Plavicki, J. S. | Search bioRxiv

bioRxiv2024

Loss of developmentally derived Irf8+ macrophages promotes hyperinnervation and arrhythmia in the adult zebrafish heart

Recent developments in cardiac macrophage biology have broadened our understanding of the critical functions of macrophages in the heart. As a result, there is further interest in understanding the independent contributions of distinct subsets of macrophage to cardiac development and function. Here, we demonstrate that genetic loss of interferon regulatory factor 8 (Irf8)-positive embryonic-derived macrophages significantly disrupts cardiac conduction, chamber function, and innervation in adult zebrafish. At 4 months post-fertilization (mpf), homozygous irf8st96/st96 mutants have significantly shortened atrial action potential duration and significant differential expression of genes involved in cardiac contraction. Functional in vivo assessments via electro- and echocardiograms at 12 mpf reveal that irf8 mutants are arrhythmogenic and exhibit diastolic dysfunction and ventricular stiffening. To identify the molecular drivers of the functional disturbances in irf8 null zebrafish, we perform single cell RNA sequencing and immunohistochemistry, which reveal increased leukocyte infiltration, epicardial activation, mesenchymal gene expression, and fibrosis. Irf8 null hearts are also hyperinnervated and have aberrant axonal patterning, a phenotype not previously assessed in the context of cardiac macrophage loss. Gene ontology analysis supports a novel role for activated epicardial-derived cells (EPDCs) in promoting neurogenesis and neuronal remodeling in vivo. Together, these data uncover significant cardiac abnormalities following embryonic macrophage loss and expand our knowledge of critical macrophage functions in heart physiology and governing homeostatic heart health.

systems biology↗

bioRxiv2022

Dysregulation of neural activity and microglia function following exposure to the global environmental contaminant perfluorooctane sulfonate (PFOS)

BackgroundPer- and polyfluoroalkyl substances (PFAS) are biopersistent pollutants that have become global contaminants as a result of their diverse applications in commerce and industry. While some in vitro and epidemiological studies have explored the neurotoxic potential of perfluorooctane sulfonate (PFOS), a prevalent PFAS congener, it is unknown how developmental exposure to PFOS affects neuronal communication and other developmentally critical neural cell types, including microglia. ObjectivesWe sought to determine the extent to which PFOS exposure disrupts brain health, neuronal activity, and neuron-microglia communication during brain development. In addition, while PFOS impairs humoral immunity, its impact on innate immune cells, including resident microglia, is unclear. As such, we aimed to determine whether microglia are cellular targets of PFOS and, if so, whether disrupted microglial development and/or function could contribute to or is influenced by PFOS-induced neural dysfunction. MethodsZebrafish were chronically exposed to either control solution (0.1% DMSO), 7 {micro}M PFOS, 14 {micro}M PFOS, 28 {micro}M PFOS, or 64 {micro}M perfluorooctanoic acid (PFOA). We used in vivo imaging and gene expression analysis to assess microglial populations in the developing brain and to determine shifts in microglial state. We functionally challenged microglial using a larval brain injury model and, to assess the neuronal signaling environment, performed functional neuroimaging experiments utilizing the photoconvertible calcium indicator CaMPARI. These studies were paired with optogenetic manipulations of neurons and microglia, an untargeted metabolome wide association study (MWAS), and larval swim behavior assessments. ResultsDevelopmental PFOS exposure resulted in a shift away from the homeostatic microglia state, as determined by functional and morphological changes, as well as transcriptional upregulation of the microglia activation gene p2ry12. PFOS-induced effects on microglia state exacerbated microglia responses to brain injury in the absence of increased cell death or inflammation. PFOS exposure also heightened neural calcium activity, and optogenetic silencing of neurons or microglia independently was sufficient to normalize microglial responses to injury. An untargeted MWAS of larval brains revealed PFOS-exposed larvae had neurochemical signatures of excitatory-inhibitory imbalance. Behaviorally, PFOS-exposed larvae also exhibited anxiety-like thigmotaxis. To test whether the neuronal and microglial phenotypes were specific to PFOS, we exposed embryos to PFOA, a known immunotoxic PFAS. PFOA did not alter thigmotaxis, neuronal activity, or microglial responses, further supporting a role for neuronal activity as a critical modifier of microglial function following PFOS exposure. DiscussionTogether, this study provides the first detailed account of the effects of PFOS exposure on neural cell types in the developing brain in vivo and adds neuronal hyperactivity as an important endpoint to assess when studying the impact of toxicant exposures on microglia function.

neuroscience↗