This work was made possible with funding from the Canadian Institutes of Health Research (CIHR) and the authors would like to thank CIHR for their funding support.

The appropriate institutional animal ethics committee should approve all of the behavioral procedures prior to starting experimentation. All animal work described here was approved by Western University Animal Use Subcommittee and follows the Canadian Council on Animal Care guidelines.&#160;These animal experiments were performed during the light phase.
1. Cylinder Task for Gross Forelimb Motor Assessment
<ol>
	<li>Equipment Setup
	<ol>
		<li>Acquire a Plexiglas cylinder with a perforated lid that will accommodate the size of the rat used for the experiment. The size of the cylinder should not allow the rat to reach the top when rearing and it should allow for 2 cm between the wall and the rat&#39;s nose and the base of the rat&#39;s tail. The standard size of a cylinder is 23 cm in diameter and 40 cm in height for a 6-month-old (400-600 g) rat.</li>
		<li>Place a mirror at a 45&#176; angle below the cylinder apparatus with the cylinder sitting on a Plexiglas stand or some other form of support approximately 30 cm above table top.</li>
		<li>Set up a video camera on a tripod at an appropriate distance to visualize the entire diameter of the cylinder in the mirror on the video camera.</li>
		<li>Turn lighting to a dimmer setting and play white noise in the room (i.e. low volume, subtle music) to reduce the effect of sudden loud noises. This should help encourage movement and prevent freezing due to loud noises.</li>
	</ol>
	</li>
	<li>Experimental Procedure
	<ol>
		<li>Move the rats into the experimental room 30 min before the start of first trial to acclimatize rats to the room with music playing and dimmer lighting.</li>
		<li>Write the corresponding animal and trial information (i.e. animal number, day in timeline and trial number) on a small white board. Place this board in front of the mirror.</li>
		<li>Press record on the video camera with the white board in front of the mirror, pick up the rat near the base of the tail, place the rat in the cylinder and secure the lid. Stand off to the side of the cylinder to avoid interfering with video recording.</li>
		<li>Remove the white board from in front of the mirror and let the video camera record the rat in the cylinder for 5 minutes (use a timer to record 5 min from when the white board is removed). This is one trial.</li>
		<li>Clean the cylinder with paper towel and water after a trial.</li>
		<li>Repeat steps 1.2.2-1.2.5 two more times to achieve a total of three trials. There should be a 20-30 min inter-trial time to decrease the chance of habituation to the cylinder. During this time, run cylinder trials for the other rats.</li>
	</ol>
	</li>
	<li>Video Analysis
	<ol>
		<li>Import video camera files into video editing program (i.e. iMovie)</li>
		<li>Compile the videos into clips for each trial. Mute the volume of the video and decrease the speed of the video to 25% of the original.</li>
		<li>Count the number of forelimb contacts with the wall of the cylinder for the left and right forelimb. For simultaneous left and right contacts, count these forelimb contacts as &#34;both&#34;. Keep in mind that the video is recorded through a mirror, what appears to be the left forelimb in the video corresponds to the animal&#39;s right forelimb in reality.</li>
		<li>Calculate the percent use of the affected forelimb (contralateral to the stroke) using the following equation: [{(affected contacts + &#189; bilateral contacts)/total number contacts} x 100].&#160;The performance of both wild type groups and the transgenic group without stroke are considered comparison groups to evaluate presentation of disease-induced problems.</li>
	</ol>
	</li>
</ol>
2. Beam-walk Task for Gross Motor Assessment
<ol>
	<li>Equipment Setup
	<ol>
		<li>Acquire a smooth sealed&#160;wooden beam that is 2 cm wide and approximately 120 cm long (optimal width for a 200-600 g rat).</li>
		<li>Place two tables or shelving units 100 cm apart. The surfaces of each unit should be approximately 40 cm above the ground.</li>
		<li>Secure both ends of the beam to the table or shelving surface using tape. Approximately 1 m of unsupported beam length should now be elevated 40 cm above the ground.</li>
		<li>Set up a video camera on a stand, capturing the entire length of the beam. To enhance contrast in the video, consider introducing a black background behind the beam when using white rats.</li>
	</ol>
	</li>
	<li>Experimental Procedure
	<ol>
		<li>Move the rats into the experimental room 30 min before the start of first trial to acclimatize rats to the room with music playing and dimmer lighting.</li>
		<li>Place the rat&#39;s home cage or environmental enrichment tubes at one end of the beam and place the rat at the other end of the beam.</li>
		<li>Conduct a few non-recorded runs two days prior to experimental trials. Let the rat explore the area and guide the rat toward the beam by holding the base of the rat&#39;s tail.</li>
		<li>Once the rat has crossed the beam, move the cage or tube to the other end of the beam and repeat. When the rat crosses the beam freely in either direction, the training session is over. Keep the number of training runs consistent among all animals.</li>
		<li>Write the corresponding animal and trial information (i.e. animal number, day in timeline and trial number) on a small white board. Tape this white board to the wall behind the beam.</li>
		<li>Place the rat&#39;s home cage or environment enrichment tubing at one end of the beam.</li>
		<li>Press record on the video camera and pick the rat up by the base of the tail and place at the end of the beam opposite to the home cage or tubing.</li>
		<li>Record the entirety of the trial, which ends when the rat has successfully completed traversing the full length of the elevated beam. If the rat pauses midway across the beam, gently scruff the rat by the base of the tail or touch the rat&#39;s tail gently to promote movement across the beam. Do not push the rat forward in any manner.</li>
		<li>Re-do trials where the rat turns sideways while on the beam, repeatedly stops walking or walks inconsistently. If a rat falls and continues to hang on to the beam, gently scoop up the rat and place it back on the beam at the position of the fall and continue the trial.</li>
		<li>Move the home cage or tubing to the other end of the beam, change the trial # on the white board and record the subsequent trial.</li>
		<li>Repeat this procedure until a total of 6 trials have been completed, with 3 trials being recorded for each direction. All 6 trials for a rat can be recorded before starting the trials for the next rat.</li>
	</ol>
	</li>
	<li>Video Analysis
	<ol>
		<li>Import video camera files into video editing program (i.e. iMovie).</li>
		<li>Compile the videos into clips for each trial and mute the volume of the video. Analyze each video clip frame by frame.</li>
		<li>Count the number of total steps the rat takes to walk the full length of the beam and the total number of left and right hindlimb and forelimb slips and total number of falls.&#160;The performance of both wild type groups and the transgenic group without stroke are considered in comparison to the co-morbid group to evaluate emergence of deficits unique to the co-morbid condition.</li>
	</ol>
	</li>
</ol>
3. Morris Water Maze for Hippocampal-dependent Spatial Learning and Reference Memory
<ol>
	<li>Equipment Setup
	<ol>
		<li>Secure a video camera positioned above the center of a circular pool (148 cm diameter and 58 cm depth). Align the four designated quadrants properly with the outline of the pool in the tracking software program.</li>
		<li>Fill the circular pool with water approximately 36 cm deep.&#160;The water should be warmed to room temperature by filling the pool a few days prior to beginning the experimental protocol.</li>
		<li>Add black non-toxic acrylic paint until the water is opaque when using white rats. Use a light color, such as white, for darker colored rats.</li>
		<li>Surround the pool with blank wall surfaces, including room dividers if needed. When in the pool, the rat should not be able to see the experimenters.</li>
		<li>Cut 4 large different shaped spatial cues from different colors of poster board and attach one shape on the wall per designated north, east, southeast and southwest pool locations. These cues should be slightly higher than the edge of the pool.</li>
		<li>Turn on a radio at low volume in the northwest quadrant to prevent the rat from being distracted by unexpected loud noises during testing.</li>
		<li>Place a circular platform (11.5 cm diameter, surface 2-3 cm below water level) in the center of the target quadrant.</li>
		<li>Turn off main room lights and turn on a floor standing lamp on the opposite side of the room divider to the pool to illuminate the area.</li>
	</ol>
	</li>
	<li>Computer Configuration
	<ol>
		<li>Use a tracking program that is designed for Morris water maze behavior tasks and set up protocol prior to starting experiments. An example of a possible program is provided in Figure 4.</li>
		<li>Set 4 consecutive days with 4 trials of 90 sec each for the spatial learning experiment.</li>
		<li>Set 2 separate probe experiments of 30 sec each at 24 hr after the last spatial learning trial and 1 week after the first probe experiment.</li>
		<li>Set cued learning trials at 4 trials per day for 2 consecutive days starting 24 hr after the second probe experiment. Each trial should be 60 sec total.</li>
		<li>For each stage, set the testing order so that there is a 20 min inter-trial interval between each trial for any given animal. Other animals can be run during this inter-trial interval, as long as every animal maintains a 20 min inter-trial interval.</li>
		<li>Ensure that the platform position is to be defined by the experimenter and that all other zones are defined by the designated platform position.</li>
		<li>Set the program to manually start and end each trial.</li>
	</ol>
	</li>
	<li>Spatial Learning Experiment
	<ol>
		<li>Place the circular platform in the southwest quadrant. It should be aligned with the circular designated &#34;platform&#34; area in the quadrant on the computer program.</li>
		<li>Start positions around the pool should be randomized for each rat. Represent all start positions in each treatment group. No rat starts from the northeast position for any spatial learning trials in the paradigm presented&#160;&#160;to allow for a novel start position during probe testing.</li>
		<li>Hold the rat at the base of its tail and gently place it in the water along the wall of the pool at the designated start position and quickly move out of the rat&#39;s sight.</li>
		<li>Have the other experimenter start the tracking software as soon as the rat is in the water. A timer should start counting up from 0 on the tracking program.</li>
		<li>If the rat locates the platform, have the other experimenter stop the trial on the computer and leave the rat on the platform for at least 30 sec before retrieving it. If the rat jumps off the platform before the trial is stopped on the computer, the trial continues.</li>
		<li>If the rat does not find the platform in the allotted trial time, guide the rat to the platform using your hand (either making the rat follow your hand or guide the rat by the base of its tail). Hold the rat on the platform for 30 sec.</li>
		<li>Remove the rat from the pool by the base of its tail onto the experimenter&#39;s arm that is covered with a towel or let the rat climb onto a portable surface.</li>
		<li>Repeat steps 3.3.3-3.3.7 for a total of 4 trials per rat. There should be an inter-trial interval of 20 min for each rat.</li>
		<li>Return the rats to their home cage under a heat lamp for at least 10 min&#160;following the rat&#8217;s final spatial learning trial.</li>
		<li>Continue the exact same spatial learning protocol for day 2 through day 4 of spatial learning.</li>
		<li>On day 2-4, do not hold the rat on the platform any longer. Allow the rat to sit on the platform without assistance for 30 sec with the experimenters out of sight. This can be done during later trials on day 1 on occasion.</li>
	</ol>
	</li>
	<li>Probe Experiment
	<ol>
		<li>Remove the platform from the pool. Ensure the previous platform position remains defined on the computer (circle in the southwest quadrant).</li>
		<li>Place the rat in the water along the wall of the pool at the northeast position and quickly move out of the rat&#39;s sight.&#160;Use of the novel northeast start position ensures that the rat recalls the platform position independent from previously trained start positions.</li>
		<li>Have the other experimenter start the tracking software as soon as the rat is in the water. A timer should start counting up from 0 on the tracking software.</li>
		<li>Retrieve the rat from the southwest quadrant of the pool by the base of its tail and hold on a towel-covered arm or let the rat climb onto a portable surface.</li>
		<li>Return the rats to their home cage under a heat lamp for at least 10 min&#160;following each rat&#8217;s probe trial.</li>
		<li>Repeat this experiment (steps 3.4.1-3.4.5) 1 week later.</li>
	</ol>
	</li>
	<li>Cued Learning Experiment
	<ol>
		<li>Use tape to secure a 4 cm diameter white spherical cue on a stand that is 8.5 cm tall on the platform (total height of cue is 12.5 cm). The top of the spherical cue will be at least 8.5 cm above water level.</li>
		<li>Remove the spatial cues from walls surrounding the pool.</li>
		<li>Randomize platform positions and start positions for each rat in each group. All platform and start positions should be represented for each treatment group.</li>
		<li>Place platform in the designated platform area and define in the tracking software.</li>
		<li>Place the rat in the water along the wall of the pool at the designated start position and quickly move out of the rat&#39;s sight.</li>
		<li>Have the other experimenter start the tracking software as soon as the rat is in the water. A timer should start counting up from 0 on the tracking software.</li>
		<li>Once the rat reaches the platform, have the other experimenter stop the trial on the computer. If the rat jumps off the platform, the trial continues. Before retrieving the rat, allow the rat to sit on the platform for 15 sec.</li>
		<li>Retrieve the rat from the pool by the base of its tail and hold on a towel-covered arm or let the rat climb onto a portable surface.</li>
		<li>Check the platform position for the next rat and move the platform into the corresponding area defined in the tracking software.</li>
		<li>Repeat steps 3.5.4-3.5.9 for the following 3 trials. There should be an inter-trial interval of 15 min.</li>
		<li>Return the rats to their home cage under a heat lamp for at least 10 min&#160;following each rat&#8217;s final cued learning trial.</li>
		<li>Continue the same protocol for day 2 of cued learning.</li>
	</ol>
	</li>
	<li>Data Analysis 
	Note: Time spent in zones, distance traveled in zones, average speed, number of entries into zones and the time to first entry into zones are often used.
	<ol>
		<li>Analyze the time and distance to reach the platform zone and the average speed per day of spatial and cued learning separately. For probe experiments, analyze the latency to first entry into the target zone as a raw time or a percent change from probe 1 to probe 2.</li>
	</ol>
	</li>
</ol>

<ol>
	<li>Auld, D., Kornecook, T., Bastianetto, S., Quirion, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Alzheimer's+disease+and+the+basal+forebrain+cholinergic+system:+relations+to+&#946;-amyloid+peptides,+cognition,+and+treatment+strategies.">Alzheimer's disease and the basal forebrain cholinergic system: relations to &#946;-amyloid peptides, cognition, and treatment strategies.</a> Prog Neurobiol. 68 (3), 209245 (2002).</li><li>Huang, Y., Mucke, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Alzheimer+Mechanisms+and+Therapeutic+Strategies.">Alzheimer Mechanisms and Therapeutic Strategies.</a> Cell. 148 (6), (2012).</li><li>Querfurth, H., LaFerla, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Alzheimer's+disease.">Alzheimer's disease.</a> New Engl J Med. 362 (4), 329-344 (2010).</li><li>Karran, E., Mercken, M., Strooper, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+amyloid+cascade+hypothesis+for+Alzheimer's+disease:+an+appraisal+for+the+development+of+therapeutics.">The amyloid cascade hypothesis for Alzheimer's disease: an appraisal for the development of therapeutics.</a> Nat Rev Drug Discov. 10 (9), 698-712 (2011).</li><li>Akiyama, 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=Inflammation+and+Alzheimer's+disease.">Inflammation and Alzheimer's disease.</a> Neurobiol Aging. 21 (3), (2000).</li><li>Butterfield, D., Drake, J., Pocernich, C., Castegna, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evidence+of+oxidative+damage+in+Alzheimer's+disease+brain:+central+role+for+amyloid+&#946;-peptide.">Evidence of oxidative damage in Alzheimer's disease brain: central role for amyloid &#946;-peptide.</a> Trends Mol Med. 7 (12), 548554 (2001).</li><li>Kalaria, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+role+of+cerebral+ischemia+in+Alzheimer's+disease.">The role of cerebral ischemia in Alzheimer's disease.</a> Neurobiol Aging. 21 (2), 321-330 (2000).</li><li>Selkoe, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Alzheimer's+disease:+genes,+proteins,+and+therapy.">Alzheimer's disease: genes, proteins, and therapy.</a> Physiol Rev. 81 (2), 741-766 (2001).</li><li>Snowdon, D. 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=Brain+infarction+and+the+clinical+expression+of+alzheimer+disease:+The+nun+study.">Brain infarction and the clinical expression of alzheimer disease: The nun study.</a> JAMA. 277 (10), 813 (1997).</li><li>Clarke, 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=Overexpression+of+APP+provides+neuroprotection+in+the+absence+of+functional+benefit+following+middle+cerebral+artery+occlusion+in+rats.">Overexpression of APP provides neuroprotection in the absence of functional benefit following middle cerebral artery occlusion in rats.</a> Eur J Neurosci. 26 (7), 1845-1852 (2007).</li><li>Whitehead, S., Hachinski, V., Cechetto, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Interaction+Between+a+Rat+Model+of+Cerebral+Ischemia+and+&#946;-Amyloid+Toxicity+Inflammatory+Responses.">Interaction Between a Rat Model of Cerebral Ischemia and &#946;-Amyloid Toxicity Inflammatory Responses.</a> Stroke. 36 (1), 107-112 (2005).</li><li>Agca, 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=Development+of+transgenic+rats+producing+human+&#946;-amyloid+precursor+protein+as+a+model+for+Alzheimer's+disease:+Transgene+and+endogenous+APP+genes+are+regulated+tissue-specifically.">Development of transgenic rats producing human &#946;-amyloid precursor protein as a model for Alzheimer's disease: Transgene and endogenous APP genes are regulated tissue-specifically.</a> BMC Neuroscience. 9 (1), 28 (2008).</li><li>Bayona, N. A., Gelb, A. W., Jiang, Z., Wilson, J. X., Urquhart, B. L., Cechetto, D. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Propofol+neuroprotection+in+cerebral+ischemia+and+its+effects+on+low-molecular-weight+antioxidants+and+skilled+motor+tasks.">Propofol neuroprotection in cerebral ischemia and its effects on low-molecular-weight antioxidants and skilled motor tasks.</a> Anesthesiology. 100 (5), 1151-1159 (2004).</li><li>Langdon, K., Clarke, J., Corbett, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Long-term+exposure+to+high+fat+diet+is+bad+for+your+brain:+exacerbation+of+focal+ischemic+brain+injury.">Long-term exposure to high fat diet is bad for your brain: exacerbation of focal ischemic brain injury.</a> Neuroscience. 182, 82-87 (2011).</li><li>Brownlow, 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=Partial+rescue+of+memory+deficits+induced+by+calorie+restriction+in+a+mouse+model+of+tau+deposition.">Partial rescue of memory deficits induced by calorie restriction in a mouse model of tau deposition.</a> Behav Brain Res. 271, 7988 (2014).</li><li>Halagappa, V., 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=Intermittent+fasting+and+caloric+restriction+ameliorate+age-related+behavioral+deficits+in+the+triple-transgenic+mouse+model+of+Alzheimer's+disease.">Intermittent fasting and caloric restriction ameliorate age-related behavioral deficits in the triple-transgenic mouse model of Alzheimer's disease.</a> Neurobiol Dis. 26 (1), (2007).</li><li>Maalouf, M., Rho, J., Mattson, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+neuroprotective+properties+of+calorie+restriction,+the+ketogenic+diet,+and+ketone+bodies.">The neuroprotective properties of calorie restriction, the ketogenic diet, and ketone bodies.</a> Behav Brain Res. 59 (2), (2008).</li><li>Tillerson, J. 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=Forced+limb-use+effects+on+the+behavioral+and+neurochemical+effects+of+6-hydroxydopamine.">Forced limb-use effects on the behavioral and neurochemical effects of 6-hydroxydopamine.</a> Neuroscience. 21 (12), 4427-4435 (2001).</li><li>Sierksma, 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=Improvement+of+spatial+memory+function+in+APPswe/PS1dE9+mice+after+chronic+inhibition+of+phosphodiesterase+type+4D.">Improvement of spatial memory function in APPswe/PS1dE9 mice after chronic inhibition of phosphodiesterase type 4D.</a> Neuropharmacology. 77, (2013).</li><li>D'Hooge, R., De Deyn, P. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Applications+of+the+Morris+water+maze+in+the+study+of+learning+and+memory.">Applications of the Morris water maze in the study of learning and memory.</a> Behav Brain Res. 36 (1), 60-90 (2001).</li><li>Vorhees, C., Williams, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Morris+water+maze:+procedures+for+assessing+spatial+and+related+forms+of+learning+and+memory.">Morris water maze: procedures for assessing spatial and related forms of learning and memory.</a> Nat Protoc. 1 (2), 848-858 (2006).</li></ol>

The authors declare that they have no competing financial interests.

The combination of stroke and Alzheimer&#39;s disease results in very distinct behavioral pathologies that can affect both motor and cognitive function depending on the severity of each condition. Thus, it is necessary to make use of a variety of behavioral tasks to determine the individual contributions of these conditions, as well as give some insight into the combined and potentially interactive effects in the co-morbid condition. The data presented demonstrate three cost-efficient, time-efficient and sensitive behavi...

Alzheimer&#39;s Disease (AD) is the most prevalent form of dementia in the elderly population and a debilitating neurodegenerative disease. Histopathologically, AD presents itself as amyloid plaques, neurofibrillary tangles and neuronal loss. Amyloid plaques consist primarily of beta-amyloid peptide (A&#946;) that has been produced through an altered proteolytic cleavage of amyloid precursor protein (APP) by &#946;-secretase and &#947;-secretase enzymes1,2,3. The cleavage product, A&#946;, deposits in the brain creating pathological amyloid plaques and has toxic effects on the brain that can lead to the characteristic learning impairments and memory loss. All of these steps together are referred to as the &#34;amyloid cascade hypothesis&#34;3,4. While this hypothesis is important when investigating AD, other cellular changes have been found to precede these plaque formations that strays from the original amyloid cascade pathway. These other cellular changes are thought to contribute to early memory loss, learning impairments and other cognitive dysfunctions involved in AD prior to plaque formation3,5,6.
With AD becoming increasingly prevalent, risk factors for developing AD are becoming an extremely important focus of research. Although age is the principal risk factor for sporadic forms of AD, other risk factors have been identified, including stroke7,8. Stroke is not only a risk factor, but it can also exacerbate already present dementias. For example, clinically, the progression of AD has been shown to be worse in patients who had previously experienced stroke9. Moreover, increased APP expression and A&#946; accumulation has been found in experimental animal models of A&#946; toxicity combined with induced stroke10,11. Since there is this important interaction between stroke and AD, it is essential that these two pathologies be further researched together in co-morbid models to better understand pathophysiology and behaviors implicated in both conditions.
To investigate co-morbid conditions, an appropriate model had to be developed in which a stroke could interact with A&#946; to produce AD-like pathology. For the first time, an APP21 transgenic rat that has a mutated human APP gene incorporated into its DNA was used to achieve an appropriate model of AD. The mutations are the Swedish double missense and Indiana single missense mutations, which have both been implicated in familial forms of AD4,8,12. In the absence of an additional insult, this rat model ages without developing the characteristic A&#946; plaques or neurofibrillary tangles12. Therefore, in an effort to induce AD-like behavioral pathology, a small stroke is introduced into the right striatum to mimic the small subcortical strokes often present in dementia patients9. The stroke in the APP21 transgenic rat embodies the co-morbid condition and allows investigation of various types of behavioral changes implicated in both disease conditions. In particular, this induction of AD-like pathology and cognitive deficits in the adult rat allows us investigate the earliest molecular and cognitive changes preceding AD.
Since the goal is to determine the first signs of behavioral changes and since both stroke and AD have very distinct behavioral pathologies, when studying the co-morbid model, behavior tasks need to assess a variety of behavioral phenotypes. There are a battery of relatively sensitive tests that can be done to analyze motor and cognitive behavior in rodent models that involve a variety of paradigms and equipment. To specifically analyze forelimb and hindlimb motor function, the cylinder task and beam-walk task have been implemented to detect motor deficits and monitor locomotion in this model. Other sensitive tasks designed to specifically assess fine forelimb motor skill (i.e. the staircase task and single pellet reaching task) require food deprivation11,13,14. To avoid any of the known effects of food deprivation on disease pathologies15,16,17, these tests have been deemed unsuitable for this study. The cylinder task assesses the spontaneous use of the rat&#39;s forelimbs during rearing in a novel environment and can detect asymmetry between forelimbs in rats with unilateral stroke10,18. A major benefit to this task is that the apparatus can be utilized for other behavior tasks, such as the Porsolt forced swim task19. Contrary to the cylinder task, the beam-walk task also allows analysis of hindlimb and forelimb motor control, in addition to locmotion10,14. Beam walking includes a locomotor component, a balance component and skilled foot placement. Both of these tests are cost-efficient, straightforward, and time-efficient and elucidate the effects of stroke and AD on differences in limb functioning.
Aside from changes in motor function, AD involves memory deficits that can present in early stages of the disease progression. When addressing AD-like pathologies in a rodent model, it is crucial that hippocampal dependent learning and memory is assessed because the hippocampus is an important brain structure largely affected in AD2. The hippocampus is an essential brain region for spatial learning and memory and its function can be tested using various maze paradigms in rodents. One of the mostly widely used maze tasks for rodent models of different diseases is the Morris water maze20. The Morris water maze utilizes spatial cues to assist the rat in locating a stationary hidden platform and tests spatial reference memory when the platform is removed. A valuable advantage of the water maze setup is that it is highly adaptable depending on the proposed research question20.
For the first time, the techniques described have been used to assess motor and cognitive function in a novel co-morbid rat model of stroke and AD. involving small strokes in an APP21 transgenic rat model.&#160;Co-morbidity was achieved by inducing vasoconstriction of the blood vessels in the striatum to produce a small stroke in APP21 transgenic rats. This stroke model has been well established as a co-morbid condition in an alternative rat model of AD11. Advancement into this novel APP21 transgenic rat model was intended to produce a more translationally valuable model. While the behavioural tasks are described using a co-morbid stroke and AD rat model, these tasks can be further applied to other models of stroke or models of other neurological diseases (i.e. Parkinson&#8217;s disease). The general methodology described will be widely applicable to these other disease states, but behavior timelines and paradigms may require alteration based on the proposed research question and model. In addition to being adaptable, the tasks described are effective in demonstrating minor deficits, while also being cost and time-efficient.&#160;

<table border="0" cellpadding="0" cellspacing="0" width="817">
	<tbody>
		<tr height="125">
			<td height="125" style="height:125px;width:216px;">Cylinder</td>
			<td style="width:115px;">Western University</td>
			<td style="width:167px;">Plexiglas Cylinder 
			Cylinder Diameter: 23 cm 
			Cylinder Height: 40 cm 
			Platform Height: 30 cm 
			Mirror Length: 35 cm 
			Mirror Width: 26.5 cm</td>
			<td style="width:320px;">This was made specifically by Western University Machine Services for our lab. Please contact your own chosen manufacturer to design this product.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Handheld Video Camera</td>
			<td style="width:115px;">JVC</td>
			<td style="width:167px;">GZ-E200</td>
			<td style="width:320px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Video Camera Tripod</td>
			<td style="width:115px;">Slik</td>
			<td style="width:167px;">F163</td>
			<td style="width:320px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">AM/FM Clock Radio</td>
			<td style="width:115px;">Sylvania</td>
			<td style="width:167px;">SCR1388</td>
			<td style="width:320px;"></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">White Board</td>
			<td style="width:115px;">Walmart</td>
			<td style="width:167px;">Width: 71.12 cm 
			Height: 55.88 cm</td>
			<td style="width:320px;">These can be purchased at any store (i.e. Walmart, University bookstores)</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Dry-erase Marker</td>
			<td style="width:115px;">Expo</td>
			<td style="width:167px;">Expo Dry-erase Original Marker</td>
			<td style="width:320px;">These can be purchased at any store (i.e. Walmart, University bookstores)</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Wooden Beam</td>
			<td style="width:115px;">Rona</td>
			<td style="width:167px;">Plywood 
			Width: 2 cm 
			Length: 100cm</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">VWR General Purpose Laboratory Tape</td>
			<td style="width:115px;">VWR Intl.</td>
			<td style="width:167px;">89097-920</td>
			<td></td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">iMovie &#39;11</td>
			<td style="width:115px;">Apple Inc.</td>
			<td style="width:167px;">Version 9.0.9 (1795)</td>
			<td style="width:320px;">This is the version used in this manuscript, but any other iMovie version or video editing software could be used.</td>
		</tr>
		<tr height="44">
			<td height="44" style="height:44px;width:216px;">Morris Water Maze Pool with Platform</td>
			<td style="width:115px;">Stoelting Co.</td>
			<td style="width:167px;">60136/60035</td>
			<td style="width:320px;">These are not the exact products used in the video, but these are essentially identical.</td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:216px;">Platform Cue</td>
			<td style="width:115px;">-</td>
			<td style="width:167px;">-</td>
			<td style="width:320px;">The platform cue used was created using a small metal stand and white spherical foam ball. These can likely be purchased at any store with home improvement materials (i.e. Walmart, Rona etc.)</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Mainstays 71&#34; Floor Lamp</td>
			<td style="width:115px;">Walmart</td>
			<td>HW-F0377SLV</td>
			<td style="width:320px;"></td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">ANY-Maze Behavioural Tracking Software</td>
			<td style="width:115px;">Stoelting Co.</td>
			<td style="width:167px;">60000</td>
			<td style="width:320px;">There are ANY-maze&#174; bundles that include the camera with or without a computer and accessories.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Compact Video Camera</td>
			<td style="width:115px;">Logitech</td>
			<td style="width:167px;">V-U0023</td>
			<td></td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Laptop</td>
			<td style="width:115px;">Hewlett-Packard</td>
			<td style="width:167px;">HP&#174; Pavilion dv6 Notebook PC</td>
			<td style="width:320px;">Laptop Specifics: AMD A6-3420M APU with Radeon&#8482; HD Graphics 1.50 GHz, 6.00 GB RAM, 64-bit operating system.</td>
		</tr>
		<tr height="62">
			<td height="62" style="height:63px;width:216px;">Americana Non-toxic Acrylic Paint</td>
			<td style="width:115px;">DecoArt</td>
			<td>DAO67-9</td>
			<td style="width:320px;">This can be ordered on the DecoArt site or purchased in store at DecoArt&#174; retailers.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Poster Board</td>
			<td style="width:115px;">Walmart</td>
			<td>PA-1961</td>
			<td style="width:320px;"></td>
		</tr>
	</tbody>
</table>

motor and hippocampal dependent spatial learning and reference memory assessment in a transgenic rat model of alzheimer's disease with stroke

Alzheimer&#39;s disease (AD) is a debilitating neurodegenerative disease that results in neurodegeneration and memory loss. While age is a major risk factor for AD, stroke has also been implicated as a risk factor and an exacerbating factor. The co-morbidity of stroke and AD results in worsened stroke-related motor control and AD-related cognitive deficits when compared to each condition alone. To model the combined condition of stroke and AD, a novel transgenic rat model of AD, with a mutated form of amyloid precursor protein (a key protein involved in the development of AD) incorporated into its DNA, is given a small unilateral striatal stroke.
For a model with the combination of both stroke and AD, behavioral tests that assess stroke-related motor control, locomotion and AD-related cognitive function must be implemented. The cylinder task involves a cost-efficient, multipurpose apparatus that assesses spontaneous forelimb motor use. In this task, a rat is placed in a cylindrical apparatus, where the rat will spontaneously rear and contact the wall of the cylinder with its forelimbs. These contacts are considered forelimb motor use and quantified during video analysis after testing. Another cost-efficient motor task implemented is the beam-walk task, which assesses forelimb control, hindlimb control and locomotion. This task involves a rat walking across a wooden beam allowing for the assessment of limb motor control through analysis of forelimb slips, hindlimb slips and falls. Assessment of learning and memory is completed with Morris water maze for this behavioral paradigm. The protocol starts with spatial learning, whereby the rat locates a stationary hidden platform. After spatial learning, the platform is removed and both short-term and long-term spatial reference memory is assessed. All three of these tasks are sensitive to behavioral differences and completed within 28 days for this model, making this paradigm time-efficient and cost-efficient.

The behavioral tasks described were used to demonstrate the effects of stroke in an APP21 transgenic rat model of Alzheimer&#39;s disease. The combination of stroke and the APP21 transgene is expected to result in greater motor deficit in the affected limbs, as well as increased memory deficits.
The cylinder task assessed gross forelimb motor function and is represented as the use of the affected forelimb. Furthermore, the beam-...

Watch this Scientific Journal Video about Motor and Hippocampal Dependent Spatial Learning and Reference Memory Assessment in a Transgenic Rat Model of Alzheimer's Disease with Stroke at JoVE.com

Motor and Hippocampal Dependent Spatial Learning and Reference Memory Assessment in a Transgenic Rat Model of Alzheimer's Disease with Stroke

To investigate the co-morbid Alzheimer&#39;s disease (AD) and stroke condition in a novel model, three behavior tasks are described that assess both motor control and cognitive behaviors. These tasks include the beam-walk task, cylinder task and Morris water maze.

Alzheimer&#39;s disease (AD) is a debilitating neurodegenerative disease that results in neurodegeneration and memory loss. While age is a major risk ...

motor-hippocampal-dependent-spatial-learning-reference-memory

Research

JoVE Journal

Medicine

10.1K Views.  Western University. To investigate the co-morbid Alzheimer's disease (AD) and stroke condition in a novel model, three behavior tasks are described that assess both motor control and cognitive behaviors. These tasks include the beam-walk task, cylinder task and Morris water maze.

Video: Motor and Hippocampal Dependent Spatial Learning and Reference Memory Assessment in a Transgenic Rat Model of Alzheimer's Disease with Stroke

Examining the Characteristics of Episodic Memory using Event-related Potentials in Patients with Alzheimer's Disease

Our laboratory uses event-related EEG potentials (ERPs) to understand and support behavioral investigations of episodic memory in patients with amnestic mild cognitive impairment (aMCI) and Alzheimer's disease (AD). Whereas behavioral data inform us about the patients' performance, ERPs allow us to record discrete changes in brain activity. Further, ERPs can give us insight into the onset, duration, and interaction of independent cognitive processes associated with memory retrieval. In patient populations, these types of studies are used to examine which aspects of memory are impaired and which remain relatively intact compared to a control population. The methodology for collecting ERP data from a vulnerable patient population while these participants perform a recognition memory task is reviewed. This protocol includes participant preparation, quality assurance, data acquisition, and data analysis. In addition to basic setup and acquisition, we will also demonstrate localization techniques to obtain greater spatial resolution and source localization using high-density (128 channel) electrode arrays.

The methodology for collecting high-density event-related potential data while patients with Alzheimer's disease perform a recognition memory task is reviewed. This protocol will include subject preparation, quality assurance, data acquisition, and data analysis.

Our laboratory uses event-related EEG potentials (ERPs) to understand and support behavioral investigations of episodic memory in patients with amnestic ...

Endothelin-1 Induced Middle Cerebral Artery Occlusion Model for Ischemic Stroke with Laser Doppler Flowmetry Guidance in Rat

Stroke is the number one cause of disability and third leading cause of death in the world, costing an estimated $70 billion in the United States in 20091, 2. Several models of cerebral ischemia have been developed to mimic the human condition of stroke. It has been suggested that up to 80% of all strokes result from ischemic damage in the middle cerebral artery (MCA) area3. In the early 1990s, endothelin-1 (ET-1) 4 was used to induce ischemia by applying it directly adjacent to the surface of the MCA after craniotomy. Later, this model was modified 5 by using a stereotaxic injection of ET-1 adjacent to the MCA to produce focal cerebral ischemia. The main advantages of this model include the ability to perform the procedure quickly, the ability to control artery constriction by altering the dose of ET-1 delivered, no need to manipulate the extracranial vessels supplying blood to the brain as well as gradual reperfusion rates that more closely mimics the reperfusion in humans5-7. On the other hand, the ET-1 model has disadvantages that include the need for a craniotomy, as well as higher variability in stroke volume8. This variability can be reduced with the use of laser Doppler flowmetry (LDF) to verify cerebral ischemia during ET-1 infusion. Factors that affect stroke variability include precision of infusion and the batch of the ET-1 used6. Another important consideration is that although reperfusion is a common occurrence in human stroke, the duration of occlusion for ET-1 induced MCAO may not closely mimic that of human stroke where many patients have partial reperfusion over a period of hours to days following occlusion9, 10. This protocol will describe in detail the ET-1 induced MCAO model for ischemic stroke in rats. It will also draw attention to special considerations and potential drawbacks throughout the procedure.

Several animal models of cerebral ischemia have been developed to simulate the human condition of stroke. This protocol describes the endothelin-1 (ET-1) induced middle cerebral artery occlusion (MCAO) model for ischemic stroke in rats. In addition, important considerations, advantages, and shortcomings of this model are discussed.

Stroke is the number one cause of disability and third leading cause of death in the world, costing an estimated $70 billion in the United States in ...

A Zebrafish Model of Diabetes Mellitus and Metabolic Memory

Diabetes mellitus currently affects 346 million individuals and this is projected to increase to 400 million by 2030. Evidence from both the laboratory and large scale clinical trials has revealed that diabetic complications progress unimpeded via the phenomenon of metabolic memory even when glycemic control is pharmaceutically achieved. Gene expression can be stably altered through epigenetic changes which not only allow cells and organisms to quickly respond to changing environmental stimuli but also confer the ability of the cell to "memorize" these encounters once the stimulus is removed. As such, the roles that these mechanisms play in the metabolic memory phenomenon are currently being examined.
We have recently reported the development of a zebrafish model of type I diabetes mellitus and characterized this model to show that diabetic zebrafish not only display the known secondary complications including the changes associated with diabetic retinopathy, diabetic nephropathy and impaired wound healing but also exhibit impaired caudal fin regeneration. This model is unique in that the zebrafish is capable to regenerate its damaged pancreas and restore a euglycemic state similar to what would be expected in post-transplant human patients. Moreover, multiple rounds of caudal fin amputation allow for the separation and study of pure epigenetic effects in an in vivo system without potential complicating factors from the previous diabetic state. Although euglycemia is achieved following pancreatic regeneration, the diabetic secondary complication of fin regeneration and skin wound healing persists indefinitely. In the case of impaired fin regeneration, this pathology is retained even after multiple rounds of fin regeneration in the daughter fin tissues. These observations point to an underlying epigenetic process existing in the metabolic memory state. Here we present the methods needed to successfully generate the diabetic and metabolic memory groups of fish and discuss the advantages of this model.

Metabolic memory is the phenomenon by which diabetic complications persist and progress unimpeded even after euglycemia is achieved pharmaceutically. Here we describe a diabetes mellitus zebrafish model which is unique in that it allows for the examination of the mitotically transmissible epigenetic components of metabolic memory in vivo.

Diabetes mellitus currently affects 346 million individuals and this is projected to increase to 400 million by 2030. Evidence from both the laboratory ...

Permanent Cerebral Vessel Occlusion via Double Ligature and Transection

Stroke is a leading cause of death, disability, and socioeconomic loss worldwide. The majority of all strokes result from an interruption in blood flow (ischemia) 1. Middle cerebral artery (MCA) delivers a great majority of blood to the lateral surface of the cortex 2, is the most common site of human stroke 3, and ischemia within its territory can result in extensive dysfunction or death 1,4,5. Survivors of ischemic stroke often suffer loss or disruption of motor capabilities, sensory deficits, and infarct. In an effort to capture these key characteristics of stroke, and thereby develop effective treatment, a great deal of emphasis is placed upon animal models of ischemia in MCA.
Here we present a method of permanently occluding a cortical surface blood vessel. We will present this method using an example of a relevant vessel occlusion that models the most common type, location, and outcome of human stroke, permanent middle cerebral artery occlusion (pMCAO). In this model, we surgically expose MCA in the adult rat and subsequently occlude via double ligature and transection of the vessel. This pMCAO blocks the proximal cortical branch of MCA, causing ischemia in all of MCA cortical territory, a large portion of the cortex. This method of occlusion can also be used to occlude more distal portions of cortical vessels in order to achieve more focal ischemia targeting a smaller region of cortex. The primary disadvantages of pMCAO are that the surgical procedure is somewhat invasive as a small craniotomy is required to access MCA, though this results in minimal tissue damage. The primary advantages of this model, however, are: the site of occlusion is well defined, the degree of blood flow reduction is consistent, functional and neurological impairment occurs rapidly, infarct size is consistent, and the high rate of survival allows for long-term chronic assessment.

Permanent Cerebral Vessel Occlusion via Double Ligature and Transection

We describe a highly reproducible method for the permanent occlusion of a rodent major cerebral blood vessel. This technique can be accomplished with very little peripheral damage, minimal blood loss, a high rate of long-term survival, and consistent infarct volume commensurate with the human clinical population.

Stroke is a leading cause of death, disability, and socioeconomic loss worldwide. The majority of all strokes result from an interruption in blood flow ...

A Murine Model of Subarachnoid Hemorrhage

In this video publication a standardized mouse model of subarachnoid hemorrhage (SAH) is presented. Bleeding is induced by endovascular Circle of Willis perforation (CWp) and proven by intracranial pressure (ICP) monitoring. Thereby a homogenous blood distribution in subarachnoid spaces surrounding the arterial circulation and cerebellar fissures is achieved. Animal physiology is maintained by intubation, mechanical ventilation, and continuous on-line monitoring of various physiological and cardiovascular parameters: body temperature, systemic blood pressure, heart rate, and hemoglobin saturation. Thereby the cerebral perfusion pressure can be tightly monitored resulting in a less variable volume of extravasated blood. This allows a better standardization of endovascular filament perforation in mice and makes the whole model highly reproducible. Thus it is readily available for pharmacological and pathophysiological studies in wild type and genetically altered mice.

A standardized mouse model of subarachnoid hemorrhage by intraluminal Circle of Willis perforation is described. Vessel perforation and subarachnoid bleeding are monitored by intracranial pressure monitoring. In addition various vital parameters are recorded and controlled to maintain physiologic conditions.

In this video publication a standardized mouse model of subarachnoid  hemorrhage (SAH) is presented. Bleeding is induced by endovascular Circle of  Willis ...

Lesion Explorer: A Video-guided, Standardized Protocol for Accurate and Reliable MRI-derived Volumetrics in Alzheimer's Disease and Normal Elderly

Obtaining in vivo human brain tissue volumetrics from MRI is often complicated by various technical and biological issues. These challenges are exacerbated when significant brain atrophy and age-related white matter changes (e.g. Leukoaraiosis) are present. Lesion Explorer (LE) is an accurate and reliable neuroimaging pipeline specifically developed to address such issues commonly observed on MRI of Alzheimer's disease and normal elderly. The pipeline is a complex set of semi-automatic procedures which has been previously validated in a series of internal and external reliability tests1,2. However, LE's accuracy and reliability is highly dependent on properly trained manual operators to execute commands, identify distinct anatomical landmarks, and manually edit/verify various computer-generated segmentation outputs.
LE can be divided into 3 main components, each requiring a set of commands and manual operations: 1) Brain-Sizer, 2) SABRE, and 3) Lesion-Seg. Brain-Sizer's manual operations involve editing of the automatic skull-stripped total intracranial vault (TIV) extraction mask, designation of ventricular cerebrospinal fluid (vCSF), and removal of subtentorial structures. The SABRE component requires checking of image alignment along the anterior and posterior commissure (ACPC) plane, and identification of several anatomical landmarks required for regional parcellation. Finally, the Lesion-Seg component involves manual checking of the automatic lesion segmentation of subcortical hyperintensities (SH) for false positive errors.
While on-site training of the LE pipeline is preferable, readily available visual teaching tools with interactive training images are a viable alternative. Developed to ensure a high degree of accuracy and reliability, the following is a step-by-step, video-guided, standardized protocol for LE's manual procedures.

Lesion Explorer (LE) is a semi-automatic, image-processing pipeline developed to obtain regional brain tissue and subcortical hyperintensity lesion volumetrics from structural MRI of Alzheimer's disease and normal elderly. To ensure a high level of accuracy and reliability, the following is a video-guided, standardized protocol for LE's manual procedures.

Obtaining in vivo human brain tissue volumetrics from MRI is often complicated by various technical and biological issues. These challenges are ...

Excitotoxic Stimulation of Brain Microslices as an In vitro Model of Stroke

Examining molecular mechanisms involved in neuropathological conditions, such as ischemic stroke, can be difficult when using whole animal systems. As such, primary or &#39;neuronal-like&#39; cell culture systems are commonly utilized. While&#160;these systems are relatively easy to work with, and are useful model systems in which various functional outcomes (such as cell death) can be readily quantified, the examined outcomes and pathways in cultured immature neurons (such as excitotoxicity-mediated cell death pathways) are not necessarily the same as those observed in mature brain, or in intact tissue. Therefore, there is the need to develop models in which cellular mechanisms in mature neural tissue can be examined. We have developed an in vitro technique that can be used to investigate a variety of molecular pathways in intact nervous tissue. The technique described herein utilizes rat cortical tissue, but this technique can be adapted to use tissue from a variety of species (such as mouse, rabbit, guinea pig, and chicken) or brain regions (for example, hippocampus, striatum, etc.). Additionally, a variety of stimulations/treatments can be used (for example, excitotoxic, administration of inhibitors, etc.). In conclusion, the brain slice model described herein can be used to examine a variety of molecular mechanisms involved in excitotoxicity-mediated brain injury.

Excitotoxic Stimulation of Brain Microslices as an In vitro Model of Stroke

We have developed a brain slice model which can be used to examine molecular mechanisms involved in excitotoxicity-mediated brain injury.&#160;This technique generates viable mature brain tissue and reduces animal numbers required for experimentation, whilst keeping the neuronal circuitry, cellular interactions, and postsynaptic compartments partly intact.

Examining molecular mechanisms involved in neuropathological conditions, such as ischemic stroke, can be difficult when using whole animal systems. As ...

Controlling Parkinson's Disease With Adaptive Deep Brain Stimulation

Adaptive deep brain stimulation (aDBS) has the potential to improve the treatment of Parkinson&#39;s disease by optimizing stimulation in real time according to fluctuating disease and medication state. In the present realization of adaptive DBS we record and stimulate from the DBS electrodes implanted in the subthalamic nucleus of patients with Parkinson&#39;s disease in the early post-operative period. Local field potentials are analogue filtered between 3 and 47 Hz before being passed to a data acquisition unit where they are digitally filtered again around the patient specific beta peak, rectified and smoothed to give an online reading of the beta amplitude. A threshold for beta amplitude is set heuristically, which, if crossed, passes a trigger signal to the stimulator. The stimulator then ramps up stimulation to a pre-determined clinically effective voltage over 250 msec and continues to stimulate until the beta amplitude again falls down below threshold. Stimulation continues in this manner with brief episodes of ramped DBS during periods of heightened beta power.
Clinical efficacy is assessed after a minimum period of stabilization (5 min) through the unblinded and blinded video assessment of motor function using a selection of scores from the Unified Parkinson&#39;s Rating Scale (UPDRS). Recent work has demonstrated a reduction in power consumption with aDBS as well as an improvement in clinical scores compared to conventional DBS. Chronic aDBS could now be trialed in Parkinsonism.

Adaptive deep brain stimulation (aDBS) is effective for Parkinson&#8217;s disease, improving symptoms and reducing power consumption compared to conventional deep brain stimulation (cDBS). In aDBS we track a local field potential biomarker (beta oscillatory amplitude) in real time and use this to control the timing of stimulation.

Adaptive deep brain stimulation (aDBS) has the potential to improve the treatment of Parkinson&#39;s disease by optimizing stimulation in real time ...

Assessment of Vascular Function in Patients With Chronic Kidney Disease

Patients with chronic kidney disease (CKD) have significantly increased risk of cardiovascular disease (CVD) compared to the general population, and this is only partially explained by traditional CVD risk factors. Vascular dysfunction is an important non-traditional risk factor, characterized by vascular endothelial dysfunction (most commonly assessed as impaired endothelium-dependent dilation [EDD]) and stiffening of the large elastic arteries. While various techniques exist to assess EDD and large elastic artery stiffness, the most commonly used are brachial artery flow-mediated dilation (FMDBA) and aortic pulse-wave velocity (aPWV), respectively. Both of these noninvasive measures of vascular dysfunction are independent predictors of future cardiovascular events in patients with and without kidney disease. Patients with CKD demonstrate both impaired FMDBA, and increased aPWV. While the exact mechanisms by which vascular dysfunction develops in CKD are incompletely understood, increased oxidative stress and a subsequent reduction in nitric oxide (NO) bioavailability are important contributors. Cellular changes in oxidative stress can be assessed by collecting vascular endothelial cells from the antecubital vein and measuring protein expression of markers of oxidative stress using immunofluorescence. We provide here a discussion of these methods to measure FMDBA, aPWV, and vascular endothelial cell protein expression.

The degree of vascular dysfunction and contributing physiological mechanisms can be assessed in patients with chronic kidney disease by measuring brachial artery flow-mediated dilation, aortic pulse-wave velocity, and vascular endothelial cell protein expression.

Patients with chronic kidney disease (CKD) have significantly increased risk of cardiovascular disease (CVD) compared to the general population, and this ...

Reproducible Motor Deficit Following Aortic Occlusion in a Rat Model Of Spinal Cord Ischemia

Spinal cord ischemia is a fatal complication following thoracoabdominal aortic aneurysm surgery. Researchers can investigate the strategies for preventing and treating this complication using experimental models of spinal cord ischemia. The model described here demonstrates varying degrees of paraplegia that relate to the length of occlusion following thoracic aortic occlusion in a rat spinal cord ischemia model.
A 2-Fr. balloon-tipped catheter was advanced through the femoral artery into the descending thoracic aorta until the catheter tip was placed at the left subclavian artery in anesthetized male Sprague-Dawley rats. Spinal cord ischemia was induced by inflating the catheter balloon. After a set period of occlusion (9, 10, or 11 min), the balloon was deflated. Neurologic assessment was performed using the motor deficit index at 24 h after surgery, and the spinal cord was harvested for histopathological examination.
Rats that underwent 9 min of aortic occlusion showed mild and reversible motor impairment in the hind limb. Rats subjected to 10 min of aortic occlusion presented with moderate but reversible motor impairment. Rats subjected to 11 min of aortic occlusion displayed complete and persistent paralysis. The motor neurons in the spinal cord sections were more preserved in rats subjected to shorter duration of aortic occlusion.
Researchers can achieve a reproducible hind limb motor deficit following thoracic aortic occlusion using this spinal cord ischemia model.

This study demonstrates the technique to make a minimally invasive and easily reproducible model of spinal cord ischemia in rats. Various degrees of hind limb motor deficit can be produced by controlling the aortic occlusion time.

Spinal cord ischemia is a fatal complication following thoracoabdominal aortic aneurysm surgery. Researchers can investigate the strategies for preventing ...