Name: getting to compliance in forced exercise in rodents: a critical standard to evaluate exercise impact in aging-related disorders and disease
Uploaded: 2014-08-22T13:00:00
Description: Watch this Scientific Journal Video about Getting to Compliance in Forced Exercise in Rodents: A Critical Standard to Evaluate Exercise Impact in Aging-related Disorders and Disease at JoVE.com

Funding for this work was provided&#160;in part&#160;by a research grant awarded to M. F. Salvatore from the National Institute on Aging (AG040261). The authors would also like to acknowledge Victoria Fields for her assistance with the behavioral experiments associated with the work presented&#160;and Mackenzie Orchard for his assistance with construction of the backstop for the treadmill.

All of the following procedures are conducted in accordance with the Institutional Animal Care and Use Committee at LSU Health Sciences Center-Shreveport, and comply with the National Institute of Health guidelines.
1. Pre-exercise Procedures
Note: A timeline of all of the relevant procedures in this forced exercise regimen is diagrammed in Figure 1.
<img alt="Figure 1" fo:content-width...

<ol>
	<li>Erickson, K. I., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Exercise+training+increases+size+of+hippocampus+and+improves+memory.">Exercise training increases size of hippocampus and improves memory.</a> Proc. Natl. Acad. Sci. U.S.A. 108 (7), 3017-3022 (2011).</li><li>Zigmond, M. J., Cameron, J. L., Hoffer, B. J., Smeyne, R. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neurorestoration+by+physical+exercise:+Moving+forward.">Neurorestoration by physical exercise: Moving forward.</a> Parkinsonis., and Related Dis. 18 (S1), S147-S150 (2012).</li><li>Savela, S. 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=Physical+activity+at+midlife+and+health-related+quality+of+life+in+older+men.">Physical activity at midlife and health-related quality of life in older men.</a> Arch. Intern. Med. 170 (13), 1171-1172 (2010).</li><li>Ridgel, A. L., Vitek, J. L., Alberts, J. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Forced,+not+voluntary,+exercise+improves+motor+function+in+Parkinson's+disease+patients.">Forced, not voluntary, exercise improves motor function in Parkinson's disease patients.</a> Neurorehabil. Neural Repair. 23 (6), 600-608 (2009).</li><li>Demirel, H. A., Serova, L., Sabban, E. L., Broxson, C. S., Powers, S. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Gene+expression+of+catecholamine+biosynthetic+enzymes+following+exercise:+modulation+by+age.">Gene expression of catecholamine biosynthetic enzymes following exercise: modulation by age.</a> Neuroscience. 103 (3), 703-711 (2001).</li><li>Petzinger, G. 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=Effects+of+treadmill+exercise+on+dopaminergic+transmission+in+the+1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-lesioned+mouse+model+of+basal+ganglia+injury.">Effects of treadmill exercise on dopaminergic transmission in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-lesioned mouse model of basal ganglia injury.</a> J. Neurosci. 27 (20), 5291-5300 (2007).</li><li>Yoon, M. 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=Treadmill+exercise+suppresses+nigrostriatal+dopaminergic+neuronal+loss+in+6-hydroxydopamine-induced+Parkinson's+rats.">Treadmill exercise suppresses nigrostriatal dopaminergic neuronal loss in 6-hydroxydopamine-induced Parkinson's rats.</a> Neurosci. Lett. 423 (1), 12-17 (2007).</li><li>Collier, T. 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=Aging-related+changes+in+the+nigrostriatal+dopamine+system+and+the+response+to+MPTP+in+nonhuman+primates:+Diminished+compensatory+mechanisms+as+a+prelude+to+parkinsonism.">Aging-related changes in the nigrostriatal dopamine system and the response to MPTP in nonhuman primates: Diminished compensatory mechanisms as a prelude to parkinsonism.</a> Neurobiol. Dis. 26 (1), 56-65 (2007).</li><li>Phelan, J. P., Austad, S. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Selecting+animal+models+of+human+aging:+inbred+strains+often+exhibit+less+biological+uniformity+than+F1+hybrids.">Selecting animal models of human aging: inbred strains often exhibit less biological uniformity than F1 hybrids.</a> J. Gerontol. 49 (1), B1-B11 (1994).</li><li>Yurek, D. M., Hipkens, S. B., Hebert, M. A., Gash, D. M., Gerhardt, G. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Age-related+decline+in+striatal+dopamine+release+and+motoric+function+in+brown+Norway/Fischer+344+hybrid+rats.">Age-related decline in striatal dopamine release and motoric function in brown Norway/Fischer 344 hybrid rats.</a> Brain Res. 791 (1-2), 246-256 (1998).</li><li>Spangler, E. L., Waggie, K. S., Hengemihle, J., Roberts, D., Hess, B., Ingram, D. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Behavioral+assessment+of+aging+in+male+Fischer+344+and+brown+Norway+rats+strains+and+their+F1+hybrid.">Behavioral assessment of aging in male Fischer 344 and brown Norway rats strains and their F1 hybrid.</a> Neurobiol. Aging. 15 (3), 319-328 (1994).</li><li>Salvatore, M. F., Pruett, B. S., Spann, S. L., Dempsey, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Aging+reveals+a+role+for+nigral+tyrosine+hydroxylase+ser31+phosphorylation+in+locomotor+activity+generation.">Aging reveals a role for nigral tyrosine hydroxylase ser31 phosphorylation in locomotor activity generation.</a> PLoS One. 4 (1), (2009).</li><li>Pruett, B. S., Salvatore, M. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=GFR+&#945;-1+receptor+expression+in+the+aging+nigrostriatal+and+mesoaccumbens+pathways.">GFR &#945;-1 receptor expression in the aging nigrostriatal and mesoaccumbens pathways.</a> J. Neurochem. 115 (3), 707-715 (2010).</li><li>Salvatore, M. F., Pruett, B. S., Dempsey, C., Fields, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comprehensive+profiling+of+dopamine+regulation+in+substantia+nigra+and+ventral+tegmental+area.">Comprehensive profiling of dopamine regulation in substantia nigra and ventral tegmental area.</a> J. Vis. Exp. (66), e4171 (2012).</li><li>Salvatore, M. F., Pruett, B. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dichotomy+of+tyrosine+hydroxylase+and+dopamine+regulation+between+somatodendritic+and+terminal+field+areas+of+nigrostriatal+and+mesoaccumbens+pathways.">Dichotomy of tyrosine hydroxylase and dopamine regulation between somatodendritic and terminal field areas of nigrostriatal and mesoaccumbens pathways.</a> PLoS One. 7 (1), e29867 (2012).</li><li>Pruett, B. S., Salvatore, M. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nigral+GFR+&#945;-1+infusion+in+aged+rats+increases+locomotor+activity,+nigral+tyrosine+hydroxylase,+and+dopamine+content+in+synchronicity.">Nigral GFR &#945;-1 infusion in aged rats increases locomotor activity, nigral tyrosine hydroxylase, and dopamine content in synchronicity.</a> Mol. Neurobiol. 47 (3), 988-999 (2013).</li><li>Abercrombie, E. D., Zigmond, M. J., Bloom, ,. F. .. E., Kupfer, D. .. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chapter+31:+Modification+of+central+categcholaminergic+systems+by+stress+and+injury:+Functional+significance+and+clinical+implications.">Chapter 31: Modification of central categcholaminergic systems by stress and injury: Functional significance and clinical implications.</a> Psychopharmacology: The Fourth Generation of Progress. 31, 355-361 (1995).</li><li>Ong, , 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=Neurobiological+consequences+of+acute+footshock+stress;+effects+on+tyrosine+hydroxylase+phosphorylation+and+activation+in+the+rat+brain+and+adrenal+medulla.">Neurobiological consequences of acute footshock stress; effects on tyrosine hydroxylase phosphorylation and activation in the rat brain and adrenal medulla.</a> J. Neurochem. , (2013).</li><li>Al-Jarrah, 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=Endurance+exercise+promotes+cardiorespiratory+rehabilitation+without+neurorestoration+in+the+chronic+mouse+model+of+parkinsonism+with+severe+neurodegeneration.">Endurance exercise promotes cardiorespiratory rehabilitation without neurorestoration in the chronic mouse model of parkinsonism with severe neurodegeneration.</a> J. Neurosci. 149 (1), 28-37 (2007).</li><li>Dishman, R. K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neurobiology+of+exercise.">Neurobiology of exercise.</a> Obesity. 14 (3), 345-356 (2006).</li></ol>

There is evidence to suggest that lifestyle strategies, such as exercise, may reduce the risk for chronic age-related diseases and their cardinal symptoms, including those of the neurodegenerative diseases like Alzheimer&#8217;s disease and PD20. The benefit of forced exercise on a treadmill, as opposed to voluntary exercise on a running wheel, is that the investigator can determine the frequency, the rate, and/or intensity of exercise, and duration of the exercise session. Thus, specific post-exercise effects...

Noninvasive lifestyle strategies that can either prevent or mitigate impairments to cognition and locomotion, both often associated with aging, are gaining traction as viable practices for maintaining health and well-being1,2. For example, moderate to rigorous exercise on a weekly and consistent basis in middle-aged men can significantly increase locomotor capabilities compared to similarly-aged peers with advancing age3. Furthermore, increasing evidence suggests that these lifestyle strategies, such as exercise, can mitigate or even reverse impairments associated with neurodegenerative diseases such as Parkinson&#8217;s disease (PD)4.
Efforts to understand the molecular mechanisms of aging-related impairments have also been directed toward identifying how noninvasive strategies, like exercise, reverse or attenuate the mechanisms that contribute to such impairments. Treadmill exercise is one such strategy being increasingly employed in models of PD5-7 and cognitive impairment1, wherein the mechanisms behind improvements in locomotor or cognitive function are still being determined. However, it is important to point out that aging is the neurobiological background of PD. Thus, for any potential translation of any exercise benefit in an animal model to be realized in the human condition it must take into consideration the neurobiological background of aging. For example, compensatory mechanisms that stave off locomotor impairment during PD progression could be impaired by the process of aging8. Thus, it stands to reason that exercise paradigms must be developed that not only consider the impact of aging in either the presence or absence of disease pathology, but that also could be initiated and maintained in aged rodents.
Therefore, in consideration of aging on the neurobiological background, the selection of the rat strain should be thoughtfully considered by the investigator. Several rat strains are available for aging studies, notably the Fischer 344 and the Brown-Norway/Fischer 344 F1 (BNF) hybrid. The commonly used Sprague Dawley rat (an outbred strain) is also amenable for such use, as it is commonly used in neurodegenerative disease models, such as the 6-hydroxydopamine PD model. Our laboratory uses both the Sprague Dawley and the BNF strains in aging and neurodegenerative disease work. In this report, we will present results highlighting our exercise protocols using both strains. For those investigators strictly focusing on age-related studies, the BNF strain offers some important advantages. First, it is comparatively less vulnerable to aging-related disorders (such as tumors) and has exceptional longevity (typical life span exceeds 30 months) in comparison to other strains. They also have less variability in a variety of physiological and behavioral outcomes9, and are also suitable to investigate approaches that intervene with the process of aging. Furthermore, experiments such as forced exercise demand considerable handling by the investigator, and the gentle disposition of the BNF strain is advantageous. Aging-related changes in striatal and midbrain dopamine tissue content, as well as locomotor activity changes are similar in BNF rats and primates10-11. Our laboratory also has extensive experience with characterizing and manipulating striatal and midbrain dopamine signaling as well as locomotor activity in the BNF strain12-16. Therefore, because the risk of other neurobiologically-based diseases increases with aging, animal models of neurodegenerative disease should consider the use of rodent strains with an extensive track record of use in aging studies.
Our protocol herein also addresses some critical issues that the investigator must consider in the interpretation of their results obtained from an exercise protocol. The rodent treadmill apparatus (the use of which we will highlight in this report), is typically equipped with electric shock coils at the back of each lane of the treadmill that can be deactivated. However when these electric shock coils are activated, a small footshock is delivered to the subject as it comes in contact with the coils. This strategy is often employed in exercise studies to facilitate compliance to treadmill exercise. This is a critical point for consideration, particularly for those investigators involved in behavioral studies that are influenced by dopamine- or norepinephrine-signaling. Electrical footshock is a physiological stressor, and its impact on both neurotransmitter systems is well-documented, with increased activation of tyrosine hydroxylase17-18. Thus, increased biosynthesis of either neurotransmitter could confound the interpretation of any exercise effect, making the investigator liable for interpreting whether any observed change in behavior after exercise is strictly due to the exercise regimen or the footshock stress. Importantly, the proposed forced exercise regimen does not employ the use of footshock at any point in the treadmill acclimation or exercise training periods.
Successful exercise regimens also require maximum compliance to exercise from test subjects. The employment of footshock in order to achieve compliance could confound interpretation of experimental outcomes when the dependent measure is related to neurotransmitter signaling that is influenced by footshock (as previously discussed). Thus, the challenge is to get rodents, and in particular aged rodents, to comply with an exercise regimen. A desirable goal of any exercise regimen is to achieve nearly 100% compliance, as this will reduce the number of animals necessary to complete the exercise regimen. Specifically, maximum compliance to exercise and interpretation of exercise regimen outcomes can be obtained in our exercise regimen as a result of several pre-exercise procedures including, 1) a reverse light-dark cycle that enables animals to exercise during their active (wake) cycle, 2) ensuring locomotor performance baselines are equal prior to segregation of control and exercise groups, and 3) introducing the test subjects to an acclimation period that gradually introduces them to the requirements of the exercise regimen. Here, the aforementioned experimental considerations and steps necessary to achieve nearly 100% compliance to treadmill exercise in an aged rodent (&#62;18 months of age) will be evaluated and presented. Finally, the environment associated with exercise may be stressful, and as such, present potential confounds to determine physiological effects specific to exercise. Our protocol also controls for the exposure to the potential stress-inducing environment of the treadmill by including the nonexercise group in every aspect of the acclimation phase (including modest treadmill running) and placing them on the stationary treadmill during the treadmill exercise training. Thus, this protocol aims to describe the measures necessary to determine the physiological impact of exercise alone.

&#160;
<table border="0" cellpadding="0" cellspacing="0" width="814">
	<tbody>
		<tr height="20">
			<td colspan="6" height="41" rowspan="2" style="height:41px;width:815px;">Table of Specific Materials/Equipment</td>
		</tr>
		<tr height="21">
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Name of Material/Equipment</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td colspan="3">Comments/Description</td>
		</tr>
		<tr height="80">
			<td height="80" style="height:80px;">Exer-4 Treadmill for Rats</td>
			<td>Columbus Instruments&#160;</td>
			<td>96157-2</td>
			<td colspan="3" style="width:321px;">Columbus has a variety of treadmills for animals with varying numbers of lanes, and other options such as metabolic measurements</td>
		</tr>
		<tr height="39">
			<td height="59" rowspan="2" style="height:59px;width:211px;">Animal Activity Meter: Opto-Varimex 4-Auto Track&#160;</td>
			<td rowspan="2">Columbus Instruments&#160;</td>
			<td rowspan="2">0170-R4</td>
			<td colspan="3" rowspan="2" style="width:321px;">Columbus has several options for activity chambers with varying arena sizes, beam spacing, and sensors</td>
		</tr>
		<tr height="20">
		</tr>
	</tbody>
</table>

getting to compliance in forced exercise in rodents: a critical standard to evaluate exercise impact in aging-related disorders and disease

There is a major increase in the awareness of the positive impact of exercise on improving several disease states with neurobiological basis; these include improving cognitive function and physical performance. As a result, there is an increase in the number of animal studies employing exercise. It is argued that one intrinsic value of forced exercise is that the investigator has control over the factors that can influence the impact of exercise on behavioral outcomes, notably exercise frequency, duration, and intensity of the exercise regimen. However, compliance in forced exercise regimens may be an issue, particularly if potential confounds of employing foot-shock are to be avoided. It is also important to consider that since most cognitive and locomotor impairments strike in the aged individual, determining impact of exercise on these impairments should consider using aged rodents with a highest possible level of compliance to ensure minimal need for test subjects. Here, the pertinent steps and considerations necessary to achieve nearly 100% compliance to treadmill exercise in an aged rodent model will be presented and discussed. Notwithstanding the particular exercise regimen being employed by the investigator, our protocol should be of use to investigators that are particularly interested in the potential impact of forced exercise on aging-related impairments, including aging-related Parkinsonism and Parkinson&#8217;s disease.

A diagram of the timeline of events in all three&#160;phases of the forced exercise regimen is shown in Figure 1. In addition to reversing the light-dark cycle, daily handling prior to any experimental testing, and stage 1 of acclimation training, where subjects sit on the stationary treadmill for 5-10 min/day for 3 consecutive days, were implemented to increase compliance to the forced exercise regimen. These procedures were added to the protocol in order to minimize stress associated with novel environ...

Watch this Scientific Journal Video about Getting to Compliance in Forced Exercise in Rodents: A Critical Standard to Evaluate Exercise Impact in Aging-related Disorders and Disease at JoVE.com

Getting to Compliance in Forced Exercise in Rodents: A Critical Standard to Evaluate Exercise Impact in Aging-related Disorders and Disease

Exercise may alleviate behavioral impairments associated with neurodegenerative diseases. Studies of the mechanisms underlying exercise-mediated effects have increased in disease models, but the neurobiological background accompanying these diseases, specifically aging, is likely not considered. Here, we illustrate measures to increase the likelihood of a successful exercise paradigm in aged rats.

There is a major increase in the awareness of the positive impact of exercise on improving several disease states with neurobiological basis; these ...

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