Distinct Defects in Synaptic Differentiation of Neocortical Neurons in Response to Prenatal Valproate Exposure

Abstract

Autism spectrum disorders (ASDs) are a heterogeneous group of neurodevelopmental disorders characterized by impairments in social interactions and stereotyped behaviors. Valproic acid (VPA) is frequently used to treat epilepsy and bipolar disorders. When taken during pregnancy, VPA increases the risk of the unborn child to develop an ASD. In rodents, in utero VPA exposure can precipitate behavioral phenotypes related to ASD in the offspring. Therefore, such rodent models may allow for identification of synaptic pathophysiology underlying ASD risk. Here, we systematically probed alterations in synaptic proteins that might contribute to autism-related behavior in the offspring of in utero VPA-exposed mice. Moreover, we tested whether direct VPA exposure of cultured neocortical neurons may recapitulate the molecular alterations seen in vivo. VPA-exposed neurons in culture exhibit a significant increase in the number of glutamatergic synapses accompanied by a significant decrease in the number of GABAergic synapses. This shift in excitatory/inhibitory balance results in substantially increased spontaneous activity in neuronal networks arising from VPA-exposed neurons. Pharmacological experiments demonstrate that the alterations in GABAergic and glutamatergic synaptic proteins and structures are largely caused by inhibition of histone deacetylases. Therefore, our study highlights an epigenetic mechanism underlying the synaptic pathophysiology in this ASD model.

Prenatal Valproate Exposure Differentially Affects Parvalbumin-Expressing Neurons and Related Circuits in the Cortex and Striatum of Mice

“…Of note, the absence of PV not only affects the output of the FSI, but also entails modifications in synaptic transmission between the cortical afferences and striatal FSI; in PV -/- mice short-term depression of EPSCs is increased in a similar time window (Wohr et al, 2015) as short-term facilitation is increased in PV-FSI (Orduz et al, 2013). We did not detect altered levels of PV in the forebrain samples of VPA mice, in contrast to recent findings by Iijima et al (2016), who reported a decrease in PV levels by >20% in lysates from neocortical cortex of PND15-21 mice. This apparent discrepancy might be related to the time point of their analyses: in the period from PND15-25, PV protein levels, the number of PV-immunoreactive neurons or of Pvalb neuron structures (e.g., boutons) increase considerably, e.g., seen in mouse or rat cortex (Alcantara et al, 1993; de Lecea et al, 1995; Huang et al, 1999), hippocampus (de Lecea et al, 1995; Chen et al, 2014), and cerebellar Pvalb neurons (Collin et al, 2005).…”

“…This decrease was also confirmed at the protein level: K v 3.1b protein levels determined by Western blot analysis were decreased to a similar extent, i.e., ∼40% reduction ( p = 0.0002) of K v 3.1b in forebrain samples of VPA mice compared to controls ( Figure 5B2 ), a finding in line with previous results from lysates of SSC of PND15-21 VPA mice (Iijima et al, 2016). We estimate that these results reflect decreased expression of K v 3.1 channels rather than loss of neurons expressing K v 3.1, since the overlap between cortical Pvalb neurons (unchanged in mPFC and SSC; Figures 3A and 4A ) and K v 3.1b is >95% (Chow et al, 1999).…”

“…Both, mRNA and protein levels of K v 3.1b were significantly decreased in forebrain extracts of VPA mice as also reported for SSC recently (Iijima et al, 2016). In the mouse cortex, K v 3.1b channels are exclusively expressed in Pvalb neurons (99% of all PV + neurons are K v 3.1b + and vice versa (Chow et al, 1999)), where they are necessary for maintaining the fast-spiking phenotype of these neurons (Chow et al, 1999; Hernandez-Pineda et al, 1999; Lien and Jonas, 2003).…”

“…Of note, our forebrain lysates of PND25 mice that were used for quantification of RNA and protein levels provide a lower spatial resolution than our results obtained by stereological counts. Interestingly in PND70-80 mice having reached adult PV expression levels with certainty, no differences existed in SSC PV levels between VPA and control mice (Iijima et al, 2016). …”

“…Interestingly, the decrease in cortical PV as well as K v 3.1 expression observed in the SSC of young (PND15–PND21) VPA mice is no longer present in adult (PND70–PND90) mice (Iijima et al, 2016). This would fit to the concept of homeostatic plasticity, which suggests the presence of adaptive/homeostatic regulatory mechanisms in the brain that aim to maintain the stability and functionality of neural circuits when challenged by environmental (e.g., VPA) or other insults during development (Turrigiano, 2011).…”