DOI: 10.5061/dryad.6q573n63k
title: RNAseq data from: Medial prefrontal cortex samples of glutamate dehydrogenase-deficient mice, stress-exposed or -naive, and their Nestin-Cre+ controls

description: Glutamate abnormalities in the medial prefrontal cortex (mPFC) are
 associated with cognitive deficits. We previously showed that homozygous
 deletion of CNS glutamate dehydrogenase 1 (Glud1), a metabolic enzyme
 critical for glutamate metabolism, leads to schizophrenia-like behavioral
 abnormalities and increased mPFC glutamate; mice heterozygous for CNS
 Glud1 deletion (C-Glud1+/- mice) showed no cognitive or molecular
 abnormalities. Here, we examined the protracted behavioral and molecular
 effects of mild injection stress on C-Glud1+/- mice. We found spatial and
 reversal learning deficits, as well as large-scale mPFC transcriptional
 changes in pathways associated with glutamate and GABA signaling, in
 stress-exposed C-Glud1+/- mice, but not in their stress-naïve or
 C-Glud1+/+ littermates. Interestingly, these effects were observed several
 weeks following stress exposure, and the expression levels of specific
 glutamatergic and GABAergic genes differentiated between high and low
 reversal learning performance. An increase in MiR203-5p expression
 immediately following stress may provide a translational regulatory
 mechanism to account for the delayed effect of stress exposure on
 cognitive function. Our findings show that chronic glutamate abnormalities
 interact with acute stress to induce cognitive deficits, and resonate with
 gene x environment theories of schizophrenia. Stress-exposed C-Glud1+/-
 mice may model a schizophrenia high-risk population, which is uniquely
 sensitive to stress-related ‘trigger’ events.  
description: RNA was extracted from 28 mPFC
 samples (n=5-9 per group,
 approximately equal numbers of males and females) as previously
 described<sup>20,90</sup>, and sent to the Technion Genome Center for
 genome-wide RNA sequencing (RNA-seq) and bioinformatical analysis. The
 experiment was comprised of two batches; the first included samples from
 three of the four groups (C-Cre+/Control, C-Cre+/Stress and
 C-<em>Glud1</em><sup>+/-</sup>/Stress), and the
 second included new samples from the three aforementioned groups, and
 added the forth group
 (C-<em>Glud1</em><sup>+/-</sup>/Control). Findings
 from both batches were combined, taking into account batch and sex
 effects. RNA was prepared using the SMARTer Stranded Total RNA-Seq Kit v2
 – Pico preparation kit according to the manufacturer’s instructions.
 RNA-seq library preparation, sequencing (using the Illumina HiSeq 2500
 sequencer for first batch; Illumina NextSeq 550 for the second), and
 data analysis was performed by the Technion Genome Center. The quality of
 the libraries was evaluated using FASTQC (v 0.11.5), quality and adapter
 trimming was conducted via trim galore (uses cutadapt v 1.10), and mapping
 was conducted via Tophat2 v 2.1.0, (uses short read aligner Bowtie2 v
 2.2.6). At the end of this process, the total reads after trimming ranged
 between 35-46 million reads per sample. Gene counting was conducted via
 HTseq-count (v0.6.1). Due to the ribosomal depletion process in the
 library preparation protocol and the expected ribosomal sequence reads,
 counting was performed with a modified annotation file which includes the
 45s ribosome annotation for better accuracy. Only counted reads without
 45s were used for the subsequent analysis. Uniquely mapped reads, aligned
 with high confidence to a single genomic location, ranged between 25-38
 million reads per sample. Differential gene expression was performed by
 DESeq2 (v1.28.1). 
description: Application and version used:
 FASTQC (v 0.11.5)