The Effect of Dietary Copper and Zinc on Fear Extinction and Motor Coordination

Abstract

Dietary manipulation incorporates not only changes in nutritional content (e.g. biometal concentration) but also alterations in base formulas. This thesis is comprised of two separate experiments that aim to elucidate the precarious process of diet selection and how dietary manipulation can impact animal behavior. The first portion of this thesis examined two “control” diets previously administered in our laboratory: a Harlan 7012 feed (7012S) and a Harlan Copper Control (CC). Animals underwent fear conditioning and extinction paradigms as well as rotarod, behavioral tests that assess associative learning and motor coordination, respectively. There were no significant differences between these control diets; however, the CC animals consistently performed worse compared to the 7012S animals. The CC diet was created in order to mimic a 7012 base and ideally induce identical behavioral phenotypes, but our findings reveal that mere manipulation of dietary base formula can produce weakened performance in animal behaviors. The second portion of the study assessed the role of biometals – particularly copper (Cu) and zinc (Zn) – on behavior by utilizing a different 7012-base (7012N) that controlled for the seasonal availability of ingredients. Copper deficiency has been under investigation due to possible causative roles in several pathological disease states that are characterized by learning and motor impairments. A lack of Cu intake in the diet can produce a direct Cu deficiency whereas excess Zn can lead to indirect Cu deficiency via competition with Cu for absorption in the gut. Previous work conducted in our laboratory revealed that Cu administered in conjunction to Zn in drinking water remediated some behavioral deficits associated with excess Zn; therefore it is suggested that Zn supplementation may be associated with Cu deficiency. Direct and indirect Cu deficiencies were studied by the respective administration of reduced Cu diets and Zn-supplemented diets. Animals were raised on the 7012N, the 7012N with reduced Cu levels (7012N-Cu), the 7012N with 10 parts per million Zn added to drinking water (7012N+Zn), or the CC diet. Fear extinction learning and open field activity were assessed at approximately four months of age, and rotarod performance was assessed at two months and four months of age. Statistical analyses indicated strong extinction-based learning throughout the three days of testing but failed to find a main effect of diet. Rotarod performance at four months of age was significantly better than performance at two months of age, and performance improved over the three days of testing. Likewise, dietary manipulation failed to contribute to our statistical models. There were significant differences in open field activity: 7012N controls and 7012N-Cu animals moved significantly more compared to 7012N+Zn and CC animals. This finding indicates that our dietary manipulation may have confounded general locomotion and weight of animals. We also believe that the recent changes to the 7012 formula base to form the 7012N version may have impacted results. Both experiments demonstrate how dietary changes in base composition and biometal concentrations can impact behavioral performance in animal models. With these data, we propose stricter control over open-access diets and suggest stronger statistical analyses that incorporate diet assignments, weight, and behavioral outcomes.