Upper cortical layer-driven network impairment in schizophrenia Mykhailo Y. Batiuk1,6, Teadora Tyler2,6, Shenglin Mei3, Rasmus Rydbirk1, Viktor Petukhov1, Dora Sedmak4, Erzsebet Frank2, Virginia Feher2, Nikola Habek4, Qiwen Hu3, Anna Igolkina3,5, Lilla Roszik2, Ulrich Pfisterer1, Zdravko Petanjek4, Istvan Adorjan2,7, Peter V. Kharchenko3,7, Konstantin Khodosevich1,7 1Biotech Research and Innovation Centre (BRIC), Faculty of Health and Medical Sciences, University of Copenhagen, 2200 Copenhagen, Denmark 2Department of Anatomy, Histology and Embryology, Semmelweis University, Budapest, H-1085, Hungary 3Department of Biomedical Informatics, Harvard Medical School, Boston, Massachusetts, 02115, USA 4Croatian Institute for Brain Research & Center of Excellence for Basic, Clinical and Translational Neuroscience, School of Medicine, University of Zagreb, Zagreb, 10000, Croatia 5St. Petersburg Polytechnical University, St. Petersburg, 195251, Russia 6These authors contributed equally 7Correspondence: email@example.com (I.A.), firstname.lastname@example.org (P.V.K.) and email@example.com (K.K.) SUMMARY Schizophrenia is one of the most wide-spread mental disorders with complex and largely unknown etiology. To characterize the impact of schizophrenia at a cellular level, we performed single nucleus RNA sequencing of >170,000 neurons from the dorsolateral prefrontal cortex of patients with schizophrenia and matched controls (7 vs 11, respectively). In addition, >100,000 neurons were analyzed topographically by immunohistochemistry in an extended cohort of cases with schizophrenia and controls (10 vs 10). Upper layer subtypes of GABAergic interneurons showed most dramatic transcriptomic changes. These results were substantiated by histological analysis, which revealed a reduction in the density of calretinin, calbindin and parvalbumin GABAergic interneurons particularly in layer 2. Common effect of schizophrenia on supragranular neuronal networks was underlined by downregulation of protein processing genes and upregulation of neuronal development/plasticity genes across supragranular subtypes of principal neurons and GABAergic interneurons. These point towards general network impairment within upper cortical layers being a core substrate associated with schizophrenia symptomatology.