The Effect of Caffeine on the Wheel Running Activity of Mus musculus

Erinne Sonnenberg

Liann Liegler



            Six Mus musculus, commonly known as house mice, were tested to see if caffeine affected activity level.  Each mouse was tested in their normal state (no caffeine) and after consumption of caffeine.  The tests were quantified by running wheel revolutions.  The tests were conducted at the same time on different days to avoid wheel habituation and activity level variances because of the time of day.  We hypothesized the wheel running activity of the mouse would increase after caffeine consumption.  Our hypothesis was supported with a p-value of 0.0104.  The caffeine consumption correlated with the wheel rotations on a linear scale of 0.742.

Keywords: Mus musculus, caffeine, wheel running.


            Caffeine can affect many different parts of the body. Caffeine is a stimulant that is often used by people to stay awake. Often times if a person consumes too much caffeine they have a high amount of energy and sometimes even feel jittery. Caffeine affects humans by blocking the adenosine receptors.  It can often times make people feel like they have more energy.  Scientists at the American Physiological Society did studies to determine caffeine and the effects it has on muscle metabolism. They used human test subjects for their studies. They gave their subjects nine milligrams for every kilogram the weighed, this ensured that all the subjects were given proportionally the same amount of caffeine. They then allowed them to cycle (ride a stationary bike) until they reached exhaustion, which they defined as eighty-percent oxygen uptake.  The control group did not consume caffeine and were instructed to ride the same stationary bike until they reached the same amount of oxygen uptake.They also kept track of how long each rider rode on the bike, so that they were able to quantify which group on average rode longer. The scientists concluded the group that was given caffeine cycled significantly longer than the group without caffeine. The caffeinated group cycled for, on average, 96.2 minutes and the control group for only 75.8 minutes (Spriet, et. al. 1992).

             Caffeine can have similar effects on animals. Scientists have looked at how caffeine can affect the house mouse, Mus musculus. Scientists in Britain did a study on how caffeine affects the adenosine receptors in central nervous system in mice. They have found that caffeine actually blocks adenosine receptors found in the cerebellum and the cortex. The scientists used two different classes of mice. The first they called the control group in which they did not introduce them to caffeine at all until the experiment. The next group they classified as the experimental group in which they habituated these mice to caffeine, by introducing small doses each day. The results that they found were; the less caffeine interaction they had the more of an effect the caffeine had on the mouse. This supported their hypothesis that caffeine does in fact have an effect on the behaviors exhibited by the mice (Yacoubi, et. al. 2000).Therefore, we hypothesized, mice will be more active, having more revolutions, when given caffeine.




            This experiment was performed on two different days at 11:00 a.m. in room TL212 at Alverno College in Milwaukee, Wisconsin. The first day was March 19th, 2009. We obtained six different house mice that were all in their designated cages. We placed each cage on the lab top tables in room TL212. Next we crushed up one 200 mg caffeine tablet, brand name Stay Awake, using a standard razor blade. We then obtained our peanut butter, House Recipe Creamy Peanut Butter, made by Sysco Corp. and measured out twenty grams.  We also measured out five grams of the crushed caffeine tablet; this would mean that each mouse received 14.25 mg of caffeine if they consumed the full dose. We then obtained a standard pipette. The peanut butter was added to the pipette tip and then dipped it in five mg of the crushed caffeine tablet ensuring to collect it all on the pipette tip.  This was done for each of the six mice. We then placed each pipette in between the metal rungs on each mouse cage; the location on the mouse cage was picked haphazardly. The mice were given twenty minutes to consume the peanut butter off of the pipette tip. We then collected the pipettes and measured the weight. This way we were able to identify how much eat mouse ate. The mice were then placed in a wheel that measures the number of times it completes a full rotation. This wheel was manufactured by the Wahmann company located in Baltimore, Maryland. The mice were allowed to run for thirty minutes and then placed back inside their cages. We recorded how many rotations the wheel had progressed. The second half of the experiment was performed on March, 27th 2009 at 11:00 a.m. in the same location. The same mice were removed from their cages and placed in the same wheels. They were then allowed to run for thirty minutes. We removed the mice and recorded how many rotations had progressed.


            We found the average wheel rotations for mice who had consumed caffeine was 273.5 with a standard deviation of 32.57.  When the mice were tested in their natural state the average wheel revolutions were 167.7 with a standard deviation of 53.31.  (Figure 1).  We used a 2-tailed paired T-test to find the p-value of 0.0104.  From this we can conclude that our results were significant and our hypothesis was supported.  We also observed the amount of the caffeine mixture consumption in comparison to the wheel rotations.  We used a linear trend line to find the positive correlation of 0.8109.

Figure 1.  Comparison of average wheel revolutions of Mus musculus after caffeine consumption and in natural state.  Mean +/- standard error.



Figure 2. Correlation of wheel rotations with amount of caffeine and peanut butter consumed.  Linear trend line used for R2 value.




            From the results of this experiment we can infer that caffeine does have an effect on the activity of small mammals, at least in the short term.  To continue this experiment we could test to see how long increased energy was present after caffeine consumption.  It would be interesting to see if there is a correlation between the time the caffeine took effect and wheel running as time progressed.  How fast does the caffeine wear off and what are the ending results?  Do the mice end with less wheel running than if they had not consumed caffeine?  If so is the wheel activity similar but spaced differently?

            When considering future experiments based on these follow up questions it might be beneficial to use a larger sample size.  For this experiment we used 6 mice which is the very minimum to statistically test.  Using a larger sample size would give us more representative results. 

            Some possible factors that may have an effect on our data include the food consumption of the mice.  When food is not available to mice there is a reduction in physical activity and play (Barber, 1991).  If the opposite is true then increased food would also lead to more energy for physical activity and play. 

As an incentive for the mouse to ingest the caffeine it was mixed with peanut butter.  The peanut butter itself may have been the cause for the increased wheel running activity.  If we were to repeat this experiment we would give the mice peanut butter with no caffeine before running them.

Some studies suggest that wheel running in itself is a rewarding behavior for mice (Gammie, et. al. 2003).  These mice were not given any opportunities to use the running wheel except during the experimental procedure.  They may have had an increased activity because of the rare opportunity.  It may be beneficial to habituate the mice to the wheel before testing them. 

After our experiment was complete we considered the possibility that we actually habituated the mice to the wheel during the first experimental period which could have affected the activity during the second experimental period.  If we were to repeat this experiment we would change the order of the tests.  In the first session we would give half the mice caffeine and half would get no caffeine.  We would then do the same thing for the second experimental period giving caffeine to the mice that had not previously had it and the mice that previously had caffeine would not get it.  Doing this would eliminate the confounding variable of previous wheel experience.  This would also be beneficial if an unknown change occurred with the mice between the two tests.  Confounding variables such as illness, change in food, or change in any other environmental factor between the two tests could have altered the results.  If we were to vary the test order these variables would not have an effect on our results.






Literature Cited

Barber, N. (1991).  Play and Energy Regulation in Mammals.  The Quarterly Review of Biology, 66, 2.  Retrieved March 29, 2009 from JSTOR database.

Gammie, S., Hasen, N., Rhodes, J., Girard, I., Garland, T. (2003).  Predatory aggression, but not maternal or intermale aggression, is associated with high voluntary wheel-running behavior in mice.  Hormones and Behavior, 44, 3, 209-221.  Retrieved March 29, 2009 from Science Direct database.

Spriet, L., MacLean, D., Dyck, D., Hultman, E., & Cederblad, G. (1992). Caffeine ingestion and muscle metabolism during prolonged exercise in humans. AJP - Endocrinology and Metabolism, 262, Retrieved March 31, 2009 from:

Yacoubi, M., Ledent, C., Ménard, J., Parmentier, M., & Costentin, J. (2000). The stimulant effects of caffeine on locomotor behaviour in mice are mediated through its blockade of adenosine A2A receptors. British Journal of Pharmacology, 129, Retrieved March 31, 2009 from: