Fecal microbiota transplantation of patients with anorexia nervosa did not alter flexible behavior in rats.

OBJECTIVE: Patients with anorexia nervosa (AN) are often anxious, display inflexible behavior and disrupted reward processing. Emerging evidence suggests that gut dysbiosis in patients contributes to the disease phenotype and progression. METHODS: In a preclinical study, we explored whether AN-derived microbiota impacts cognitive flexibility, anxiety, and dopamine signaling using fecal microbiota transplantation (FMT) in tyrosine hydroxylase-cre rats. We performed probabilistic reversal learning task (PRLT) at the baseline, after antibiotic treatment, and following FMT from patients with AN and controls. We assessed flexible behavior, task engagement, and ventral tegmental area (VTA) dopamine signaling during and in the absence of reward. Furthermore, anxiety-like behavior was evaluated with open field (OF) and elevated plus maze (EPM) tests. RESULTS: Neither antibiotic-induced dysbiosis nor AN FMT led to significant alterations in the number of reversals or lever press strategies after reinforced or nonreinforced lever presses (win and lose-stay) in the PRLT. However, the number of initiated trials decreased after antibiotic treatment while remaining unchanged after FMT. No significant differences were observed in VTA dopamine activity, anxiety measures in the OF and EPM tests. Microbiome analysis revealed limited overlap between the microbiota of the donors and recipients. DISCUSSION: No evidence was found that the microbiota of patients compared to controls, nor a depleted microbiome impacts cognitive flexibility. Nonetheless, antibiotic-induced dysbiosis resulted in reduced task engagement during the PRLT. The relatively low efficiency of the FMT is a limitation of our study and highlights the need for improved protocols to draw robust conclusions in future studies. PUBLIC SIGNIFICANCE: While our study did not reveal direct impacts of AN-associated gut microbiota on cognitive flexibility or anxiety behaviors in our preclinical model, we observed a decrease in task engagement after antibiotic-induced dysbiosis, underscoring that the presence of a gut microbiome matters. Our findings underscore the need for further refinement in FMT protocols to better elucidate the complex interplay between gut microbiota and behaviors characteristic of anorexia nervosa.

Cytokine and Microbiome Changes in Adolescents with Anorexia Nervosa at Admission, Discharge, and One-Year Follow-Up.

Anorexia nervosa (AN) is a severe eating disorder that predominantly affects females and typically manifests during adolescence. There is increasing evidence that serum cytokine levels are altered in individuals with AN. Previous research has largely focused on adult patients, assuming a low-grade pro-inflammatory state. The serum levels of the cytokine tumour necrosis factor-alpha (TNF-α), interleukin (IL)-1ß, IL-6 and IL-15, which are pro-inflammatory, were examined in 63 female adolescents with AN and 41 age-matched healthy controls (HC). We included three time points (admission, discharge, and 1-year follow-up) and investigated the clinical data to assess whether the gut microbiota was associated with cytokine alterations. Relative to the HC group, serum levels of IL-1ß and IL-6 were significantly lower during the acute phase (admission) of AN. IL-1ß expression was normalised to control levels after weight recovery. TNF-α levels were not significantly different between the AN and HC groups. IL-15 levels were significantly elevated in patients with AN at all time points. We found associations between cytokines and bodyweight, illness duration, depressive symptoms, and the microbiome. In contrast to most findings for adults, we observed lower levels of the pro-inflammatory cytokines IL-1ß and IL-6 in adolescent patients, whereas the level of IL-15 was consistently increased. Thus, the presence of inflammatory dysregulation suggests a varied rather than uniform pro-inflammatory state.

Epigenetic alterations in patients with anorexia nervosa-a systematic review.

Anorexia nervosa (AN) is a complex metabolic and psychological disorder that is influenced by both heritable genetic components and environmental factors. Exposure to various environmental influences can lead to epigenetically induced changes in gene expression. Epigenetic research in AN is still in its infancy, and studies to date are limited in determining clear, valid links to disease onset and progression are limited. Therefore, the aim of this systematic review was to compile and critically evaluate the available results of epigenetic studies specifically in AN and to provide recommendations for future studies. In accordance with the PRISMA guidelines, a systematic literature search was performed in three different databases (PubMed, Embase, and Web of Science) through May 2023. Twenty-three original papers or conference abstracts on epigenetic studies in AN were collected. Epigenome-wide association studies (EWASs), which analyze DNA methylation across the genome in patients with AN and identify potential disease-relevant changes in promoter/regulatory regions of genes, are the most promising for future research. To date, five EWASs on AN have been published, suggesting a potential reversibility of malnutrition-induced epigenetic changes once patients recover. Hence, determining differential DNA methylation levels could serve as a biomarker for disease status or early diagnosis and might be involved in disease progression or chronification. For future research, EWASs with a larger sample size, longitudinal study design and uniform methods should be performed to contribute to the understanding of the pathophysiology of AN, the development of individual interventions and a better prognosis for affected patients.

Reversibility of Endoplasmic Reticulum Stress Markers During Long-Term Glucose Starvation in Astrocytes.

Previous studies have demonstrated a brain volume decrease linked to long-term starvation in patients with anorexia nervosa (AN). Food intake is critically diminished in this disorder, leading to one of the highest mortality rates within the psychiatric disease spectrum. As reported in animal models, astrocytes seem to be the most affected cell type in AN. In a recently established primary cell culture model, an elevated unfolded protein response (UPR) was observed in long-term glucose semi-starved astrocytes. A well-functioning protein machinery is essential for every cell, and prolonged UPR will lead to cell death. As a nucleic acid stress-sensing pathway with the activator located in the endoplasmic reticulum, the regulation of the cGAS-STING pathway (cyclic GMP-AMP synthase/stimulator of interferon genes) was additionally investigated in the starvation context. In the current study, a glucose semi-starvation protocol of 15 days, during which cells were supplied with 2 mM glucose in the medium, was prolonged with an additional 6-day long recovery period. Our findings showed that increased UPR mRNA expression was reversible after re-establishing the standard glucose concentration of 25 mM. Furthermore, we were able to verify the presence of cGAS and STING in astrocytes with a characteristic presence of cGAS in the astrocyte nucleus during starvation. A correlation between STING and the glial fibrillary acidic protein (GFAP) could be established, hinting at a conditional presence of STING with a specific astrocyte phenotype.

MiRNA research-The potential for understanding the multiple facets of anorexia nervosa.

Anorexia nervosa (AN) has a multifaceted and complex pathology, yet major gaps remain in our understanding of factors involved in AN pathology. MicroRNAs (miRNAs) play a regulatory role in translating genes into proteins and help understand and treat diseases. An extensive literature review on miRNAs with AN and comorbidities has uncovered a significant lack in miRNA research. To demonstrate the importance of understanding miRNA deregulation, we surveyed the literature on depression and obesity providing examples of relevant miRNAs. For AN, no miRNA sequencing or array studies have been found, unlike other psychiatric disorders. For depression and obesity, screenings and mechanistic studies were conducted, leading to clinical studies to improve understanding of their regulatory influences. MiRNAs are promising targets for studying AN due to their role as signaling molecules, involvement in psychiatric-metabolic axes, and potential as biomarkers. These characteristics offer valuable insights into the disease's etiology and potential new treatment options. The first miRNA-based treatment for rare metabolic disorders has been approved by the FDA and it is expected that these advancements will increase in the next decade. MiRNA research in AN is essential to examine its role in the development, manifestation, and progression of the disease. PUBLIC SIGNIFICANCE: The current understanding of the development and treatment of AN is insufficient. miRNAs are short regulatory sequences that influence the translation of genes into proteins. They are the subject of research in various diseases, including both metabolic and psychiatric disorders. Studying miRNAs in AN may elucidate their causal and regulatory role, uncover potential biomarkers, and allow for future targeted treatments.

Gut Microbiota and Brain Alterations after Refeeding in a Translational Anorexia Nervosa Rat Model.

The gut microbiota composition is causally involved in the regulation of body weight. Through the gut-brain axis, microbiota play a role in psychiatric disorders including anorexia nervosa (AN). Previously, we showed microbiome changes to be associated with brain volume and astrocyte reductions after chronic starvation in an AN animal model. Here, we analyzed whether these alterations are reversible after refeeding. The activity-based anorexia (ABA) model is a well-established animal model that mimics several symptoms of AN. Fecal samples and the brain were analyzed. Like previous results, significant alterations in the microbiome were observed after starvation. After refeeding, including the normalization of food intake and body weight, α- and ß-diversity, as well as the relative abundance of specific genera, were largely normalized in starved rats. Brain parameters appeared to normalize alongside microbial restitution with some aberrations in the white matter. We confirmed our previous findings of microbial dysbiosis during starvation and showed a high degree of reversibility. Thus, microbiome alterations in the ABA model appear to be mostly starvation-related. These findings support the usefulness of the ABA model in investigating starvation-induced effects on the microbiota-gut-brain axis to help comprehend the pathomechanisms of AN and potentially develop microbiome-targeted treatments for patients.

Gut-Associated Lymphatic Tissue in Food-Restricted Rats: Influence of Refeeding and Probiotic Supplementation.

Anorexia nervosa (AN) is a severe and often chronic eating disorder that leads to alterations in the gut microbiome, which is known to influence several processes, such as appetite and body weight regulation, metabolism, gut permeability, inflammation, and gut-brain interactions. Using a translational activity-based anorexia (ABA) rat model, this study examined the effect of chronic food starvation, as well as multistrain probiotic supplementation and refeeding, on the structure of the gut and gut-associated lymphatic tissue (GALT). Our results indicated that ABA had an atrophic influence on intestinal morphology and increased the formation of GALT in the small bowel and colon. Higher formation of GALT in ABA rats appeared to be reversible upon application of a multistrain probiotic mixture and refeeding of the starved animals. This is the first time that increased GALT was found following starvation in the ABA model. Our results underscore a potential role of gut inflammatory alterations in the underlying pathophysiology of AN. Increased GALT could be linked to the gut microbiome, as probiotics were able to reverse this finding. These results emphasize the role of the microbiome-gut-brain axis in the pathomechanisms of AN and point to probiotics as potentially beneficial addendum in the treatment of AN.

The role of the brain-derived neurotrophic factor (BDNF) in anorexia nervosa.

The brain-derived neurotrophic factor (BDNF) is a growth factor belonging to the neurotrophin family which plays a pivotal role in the differentiation, survival, and plasticity of neurons in the central nervous system. Evidence suggests that BDNF is an important signal molecule in the regulation of energy balance and thus implicated in body weight control. The discovery of BDNF-expressing neurons in the paraventricular hypothalamus which is important in the regulation of energy intake, physical activity, and thermogenesis gives more evidence to the suggested participation of BDNF in eating behavior. Until now it remains questionable whether BDNF can be used as a reliable biomarker for eating disorders such as anorexia nervosa (AN) as available findings on BDNF levels in patients with AN are ambiguous. AN is an eating disorder characterized by a pathological low body weight in combination with a body image disturbance typically developing during adolescence. A severe drive for thinness leads to restrictive eating behavior often accompanied by physical hyperactivity. During therapeutic weight restoration an increase of BDNF expression levels seems desirable as it might improve neuronal plasticity and survival which is essential for learning processes and thereby essential for the success of the psychotherapeutic treatment of patients. On the contrary, the well-known anorexigenic effect of BDNF might favor relapse in patients as soon as the BDNF levels significantly increase during weight rehabilitation. The present review summarizes the association between BDNF and general eating behavior and especially focuses on the eating disorder AN. In this regard findings from preclinical AN studies (activity-based anorexia model) are outlined as well.

[Fecal Microbiota Transplants in the Context of (Child and Adolescent) Psychiatric Disorders]. / Fäkale Mikrobiotatransplantationen im Zusammenhang mit (kinder- und jugend-)psychiatrischen Erkrankungen.

Fecal Microbiota Transplants in the Context of (Child and Adolescent) Psychiatric Disorders Abstract: There has recently been a significant increase in interest in gut microbiota and its interaction with the brain (gut-brain axis). Not only are the findings of microbiome research interesting for basic scientists, they also offer relevant insights for clinical practice. A causal relationship between gut microbiome and various somatic diseases such as diabetes mellitus, inflammatory bowel diseases, and obesity as well as psychiatric diseases such as major depression, anxiety disorders, and eating disorders seems plausible. To study the causal relationship of intestinal bacteria with individual phenotypes, researchers apply so-called stool transplantations (fecal microbiota transplantations) in the preclinical context. For this purpose, they transfer microbiota samples from patients into laboratory animals to observe possible changes in phenotype. In the clinical context, fecal microbiota transplantation is already being used with therapeutic intentions for selected diseases, for example, recurrent infections with Clostridioides difficile or inflammatory bowel diseases; they have already become part of the official clinical guidelines for C. difficile. For many other diseases, however, including mental illnesses, the potential of using fecal transplantations for therapeutic purposes is still being explored. Previous findings suggest that the intestinal microbiome, particularly fecal microbiota transplantations, represent a promising starting point for new therapeutic approaches.

The effects of polyunsaturated fatty acid (PUFA) administration on the microbiome-gut-brain axis in adolescents with anorexia nervosa (the MiGBAN study): study protocol for a longitudinal, double-blind, randomized, placebo-controlled trial.

BACKGROUND: Anorexia nervosa (AN) is a severe psychiatric disease that often takes a chronic course due to insufficient treatment options. Emerging evidence on the gut-brain axis offers the opportunity to find innovative treatments for patients with psychiatric disorders. The gut microbiome of patients with AN shows profound alterations that do not completely disappear after weight rehabilitation. In previous studies, the administration of polyunsaturated fatty acids (PUFA) resulted in effects that might be beneficial in the treatment of AN, affecting the microbiome, body weight and executive functions. Therefore, the MiGBAN study aims to examine the effects of a nutritional supplementation with PUFA on the gut microbiome and body mass index (BMI) in patients with AN. METHODS: This is a longitudinal, double-blind, randomized, placebo-controlled trial. Within 2 years, 60 adolescent patients aged 12 to 19 years with AN will receive either PUFA or placebo for 6 months additional to treatment as usual. After 1 year, the long-term effect of PUFA on the gut microbiome and consecutively on BMI will be determined. Secondary outcomes include improvement of gastrointestinal symptoms, eating disorder psychopathology, and comorbidities. Additionally, the interaction of the gut microbiome with the brain (microbiome-gut-brain axis) will be studied by conducting MRI measurements to assess functional and morphological changes and neuropsychological assessments to describe cognitive functioning. Anti-inflammatory effects of PUFA in AN will be examined via serum inflammation and gut permeability markers. Our hypothesis is that PUFA administration will have positive effects on the gut microbiota and thus the treatment of AN by leading to a faster weight gain and a reduction of gastrointestinal problems and eating disorder psychopathology. DISCUSSION: Due to previously heterogeneous results, a systematic and longitudinal investigation of the microbiome-gut-brain axis in AN is essential. The current trial aims to further analyse this promising research field to identify new, effective therapeutic tools that could help improve the treatment and quality of life of patients. If this trial is successful and PUFA supplementation contributes to beneficial microbiome changes and a better treatment outcome, their administration would be a readily applicable additional component of multimodal AN treatment. TRIAL REGISTRATION: German Clinical Trials Register DRKS00017130 . Registered on 12 November 2019.

Fear and food: Anxiety-like behavior and the susceptibility to weight loss in an activity-based anorexia rat model.

Anorexia nervosa (AN) is a severe psychiatric disorder characterized by energy restriction, low body weight, a fear of gaining weight, and often excessive physical activity. Anxiety disorders appear to constitute a major risk factor for developing AN and are the most frequent comorbidity. Here, the influence of anxiety-like behavior prior to food restriction on increased physical activity, leading to greater susceptibility to weight loss, was tested in rats. Furthermore, the possible anxiolytic effect of starvation itself was analyzed. A chronic starvation model activity-based anorexia (ABA) was applied to mimic physiological and behavioral characteristics of AN. During the induction of starvation and acute starvation, food intake was reduced by 70% and the rats lost 25% of their body weight, which was kept stable to imitate chronic starvation. Anxiety-like behavior was quantified before and after chronic starvation using the elevated plus maze, based on rodents' aversion to open spaces. Anxiety-related behavior before food restriction was associated with increased running-wheel activity during habituation and during the induction of starvation, and predicted faster weight loss in ABA rats. Additionally, food-restricted animals showed less anxiety-like behavior after chronic starvation. Animals showing more anxiety-like behavior appear to be more susceptible to weight loss, partially mediated by increased physical activity. Anxiety-related behavior was associated with increased physical activity, which in turn was associated with more rapid weight loss. Our data let us assume that food restriction has an anxiolytic effect. These findings demonstrate the importance of considering anxiety disorders in patients with AN.

Brain Volume Loss, Astrocyte Reduction, and Inflammation in Anorexia Nervosa.

Anorexia nervosa is the third most common chronic disease in adolescence and is characterized by low body weight, body image distortion, weight phobia, and severe somatic consequences. Among the latter, marked brain volume reduction has been linked to astrocyte cell count reduction of about 50% in gray and white matter, while neuronal and other glial cell counts remain normal. Exact underlying mechanisms remain elusive; however, first results point to important roles of the catabolic state and the very low gonadal steroid hormones in these patients. They also appear to involve inflammatory states of "hungry astrocytes" and interactions with the gut microbiota. Functional impairments could affect the role of astrocytes in supporting neurons metabolically, neurotransmitter reuptake, and synapse formation, among others. These could be implicated in reduced learning, mood alterations, and sleep disturbances often seen in patients with AN and help explain their rigidity and difficulties in relearning processes in psychotherapy during starvation.

[The Gut Microbiome and Its Clinical Implications in Anorexia Nervosa]. / Das Darmmikrobiom und seine klinischen Implikationen im Kontext der Anorexia nervosa.

The Gut Microbiome and Its Clinical Implications in Anorexia Nervosa Abstract. The diverse interactions of the gut microbiome with the metabolism, the immune system, and the brain of the host are increasingly becoming to the forefront of relevant research. Studies suggest a connection between an altered intestinal microbiome and somatic diseases, such as colitis ulcerosa, Crohn's disease, and diabetes, as well as mental illnesses such as anxiety and depression. Patients with anorexia nervosa (AN) also show significant changes in their gut microbiome which seem to be associated, among other things, with a different energy uptake from food, immunological and inflammatory processes, genetic predisposition, hormonal changes, and possibly increased intestinal permeability. In rats, stool transplantation from patients with AN resulted in decreased appetite and weight as well as anxious and compulsive behavior. In this review, we summarize the possible mechanisms of interaction between the microbiome and the host, and present initial findings on the microbiome in AN. Research on nutritional interventions, for example, with prebiotics and probiotics or nutritional supplements such as omega-3 fatty acids, which aim to positively influence the intestinal microbiome, could lead to additional treatment options in the therapy of patients with AN.

Long-Term Glucose Starvation Induces Inflammatory Responses and Phenotype Switch in Primary Cortical Rat Astrocytes.

Astrocytes are the most abundant cell type in the brain and crucial to ensure the metabolic supply of neurons and their synapse formation. Overnutrition as present in patients suffering from obesity causes astrogliosis in the hypothalamus. Other diseases accompanied by malnutrition appear to have an impact on the brain and astrocyte function. In the eating disorder anorexia nervosa (AN), patients suffer from undernutrition and develop volume reductions of the cerebral cortex, associated with reduced astrocyte proliferation and cell count. Although an effect on astrocytes and their function has already been shown for overnutrition, their role in long-term undernutrition remains unclear. The present study used primary rat cerebral cortex astrocytes to investigate their response to chronic glucose starvation. Cells were grown with a medium containing a reduced glucose concentration (2 mM) for 15 days. Long-term glucose starvation increased the expression of a subset of pro-inflammatory genes and shifted the primary astrocyte population to the pro-inflammatory A1-like phenotype. Moreover, genes encoding for proteins involved in the unfolded protein response were elevated. Our findings demonstrate that astrocytes under chronic glucose starvation respond with an inflammatory reaction. With respect to the multiple functions of astrocytes, an association between elevated inflammatory responses due to chronic starvation and alterations found in the brain of patients suffering from undernutrition seems possible.

Gut microbiota and brain alterations in a translational anorexia nervosa rat model.

Anorexia nervosa (AN) is an eating disorder that leads to brain volume reduction and is difficult to treat since the underlying pathophysiology is poorly understood. The human gut microbiota is known to be involved in host metabolism, appetite- and bodyweight regulation, gut permeability, inflammation and gut-brain interactions. In this study, we used a translational activity-based anorexia (ABA) rat model including groups with food restriction, running-wheel access and a combination to disentangle the influences on the gut microbiota and associated changes in brain volume parameters. Our data demonstrated that chronic food restriction but not running-wheel activity had a major influence on the gut microbiota diversity and composition and reduced brain volume. Negative correlations were found between global brain weight and α-diversity, and astrocyte markers and relative abundances of the genera Odoribacter and Bifidobacterium. In contrast, the presence of lactobacilli was positively associated with white and grey brain matter volume. ABA and food-restricted rats are an interesting pre-clinical model to assess the causal influence of starvation on the gut microbiome and gut-brain interactions and can help to dissect the underlying pathophysiologic mechanisms relevant to AN.

The Role of Glial Cells in Regulating Feeding Behavior: Potential Relevance to Anorexia Nervosa.

Eating behavior is controlled by hypothalamic circuits in which agouti-related peptide-expressing neurons when activated in the arcuate nucleus, promote food intake while pro-opiomelanocortin-producing neurons promote satiety. The respective neurotransmitters signal to other parts of the hypothalamus such as the paraventricular nucleus as well as several extra-hypothalamic brain regions to orchestrate eating behavior. This complex process of food intake may be influenced by glia cells, in particular astrocytes and microglia. Recent studies showed that GFAP+ astrocyte cell density is reduced in the central nervous system of an experimental anorexia nervosa model. Anorexia nervosa is an eating disorder that causes, among the well-known somatic symptoms, brain volume loss which was associated with neuropsychological deficits while the underlying pathophysiology is unknown. In this review article, we summarize the findings of glia cells in anorexia nervosa animal models and try to deduce which role glia cells might play in the pathophysiology of eating disorders, including anorexia nervosa. A better understanding of glia cell function in the regulation of food intake and eating behavior might lead to the identification of new drug targets.

The reduction of astrocytes and brain volume loss in anorexia nervosa-the impact of starvation and refeeding in a rodent model.

Anorexia nervosa (AN) is an often chronic, difficult to treat illness that leads to brain volume reductions in gray and white matter. The underlying pathophysiology is poorly understood, despite its potential importance in explaining the neuropsychological deficits and clinical symptoms associated with the illness. We used the activity-based anorexia model (ABA), which includes food reduction and running wheel access in female rats to study brain changes after starvation and refeeding. Longitudinal animal MRI and post-mortem brain sections confirmed a reduction in the mean brain volumes of ABA animals compared to controls. In addition, the mean number of astrocytes was reduced by over 50% in the cerebral cortex and corpus callosum, while the mean number of neurons was unchanged. Furthermore, mean astrocytic GFAP mRNA expression was similarly reduced in the ABA animals, as was the mean cell proliferation rate, whereas the mean apoptosis rate did not increase. After refeeding, the starvation-induced effects were almost completely reversed. The observation of the astrocyte reduction in our AN animal model is an important new finding that could help explain starvation-induced neuropsychological changes in patients with AN. Astrocyte-targeted research and interventions could become a new focus for both AN research and therapy.

The Microbiome and Eating Disorders.

Growing interest exists in the association of gut bacteria with diseases, such as diabetes, obesity, inflammatory bowel disease, and psychiatric disorders. Gut microbiota influence the fermentation of nutrients, body-weight regulation, gut permeability, hormones, inflammation, immunology, and behavior (gut-brain axis). Regarding anorexia nervosa (AN), altered microbial diversity and taxa abundance were found and associated with depressive, anxious, and eating disorder symptoms. Potential mechanisms involve increased gut permeability, low-grade inflammation, autoantibodies, and reduced brain cell neogenesis and learning. Gut microbiome is strongly influenced by refeeding practices. Microbiota-modulating strategies like nutritional interventions or psychobiotics application could become relevant additions to AN treatment.

Establishment of a chronic activity-based anorexia rat model.

BACKGROUND: Anorexia nervosa (AN) is often a chronic eating disorder characterised by body image disturbance and low body weight often associated with starvation-induced amenorrhoea and excessive exercise. Activity-based anorexia (ABA) is an animal model representing many somatic aspects of this psychiatric illness. We systematically manipulated the extent and length of starvation and animal age to find the optimal parameters to study chronic starvation. NEW METHODS: Wistar rats had 24h/day running wheel access and received 40% of their baseline food intake until a 20% or 25% weight reduction was reached (acute starvation). This body weight was then maintained for two weeks (chronic starvation). The rats of different ages of 4 or 8 weeks were used to represent early and late adolescent animals, respectively. The complete absence of a menstrual cycle was defined as the primary outcome parameter. RESULTS: Acute starvation caused a disruption of the oestrous cycle in 58% of the animals. During chronic starvation, a complete loss of the oestrous cycle could be found. Furthermore, 4-week-old rats exhibited higher levels of hyperactivity and amenorrhoea than 8-week-old animals. A 20% starvation level led to 90% loss of cycle, while a 25% starvation level triggered complete loss. COMPARISON WITH EXISTING METHODS: Most current ABA models focus on acute starvation, while most patients are chronically ill. CONCLUSIONS: The optimal parameters to achieve complete amenorrhoea included early adolescence, chronic starvation and 25% weight loss. The new ABA model allows studying the effects of chronic AN on underlying behavioural, hormonal and brain pathobiology.