Rapid, automated, and experimenter-free touchscreen testing reveals reciprocal interactions between cognitive flexibility and activity-based anorexia in female rats.

Anorexia nervosa has among the highest mortality rates of any psychiatric disorder and is characterized by cognitive inflexibility that persists after weight recovery and contributes to the chronic nature of the condition. What remains unknown is whether cognitive inflexibility predisposes individuals to anorexia nervosa, a question that is difficult to address in human studies. Our previous work using the most well-established animal model of anorexia nervosa, known as activity-based anorexia (ABA) identified a neurobiological link between cognitive inflexibility and susceptibility to pathological weight loss in female rats. However, testing flexible learning prior to exposure to ABA in the same animals has been thus far impossible due to the length of training required and the necessity of daily handling, which can itself influence the development of ABA. Here, we describe experiments that validate and optimize the first fully-automated and experimenter-free touchscreen cognitive testing system for rats and use this novel system to examine the reciprocal links between reversal learning (an assay of cognitive flexibility) and weight loss in the ABA model. First, we show substantially reduced testing time and increased throughput compared to conventional touchscreen testing methods because animals engage in test sessions at their own direction and can complete multiple sessions per day without experimenter involvement. We also show that, contrary to expectations, cognitive inflexibility measured by this reversal learning task does not predispose rats to pathological weight loss in ABA. Instead, rats that were predisposed to weight loss in ABA were more quickly able to learn this reversal task prior to ABA exposure. Intriguingly, we show reciprocal links between ABA exposure and cognitive flexibility, with ABA-exposed (but weight-recovered) rats performing much worse than ABA naïve rats on the reversal learning task, an impairment that did not occur to the same extent in rats exposed to food restriction conditions alone. On the other hand, animals that had been trained on reversal learning were better able to resist weight loss upon subsequent exposure to the ABA model. We also uncovered some stable behavioral differences between ABA susceptible versus resistant rats during touchscreen test sessions using machine learning tools that highlight possible predictors of anorectic phenotypes. These findings shed new light on the relationship between cognitive inflexibility and pathological weight loss and provide targets for future studies using the ABA model to investigate potential novel pharmacotherapies for anorexia nervosa.

Evaluating anhedonia in the activity-based anorexia (ABA) rat model.

Patients suffering anorexia nervosa (AN) become anhedonic, in other words, unable or unwilling to derive normal pleasures and avoid rewarding outcomes, most profoundly in food intake. The neurobiological underpinnings of anhedonia are likely to involve mesolimbic reward circuitry. We propose here that this circuitry and its involvement in AN can be investigated using the activity-based anorexia (ABA) rodent model that recapitulates many of the characteristics of the human condition, most notably rapid weight loss. Preference for sweetened water was used to assay hedonic processing in female Sprague-Dawley rats exposed to the ABA protocol, which involves free access to running wheels paired with time-limited access to food. This protocol uncovered a transient anhedonia in only one quarter of cases; however, exposure to running wheels alone was associated with a rapid aversion to sweetened water (F1.833, 20.17 = 78.29, p < .0001), and time-limited food access alone did not impact preference (F2.205, 24.25 = 0.305, p = .761). High levels of running wheel activity prior to the onset of food restriction increased susceptibility to body weight loss in ABA (F10,196.129 = 2.069, p = .029) and food anticipatory activity predicted subsequent food intake only for rats that were resistant to body weight loss (r = 0.44, p = .001). These data are inconsistent with the hypothesis that anhedonia underscores the precipitous weight loss in ABA, however, they highlight the predictive nature of hyperactivity in susceptibility to the ABA paradigm. These results will help inform the neurobiological framework of ABA and provide insight into the mechanisms of reward relevant to feeding and weight loss.

The Role of Mesolimbic Reward Neurocircuitry in Prevention and Rescue of the Activity-Based Anorexia (ABA) Phenotype in Rats.

Patients suffering from anorexia nervosa (AN) become anhedonic; unable or unwilling to derive normal pleasures and avoid rewarding outcomes, most profoundly in food intake. The activity-based anorexia (ABA) model recapitulates many of the characteristics of the human condition, including anhedonia, and allows investigation of the underlying neurobiology of AN. The potential for increased neuronal activity in reward/hedonic circuits to prevent and rescue weight loss is investigated in this model. The mesolimbic pathway extending from the ventral tegmental area (VTA) to the nucleus accumbens (NAc) was activated using a dual viral strategy, involving retrograde transport of Cre (CAV-2-Cre) to the VTA and coincident injection of DREADD receptors (AAV-hSyn-DIO-hM3D(Gq)-mCherry). Systemic clozapine-n-oxide (CNO; 0.3 mg/kg) successfully recruited a large proportion of the VTA-NAc dopaminergic projections, with activity evidenced by colocalization with elevated levels of Fos protein. The effects of reward circuit activation on energy balance and predicted survival was investigated in female Sprague-Dawley rats, where free access to running wheels was paired with time-limited (90 min) access to food, a paradigm (ABA) which will cause anorexia and death if unchecked. Excitation of the reward pathway substantially increased food intake and food anticipatory activity (FAA) to prevent ABA-associated weight loss, while overall locomotor activity was unchanged. Similar activation of reward circuitry, delayed until establishment of the ABA phenotype, rescued rats from their precipitous weight loss. Although these data are consistent with shifts primarily in food intake, the contribution of mechanisms including energy expenditure to survival remains to be determined. These results will inform the neurobiological underpinnings of AN, and provide insight into the mechanisms of reward circuitry relevant to feeding and weight loss.