Light Cycle Phase Effects on Behavioral Responses to Stress Exposure in Rats

Animal models enable investigation of the impact of stress on emotional-behavioral performance to facilitate understanding of posttraumatic stress symptoms experienced in humans. Refinement of animal stress models could lead to a reduction in the number of subjects needed to detect statistically significant stress effects, in accordance with Russel and Burch's three Rs of research. We assessed whether performance of experimental procedures (that is, stress exposure and poststress behavioral testing) during the dark or light phases of the 12-h light/12 h-dark cycle is a refinement that could accomplish this reduction. At 3 h into either the light or dark phase, male and female adult Sprague-Dawley rats underwent a single-day traumatic stress exposure. Rats then underwent behavioral testing for exploratory behaviors, startle responses, and conditioned fear responses at 2 h, 1 d, and 9 d after stress exposure. Distance traveled in the elevated plus maze (EPM) by both male and female rats was significantly reduced in the dark phase compared with the light phase. Male rats of the dark phase group also spent less time in the open arms of the EPM, and traveled less, spent less time in the center, and spent more cumulative time freezing in the open field. Female rats of the dark phase group spent more cumulative time freezing in the EPM and exhibited significantly more tone-cued conditioned freezing responses. Our results suggest that performing experimental procedures during the dark phase of the light cycle may be a useful refinement mechanism, as procedures performed during this period had the greatest effect on behavioral outcomes in both males and females. Light cycle phase is an experimental variable that should be considered when designing experiments to maximize behavioral effects, including those in response to stress.