Biology subjects

Thome, C.

Publications and source records attributed to Thome, C..

7 recordsLinked to original sources

bioRxiv2025

HIF1α-dependent induction of the T-Type calcium channel CaV3.2 mediates hypoxia-induced neuronal hyperexcitability

Post-stroke epilepsy (PSE) is a leading cause of acquired epilepsy in adults, yet the molecular mechanisms linking post-ischemic hypoxia to neuronal hyperexcitability remain poorly understood. The transcription factor hypoxia-inducible factor 1 (HIF1) is a central mediator of the cellular response to hypoxia and may contribute to epileptogenesis by regulating ion channel expression. Here, we identify the T-type calcium channel CaV3.2 (encoded by Cacna1h) as a direct transcriptional target of HIF1 and demonstrate its role in hypoxia-induced network hyperexcitability. Oxygen-glucose deprivation followed by reoxygenation (OGD/R) induced a persistent increase in neuronal firing rate. In murine and human organotypic brain slices, hypoxia led to a marked increase in HIF1 and Cacna1h expression at both transcript and protein levels. Using neuronal cell lines and primary cortical neurons we show that HIF1 activation through HIF1 overexpression, consistently increases Cacna1h expression. In NS20Y cells, overexpression of a normoxia-stable HIF1 variant increased Cacna1h promoter activity in both fluorescent and dual-luciferase reporter assays. The same effect was observed in primary cortical neurons, where HIF1 overexpression also elevated network activity measured by multielectrode array recordings, replicating the effect of the OGD/R model. Together, these results establish HIF1 as a key transcriptional regulator of CaV3.2 in neurons and reveal a conserved hypoxia-HIF1-CaV3.2 pathway that enhances neuronal excitability. This mechanism may underlie hypoxia-induced network hyperactivity and contribute to the pathogenesis of post-ischemic epileptogenesis, offering a potential molecular target for intervention.

neuroscience↗

bioRxiv2025

Defined human tri-lineage brain microtissues

Microglia are the immune cells of the central nervous system and are thought to be key players in both physiological and disease conditions. Several microglial features are poorly conserved between mice and human, such as the function of the neurodegeneration-associated immune receptor Trem2. Induced pluripotent stem cell (iPSC)-derived microglia offer a powerful opportunity to generate and study human microglia. However, human iPSC-derived microglia often exhibit activated phenotypes in vitro, and assessing their impact on other brain cell types remains challenging due to limitations in current co-culture systems. Here, we developed fully defined brain microtissues, composed of human iPSC-derived neurons, astrocytes, and microglia, co-cultured in 2D or 3D formats. Our microtissues are stable and self-sufficient over time, requiring no exogenous cytokines or growth factors. All three cell types exhibit morphologies characteristic of their in vivo environment and show functional properties. Co-cultured microglia develop more homeostatic phenotypes compared to microglia exposed to exogenous cytokines. Hence, these tri-cultures provide a unique approach to investigate cell-cell interactions between brain cell types. We found that astrocytes and not neurons are sufficient for microglial survival and maturation, and that astrocyte-derived M-CSF is essential for microglial survival. Single-cell and single-nucleus RNA sequencing analyses nominated a network of reciprocal communication between cell types. Brain microtissues faithfully recapitulated pathogenic -synuclein seeding and aggregation, suggesting their usefulness as human cell models to study not only normal but also pathological cell biological processes.

neuroscience↗

bioRxiv2024

Variability of axon initial segment geometry and its impact on hippocampal pyramidal cell function

Action potentials, the primary information units of the nervous system, are usually generated at the axon initial segment (AIS). Changes in the length and position of the AIS are associated with alterations in neuronal excitability but there is only limited information about the baseline structural variability of the AIS. This work provides a comprehensive atlas of the diversity of proximal cell geometries across all anatomical axes of the murine hippocampus, encompassing dorsal-ventral, superficial-deep, and proximal-distal regions. We analyzed the morphology of 3,936 hippocampal pyramidal neurons in 12 animals of both sexes, focusing on AIS length, position, and their association with proximal cellular features such as the soma and dendritic geometries. Notably, neurons with axon-carrying dendrites were significantly more common in ventral compared to dorsal hippocampal areas, suggesting a functional adaptation to regional demands. Validation of this finding in human samples confirms the translational relevance of our murine model. We employed NEURON simulations to assess the functional implications of this variability. Here, variation in proximal geometry only minimally contributed to neuronal homeostasis, but instead increased heterogeneity of response patterns across neurons.

neuroscience↗

bioRxiv2023

Axon onset remodeling in response to network activity in mouse cortical neurons

In certain subtypes of pyramidal neurons, axons originate from basal dendrites, resulting in an axon-carrying dendrite branch with unique functional characteristics (AcD cells). The number of AcD cells increases during development, however, it is so far unclear whether neurons remodel their axon emergence throughout their lifetime in response to changes in network activity. To test our hypothesis of such large-scale morphological plasticity, we utilized in vitro and in vivo strategies in mouse primary somatosensory cortex to test whether network activity impacts axon onset. Based on data obtained by immunofluorescence, confocal microscopy and/or live-cell imaging, we show that neurons are indeed capable of changing the onset of their axon origin from somatic to dendritic and vice versa within a few days in vitro and that peripheral whisker manipulation and thus changes in sensory input drives large-scale morphological plasticity in vivo.

neuroscience↗

bioRxiv2023

Live imaging of excitable axonal microdomains in ankyrin-G-GFP mice

The axon initial segment (AIS) constitutes not only the site of action potential initiation, but also a hub for activity-dependent modulation of output generation. Recent studies shedding light on AIS function used predominantly post-hoc approaches since no robust murine in vivo live reporters exist. Here, we introduce a reporter line in which the AIS is intrinsically labeled by an ankyrin-G-GFP fusion protein activated by Cre recombinase, tagging the native Ank3 gene. Using confocal, superresolution, and two-photon microscopy as well as whole-cell patch-clamp recordings in vitro, ex vivo, and in vivo, we confirm that the subcellular scaffold of the AIS and electrophysiological parameters of labeled cells remain unchanged. We further uncover rapid AIS remodeling following increased network activity in this model system, as well as highly reproducible in vivo labeling of AIS over weeks. This novel reporter line allows longitudinal studies of AIS modulation and plasticity in vivo in real-time and thus provides a unique approach to study subcellular plasticity in a broad range of applications.

neuroscience↗

bioRxiv2022

Microglia integration into human midbrain organoids leads to increased neuronal maturation and functionality.

The human brain is a complex, three-dimensional structure. To better recapitulate brain complexity, recent efforts have focused on the development of human specific midbrain organoids. Human iPSC-derived midbrain organoids consist of differentiated and functional neurons, which contain active synapses, as well as astrocytes and oligodendrocytes. However, the absence of microglia, with their ability to remodel neuronal networks and phagocytose apoptotic cells and debris, represents a major disadvantage for the current midbrain organoid systems. Additionally, neuro-inflammation related disease modeling is not possible in the absence of microglia. So far, no studies about the effects of human iPSC-derived microglia on midbrain organoid neural cells have been published. Here we describe an approach to derive microglia from human iPSCs and integrate them into iPSC-derived midbrain organoids. Using single nuclear RNA Sequencing, we provide a detailed characterization of microglia in midbrain organoids as well as the influence of their presence on the other cells of the organoids. Furthermore, we describe the effects that microglia have on cell death and oxidative stress- related gene expression. Finally, we show that microglia in midbrain organoids affect synaptic remodeling and increase neuronal excitability. Altogether, we show a more suitable system to further investigate brain development, as well as neurodegenerative diseases and neuro- inflammation. Main Points- Macrophage precursors can be efficiently co-cultured with midbrain organoids, they integrate into the tissue and differentiate into microglia in 3D. - Organoids containing microglia have a smaller size and show a down-regulation of oxidative stress-related genes. - Organoids co-cultured with microglia show differences in genes related to synaptic remodeling and action potential, as well as a more spontaneous action potential firing.

neuroscience↗

bioRxiv2017

The DNA methylation landscape of glioblastoma disease progression shows extensive heterogeneity in time and space

Glioblastoma is characterized by widespread genetic and transcriptional heterogeneity, yet little is known about the role of the epigenome in glioblastoma disease progression. Here, we present genome-scale maps of the DNA methylation dynamics in matched primary and recurring glioblastoma tumors, based on a national population registry and a comprehensively annotated clinical cohort. We demonstrate the feasibility of DNA methylation mapping in a large set of routinely collected formalin-fixed paraffin-embedded (FFPE) samples, and we validate bisulfite sequencing as a multi-purpose assay that allowed us to infer a range of different genetic, epigenetic, and transcriptional tumor characteristics. Based on these data, we identified characteristic differences between primary and recurring tumors, links between DNA methylation and the tumor microenvironment, and an association of epigenetic tumor heterogeneity with patient survival. In summary, this study provides a resource for dissecting DNA methylation heterogeneity in genetically diverse and heterogeneous tumors, and it demonstrates the feasibility of integrating epigenomics, radiology, and digital pathology in a representative national cohort, leveraging samples and data collected as part of routine clinical practice.

cancer biology↗