Molecular profiles of pyramidal neurons in the superior temporal cortex in schizophrenia

Abstract

Disrupted synchronized oscillatory firing of pyramidal neuronal networks in the cerebral cortex in the gamma frequency band (i.e., 30-100 Hz) mediates many of the cognitive deficits and symptoms of schizophrenia. In fact, the density of dendritic spines and the average somal area of pyramidal neurons in layer 3 of the cerebral cortex, which mediate both long-range (associational) and local (intrinsic) corticocortical connections, are decreased in subjects with this illness. To explore the molecular pathophysiology of pyramidal neuronal dysfunction, we extracted ribonucleic acid (RNA) from laser-captured pyramidal neurons from layer 3 of Brodmann's area 42 of the superior temporal gyrus (STG) from postmortem brains from schizophrenia and normal control subjects. We then profiled the messenger RNA (mRNA) expression of these neurons, using microarray technology. We identified 1331 mRNAs that were differentially expressed in schizophrenia, including genes that belong to the transforming growth factor beta (TGF-β) and the bone morphogenetic proteins (BMPs) signaling pathways. Disturbances of these signaling mechanisms may in part contribute to the altered expression of other genes found to be differentially expressed in this study, such as those that regulate extracellular matrix (ECM), apoptosis, and cytoskeletal and synaptic plasticity. In addition, we identified 10 microRNAs (miRNAs) that were differentially expressed in schizophrenia; enrichment analysis of their predicted gene targets revealed signaling pathways and gene networks that were found by microarray to be dysregulated, raising an interesting possibility that dysfunction of pyramidal neurons in schizophrenia may in part be mediated by a concerted dysregulation of gene network functions as a result of the altered expression of a relatively small number of miRNAs. Taken together, findings of this study provide a neurobiological framework within which specific hypotheses about the molecular mechanisms of pyramidal cell dysfunction in schizophrenia can be formulated.

Keywords: cerebral cortex; gene expression profiling; laser-capture microdissection; microRNA; schizophrenia.

Figure 1.

Identification of pyramidal neurons and LCM procedure. Photomicrograph of a pyramidal neuron in the STG visualized with Histogene stain (A). After identification (B), pyramidal neurons are captured (C) onto the LCM cap (D). (E) Representative virtual gels showing the distribution of sizes of amplified products after two rounds of linear amplification of RNA extracted from ˜500 pyramidal neurons from a normal control (C) and a schizophrenia (S) subject. L = ladder. (F) Heatmap of 1331 differentially expressed genes identified based on the stringency criteria of fold-change > 1.1 and FDR-corrected p < 0.05. Scale bars = 25 μm.

Figure 2.

Correlation analysis comparing fold-changes of selected genes determined by microarray and qRT-PCR. Comparison of fold-changes of differentially expressed genes within signaling pathways identified as dysregulated in schizophrenia (N = 6) and several randomly selected genes (N = 4) determined by microarray and qRT-PCR. BMP7 = bone morphogenetic protein 7; BMPR1A = bone morphogenetic protein receptor type IA; CLU = clusterin; HLA-A = major histocompatibility complex (MHC), class I, A; HPRT = hypoxanthine guanine phosphoribosyl transferase; P2RY14 = purinergic receptor P2Y, G-protein coupled, 14; MCPH1 = microcephalin 1; SMAD4 = mothers against decapentaplegic homolog 4; SMAD5 = mothers against decapentaplegic homolog 5; VCAN = versican.

Figure 3.

Representative diagram of TGF-β and BMP signaling pathways. Both TGF-β and BMP7 activate SMAD4 and CREB-binding protein, a transcription co-activator of SMAD, leading to the transcription of target genes that influence extracellular matrix composition, apoptosis, and synaptic/cytoskeletal plasticity. BMPR1A = bone morphogenetic protein receptor type IA; CBP = CREB-binding protein; SARA (ZFYVE9) = zinc finger, FYVE domain containing 9; SMAD4 = mothers against decapentaplegic homolog 4. Genes up-regulated in schizophrenia are depicted by red arrows.

Figure 4.

Hypothetical model of pyramidal cell dysfunction in schizophrenia. It is hypothesized that as a result of disinhibition-induced excitotoxic insult (see text for details), TGFβ signaling within pyramidal neurons is up-regulated, which may contribute to extracellular matrix abnormalities, dysregulated apoptosis, and impaired synaptic and cytoskeletal plasticity. Shown in this highly simplified diagram are some of the hub genes that can be targeted in future experiments in the testing of this hypothesis.

Figure 5.

Wisteria floribunda agglutinin (WFA) histochemical labeling of chondroitin sulfate proteoglycans (CSPGs) in the human cerebral cortex. ( A) CSPG-rich perineuronal nets (arrowheads). (B) WFA also labeled intracellular CSPGs (arrowhead). Scale bars = 5 μm (A and B).