From synaptic ultrastructure to circuit connectivity: Conventional and Correlative EM to study neuronal dysfunction in Angelman Syndrome

Dendritic spines represent the post-synaptic compartment of most excitatory synapses, whose size and shape are finely regulated during development and experience-dependent remodelling of the neuronal network. Early and late onset neurological conditions often derive from mutations in genes accounting for synapse maturation and plasticity. Indeed, genomic alterations leading to unbalanced expression of the Ubiquitin Ligase-E3A (UBE3A) cause defects in brain development, resulting in neurodevelopmental disorders. Evidences for the importance of proper UBE3A dosage in the brain comes from findings showing that elevated UBE3A expression or activity in neurons is linked with the appearance of autistic-like phenotypes, whereas its reduction, due to the loss of the maternal copy of UBE3A gene (paternally imprinted in neurons), is causing Angelman Syndrome (AS)1,2. AS patients present with severe developmental delay, learning difficulties, little or no speech, movement and balance impairments, seizures, sleep disorders and autism-like behaviour. To explore whether the neuronal dysfunction in AS has a cell-autonomous origin, we in-utero electroporated CRISPR/Cas9 constructs to silence the endogenous UBE3A expression in a subset of layer 2/3 pyramidal neurons of the mouse somatosensory cortex. Furthermore, we exploited conventional EM to examine excitatory and inhibitory synapses structure in the cortical circuit of a classic AS model pan-neuronally depleted from UBE3A expression