Conference Agenda

An increasing number of mutations associated with Autism Spectrum Disorders (ASDs) have been reported for several synaptic cell adhesion molecules such as the post-synaptic Neuroligins. Specifically, the substitution Arg451Cys (R451C) in Neuroligin3 affects folding of the extracellular domain of the protein that, as a consequence, is retained in the endoplasmic reticulum rather than being trafficked to the cell surface and degraded by the proteasome. The resulting cellular phenotype is characterized by impaired synaptic function, activation of the unfolded protein response and by altered social behaviors in the knock-in mouse expressing the human mutation in the endogenous gene.

In order to rescue the cellular and functional phenotype caused by the mutation in Neuroligin3, we have selected a class of compounds belonging to the glucocorticoid family for their ability to promote the exit of the mutant protein from the endoplasmic reticulum and to improve the exposure of R451C Neuroligin3 on the cell membrane in the HEK-293 cell line. In particular we show that the treatment with dexamethasone promotes the formation of artificial synapses, between HEK-293 expressing the R451C mutant and primary hippocampal neurons. These results were then validated in a more physiological cellular system, consisting of neural stem cells that can be differentiated in either astrocytes or mixed neurons/astrocytes cultures, derived from the hippocampus of WT or R451C Neuroligin3 mice. Our results show that the treatment promotes the recovery of endogenous levels of mutant Neuroligin3, the synaptic functional impairments and lowers the activation of the unfolded protein response.

Sensory abnormalities are a common feature in autism spectrum disorders (ASD). Similarly, sensory deficits have been described in mice lacking ASD-associated genes. In our laboratory, we investigate somatosensory abnormalities in Cntnap2 and Shank3b mutant mice, two well-characterized mouse models of ASD. When compared to controls, both strains of mutant mice displayed impaired whisker-dependent discrimination in the textured novel object recognition test (tNORT). Shank3b but not Cntnap2 mutant mice also showed avoidance behavior responses to repetitive whisker stimulation. Impaired whisker-dependent behaviors were accompanied by altered c-fos mRNA induction following whisker stimulation, with Cntnap2 and Shank3b mutants showing c-fos mRNA up- and down-regulation within the primary somatosensory cortex (S1). The different c-fos mRNA induction profiles observed in the two mutant strains were paralleled by different connectivity within S1: resting-state fMRI revealed S1 hyper- and hypo-connectivity in Cntnap2 and Shank3b mutant mice, respectively. Preliminary data suggest that somatosensory abnormalities observed in Cntnap2 and Shank3b mutant mice are associated to neuroimmune dysfunction. Our data indicate that are Cntnap2 and Shank3b mutant mice are reliable models to investigate somatosensory abnormalities that characterize ASD.

Social memory, meaning the ability to discriminate between novel and familiar subjects is altered in ASD. Cholinergic (ChAT) neurons are required for this cognitive function and are altered in ASD.

Some forms of ASD are associated with mutations/deletions in genes encoding for synaptic proteins including the neuroligin 3 (NLG3). NLG3 is a postsynaptic adhesion molecule that binds its presynaptic partner neurexin and stabilizes both excitatory and inhibitory synapses. Here, I will discuss data showing that cholinergic dysfunction is associated to social memory deficits observed in NLG3-lacking (KO) mice, a model of ASD. Similar results were obtained also by conditional suppression of Nlg3 expression in ChAT neurons, using a shRNA-based viral strategy, corroborating the evidence that a cholinergic dysfunction may cause social memory deficits in ASD.

Autism and related developmental disorders encompass a wide range of heterogeneous conditions characterized by social and communicative impairment, as well as restricted and repetitive behaviors. While the etiology of these disorders is strongly influenced by genetics, how diverse genetic alterations converge to produce the behavioral and etiopathological manifestations remains unclear. A popular theory posits that an imbalance between excitatory and inhibitory (E-I) activity as a result of genetic or developmental insult might contribute (or underlie) to the etiology of these pathologies, and the related alteration in social interactions. However, controversy exists about whether E-I imbalance observed in patients and animal models is a direct, causal contributor to underlying pathology, or if it is instead a compensatory, adaptive epiphenomenal phenotype of limited etiological relevance. In an attempt to reconcile these views, here we test the hypothesis that transient E-I imbalance during early development is sufficient to alter the developmental trajectory of the brain, leading to long-lasting social phenotypes.

Cell-type specific increase of neuronal excitability was obtained with the use of intersectional genetics to express excitatory DREADD receptors in Vglut1-cre mice and the following chronic CNO treatment during the first two postnatal weeks. Longitudinal behavioral tests, resting state fMRI and RNA- sequencing were performed at multiple developmental stages.

Notably, chemogenetically increasing E-I balance during early development resulted in robustly impaired sociability that lasted throughout adulthood. Social alterations were not associated with anxiety-like phenotypes, impairments in olfactory abilities, tactile sensitivity, motor coordination, working or long-term memory. The same animals also exhibited disrupted fronto-hippocampal functional connectivity as measured with fMRI and patterns of altered gene expression, with a prominent dysregulation of ribosomal and synaptic related transcripts. These findings support a chronobiological reconceptualization of the E-I imbalance theory whereby early developmental insult is sufficient to produce long-lasting developmental derailment of relevance for these disorders.