Heterogeneous brain region-specific responses to astrocytic mitochondrial DNA damage in mice.

Astrocytes display context-specific diversity in their functions and respond to noxious stimuli between brain regions. Astrocytic mitochondria have emerged as key players in governing astrocytic functional heterogeneity, given their ability to dynamically adapt their morphology to regional demands on ATP generation and Ca buffering functions. Although there is reciprocal regulation between mitochondrial dynamics and mitochondrial Ca signaling in astrocytes, the extent of this regulation in astrocytes from different brain regions remains unexplored.

Heterogeneous brain region-specific responses to astrocytic mitochondrial DNA damage in mice.

Astrocytes use Ca signals to regulate multiple aspects of normal and pathological brain function. Astrocytes display context-specific diversity in their functions, and in their response to noxious stimuli between brain regions. Indeed, astrocytic mitochondria have emerged as key players in governing astrocytic functional heterogeneity, given their ability to dynamically adapt their morphology to regional demands on their ATP generation and Ca buffering functions.

Validation of terminal Schwann cell gene marker expression by fluorescent in situ hybridization using RNAscope.

Recent RNA-seq studies have generated a new crop of putative gene markers for terminal Schwann cells (tSCs), non-myelinating glia that cap axon terminals at the vertebrate neuromuscular junction (NMJ). While compelling, these studies did not validate the expression of the novel markers using in situ hybridization techniques. Here, we use RNAscope technology to study the expression of top candidates from recent tSC and non-myelinating Schwann cell marker RNA-seq studies.

Extracellular signal-regulated kinases 1 and 2 regulate neuromuscular junction and myofiber phenotypes in mammalian skeletal muscle.

The neuromuscular junction is the synapse between a motor neuron of the spinal cord and a skeletal muscle fiber in the periphery. Reciprocal interactions between these excitable cells, and between them and others cell types present within the muscle tissue, shape the development, homeostasis and plasticity of skeletal muscle. An important aim in the field is to understand the molecular mechanisms underlying these cellular interactions, which include identifying the nature of the signals and receptors involved but also of the downstream intracellular signaling cascades elicited by them.

Nerve sprouting capacity in a pharmacologically induced mouse model of spinal muscular atrophy.

Spinal muscular atrophy (SMA) is caused by loss-of-function mutations in the survival of motoneuron gene 1 (SMN1). SMA is characterized by motoneuron death, skeletal muscle denervation and atrophy. Disease severity inversely correlates with copy number of a second gene (SMN2), which harbors a splicing defect that causes the production of inadequate levels of functional SMN protein.

Defective Acetylcholine Receptor Subunit Switch Precedes Atrophy of Slow-Twitch Skeletal Muscle Fibers Lacking ERK1/2 Kinases in Soleus Muscle.

To test the role of extracellular-signal regulated kinases 1 and 2 (ERK1/2) in slow-twitch, type 1 skeletal muscle fibers, we studied the soleus muscle in mice genetically deficient for myofiber ERK1/2. Young adult mutant soleus was drastically wasted, with highly atrophied type 1 fibers, denervation at most synaptic sites, induction of "fetal" acetylcholine receptor gamma subunit (AChRγ), reduction of "adult" AChRε, and impaired mitochondrial biogenesis and function. In weanlings, fiber morphology and mitochondrial markers were mostly normal, yet AChRγ upregulation and AChRε downregulation were observed.

Muscle-derived extracellular signal-regulated kinases 1 and 2 are required for the maintenance of adult myofibers and their neuromuscular junctions.

The Ras-extracellular signal-regulated kinase 1 and 2 (ERK1/2) pathway appears to be important for the development, maintenance, aging, and pathology of mammalian skeletal muscle. Yet no gene targeting of Erk1/2 in muscle fibers in vivo has been reported to date. We combined a germ line Erk1 mutation with Cre-loxP Erk2 inactivation in skeletal muscle to produce, for the first time, mice lacking ERK1/2 selectively in skeletal myofibers.

Improvement of neuromuscular synaptic phenotypes without enhanced survival and motor function in severe spinal muscular atrophy mice selectively rescued in motor neurons.

In the inherited childhood neuromuscular disease spinal muscular atrophy (SMA), lower motor neuron death and severe muscle weakness result from the reduction of the ubiquitously expressed protein survival of motor neuron (SMN). Although SMA mice recapitulate many features of the human disease, it has remained unclear if their short lifespan and motor weakness are primarily due to cell-autonomous defects in motor neurons. Using Hb9(Cre) as a driver, we selectively raised SMN expression in motor neurons in conditional SMAΔ7 mice.

Muscles in a mouse model of spinal muscular atrophy show profound defects in neuromuscular development even in the absence of failure in neuromuscular transmission or loss of motor neurons.

A mouse model of the devastating human disease "spinal muscular atrophy" (SMA) was used to investigate the severe muscle weakness and spasticity that precede the death of these animals near the end of the 2nd postnatal week. Counts of motor units to the soleus muscle as well as of axons in the soleus muscle nerve showed no loss of motor neurons. Similarly, neither immunostaining of neuromuscular junctions nor the measurement of the tension generated by nerve stimulation gave evidence of any significant impairment in neuromuscular transmission, even when animals were maintained up to 5days longer via a supplementary diet.

Emerging roles for MAP kinases in agrin signaling.

Information between neurons and the target cells they innervate passes through sites of functional contact called synapses. How synapses form and are altered by sensory or cognitive experience is central to understand nervous system function. Studies of synapse formation and plasticity have concentrated on a few "model" synapses.

Modulation of agrin-induced acetylcholine receptor clustering by extracellular signal-regulated kinases 1 and 2 in cultured myotubes.

Agrin released by motoneurons induces and/or maintains acetylcholine receptor (AChR) clustering and other aspects of postsynaptic differentiation at the vertebrate neuromuscular junction. Agrin acts by binding and activating a receptor complex containing LDL receptor protein 4 (Lrp4) and muscle-specific kinase (MuSK). Two critical downstream components of this signaling cascade, Dox-7 and rapsyn, have been identified.

CUGBP1 overexpression in mouse skeletal muscle reproduces features of myotonic dystrophy type 1.

The neuromuscular disease myotonic dystrophy type I (DM1) affects multiple organ systems with the major symptoms being severe muscle weakness, progressive muscle wasting and myotonia. The causative mutation in DM1 is a CTG repeat expansion in the 3'-untranslated region of the DM protein kinase (DMPK) gene. RNA transcribed from the expanded allele contains the expanded CUG repeats and leads to the nuclear depletion of Muscleblind-like 1 (MBNL1) and to the increased steady-state levels of CUG-binding protein 1 (CUGBP1).

Evidence for muscle-dependent neuromuscular synaptic site determination in mammals.

Recent evidence challenges the prevalent view that neural factors induce the formation of a de novo postsynaptic apparatus during development of the vertebrate neuromuscular junction. The latest experiments suggest an alternative model in which the muscle fiber induces a nascent postsynaptic apparatus and sets the location of the future synapse. On axonal contact, these sites, laid out in a prepattern in the central area of developing muscle fibers, mature into synapses by the combined action of neural factors such as agrin and ACh.

Regulation of the intermediate filament protein nestin at rodent neuromuscular junctions by innervation and activity.

The intermediate filament nestin is localized postsynaptically at rodent neuromuscular junctions. The protein forms a filamentous network beneath and between the synaptic gutters, surrounds myofiber nuclei, and is associated with Z-discs adjacent to the junction. In situ hybridization shows that nestin mRNA is synthesized selectively by synaptic myonuclei.

Neuregulins at the neuromuscular synapse: past, present, and future.

At the developing vertebrate neuromuscular junction, neuregulins are growth/differentiation factors essential for terminal Schwann cell survival. Neuregulins have also been thought as the critical signals responsible for the increased transcription of acetylcholine receptor subunit genes at the neuromuscular synapse. This latter role is now highly controversial.

Schwann cell-derived neuregulin-2alpha can function as a cell-attached activator of muscle acetylcholine receptor expression.

Here we show that neuregulin-2 (Nrg-2) alpha- and beta-isoforms can activate acetylcholine receptor (AChR) transcription as surface-attached ligands. More importantly, we demonstrate that Schwann cells that express Nrg-2alpha on their cell surface, the same Nrg-2 isoform expressed by terminal Schwann cells at the neuromuscular junction, can induce AChR expression if brought into cell-to-cell contact with myotubes specifically expressing ErbB4. These Schwann cells, the D6P2T cell line, induce AChR expression apparently as well as 293T cells transfected with Nrg-2beta, the isoform with the highest AChR-inducing activity when presented in a soluble form.

Induction of neuregulin signaling in mouse schwann cells in vivo mimics responses to denervation.

Neuregulins play crucial roles in early development of Schwann cells (SCs), but their roles in the activities of SCs during denervation and reinnervation of muscle are less clear. In the present study, the Tet-On system has been used in transgenic mice to enable inducible expression of a mutant, constitutively active neuregulin receptor (ErbB2) in SCs. This induction simulates neuregulin signaling to these cells.

Defective neuromuscular synaptogenesis in mice expressing constitutively active ErbB2 in skeletal muscle fibers.

We overexpressed a constitutively active form of the neuregulin receptor ErbB2 (CAErbB2) in skeletal muscle fibers in vivo and in vitro by tetracycline-inducible expression. Surprisingly, CAErbB2 expression during embryonic development was lethal and impaired synaptogenesis yielding a phenotype with loss of synaptic contacts, extensive axonal sprouting, and diffuse distribution of acetylcholine receptor (AChR) transcripts, reminiscent of agrin-deficient mice. CAErbB2 expression in cultured myotubes inhibited the formation and maintenance of agrin-induced AChR clusters, suggesting a muscle- and not a nerve-origin for the defect in CAErbB2-expressing mice.

Neuregulin-1 immunoglobulin-like domain mutant mice: clozapine sensitivity and impaired latent inhibition.

Genetic and behavioral studies in humans and mouse mutants have implicated the gene encoding neuregulin-1 (Nrg-1) as a candidate susceptibility gene for schizophrenia. We examined the behavior of mice heterozygous for a mutation in neuregulin-1's immunoglobulin (Ig)-like domain (Ig-nrg-1 mice). We found that these animals displayed behaviors related to a schizophrenia-like phenotype, such as clozapine suppression of open-field and running wheel activity and impaired latent inhibition.

Neuregulin-2 is synthesized by motor neurons and terminal Schwann cells and activates acetylcholine receptor transcription in muscle cells expressing ErbB4.

Acetylcholine receptor (AChR) genes are transcribed selectively in synaptic nuclei of skeletal muscle fibers, leading to accumulation of the mRNAs encoding AChR subunits at synaptic sites. The signals that regulate synapse-specific transcription remain elusive, though Neuregulin-1 is considered a favored candidate. Here, we show that motor neurons and terminal Schwann cells express neuregulin-2, a neuregulin-1-related gene.

Neuregulins: primary or secondary signals for the control of synapse-specific gene expression.

The selective transcription of acetylcholine receptor (AChR) subunit genes in synaptic myonuclei leads to the accumulation of AChR subunit mRNAs at the neuromuscular junction (NMJ). This mechanism contributes to the concentration of AChRs at the postsynaptic sarcolemma, and its physiological significance is underscored by the cases of human congenital myasthenias caused by mutation in a cis-regulatory element of the AChRepsilon-subunit promoter, which is necessary for its synaptic expression. The signal(s) that drives synapse-specific expression is unknown but neuregulin-1 (Nrg-1), a group of growth-factor-like polypeptides encoded by the nrg-1 gene, has been a favorite candidate.