Astrocyte glypican 5 regulates synapse maturation and stabilization.

The maturation and stabilization of appropriate synaptic connections is a vital step in neural circuit development; however, the molecular signals underlying these processes are not fully understood. We show that astrocytes, through production of glypican 5 (GPC5), are required for maturation and refinement of synapses in the mouse cortex during the critical period. In the absence of astrocyte GPC5, thalamocortical synapses show structural immaturity, including smaller presynaptic terminals, decreased postsynaptic density area, and presence of more postsynaptic partners at multisynaptic connections.

Dissecting the functional heterogeneity of glutamatergic synapses with high-throughput optical physiology.

Fluorescent reporters for glutamate release and postsynaptic Ca signaling are essential tools for quantifying synapse functional heterogeneity across neurons and circuits. However, leveraging these probes for neuroscience requires scalable experimental frameworks. Here, we devised a high-throughput approach to efficiently collect and analyze hundreds of optical recordings of glutamate release activity at presynaptic boutons in cultured rat hippocampal neurons.

A trainee-informed model for undergraduate neuroscience research programs serving marginalized students.

Undergraduate research programs improve career outcomes for historically marginalized students in the US, but low retention rates in postgraduate research persist. As graduate students and postdocs, we present a combination of trainee-informed approaches for tailoring summer research programs to these students’ needs and share key materials to facilitate adoption of these approaches at other institutions.

Protocol for the purification and transcriptomic analysis of mouse astrocytes using GFAT.

Astrocytes are glial cells of the central nervous system that modulate neuronal function. Here, we present glyoxal-fixed astrocyte nuclei transcriptomics (GFAT), a protocol for the purification and transcriptomic analysis of astrocyte nuclei from the cortex and cerebellum of adult and aged fresh mouse brain. We describe steps for tissue dissection, glyoxal fixation, homogenization, nuclei isolation, antibody staining, fluorescence-activated cell sorting, and RT-qPCR or bulk RNA sequencing.

Phosphorylation of CREB at Serine 142 and 143 Is Essential for Visual Cortex Plasticity.

The transcription factor cAMP response element-binding protein (CREB) is involved in a myriad of cellular functions in the central nervous system. For instance, the role of CREB via phosphorylation at the amino-acid residue Serine (Ser)133 in expressing plasticity-related genes and activity-dependent neuronal plasticity processes has been extensively demonstrated. However, much less is known about the role of CREB phosphorylation at Ser142 and Ser143.

Voltage sensor movements of Ca1.1 during an action potential in skeletal muscle fibers.

The skeletal muscle L-type Ca channel (Ca1.1) works primarily as a voltage sensor for skeletal muscle action potential (AP)-evoked Ca release. Ca1.

Glia as sculptors of synaptic plasticity.

Glial cells are non-neuronal cells in the nervous system that are crucial for proper brain development and function. Three major classes of glia in the central nervous system (CNS) include astrocytes, microglia and oligodendrocytes. These cells have dynamic morphological and functional properties and constantly surveil neural activity throughout life, sculpting synaptic plasticity.

Bi-allelic Variants in METTL5 Cause Autosomal-Recessive Intellectual Disability and Microcephaly.

Intellectual disability (ID) is a genetically and clinically heterogeneous disorder, characterized by limited cognitive abilities and impaired adaptive behaviors. In recent years, exome sequencing (ES) has been instrumental in deciphering the genetic etiology of ID. Here, through ES of a large cohort of individuals with ID, we identified two bi-allelic frameshift variants in METTL5, c.

Long-Term Potentiation Requires a Rapid Burst of Dendritic Mitochondrial Fission during Induction.

Synaptic transmission is bioenergetically demanding, and the diverse processes underlying synaptic plasticity elevate these demands. Therefore, mitochondrial functions, including ATP synthesis and Ca handling, are likely essential for plasticity. Although axonal mitochondria have been extensively analyzed, LTP is predominantly induced postsynaptically, where mitochondria are understudied.

Elevated extracellular glucose and uncontrolled type 1 diabetes enhance NFAT5 signaling and disrupt the transverse tubular network in mouse skeletal muscle.

The transcription factor nuclear factor of activated T-cells 5 (NFAT5) is a key protector from hypertonic stress in the kidney, but its role in skeletal muscle is unexamined. Here, we evaluate the effects of glucose hypertonicity and hyperglycemia on endogenous NFAT5 activity, transverse tubular system morphology and Ca(2+) signaling in adult murine skeletal muscle fibers. We found that exposure to elevated glucose (25-50 mmol/L) increased NFAT5 expression and nuclear translocation, and NFAT-driven transcriptional activity.

DNA binding sites target nuclear NFATc1 to heterochromatin regions in adult skeletal muscle fibers.

We have previously demonstrated that Ca²+/calcineurin-dependent dephosphorylation of the transcription factor nuclear factor of activated T cells subtype 1 (NFATc1) during repetitive skeletal muscle activity causes NFAT nuclear translocation and concentration in subnuclear NFAT foci. We now show that NFAT nuclear foci colocalize with heterochromatin regions of intense staining by DAPI or TO-PRO-3 that are present in the nucleus prior to NFATc1 nuclear entry. Nuclear NFATc1 also colocalizes with the heterochromatin markers trimethyl-histone H3 (Lys9) and heterochromatin protein 1α.

Augmentation of Cav1 channel current and action potential duration after uptake of S100A1 in sympathetic ganglion neurons.

S100A1, a 21-kDa dimeric Ca2+-binding protein of the EF-hand type, is expressed in cardiomyocytes and is an important regulator of heart function. During ischemia, cardiomyocytes secrete S100A1 to the extracellular space. Although the effects of extracellular S100A1 have been documented in cardiomyocytes, it is unclear whether S100A1 exerts modulatory effects on other tissues in proximity with cardiac cells.

Alpha-adrenergic signalling activates protein kinase D and causes nuclear efflux of the transcriptional repressor HDAC5 in cultured adult mouse soleus skeletal muscle fibres.

The protein kinase PKD1 has recently been linked to slow fibre-type gene expression in fast skeletal muscle through phosphorylation of class II histone deacetylase (HDAC) molecules, resulting in nuclear efflux of HDAC and consequent activation of the transcription factor MEF2. However, possible upstream activators of PKD, and the time course and signalling pathway of downstream effectors have not been determined in skeletal muscle. Using fluorescent fusion proteins HDAC5-green fluorescent protein (GFP) and PKD1-mPlum expressed in fibres isolated from predominantly slow soleus muscle and maintained for 4 days in culture, we now show that alpha-adrenergic receptor activation by phenylephrine causes a transient, PKD-dependent HDAC5-GFP nuclear efflux.

Ca2+ signal summation and NFATc1 nuclear translocation in sympathetic ganglion neurons during repetitive action potentials.

NFATc-mediated gene expression constitutes a critical step during neuronal development and synaptic plasticity. Although considerable information is available regarding the activation and functionality of specific NFATc isoforms, in neurons little is known about how sensitive NFAT nuclear translocation is to specific patterns of electrical activity. Here we used high-speed fluo-4 confocal imaging to monitor action potential (AP)-induced cytosolic Ca2+ transients in rat sympathetic neurons.