Biology subjects

Schmied, K.

Publications and source records attributed to Schmied, K..

3 recordsLinked to original sources

bioRxiv2024

Temporal Dynamics of Neocortical Development in Organotypic Mouse Cultures: A Comprehensive Analysis

Murine organotypic brain slice cultures have been widely used in neuroscientific research and are offering the opportunity to study neuronal function under normal and disease conditions. Despite the brought application, the mechanisms governing the maturation of immature cortical circuits in vitro are not well understood. In this study, we present a detailed investigation into the development of the neocortex in vitro. Utilizing a holistic approach, we studied organotypic whole-hemisphere brain slice cultures from postnatal mice and tracked the development of the somatosensory area over a five-week period. Our analysis revealed the maturation of passive and active intrinsic properties of pyramidal cells together with their morphology, closely resembling in vivo development. Detailed Multi-electrode array (MEA) electrophysiological assessments and RNA expression profiling demonstrated stable network properties by two weeks in culture, followed by the transition of spontaneous activity towards more complex patterns including high-frequency oscillations. However, weeks 4 and 5 exhibited increased variability and initial signs of neuronal loss, highlighting the importance of considering developmental stages in experimental design. This comprehensive characterization is vital for understanding the temporal dynamics of the neocortical development in vitro, with implications for neuroscientific research methodologies, particularly in the investigation of diseases such as epilepsy and other neurodevelopmental disorders.

neuroscience↗

bioRxiv2023

Long-term human organotypic brain slice cultures: a detailed protocol to provide a comprehensive framework for single-neuron and neuronal network investigations

BackgroundThe investigation of the human brain at cellular and microcircuit level remains challenging due to the fragile viability of neuronal tissue, inter- and intra-variability of the samples and limited availability of human brain material. New methodHere, we present an optimized work-up to use resected tissue from brain surgeries for live cell experiments in vitro. Comparison with existing methodsWe provide a reworked, detailed protocol of the production, culturing and viral transduction of human organotypic brain slice cultures for research purposes. ResultsWe highlight the critical pitfalls of the culturing process of the human brain tissue and present results on viral expression, single-cell Patch-Clamp recordings, as well as multi-electrode array recordings over a prolonged period of time. Additionally, our statistics show that brain tissue from patients of any age and morbidity can be used for organotypic brain slice cultures if carefully selected. ConclusionsOrganotypic brain slice cultures are of great value for basic neuroscience and disease modeling over a time course of three weeks. HighlightsO_LILong-term human organotypic brain slice cultures are viable for 2-3 weeks and provide a framework for basic neuroscience and disease modeling C_LIO_LIWe provide a reworked, detailed protocol for organotypic human brain slice culture production and maintenance C_LIO_LIWe show results of long-term culturing of human organotypic brain slice cultures in terms of viral transduction, whole-cell Patch-Clamp recordings and multi-electrode array recordings C_LIO_LIStatistics of 16 surgeries show the correlation between the overall success rates of efficient culturing and viral transduction with the patient age and morbidity C_LI O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=139 SRC="FIGDIR/small/561508v1_fig1.gif" ALT="Figure 1"> View larger version (28K): org.highwire.dtl.DTLVardef@17c985aorg.highwire.dtl.DTLVardef@f651fdorg.highwire.dtl.DTLVardef@180d32eorg.highwire.dtl.DTLVardef@a896b5_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOFigure 1, Graphical abstract.C_FLOATNO Schematic overview of preparation, maintenance and experimental workup of human organotypic brain slice cultures. After surgical resection, human brain tissue is prepared, sliced and cultured at air-liquid-interface between human cerebrospinal fluid and defined incubator atmosphere, allowing for week-long viability. This enables extensive experimental workup such as viral transduction, single cell and multi-electrode array electrophysiological recordings. C_FIG

neuroscience↗

bioRxiv2022

The spike gene is a major determinant for the SARS-CoV-2 Omicron-BA.1 phenotype

Variant of concern (VOC) Omicron-BA1 has achieved global predominance in early 2022. Therefore, surveillance and comprehensive characterization of Omicron-BA.1 in advanced primary cell culture systems and multiple animal models is urgently needed. Here, we characterized Omicron-BA.1 and recombinant Omicron-BA.1 spike gene mutants in comparison with VOC Delta in well-differentiated primary human nasal and bronchial epithelial cells in vitro, followed by in vivo fitness characterization in naive hamsters, ferrets and hACE2-expressing mice, and in immunized hACE2-mice. We demonstrate a spike-mediated enhancement of early replication of Omicron-BA.1 in nasal epithelial cultures, but limited replication in bronchial epithelial cultures. In Syrian hamsters, Delta showed dominance over Omicron-BA.1 and in ferrets, Omicron-BA.1 infection was abortive. In mice expressing the authentic hACE2-receptor, Delta and a Delta spike clone also showed dominance over Omicron-BA.1 and an Omicron-BA.1 spike clone, respectively. Interestingly, in naive K18-hACE2 mice, we observed Delta spike-mediated increased replication and pathogenicity and Omicron-BA.1 spike-mediated reduced replication and pathogenicity, suggesting that the spike gene is a major determinant of both Delta and Omicron-BA.1 replication and pathogenicity. Finally, the Omicron-BA.1 spike clone was less well controlled by mRNA-vaccination in K18-hACE2-mice and became more competitive compared to the progenitor and Delta spike clones, suggesting that spike gene-mediated immune evasion is another important factor that led to Omicron-BA.1 dominance.

microbiology↗