Telencephalon Organoids Derived from an Individual with ADHD Show Altered Neurodevelopment of Early Cortical Layer Structure.

Attention-deficit/hyperactivity disorder (ADHD) is a neurodevelopmental disorder that occurs in early childhood and can persist to adulthood. It can affect many aspects of a patient's daily life, so it is necessary to explore the mechanism and pathological alterations. For this purpose, we applied induced pluripotent stem cell (iPSC)-derived telencephalon organoids to recapitulate the alterations occurring in the early cerebral cortex of ADHD patients. We found that telencephalon organoids of ADHD showed less growth of layer structures than control-derived organoids. On day 35 of differentiation, the thinner cortex layer structures of ADHD-derived organoids contained more neurons than those of control-derived organoids. Furthermore, ADHD-derived organoids showed a decrease in cell proliferation during development from day 35 to 56. On day 56 of differentiation, there was a significant difference in the proportion of symmetric and asymmetric cell division between the ADHD and control groups. In addition, we observed increased cell apoptosis in ADHD during early development. These results show alterations in the characteristics of neural stem cells and the formation of layer structures, which might indicate key roles in the pathogenesis of ADHD. Our organoids exhibit the cortical developmental alterations observed in neuroimaging studies, providing an experimental foundation for understanding the pathological mechanisms of ADHD.

Self-organizing cortex generated from human iPSCs with combination of FGF2 and ambient oxygen.

Human brain development has generally been studied through the analysis of postmortem tissues because of limited access to fetal brain tissues. This approach, however, only provides information from the perspective of long-term development. To investigate the pathophysiology of neurodevelopmental disorders, it is necessary to understand the detailed mechanisms of human brain development. Recent advances in pluripotent stem cell (PSC) technologies enable us to establish in vitro brain models from human induced PSCs (hiPSCs), which can be used to examine the pathophysiological mechanisms of neurodevelopmental disorders. We previously demonstrated that self-organized cerebral tissues can be generated from human PSCs in a three-dimensional (3D) culture system. Here, we describe the cerebral tissues differentiated from hiPSCs in a further-optimized 3D culture. We found that treatment with FGF2 is helpful to form iPSC aggregates with efficiency. Neuroepithelial structures spontaneously formed with apico-basal polarity in the aggregates expressing forebrain marker FOXG1. The neuroepithelium self-forms a multilayered structure including progenitor zones (ventricular and subventricular zones) and neuronal zone (cortical plate). Furthermore, with the same level of oxygen (O2) as in ambient air (20% O2), we found that self-formation of cortical structures lasted for 70 days in culture. Thus, our optimized 3D culture for the generation of cortical structure from hiPSCs is a simple yet effective method.

Slow synaptic transmission mediated by TRPV1 channels in CA3 interneurons of the hippocampus.

Metabotropic glutamate receptors (mGluRs) modulate various neuronal functions in the central nervous system. Many studies reported that mGluRs have linkages to neuronal disorders such as schizophrenia and autism related disorders, indicating that mGluRs are involved in critical functions of the neuronal circuits. To study this possibility further, we recorded mGluR-induced synaptic responses in the interneurons of the CA3 stratum radiatum using rat hippocampal organotypic slice cultures. Electrical stimulation in the CA3 pyramidal cell layer evoked a slow inward current in the interneurons at a holding potential of -70mV in the presence of antagonists for AMPA/kainate receptors, NMDA receptors, GABAA receptors and GABAB receptors. The slow inward current was blocked in the absence of extracellular calcium, suggesting that this was a synaptic response. The slow excitatory postsynaptic current (EPSC) reversed near 0mV, reflecting an increase in a non-selective cationic conductance. The slow EPSC is mediated by group I mGluRs, as it was blocked by AP3, a group I mGluR antagonist. Neither a calcium chelator BAPTA nor a phospholipase C (PLC) inhibitor U73122 affected the slow EPSC. La(3+), a general TRP channel blocker or capsazepine, a selective TRPV1 channel antagonist significantly suppressed the slow EPSC. DHPG, a selective group I mGluRs agonist induced an inward current, which was suppressed by capsazepine. These results indicate that in the interneurons of the hippocampal CA3 stratum radiatum group I mGluRs activate TRPV1 channels independently of PLC and intracellular Ca(2+), resulting in the slow EPSC in the interneurons.

Association analysis of putative cis-acting polymorphisms of interleukin-19 gene with schizophrenia.

BACKGROUND: Genome-wide association studies (GWAS) and gene expression analyses have revealed that single nucleotide polymorphisms (SNPs) associated with multifactorial diseases, such as schizophrenia, are significantly more likely to be associated with expression quantitative trait loci (eQTL). It was recently suggested that an immune system imbalance plays an important role in the pathogenesis of schizophrenia. Interleukin-19 is a novel cytokine that may play multiple roles in immune regulation and various diseases. METHOD: We selected eight tag SNPs in the eQTL of the IL-19 gene. Seven of the SNPs are putative cis-acting SNPs. Then, we conducted a case-control study using two independent samples. The first sample comprised 567 schizophrenia patients and 710 controls, and the second sample comprised 677 schizophrenia patients and 667 controls. RESULT: We identified the TGAA haplotype as being significantly associated with schizophrenia (p=0.0036 and corrected p=0.0264), although a combined analysis of the TGAA haplotype with the replication samples exhibited a nominally significant difference (p=0.022 and corrected p=0.235). CONCLUSIONS: These results suggest that the IL-19 gene might slightly contribute to the genetic risk of schizophrenia. Thus, further research on the association of eQTL SNPs with schizophrenia is warranted.

A missense mutation in the ITGA8 gene, a cell adhesion molecule gene, is associated with schizophrenia in Japanese female patients.

BACKGROUND: Cell adhesion molecules (CAMs) play pivotal role in the development of the central nervous system (CNS) and have also been reported to play role in the pathophysiology of schizophrenia. Missense mutations in the CAMs genes might alter the binding of their ligands, increasing the vulnerability to develop schizophrenia. METHODS: We selected 15 missense mutations in the CAMs genes of the CNS reported in the Kyoto Encyclopedia of Genes and Genomes (KEGG) and examined the association between these mutations and schizophrenia in 278 patients and 284 control subjects (first batch). We also genotyped the positive single nucleotide polymorphisms (SNPs) in 567 patients and 710 control subjects (second batch) and in 635 patients and 639 control subjects (replication samples). RESULTS: Genotypic and allelic distributions of rs2298033 in the ITGA8 gene between the schizophrenia and control groups were significantly different in the first batch (p=0.005 and 0.007, respectively). Gender-based analysis revealed that the allelic and genotypic distributions of rs2298033 in the ITGA8 were significantly different between the schizophrenia and control groups among females in both batches (p=0.010, 0.011 and 0.0086, 0.010, respectively) but not among males. Combine analysis of rs2298033 with the replication samples revealed a more significant differences (p=0.0032; 0.0035 in the overall subjects and p=0.0024; 0.0025 in the female subjects, respectively). The significant differences for rs2802808 of the NFASC gene were only observed in the female subgroup of the first batch. CONCLUSION: These results suggest that the ITGA8 gene might have gender-specific roles in the development of schizophrenia. Further replication and functional studies are required to confirm these findings.

Haplotypes in the expression quantitative trait locus of interleukin-1ß gene are associated with schizophrenia.

Recent genome-wide association study (GWAS) and gene expression analyses have revealed that single nucleotide polymorphisms (SNPs) associated with complex diseases such as schizophrenia are significantly more likely to be associated with expression quantitative trait loci (eQTL). The interleukin-1ß (IL1B) gene has been strongly implicated in the susceptibility to schizophrenia. In order to test this association, we selected five tag SNPs in the eQTL of the IL1B gene and conducted a case-control study using two independent samples. The first sample comprised 528 schizophrenic patients and 709 controls and the second sample comprised 576 schizophrenic patients and 768 controls. We identified two SNPs and several haplotypes as being significantly associated with schizophrenia. Previous reports indicated that one major haplotype that was protective against schizophrenia reduced IL1B transcription, while two risk haplotypes for schizophrenia enhanced IL1B transcription. Therefore, we measured IL1B mRNA expression in PAXgene-stabilized whole blood from 40 schizophrenic patients and 40 controls to explore the possibility of using five tag SNPs as schizophrenic trait markers. A multiple regression analysis taking confounding factors into account revealed that the T allele of rs4848306 SNP, which is a protective allele for schizophrenia, predicted reduced change in IL1B mRNA expression, regardless of phenotype. Our results appear to support the previous hypothesis that IL1B contributes to the genetic risk of schizophrenia and warrant further research on the association of eQTL SNPs with schizophrenia.