Effects of Isoflurane Exposure on Spatial Memory Performance and Neuronal Density in Mice: Evaluating a Potential Confound

Samuel Neff

Advisor: Craig McDonald, PhD, Department of Psychology

Committee Members: Jane Flinn, Geraldine Grant, Erin Murdoch

Johnson Center, #325, Meeting Room A
November 24, 2025, 09:00 AM to 11:00 AM

Abstract:

Mild traumatic brain injury, a form of traumatic brain injury (TBI) accounting for over 1 million of the reported TBI’s annually in the United States, is underreported in literature because of the transient effects and lack of targeted treatments. Although a single mild TBI may resolve easily without treatment, repeated mild traumatic brain injury (rmTBI) can prolong recovery time and increase the risk of long-term neurodegeneration. To investigate these effects, animal models have been developed to replicate rmTBI, with a vast majority of those techniques relying on the use of anesthetization in the form of isoflurane while administering each TBI to minimize pain and suffering. The administration of anesthetic, however, has been associated with many of the same behavioral and morphological effects observed from rmTBI administration, including neuronal death and spatial memory deficits, potentially confounding scientific outcomes. To assess whether isoflurane exposure influences rmTBI research findings in mice, two studies were conducted. Study 1 used a machine learning pipeline (DeepLabCut and a multilayer perceptron network) to classify swim strategies in the Morris Water Maze (MWM) among anesthetized groups and a non-anesthetized control group. Analyses revealed no significant differences across groups in latency to find the platform, distance travelled, or cognitive performance derived from swim strategy complexity. Study 2 investigated neuronal density among anesthetized groups and a non-anesthetized control group in 6 brain regions (CA1, CA3, PAC, IL, EC) using cresyl-violet staining. Analyses revealed no significant group differences in any of the brain regions analyzed. These results suggest that isoflurane does not appear to exert a strong confounding effect on behavioral or histological outcomes in rmTBI mouse models. However, the lack of observable differences between rmTBI and sham mouse groups highlights possible limitations in the injury model itself. Future research should be performed to better understand the relationships between rmTBI, anesthetization, behavior, and brain morphology, as well as to refine the rmTBI mouse model used in this study.