Serum response factor Is required for cortical axon growth but Is dispensable for neurogenesis and neocortical lamination

Previous studies have shown that neuron-specific deletion of serum response factor (SRF) results in deficits in tangential cell migration, guidance-dependent circuit assembly, activity-dependent gene expression, and synaptic plasticity in the hippocampus. Furthermore, SRF deletion in mouse embryonic stem cells causes cell deathin vitro. However, the requirement of SRF for early neuronal development including neural stem cell homeostasis, neurogenesis, and axonal innervations remains unknown. Here, we report that SRF is critical for development of major axonal tracts in the forebrain. Conditional mutant mice lacking SRF in neural progenitor cells (Srf-Nestin-cKO) exhibit striking deficits in cortical axonal projections including corticostriatal, corticospinal, and corticothalamic tracts, and they show a variable loss of the corpus callosum. Neurogenesis and interneuron specification occur normally in the absence of SRF and the deficits in axonal projections were not due to a decrease or loss in cell numbers. Radial migration of neurons and neocortical lamination were also not affected. No aberrant cell death was observed during development, whereas there was an increase in the number of proliferative cells in the ventricular zone from embryonic day 14 to day 18. Similar axonal tract deficits were also observed in mutant mice lacking SRF in the developing excitatory neurons of neocortex and hippocampus (Srf-NEX-cKO). Together, these findings suggest distinct roles for SRF during neuronal development; SRF is specifically required in a cell-autonomous manner for axonal tract development but is dispensable for cell survival, neurogenesis, neocortical lamination, and neuronal differentiation.</jats:p

SRF is required for maintenance of astrocytes in non-reactive state in the mammalian brain

Astrocytes play several critical roles in the normal functioning of the mammalian brain, including ion homeostasis, synapse formation, and synaptic plasticity. Following injury and infection or in the setting of neurodegeneration, astrocytes become hypertrophic and reactive, a process termed astrogliosis. Although acute reactive gliosis is beneficial in limiting further tissue damage, chronic gliosis becomes detrimental for neuronal recovery and regeneration. Several extracellular factors have been identified that generate reactive astrocytes; however, very little is known about the cell-autonomous transcriptional mechanisms that regulate the maintenance of astrocytes in the normal non-reactive state. Here, we show that conditional deletion of the stimulus-dependent transcription factor, serum response factor (SRF) in astrocytes

SRF-deficient astrocytes provide neuroprotection in mouse models of excitotoxicity and neurodegeneration

Reactive astrogliosis is a common pathological hallmark of CNS injury, infection, and neurodegeneration, where reactive astrocytes can be protective or detrimental to normal brain functions. Currently, the mechanisms regulating neuroprotective astrocytes and the extent of neuroprotection are poorly understood. Here, we report that conditional deletion of serum response factor (SRF) in adult astrocytes causes reactive-like hypertrophic astrocytes throughout the mouse brain. Thes

DataSheet1_Region-specific heterogeneity in neuronal nuclear morphology in young, aged and in Alzheimer’s disease mouse brains.PDF

Neurons in the mammalian brain exhibit enormous structural and functional diversity across different brain regions. Compared to our understanding of the morphological diversity of neurons, very little is known about the heterogeneity of neuronal nuclear morphology and how nuclear size changes in aging and diseased brains. Here, we report that the neuronal cell nucleus displays differences in area, perimeter, and circularity across different anatomical regions in the mouse brain. The pyramidal neurons of the hippocampal CA3 region exhibited the largest area whereas the striatal neuronal nuclei were the smallest. These nuclear size parameters also exhibited dichotomous changes with age across brain regions–while the neocortical and striatal neurons showed a decrease in nuclear area and perimeter, the CA3 neurons showed an increase with age. The nucleus of parvalbumin- and calbindin-positive interneurons had comparable morphological features but exhibited differences between brain regions. In the context of activity-dependent transcription in response to a novel environment, there was a decrease in nuclear size and circularity in c-Fos expressing neurons in the somatosensory cortex and hippocampal CA1 and CA3. In an APP/PS1 mutant mouse model of Alzheimer’s disease (AD), the neuronal nuclear morphology varies with plaque size and with increasing distance from the plaque. The neuronal nuclear morphology in the immediate vicinity of the plaque was independent of the plaque size and the morphology tends to change away from the plaque. These changes in the neuronal nuclear size and shape at different ages and in AD may be attributed to changes in transcriptional activity. This study provides a detailed report on the differences that exist between neurons in nuclear morphology and can serve as a basis for future studies.</p

Quantifying molecular aggregation by super resolution microscopy within an excitatory synapse from mouse hippocampal neurons

Summary: Super-resolution microscopy (SRM) has been widely adopted to probe molecular distribution at excitatory synapses. We present an SRM paradigm to evaluate the nanoscale organization heterogeneity between neuronal subcompartments. Using mouse hippocampal neurons, we describe the identification of the morphological characteristics of nanodomains within functional zones of a single excitatory synapse. This information can be used to correlate structure and function at molecular resolution in single synapses. The protocol can be applied to immunocytochemical/histochemical samples across different imaging paradigms.For complete details on the use and execution of this protocol, please refer to Kedia et al. (2021)

Centrosome Inheritance Does Not Regulate Cell Fate in Granule Neuron Progenitors of the Developing Cerebellum

An inherent asymmetry exists between the two centrosomes of a dividing cell. One centrosome is structurally more mature (mother centrosome) than the other (daughter centrosome). Post division, one daughter cell inherits the mother centrosome while the other daughter cell inherits the daughter centrosome. Remarkably, the kind of centrosome inherited is associated with cell fate in several developmental contexts such as in radial glial progenitors in the developing mouse cortex, Drosophila neuroblast divisions and in Drosophila male germline stem cells. However, the role of centrosome inheritance in granule neuron progenitors in the developing cerebellum has not been investigated. Here, we show that mother and daughter centrosomes do exist in these progenitors, and the amount of pericentriolar material (PCM) each centrosome possesses is different. However, we failed to observe any correlation between the fate adopted by the daughter cell and the nature of centrosome it inherited

SRF Phosphorylation by Glycogen Synthase Kinase-3 Promotes Axon Growth in Hippocampal Neurons

The growth of axons is an intricately regulated process involving intracellular signaling cascades and gene transcription. We had previously shown that the stimulus-dependent transcription factor, serum response factor (SRF), plays a critical role in regulating axon growth in the mammalian brain. However, the molecular mechanisms underlying SRF-dependent axon growth remains unknown. Here we report that SRF is phosphorylated and activated by GSK-3 to promote axon outgrowth in mouse hippocampal neurons. GSK-3 binds to and directly phosphorylates SRF on a highly conserved serine residue. This serine phosphorylation is necessary for SRF activity and for its interaction with MKL-family cofactors, MKL1 and MKL2, but not with TCF-family cofactor, ELK-1. Axonal growth deficits caused by GSK-3 inhibition could be rescued by expression of a constitutively active SRF. The SRF target gene and actin-binding protein, vinculin, is sufficient to overcome the axonal growth deficits of SRF-deficient and GSK-3-inhibited neurons. Furthermore, short hairpin RNA-mediated knockdown of vinculin also attenuated axonal growth. Thus, our findings reveal a novel phosphorylation and activation of SRF by GSK-3 that is critical for SRF-dependent axon growth in mammalian central neurons

Genetic rescue of functional senescence in synaptic and behavioral plasticity

STUDY OBJECTIVES: Aging has been linked with decreased neural plasticity and memory formation in humans and in laboratory model species such as the fruit fly, Drosophila melanogaster. Here, we examine plastic responses following social experience in Drosophila as a high-throughput method to identify interventions that prevent these impairments. PATIENTS OR PARTICIPANTS: Wild-type and transgenic Drosophila melanogaster. DESIGN AND INTERVENTIONS: Young (5-day old) or aged (20-day old) adult female Drosophila were housed in socially enriched (n = 35-40) or isolated environments, then assayed for changes in sleep and for structural markers of synaptic terminal growth in the ventral lateral neurons (LNVs) of the circadian clock. MEASUREMENTS AND RESULTS: When young flies are housed in a socially enriched environment, they exhibit synaptic elaboration within a component of the circadian circuitry, the LNVs, which is followed by increased sleep. Aged flies, however, no longer exhibit either of these plastic changes. Because of the tight correlation between neural plasticity and ensuing increases in sleep, we use sleep after enrichment as a high-throughput marker for neural plasticity to identify interventions that prolong youthful plasticity in aged flies. To validate this strategy, we find three independent genetic manipulations that delay age-related losses in plasticity: (1) elevation of dopaminergic signaling, (2) over-expression of the transcription factor blistered (bs) in the LNVs, and (3) reduction of the Imd immune signaling pathway. These findings provide proof-of-principle evidence that measuring changes in sleep in flies after social enrichment may provide a highly scalable assay for the study of age-related deficits in synaptic plasticity. CONCLUSIONS: These studies demonstrate that Drosophila provides a promising model for the study of age-related loss of neural plasticity and begin to identify genes that might be manipulated to delay the onset of functional senescence

Nanoscale regulation of Ca2+ dependent phase transitions and real-time dynamics of SAP97/hDLG

SAP97/hDLG is a ubiquitous, alternatively spliced, and conserved modular scaffolding protein involved in the organization cell junctions and excitatory synapses. Here, authors confirm that SAP97/hDLG condenses in to nanosized molecular domains in both heterologous cells and hippocampal pyramidal neurons. Authors demonstrate that in vivo and in vitro condensation, molecular signatures of nanoscale condensates and exchange kinetics of SAP97/hDLG is modulated by the local availability of alternatively spliced isoforms. Additionally, SAP97/hDLG isoforms exhibits a differential sensitivity to Ca2+ bound Calmodulin, resulting in altered properties of nanocondensates and their real-time regulatio