Integrity and Functionality of the Hypothalamic Oxytocin System and the Effect of Oxytocin Treatment in Two Rat Models for Autism

Background: Social behavior deficits are a core symptom in autism spectrum disorder (ASD), which up to date have no pharmacological treatment. The hypothalamic oxytocin (OXT) system is a well-known modulator of social behavior, which brought interest in using the OXT peptide to treat social behavioral deficits. Our studies in a rat model for ASD, the Shank3-deficient rat, demonstrated that intracerebroventricular (I.C.V) OXT administration ameliorates attentional and social memory deficits. Clinical trials of OXT in ASD produced equivocal results raising questions about (1) whether this equivocally is driven by the heterogeneity in ASD, where some ASD subgroups may benefit better than others and (2) if the efficacy of OT treatment is dependent on the functionality of the OXT system and/or is sensitive to specific developmental window. Addressing these questions is challenging due to the lack of predictive biomarkers to identity relevant subgroups that might be most helped by OXT, and the lack of sufficient and diverse postmortem samples to determine how OXT-system is affected in different ASD subjects. Genetic animal models with mutations in ASD-associated genes are powerful tool to help overcome these limitations, yet to date only few studies have employed these models to tackle these limitations.

Objectives: We aim to determine the effect of ASD-associated mutations, specifically, Shank3 and Fmr1, on the integrity and functionality of the OXT hypothalamic system and to assess the effect of OXT treatment on behavioral deficits, during different developmental stages, using rat models with Shank3 and Fmr1 mutations.

Methods: To assess the integrity of the OXT system, we are applying diaminobenzidine-based method and using anti-OXT antibodies to visualize oxytocin-expressing cells within the hypothalamic periventricular nucleus (PVN) and their axonal projections to their target sites in the brain. To assess the functionality of the OXT system, we are testing the levels of OXT in the peripheral blood, brain and, cerebrospinal fluid, using micor-dialysis, ELISA, and mass spectrometry techniques. To study the effect of OXT administration on behavior, we use the social discrimination task and assess social recognition memory, which we found to be impaired in Shank3-deficits rats. We test performance of the rats on this task, before and after oxytocin intracerebroventricular (I.C.V) or intranasal administration, during different developmental periods.

Results: Our morphological analysis demonstrates that OXT-expressing neurons show increased OT immunoreactivity and dendritic swelling in Shank3-knockout (KO) rats, and decreased number of OXT-expressing cells in Fmr1-KO rats. We see no changes in OXT peripheral blood levels in Shank3-deficits rats. Behaviorally, we found that OXT I.C.V treatment during adulthood improves social-recognition memory deficits in Shank3-deficinet rats.

Conclusions: Our findings show that mutations in the Shank3 and Fmr1 genes disturb the integrity of the OXT hypothalamic system. Findings from Shank3-KO rats suggest that OXT release is impaired and that this impairment is specific to oxytocin release within the brain. Altogether, this implies that treatment with OXT may be beneficial in at least in two subset of individuals with ASD, with Shank3 and Fmr1 mutations, where the integrity of the OXT system could be potentially disturbed.