Epilepsy-Associated SCN2A-L1342P Mutation Drives Network Hyperexcitability and Widespread Transcriptomic Changes in Human Cortical Organoids
Year: 2025
Type: preprint
Abstract: Objective SCN2A pathogenic mutations, such as the recurrent heterozygous Nav1.2-L1342P, are monogenic causes of epilepsy. In this human-induced pluripotent stem cell model system, we aim to investigate the molecular and cellular mechanisms underlying the SCN2A-L1342P-associated pathology. Methods Using a human male iPSC reference line (KOLF) carrying the Nav1.2-L1342P mutation, we generated 3D cortical organoids for functional studies. Patch clamp and multi-electrode array (MEA) recordings, immunocytochemistry, and RNA sequencing were used to characterize the disease phenotypes. Results Nav1.2-L1342P organoid neurons displayed increased intrinsic excitability, and amplified excitatory post-synaptic currents, which are consistent with an increase in excitatory synapse formation revealed by PSD95/SYN1 immunostaining. Moreover, elevated network firing activity, as demonstrated by MEA, indicates a pronounced network hyperexcitability. Transcriptomic profiling of organoids carrying the Nav1.2-L1342P mutation further revealed significant alterations in synaptic, glutamatergic, and developmental pathways. Significance Our findings demonstrate that the Nav1.2-L1342P mutation drives a multifaceted disease phenotype, including network hyperexcitability and disruption of pathways related to neuronal and synaptic functions. These results advance our understanding of SCN2A-related Developmental and Epileptic Encephalopathy (DEE), laying a foundation for personalized interventions. Key Points Key Point 1 : Human neurons carrying epilepsy-causing SCN2A -L1342P display intrinsic hyperexcitability in cortical organoids. Key Point 2 : Synaptic transmission is enhanced in organoids carrying the SCN2A -L1324P mutation, consistent with the presence of elevated excitatory synapses, which leads to increased network hyperexcitability. Key Point 3: Transcriptomics analysis reveals that the SCN2A -L1342P mutation causes significant differential changes in forebrain developmental genes, synaptic and glutamatergic signaling pathways, and enhances cellular senescence and apoptosis.
Authors: Maria I. Olivero-Acosta, Morgan Robinson, Zhefu Que, Zaiyang Zhang, Hope Elizabeth Harlow, Vinayak Shankar, Seoyong Hong, Muhan Wang, Conrad M. Otterbacher, Hina Kadono, Manasi Halurkar, Harish Kothandaraman, Nadia Lanman, Trang Nguyen, Kyle Wettschurack, Benjamin Zirkle, Layan Yunis, Ningren Cui, Xiaoling Chen, Jingliang Zhang, Jiaxiang Wu, William C. Skarnes, Chongli Yuan, Feng Guo, Yang Yang
Language: English
Subfield: Psychiatry and Mental health
Field: Medicine
Domain: Health Sciences
Open Access status: green
OpenAlex ID: W4413306485