion which combines force and rotation with in vivo cerebral microdialysis, the acute extracellular glutamate and GABA changes over time following a concussion or sham injury were investigated in 21 male, adult, Sprague-Dawley rats by the implantation of a guide cannula in the CA1 region of the hippocampus.
Histological verification of probe placement and injury 
No morphological changes such as massive intracerebral hemorrhages or contusions were reported following the histological verification of hippocampus tissue damage on sections stained with cresyl violet. Guide cannula implantation and microdialysis probe insertion induced minor and similar damages between injured and sham cases. Moreover, not removing the probe right before sham injury or concussion induction did not yield any distinguishable hippocampus tissue damage as seen under a microscope (Figure 2B,C, respectively), with the membrane of the probe still intact afterwards (Figure 2D,E). Concussion and sham injury brains perfused with paraformaldehyde (4%) 1 month following microdialysis procedures are indistinguishable upon visual inspection (Figure 2F,G).
Righting reflex time 
Animals from the injured group had a significantly increased righting time on average versus sham cases (Student&#8217;s t-test, p = 0.042801) (Figure 4) and appeared stunned upon regaining consciousness. Of the 10 cases from the concussion group, a single animal showed minor signs of bleeding under the impact site following the weight-drop. No other signs of skull fracture or intracranial bleeding were observed.
In vivo cerebral microdialysis 
To act as representative results of our method, fifteen 10 &#956;L samples of dialysate were extracted from the hippocampus, in vivo, at intervals of 10 min and a flow rate of 1 &#956;L/min. Extracellular levels of glutamate and GABA were measured from 6 samples during baseline (60 min) and from 9 samples following induction of sham injury or concussion (90 min).
Extracellular concentrations of glutamate 
Significant increases in extracellular glutamate concentrations were observed in the CA1 region of the hippocampus during the first 10 min following induction of trauma compared to sham injury (Mann-Whitney U Test, p = 0.009175) (Figure 5). No other difference in glutamate concentrations were observed between groups at any other time point.
Extracellular concentrations of GABA 
No significant change in GABA concentrations were observed in the CA1 region of the hippocampus during the first 10 min following induction of trauma compared to sham injury (Mann-Whitney U Test, p = 0.943861) (Figure 6). There was no other significant difference in GABA concentrations at any other time point between concussion cases and sham injury cases.

<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/59585/59585fig1.jpg" /> 
Figure 1: Schematic outline of the research protocol. This figure has been modified from IO Masse 2018. <a href="https://www.jove.com/files/ftp_upload/59585/59585fig1large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/59585/59585fig2.jpg" /> 
Figure 2: Histological verification of probe placement and injury. (A)&#160;Coronal view of the microdialysis probe and guide cannula placement site in the hippocampus using the stereotaxic atlas of Paxinos and Watson. (B)&#160;Representative photomicrograph of hippocampus tissue damage (cresyl violet) produced by a microdialysis probe and guide cannula from a sham injury case. (C)&#160;Representative photomicrograph of hippocampus tissue damage (cresyl violet) produced by a microdialysis probe and guide cannula from a concussion case. (D)&#160;Representative photomicrograph of a microdialysis probe before induction of concussion. (E)&#160;Representative photomicrograph of a microdialysis probe after induction of concussion. The membrane is still intact. (F-G). Representative photomicrograph of a sham (F) and concussion (G) injured brain following perfusion with 4% paraformaldehyde at 1-month after sham injury or concussion procedure. Upon visual inspection, the 2 brains are indistinguishable. This figure has been modified from IO Masse 2018. <a href="https://www.jove.com/files/ftp_upload/59585/59585fig2large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 3" class="xfigimg" src="/files/ftp_upload/59585/59585fig3.jpg" /> 
Figure 3: Concussion apparatus and microdialysis instruments essential components depictions. (A)&#160;A photograph of the entire assembly comprised of a vertical polyvinyl chloride (PVC) guide tube for the falling weight situated above the rat stage, Plexiglas frame, foam cushion, computer-controlled microinfusion pump, gastight syringes, liquid swivels, and side-by-side fused silica inlet-outlet lines. (B)&#160;Schematic representation of the Plexiglas frame and foam cushion with all pertinent dimensions. (C)&#160;A photograph of the slotted piece of aluminum foil that serves as the rat stage above the foam cushion. (D)&#160;A photograph showing the positioning of the rat on the stage immediately prior to head impact by the falling weight. (E)&#160;A photograph showing the rat after head impact, illustrating the 180&#176; horizontal rotation of the body of the rat after the head impact and ensuing acceleration and rotation. This figure has been modified from IO Masse 2018. <a href="https://www.jove.com/files/ftp_upload/59585/59585fig3large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 4" class="xfigimg" src="/files/ftp_upload/59585/59585fig4.jpg" /> 
Figure 4: Righting time. Histogram representations of the time taken by rats to wake from the anesthetic and flip from the supine position to the prone position or begin walking following concussion (red diamonds, n = 10) or sham injury (blue squares, n = 11). Rats from the concussion group took significantly longer to right themselves compared to the sham injury group. Mean values are represented as a horizontal line in each graph. * p &#60; 0.05, ** p &#60; 0.01, *** p &#60; 0.001. This figure has been modified from IO Masse 2018. <a href="https://www.jove.com/files/ftp_upload/59585/59585fig4large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 5" class="xfigimg" src="/files/ftp_upload/59585/59585fig5.jpg" /> 
Figure 5: Extracellular concentrations of glutamate. Mean extracellular concentrations of glutamate (&#956;g/mL) measured by microdialysis in the hippocampus during baseline (60 min) and after concussion (red diamonds, n = 10) or sham injury (blue squares, n = 11) conditions (90 min). Error bars represent the standard error of mean. * P &#60; 0.05, ** P &#60; 0.01, *** P &#60; 0.001. This figure has been modified from IO Masse 2018. <a href="https://www.jove.com/files/ftp_upload/59585/59585fig5large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 6" class="xfigimg" src="/files/ftp_upload/59585/59585fig6.jpg" /> 
Figure 6: Extracellular concentrations of GABA. Mean extracellular concentrations of GABA (&#956;g/mL) measured by microdialysis in the hippocampus during baseline (60 min) and after concussion (red diamonds, n = 10) or sham injury (blue squares, n = 11) conditions (90 min). Error bars represent the standard error of mean. * P &#60; 0.05, ** P &#60;&#160;0.01, *** P &#60; 0.001. This figure has been modified from IO Masse 2018. <a href="https://www.jove.com/files/ftp_upload/59585/59585fig6large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>

Watch this Scientific Journal Video about A Novel and Translational Rat Model of Concussion Combining Force and Rotation with In Vivo Cerebral Microdialysis at JoVE.com

A Novel and Translational Rat Model of Concussion Combining Force and Rotation with In Vivo Cerebral Microdialysis

Neurotransmitter alteration is a mechanism of neural dysfunction that occurs after concussion and contributes to the sometimes-catastrophic long-term consequences. This rat model combines microdialysis, allowing in vivo neurotransmitter quantification, with a weight-drop technique exerting rapid acceleration and deceleration of the head and torso, an important factor of human craniocerebral trauma.

Persistent cognitive and motor symptoms are known consequences of concussions/mild traumatic brain injury (mTBIs) that can be partly attributable to ...

a-novel-translational-rat-model-concussion-combining-force-rotation

Research

JoVE Journal

Neuroscience

8.7K Views.  Hôpital du Sacré-Cœur de Montréal. Neurotransmitter alteration is a mechanism of neural dysfunction that occurs after concussion and contributes to the sometimes-catastrophic long-term consequences. This rat model combines microdialysis, allowing in vivo neurotransmitter quantification, with a weight-drop technique exerting rapid acceleration and deceleration of the head and torso, an important factor of human craniocerebral trauma.

Video: A Novel and Translational Rat Model of Concussion Combining Force and Rotation with In Vivo Cerebral Microdialysis

Focal Cerebral Ischemia Model by Endovascular Suture Occlusion of the Middle Cerebral Artery in the Rat

Stroke is the leading cause of disability and the third leading cause of death in adults worldwide1. In human stroke, there exists a highly variable clinical state; in the development of animal models of focal ischemia, however, achieving reproducibility of experimentally induced infarct volume is essential. The rat is a widely used animal model for stroke due to its relatively low animal husbandry costs and to the similarity of its cranial circulation to that of humans2,3. In humans, the middle cerebral artery (MCA) is most commonly affected in stroke syndromes and multiple methods of MCA occlusion (MCAO) have been described to mimic this clinical syndrome in animal models. Because recanalization commonly occurs following an acute stroke in the human, reperfusion after a period of occlusion has been included in many of these models. In this video, we demonstrate the transient endovascular suture MCAO model in the spontaneously hypertensive rat (SHR). A filament with a silicon tip coating is placed intraluminally at the MCA origin for 60 minutes, followed by reperfusion. Note that the optimal occlusion period may vary in other rat strains, such as Wistar or Sprague-Dawley. Several behavioral indicators of stroke in the rat are shown. Focal ischemia is confirmed using T2-weighted magnetic resonance images and by staining brain sections with 2,3,5-triphenyltetrazolium chloride (TTC) 24 hours after MCAO.

Surgical induction of ischemic brain damage in the rat is a widely used model for stroke research. Here we demonstrate the induction of focal cerebral ischemia by occlusion of the middle cerebral artery. Visualization of the resulting infarct by histological staining and magnetic resonance imaging is also shown.

Stroke is the leading cause of disability and the third leading cause of death in adults worldwide1. In human stroke, there exists a highly variable ...

Combining Behavioral Endocrinology and Experimental Economics: Testosterone and Social Decision Making

Behavioral endocrinological research in humans as well as in animals suggests that testosterone plays a key role in social interactions. Studies in rodents have shown a direct link between testosterone and aggressive behavior1 and folk wisdom adapts these findings to humans, suggesting that testosterone induces antisocial, egoistic or even aggressive behavior2. However, many researchers doubt a direct testosterone-aggression link in humans, arguing instead that testosterone is primarily involved in status-related behavior3,4. As a high status can also be achieved by aggressive and antisocial means it can be difficult to distinguish between anti-social and status seeking behavior.
We therefore set up an experimental environment, in which status can only be achieved by prosocial means. In a double-blind and placebo-controlled experiment, we administered a single sublingual dose of 0.5 mg of testosterone (with a hydroxypropyl-&beta;-cyclodextrin carrier) to 121 women and investigated their social interaction behavior in an economic bargaining paradigm. Real monetary incentives are at stake in this paradigm; every player A receives a certain amount of money and has to make an offer to another player B on how to share the money. If B accepts, she gets what was offered and player A keeps the rest. If B refuses the offer, nobody gets anything. A status seeking player A is expected to avoid being rejected by behaving in a prosocial way, i.e. by making higher offers.
The results show that if expectations about the hormone are controlled for, testosterone administration leads to a significant increase in fair bargaining offers compared to placebo. The role of expectations is reflected in the fact that subjects who report that they believe to have received testosterone make lower offers than those who say they believe that they were treated with a placebo. These findings suggest that the experimental economics approach is sensitive for detecting neurobiological effects as subtle as those achieved by administration of hormones. Moreover, the findings point towards the importance of both psychosocial as well as neuroendocrine factors in determining the influence of testosterone on human social behavior.

The procedure described in this protocol shows that testosterone administration and folk beliefs about testosterone may be associated with directly opposed social behaviors.

Behavioral endocrinological research in humans as well as in animals suggests that testosterone plays a key role in social interactions. Studies in ...

Dual Electrophysiological Recordings of Synaptically-evoked Astroglial and Neuronal Responses in Acute Hippocampal Slices

Astrocytes form together with neurons tripartite synapses, where they integrate and modulate neuronal activity. Indeed, astrocytes sense neuronal inputs through activation of their ion channels and neurotransmitter receptors, and process information in part through activity-dependent release of gliotransmitters. Furthermore, astrocytes constitute the main uptake system for glutamate, contribute to potassium spatial buffering, as well as to GABA clearance. These cells therefore constantly monitor synaptic activity, and are thereby sensitive indicators for alterations in synaptically-released glutamate, GABA and extracellular potassium levels. Additionally, alterations in astroglial uptake activity or buffering capacity can have severe effects on neuronal functions, and might be overlooked when characterizing physiopathological situations or knockout mice. Dual recording of neuronal and astroglial activities is therefore an important method to study alterations in synaptic strength associated to concomitant changes in astroglial uptake and buffering capacities. Here we describe how to prepare hippocampal slices, how to identify stratum radiatum astrocytes, and how to record simultaneously neuronal and astroglial electrophysiological responses. Furthermore, we describe how to isolate pharmacologically the synaptically-evoked astroglial currents.

The preparation of acute brain slices from isolated hippocampi, as well as the simultaneous electrophysiological recordings of astrocytes and neurons in stratum radiatum during stimulation of schaffer collaterals is described. The pharmacological isolation of astroglial potassium and glutamate transporter currents is demonstrated.

Astrocytes form together with neurons tripartite synapses, where they integrate and modulate neuronal activity. Indeed, astrocytes sense neuronal inputs ...

Hippocampal Insulin Microinjection and In vivo Microdialysis During Spatial Memory Testing

Glucose metabolism is a useful marker for local neural activity, forming the basis of methods such as 2-deoxyglucose and functional magnetic resonance imaging. However, use of such methods in animal models requires anesthesia and hence both alters the brain state and prevents behavioral measures. An alternative method is the use of in vivo microdialysis to take continuous measurement of brain extracellular fluid concentrations of glucose, lactate, and related metabolites in awake, unrestrained animals. This technique is especially useful when combined with tasks designed to rely on specific brain regions and/or acute pharmacological manipulation; for example, hippocampal measurements during a spatial working memory task (spontaneous alternation) show a dip in extracellular glucose and rise in lactate that are suggestive of enhanced glycolysis1-3,4-5, and intrahippocampal insulin administration both improves memory and increases hippocampal glycolysis6. Substances such as insulin can be delivered to the hippocampus via the same microdialysis probe used to measure metabolites. The use of spontaneous alternation as a measure of hippocampal function is designed to avoid any confound from stressful motivators (e.g. footshock), restraint, or rewards (e.g. food), all of which can alter both task performance and metabolism; this task also provides a measure of motor activity that permits control for nonspecific effects of treatment. Combined, these methods permit direct measurement of the neurochemical and metabolic variables regulating behavior.

Hippocampal Insulin Microinjection and In vivo Microdialysis During Spatial Memory Testing

Modulation of hippocampally-dependent spatial working memory by direct intrahippocampal microinjection, accompanied and followed by in vivo microdialysis for metabolites in conscious, behaving animals.

Glucose metabolism is a useful marker for local neural activity, forming the basis of methods such as 2-deoxyglucose and functional magnetic resonance ...

Utilizing 3D Printing Technology to Merge MRI with Histology: A Protocol for Brain Sectioning

Magnetic resonance imaging (MRI) allows for the delineation between normal and abnormal tissue on a macroscopic scale, sampling an entire tissue volume three-dimensionally. While MRI is an extremely sensitive tool for detecting tissue abnormalities, association of signal changes with an underlying pathological process is usually not straightforward. In the central nervous system, for example, inflammation, demyelination, axonal damage, gliosis, and neuronal death may all induce similar findings on MRI. As such, interpretation of MRI scans depends on the context, and radiological-histopathological correlation is therefore of the utmost importance. Unfortunately, traditional pathological sectioning of brain tissue is often imprecise and inconsistent, thus complicating the comparison between histology sections and MRI. This article presents novel methodology for accurately sectioning primate brain tissues and thus allowing precise matching between histology and MRI. The detailed protocol described in this article will assist investigators in applying this method, which relies on the creation of 3D printed brain slicers. Slightly modified, it can be easily implemented for brains of other species, including humans.

The overall goal of this protocol is to accurately align magnetic resonance imaging (MRI) image volumes with histology sections via the creation of customized 3D-printed brain holders and slicer boxes.

Magnetic resonance imaging (MRI) allows for the delineation between normal and abnormal tissue on a macroscopic scale, sampling an entire tissue volume ...

A Rat Model of Mild Intrauterine Hypoperfusion with Microcoil Stenosis

Intrauterine hypoperfusion/ischemia is one of the major causes of intrauterine/fetal growth restriction, preterm birth, and low birth weight. Most studies of this phenomenon have been performed in either models with severe intrauterine ischemia or models with gradient degree of intrauterine hypoperfusion. No study has been performed in a model on uniform mild intrauterine hypoperfusion (MIUH). Two models have been used for studies of MIUH: a model based on suture ligation of either side of the arterial arcade formed with the uterine and ovarian arteries, and a transient model based on clipping the bilateral ovarian arteries and aorta having patency. Those two rodent models of MIUH have some limitations, e.g., not all fetuses are subjected to MIUH, depending on their position in the uterine horn. In our MIUH model, all fetuses are subjected to a comparable level of intrauterine hypoperfusion. MIUH was achieved by mild stenosis of all four arteries feeding the uterus, i.e., the bilateral uterine and ovarian arteries.
Arterial stenosis was induced by metal microcoils wrapped around the feeding arteries. Producing arterial stenosis with microcoils allowed us to control, optimize, and reproduce decreased blood flow with very little inter-animal variability and a low mortality rate, thus enabling accurate evaluation. When microcoils with an inner diameter of 0.24 mm were used, the blood flow in both the placenta and fetus was mildly decreased (approximately 30% from the pre-stenosis level in the placenta). The offspring of our MIUH model clearly demonstrates long-lasting alterations in neurological, neuroanatomical and behavioral test results.

Mild intrauterine hypoperfusion was produced by artery stenosis with metal microcoils wrapped around the uterine and ovarian arteries in rats at embryonic day 17. This procedure produced prenatal hypoperfusion and intrauterine growth restriction.

Intrauterine hypoperfusion/ischemia is one of the major causes of intrauterine/fetal growth restriction, preterm birth, and low birth weight. Most studies ...

A Novel In Vitro Model of Blast Traumatic Brain Injury

Traumatic brain injury is a leading cause of death and disability in military and civilian populations. Blast traumatic brain injury results from the detonation of explosive devices, however, the mechanisms that underlie the brain damage resulting from blast overpressure exposure are not entirely understood and are believed to be unique to this type of brain injury. Preclinical models are crucial tools that contribute to better understand blast-induced brain injury. A novel in vitro blast TBI model was developed using an open-ended shock tube to simulate real-life open-field blast waves modelled by the Friedlander waveform. C57BL/6N mouse organotypic hippocampal slice cultures were exposed to single shock waves and the development of injury was characterized up to 72 h using propidium iodide, a well-established fluorescent marker of cell damage that only penetrates cells with compromised cellular membranes. Propidium iodide fluorescence was significantly higher in the slices exposed to a blast wave when compared with sham slices throughout the duration of the protocol. The brain tissue injury is very reproducible and proportional to the peak overpressure of the shock wave applied.

A Novel In Vitro Model of Blast Traumatic Brain Injury

This paper describes a novel model of primary blast traumatic brain injury. A compressed-air driven shock tube is used to expose in vitro mouse hippocampal slice cultures to a single shock wave. This is a simple and rapid protocol generating a reproducible brain tissue injury with a high throughput.

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

Combining Behavior and EEG to Study the Effects of Mindfulness Meditation on Episodic Memory

Although there has been recent interest in how mindfulness meditation can affect episodic memory as well as brain structure and function, no study has examined the behavioral and neural effects of mindfulness meditation on episodic memory. Here we present a protocol that combines mindfulness meditation training, an episodic memory task, and EEG to examine how mindfulness meditation changes behavioral performance and the neural correlates of episodic memory. Subjects in a mindfulness meditation experimental group were compared to a waitlist control group. Subjects in the mindfulness meditation experimental group spent four weeks training and practicing mindfulness meditation. Mindfulness was measured before and after training using the Five Facet Mindfulness Questionnaire (FFMQ). Episodic memory was measured before and after training using a source recognition task. During the retrieval phase of the source recognition task, EEG was recorded. The results showed that mindfulness, source recognition behavioral performance, and EEG theta power in right frontal and left parietal channels increased following mindfulness meditation training. In addition, increases in mindfulness correlated with increases in theta power in right frontal channels. Therefore, results obtained from combining mindfulness meditation training, an episodic memory task, and EEG reveal the behavioral and neural effects of mindfulness meditation on episodic memory.

Here we present a protocol for combining mindfulness meditation training, an episodic memory task, and EEG to understand the behavioral and neural effects of mindfulness meditation on episodic memory.

Although there has been recent interest in how mindfulness meditation can affect episodic memory as well as brain structure and function, no study has ...

A Rat Model of EcoHIV Brain Infection

It has been well studied that the EcoHIV infected mouse model is of significant utility in investigating HIV associated neurological complications. Establishment of the EcoHIV infected rat model for studies of drug abuse and neurocognitive disorders, would be beneficial in the study of neuroHIV and HIV-1 associated neurocognitive disorders (HAND). In the present study, we demonstrate the successful creation of a rat model of active HIV infection using chimeric HIV (EcoHIV). First, the lentiviral construct of EcoHIV was packaged in cultured 293 FT cells for 48 hours. Then, the conditional medium was concentrated and titered. Next, we performed bilateral stereotaxic injections of the EcoHIV-EGFP into F344/N rat brain tissue. One week after infection, EGFP fluorescence signals were detected in the infected brain tissue, indicating that EcoHIV successfully induces an active HIV infection in rats. In addition, immunostaining for the microglial cell marker, Iba1, was performed. The results indicated that microglia were the predominant cell type harboring EcoHIV. Furthermore, EcoHIV rats exhibited alterations in temporal processing, a potential underlying neurobehavioral mechanism of HAND as well as synaptic dysfunction eight weeks after infection. Collectively, the present study extends the EcoHIV model of HIV-1 infection to the rat offering a valuable biological system to study HIV-1 viral reservoirs in the brain as well as HAND and associated comorbidities such as drug abuse.

Here, we present a protocol to establish a new rat model of active HIV infection using chimeric HIV (EcoHIV), which is critical for enhancing our understanding of HIV-1 viral reservoirs in the brain and offering a system to study HIV-associated neurocognitive disorders and associated comorbidities (i.e., drug abuse).

It has been well studied that the EcoHIV infected mouse model is of significant utility in investigating HIV associated neurological complications. ...

Construction and Implementation of Carbon Fiber Microelectrode Arrays for Chronic and Acute In Vivo Recordings

Multichannel electrode arrays offer insight into the working brain and serve to elucidate neural processes at the single-cell and circuit levels. Development of these tools is crucial for understanding complex behaviors and cognition and for advancing clinical applications. However, it remains a challenge to densely record from cell populations stably and continuously over long time periods. Many popular electrodes, such as tetrodes and silicon arrays, feature large cross-diameters that produce damage upon insertion and elicit chronic reactive tissue responses associated with neuronal death, hindering the recording of stable, continuous neural activity. In addition, most wire bundles exhibit broad spacing between channels, precluding simultaneous recording from a large number of cells clustered in a small area. The carbon fiber microelectrode arrays described in this protocol offer an accessible solution to these concerns. The study provides a detailed method for fabricating carbon fiber microelectrode arrays that can be used for both acute and chronic recordings in vivo. The physical properties of these electrodes make them ideal for stable and continuous long-term recordings at high cell densities, enabling the researcher to make robust, unambiguous recordings from single units across months.

Construction and Implementation of Carbon Fiber Microelectrode Arrays for Chronic and Acute In Vivo Recordings

This protocol describes a procedure for constructing carbon fiber microelectrode arrays for chronic and acute in vivo electrophysiological recordings in mouse (Mus musculus) and ferret (Mustela putorius furo) from multiple brain regions. Each step, following the purchase of raw carbon fibers to microelectrode array implantation, is described in detail, with emphasis on microelectrode array construction.