Biology subjects

Habela, C. W.

Publications and source records attributed to Habela, C. W..

6 recordsLinked to original sources

bioRxiv2025

Development of a Human iPSC-Derived "Corticospinal Tract-on-a-Chip" for Neurodegenerative Disease Research

Degeneration of the corticospinal tract is a central feature in a number of neurodegenerative disorders and leads to significant disability. However, modeling corticospinal neuron (CSN) pathology and corticospinal connectivity in neurological disorders is particularly challenging. While rodent models are important for understanding early degeneration of CSN, interspecies differences in corticospinal connectivity and challenges of in vivo study suggest that human in vitro models of corticospinal biology may be ripe for development. Human induced pluripotent stem cells (hiPSC) are promising tools for overcoming intrinsic limitations that arise from physiological differences between rodents and humans. We have developed an innovative hiPSC-based microfluidic platform for modeling human CSN and spinal motor neuron (SpMN) connectivity. The incorporation of regionally specific astrocyte subtypes (cortical and spinal) in addition to CSNs and SpMNs in this newly designed system allows for the modeling of both regional and neural cell-subtype interactions. Using this model, multielectrode array electrophysiology reveals the maturation of both cortical and spinal motor neurons over the time course of 12 weeks. Retrograde labeling methods demonstrate synaptic connectivity between corticospinal and spinal motor neurons. Optogenetic strategies to selectively activate excitatory CNs attenuated by glutamate receptor antagonism confirms the functional relevance of the model. Incorporating morphological, electrophysiological and physiological measures of corticospinal connectivity, this platform is a versatile model for use in neurodegenerative disease research and for the future development of targeted CSN therapies. Significance StatementDegeneration of the corticospinal tract is a key feature of numerous neurodegenerative diseases, yet current in vitro models lack the anatomical and functional fidelity to study this system. We developed a human iPSC-derived "Corticospinal Tract-on-a-Chip" using a multielectrode array platform that incorporates regionally patterned cortical and spinal neurons and astrocytes. This model demonstrates structural and functional synaptic connectivity and enables longitudinal electrophysiological recordings. Critically, it supports compartment-specific manipulation and real-time analysis of CST network dynamics, capabilities lacking in existing systems. By mimicking human corticospinal physiology in vitro, this platform offers a novel tool for mechanistic investigation and preclinical testing of CST-targeted therapies. It holds broad relevance for studying disorders such as ALS, hereditary spastic paraplegia, and primary lateral sclerosis.

neuroscience↗

bioRxiv2024

Spatiotemporal coordination of actin regulators generates invasive protrusions in cell-cell fusion

Invasive membrane protrusions play a central role in a variety of cellular processes. Unlike filopodia, invasive protrusions are mechanically stiff and propelled by branched actin polymerization. However, how branched actin filaments are organized to create finger-like invasive protrusions is unclear. Here, by examining the mammalian fusogenic synapse, where invasive protrusions are generated to promote cell membrane juxtaposition and fusion, we have uncovered the mechanism underlying invasive protrusion formation. We show that two nucleation promoting factors (NPFs) for the Arp2/3 complex, WAVE and N-WASP, exhibit different localization patterns in the protrusions. While WAVE is closely associated with the plasma membrane at the leading edge of the protrusive structures, N-WASP is enriched with WIP along the actin bundles in the shafts of the protrusions. During protrusion initiation and growth, the Arp2/3 complex nucleates branched actin filaments to generate low density actin clouds, in which the large GTPase dynamin organizes the new branched actin filaments into bundles, followed by actin-bundle stabilization by WIP, the latter functioning as a novel actin-bundling protein. Disrupting any of these components results in defective protrusions and failed myoblast fusion in cultured cells and mouse embryos. Taken together, our study has revealed the intricate spatiotemporal coordination between two NPFs and two actin-bundling proteins in building invasive protrusions at the mammalian fusogenic synapse, and has general implications in understanding invasive protrusion formation in cellular processes beyond cell-cell fusion.

cell biology↗

bioRxiv2024

Branched actin polymerization drives invasive protrusion formation to promote myoblast fusion during skeletal muscle regeneration

Skeletal muscle regeneration is a multistep process involving the activation, proliferation, differentiation, and fusion of muscle stem cells, known as satellite cells. The fusion of satellite cell-derived mononucleated muscle cells (SCMs) is indispensable for the generation of multinucleated, contractile myofibers during muscle repair. However, the molecular and cellular mechanisms underlying SCM fusion during muscle regeneration remain poorly understood. In this study, we uncovered an essential role for branched actin polymerization in SCM fusion. Using conditional knockouts of the Arp2/3 complex and its actin nucleation-promoting factors, N-WASP and WAVE, we demonstrated that branched actin polymerization is required for the SCM fusion, but not for satellite cell proliferation, differentiation, and migration. We showed that the N-WASP and WAVE complexes have partially redundant functions in regulating SCM fusion. Furthermore, we revealed that branched actin polymerization is essential for generating invasive protrusions at the fusogenic synapses in SCMs. Taken together, our study has identified new components of the myoblast fusion machinery in skeletal muscle regeneration and demonstrated a critical role for branched actin-propelled invasive protrusions in this process.

developmental biology↗

bioRxiv2024

Altered development and network connectivity in a human neuronal model of 15q11.2 deletion-related neurodevelopmental disorders

The chromosome 15q11.2 locus is deleted in 1.5% of patients with genetic epilepsy and confers a risk for intellectual disability and schizophrenia. Individuals with this deletion demonstrate increased cortical thickness, decreased cortical surface area and white matter abnormalities. Human induced pluripotent stem cell (iPSC)-derived neural progenitor cells (NPC) from 15q11.2 deletion individuals exhibit early adhesion junction and migration abnormalities, but later neuronal development and function have not been fully assessed. Imaging studies indicating altered structure and network connectivity in the anterior brain regions and the cingulum suggest that in addition to alterations in progenitor dynamics, there may also be structural and functional changes within discrete networks of mature neurons. To explore this, we generated human forebrain cortical neurons from iPSCs derived from individuals with or without 15q11.2 deletion and used longitudinal imaging and multielectrode array analysis to evaluate neuronal development over time. 15q11.2 deleted neurons exhibited fewer connections and an increase in inhibitory neurons. Individual neurons had decreased neurite complexity and overall decreased neurite length. These structural changes were associated with a reduction in multiunit action potential generation, bursting and synchronization. The 15q11.2 deleted neurons also demonstrated specific functional deficits in glutamate and GABA mediated network activity and synchronization with a delay in the maturation of the inhibitory response to GABA. These data indicate that deletion of the 15q11.2 region is sufficient to impair the structural and functional maturation of cortical neuron networks which likely underlies the pathologic changes in humans with the 15q11.2 deletion.

neuroscience↗

bioRxiv2023

Phase-Amplitude Coupling Detection and Analysis of Human 2-Dimensional Neural Cultures in Multi-well Microelectrode Array in Vitro

Human induced pluripotent stem cell (hiPSC)- derived neurons offer the possibility of studying human-specific neuronal behaviors in physiologic and pathologic states in vitro. However, it is unclear whether these cultured neurons can achieve the fundamental network behaviors that are required to process information in the human brain. Investigating neuronal oscillations and their interactions, as occurs in cross-frequency coupling (CFC), is potentially a relevant approach. Microelectrode array culture plates provide a controlled framework to study populations of hiPSC-derived cortical neurons (hiPSC-CNs) and their electrical activity. Here, we examined whether networks of two-dimensional cultured hiPSC-CNs recapitulate the CFC that is present in networks in vivo. We analyzed the electrical activity recorded from hiPSC-CNs grown in culture with hiPSC-derived astrocytes. We employed the modulation index method for detecting phase-amplitude coupling (PAC) and used an offline spike sorting method to analyze the contribution of a single neurons spiking activities to network behavior. Our analysis demonstrates that the degree of PAC is specific to network structure and is modulated by external stimulation, such as bicuculine administration. Additionally, the shift in PAC is not driven by a single neurons properties but by network-level interactions. CFC analysis in the form of PAC explores communication and integration between groups of nearby neurons and dynamical changes across the entire network. In vitro, it has the potential to capture the effects of chemical agents and electrical or ultrasound stimulation on these interactions and may provide valuable information for the modulation of neural networks to treat nervous system disorders in vivo. SignificancePhase amplitude coupling (PAC) analysis demonstrates that the complex interactions that occur between neurons and network oscillations in the human brain, in vivo, are present in 2-dimensional human cultures. This coupling is implicated in normal cognitive function as well as disease states. Its presence in vitro suggests that PAC is a fundamental property of neural networks. These findings offer the possibility of a model to understand the mechanisms and of PAC more completely and ultimately allow us to understand how it can be modulated in vivo to treat neurologic disease.

neuroscience↗

bioRxiv2019

Persistent Cyfip1 expression is required to maintain the adult subventricular zone neurogenic niche

Neural stem cells (NSCs) persist throughout life in the subventricular zone (SVZ) niche of the lateral ventricles as B1 cells. Maintaining this population of NSCs depends on the balance between quiescence and self-renewing or self-depleting proliferation. Interactions between B1 cells and the surrounding niche are important in regulating this balance, but the mechanisms governing these processes have not been fully elucidated in adult mammals. The cytoplasmic FMRP-interacting protein (CYFIP1) regulates apical-basal polarity in the embryonic brain. Loss of Cyfip1 during embryonic development in mice disrupts the embryonic niche and affects cortical neurogenesis. However, a direct role for Cyfip1 in the regulation of adult NSCs has not been established. Here, we demonstrate that Cyfip1 expression is preferentially localized to B1 cells in the adult SVZ. Loss of Cyfip1 in the embryonic mouse brain results in altered adult SVZ architecture and expansion of the adult B1 cell population at the ventricular surface. Furthermore, acute deletion of Cyfip1 in adult NSCs results in a rapid change in adherens junction proteins as well as increased proliferation and the number of B1 cells at the ventricular surface. Together, these data indicate that CYFIP1 plays a critical role in the formation and maintenance of the adult SVZ niche and, furthermore, deletion of Cyfip1 unleashes the capacity of adult B1 cells for symmetric renewal to increase the adult NSC pool.\n\nSIGNIFICANCENeural stem cells (NSCs) persist in the subventricular zone (SVZ) of the lateral ventricles in adult mammals and their population is determined by the balance between quiescence and self-depleting or renewing cell division. The mechanisms regulating their biology are not fully understood. This study establishes that the cytoplasmic FMRP interacting protein 1 (Cyfip1) regulates NSC fate decisions in the adult SVZ and NSCs that are quiescent or typically undergo self-depleting divisions retain the ability to self-renew in the adult. This contributes to our understanding of how adult NSCs are regulated throughout life and has potential implications for human brain disorders.

developmental biology↗