Stem cell-derived neurons from autistic individuals with SHANK3 mutation show morphogenetic abnormalities during early development.

Shank3 is a structural protein found predominantly at the postsynaptic density. Mutations in the SHANK3 gene have been associated with risk for autism spectrum disorder (ASD). We generated induced pluripotent stem cells (iPSCs) from control individuals and from human donors with ASD carrying microdeletions of SHANK3. In addition, we used Zinc finger nucleases to generate isogenic SHANK3 knockout human embryonic stem (ES) cell lines. We differentiated pluripotent cells into either cortical or olfactory placodal neurons. We show that patient-derived placodal neurons make fewer synapses than control cells. Moreover, patient-derived cells display a developmental phenotype: young postmitotic neurons have smaller cell bodies, more extensively branched neurites, and reduced motility compared with controls. These phenotypes were mimicked by SHANK3-edited ES cells and rescued by transduction with a Shank3 expression construct. This developmental phenotype is not observed in the same iPSC lines differentiated into cortical neurons. Therefore, we suggest that SHANK3 has a critical role in neuronal morphogenesis in placodal neurons and that early defects are associated with ASD-associated mutations.

Differential regulation of CaMKII inhibitor beta protein expression after exposure to a novel context and during contextual fear memory formation.

Understanding of the molecular basis of long-term fear memory (fear LTM) formation provides targets in the treatment of emotional disorders. Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) is one of the key synaptic molecules involved in fear LTM formation. There are two endogenous inhibitor proteins of CaMKII, CaMKII N alpha and N beta, which can regulate CaMKII activity in vitro. However, the physiological role of these endogenous inhibitors is not known. Here, we have investigated whether CaMKII N beta protein expression is regulated after contextual fear conditioning or exposure to a novel context. Using a novel CaMKII N beta-specific antibody, CaMKII N beta expression was analysed in the naïve mouse brain as well as in the amygdala and hippocampus after conditioning and context exposure. We show that in naïve mouse forebrain CaMKII N beta protein is expressed at its highest levels in olfactory bulb, prefrontal and piriform cortices, amygdala and thalamus. The protein is expressed both in dendrites and cell bodies. CaMKII N beta expression is rapidly and transiently up-regulated in the hippocampus after context exposure. In the amygdala, its expression is regulated only by contextual fear conditioning and not by exposure to a novel context. In conclusion, we show that CaMKII N beta expression is differentially regulated by novelty and contextual fear conditioning, providing further insight into molecular basis of fear LTM.