MODELLO SPERIMENTALE DI ANORESSIA NERVOSA “ACTIVITY-BASED ANOREXIA”: MODIFICAZIONI EPIGENETICHE DEL SISTEMA ENDOCANNABINOIDE, ALTERAZIONI DEI LIVELLI DELLE MONOAMINE NEL SISTEMA NERVOSO CENTRALE E DEGLI ACIDI GRASSI NEL PLASMA.

The hallmark signs of anorexia nervosa (AN) are abnormal eating patterns and a strong control over body weight accompanied by physical hyperactivity. This pathology is characterized by an high chronicity rate and frequent relapse. There have been significant advances in understanding biological factors that contribute to the onset and progression of AN. Several research pointed out a deregulation in many central (neurotransmitters) and peripheral (fatty acids) molecules implicated in the hedonic and homeostatic aspects of feeding behavior in AN condition. Number of evidence has shed light on the role of the endocannabinoid system (ECs) in AN. Anorexic patients displayed increased anandamide levels and alteration in the mRNA of CB1 receptor. Moreover, our previous study demonstrated the efficacy of the modulation of ECs in ameliorating anorexic-like traits (e.g., body weight loss, hyperactivity, endocrine deregulations) in female rats subjected to activity-based anorexia (ABA) rodent model of AN. On this basis, the first goal of our study was to evaluate the transcriptional regulation of genes coding for different components of the ECs in female ABA rats exposed to food restriction and having free access to a running wheel, at early (3 days) and late (6 days) stages of the ABA induction period. Our data evidenced a down-regulation of cannabinoid type-1 receptor gene (Cnr1) in the Hypothalamus and Nucleus Accumbens (NAcc) at the late stage of ABA induction in ABA rats. Moreover, pyrosequencing showed an increase in DNA methylation levels at Cnr1 promoter in the NAcc of ABA rats at day 6 of the induction phase. Another neurotransmitter system that is altered both in human and rodents AN conditions is the dopaminergic system. Many studies showed an imbalance in dopamine (DA) levels and and DA type 2 receptor (D2) receptor in AN patients. Also, administration of olanzapine, D2/3 receptor antagonist, ameliorate ABA traits in rats. Considering this, the second aim of our study was to evaluate in ABA rats, possible alterations in the levels of DA, its metabolites (e.g., 3,4-Dihydroxyphenylacetic acid and omovanillic acid ) and D2 in brain areas involved in reward and homeostatic aspects of feeding behavior. We observed, in ABA rats, increased DA levels in Prefrontal Cortex and NAcc and this enhancement persisted, only in the NAcc, after a period of recovery from the ABA condition, in which animals were fed ad libitum. D2 receptor levels were found raised in the Caudato Putamen and in the Shell of NAcc at the end of the induction phase, and this condition was reverted after the restoration period. Several lines of evidence demonstrate that AN patients display a deficit in plasma fatty acids (FA) composition due to the caloric restriction. FA are important for cellular membrane fluidity and their deficiency can lead to worsening AN comorbidities. Also, a recent study in our laboratory highlighted impairments in the brain content of FA derivates (e.g., prostaglandins, tromboxanes, hydroxyeicosatetraenoic acids) in ABA rats. In light of this, the third goal of our study was to evaluate plasma levels of their precursors in a modified ABA protocol that reproduces in animals a chronic condition similar to human. Results showed an imbalance in some of the most important polinsaturated fatty acids like arachidonic, α-linoleic and linoleic acids in the plasma of ABA rats. To conclude, our work highlights profound impairments of ECs and dopaminergic systems, as well as of the analyzed lipid mediators in female rats undergoing acute and chronic protocols of the ABA paradigm. Our results help to understand that these systems could represent pharmacological targets for a therapeutic action in the prevention and treatment of AN.