Abstract Title Hyperexcitation of Mediobasal Hypothalamic Glutamatergic Neurons Induces a Phenotype Resembling Anorexia Nervosa

Background Anorexia Nervosa (AN) is an eating disorder characterized by a voluntary restriction in food intake resulting in a sustained critically low body weight, hyperactivity, and increased anxiety. AN is among the deadliest psychiatric disorders, and the primary method of assessing its neurobiological basis is with animal models. However, current animal models restrict feeding, exogenously induce stress, and/or show unrelated phenotypes like normoactivity, dissimilar from the hyperactivity and self-induced starvation seen in AN. The Mediobasal hypothalamus (MBH) is known to be involved in the regulation of feeding and anxiety-related behaviors. In particular, once activated, glutamatergic neurons within the MBH promote these behaviors. However, these approaches acutely activated the neurons, and the behavioral changes were usually transient. Whether chronic activation of glutamatergic MBH neurons is involved in AN is unknown. Previous studies identified a bacterial sodium channel (NachBac), which is more readily activated and takes more time to inactivate than mammalian counterparts. This leads to constantly increased activity when expressed in mammalian neurons. Recent treatment-based rationale suggested chronic glutamatergic signaling may contribute to AN. Based on these observations, we hypothesized that chronic activation of glutamatergic neurons in the MBH will recapitulate the self-induced starvation observed in AN.

Methods We used the Cre-LoxP technique to achieve neuron-specific gene expression, where genes flanked with LoxP were expressed in the presence of Cre recombinase. We used mice expressing Cre driven by the promoter of vesicular glutamate transporter 2 (Vglut2, a marker gene for glutamatergic neurons), Vglut2-Cre, to target hypothalamic glutamatergic neurons. We stereotaxically injected an Adeno-Associated Virus with a FLip-EXcision vector, AAV-FLEX-NaChBac-GFP, into the MBH to express NaChBac in a Cre-dependent manner.

Results Interestingly, not only did we observe decreased feeding behavior, but we also observed increased locomotion, increased anxiety, and drastically decreased body weight, classically resembling AN.

Conclusion These results demonstrate that constitutive activation of glutamatergic MBH neurons recapitulates the AN phenotype. In contrast to the most-used AN mouse models, our model exhibits AN phenotype without feeding restriction or stress induction. Importantly, these results complement the growing rationale of increased glutamatergic signaling in the pathophysiology of AN, based on the use of the glutamate receptor antagonist, ketamine, in treating AN in mice and humans. These findings reveal neurobiological evidence of glutamatergic signaling hyperexcitation contributing to the AN phenotype and provide a more clinically relevant animal model to examine mechanistic insights on AN pathogenesis.