Astrocytes mediate synapse elimination through MEGF10 and MERTK pathways

Mutations in the human X-linked cyclin-dependent kinase-like 5 (CDKL5) gene have been identified in patients with Rett syndrome (RTT), West syndrome, and X-linked infantile spasms, sharing the common feature of mental retardation and early seizures. CDKL5 is a rather uncharacterized kinase, but its involvement in RTT seems to be explained by the fact that it works upstream of MeCP2, the main cause of Rett syndrome. To understand the role of this kinase for nervous system functions and to address if molecular mechanisms are involved in regulating its distribution and activity, we studied the ontogeny of CDKL5 expression in developing mouse brains by immunostaining and Western blotting. The expression profile of CDKL5 was compared with that of MeCP2. The two proteins share a general expression profile in the adult mouse brain, but CDKL5 levels appear to be highly modulated at the regional level. Its expression is strongly induced in early postnatal stages, and in the adult brain CDKL5 is present in mature neurons, but not in astroglia. Interestingly, the presence of CDKL5 in the cell nucleus varies at the regional level of the adult brain and is developmentally regulated. CDKL5 shuttles between the cytoplasm and the nucleus and the C-terminal tail is involved in localizing the protein to the cytoplasm in a mechanism depending on active nuclear export. Accordingly, Rett derivatives containing disease-causing truncations of the C terminus are constitutively nuclear, suggesting that they might act as gain of function mutations in this cellular compartment.

Low Current-driven Micro-electroporation Allows Efficient In Vivo Delivery of Nonviral DNA into the Adult Mouse Brain

Receptor protein tyrosine phosphatase σ binds to neurons in the adult mouse brain

Recent studies show that type II transmembrane serine proteases play important roles in diverse cellular activities and pathological processes. Their expression and functions in the central nervous system, however, are largely unexplored. In this study, we show that the expression of one such member, matriptase (MTP), was cell type-restricted and primarily expressed in neural progenitor (NP) cells and neurons. Blocking MTP expression or MTP activity prevented NP cell traverse of reconstituted basement membrane, whereas overexpression of MTP promoted it. The NP cell mobilization induced by either vascular endothelial growth factor or hepatocyte growth factor was also impaired by knocking down MTP expression. MTP acts upstream of matrix metalloproteinase 2 in promoting NP cell mobility. In embryonic stem cell differentiation to neural cells, MTP knockdown had no effect on entry of embryonic stem cells into the neural lineage. High MTP expression or activity, however, shifts the population dynamics from NP cells toward neurons to favor neuronal differentiation. This is the first report to demonstrate the direct involvement of type II transmembrane serine protease in NP cell function.

Genetic analysis of Ntn1 in adult mouse midbrain neurons reveals its function in maintaining excitatory synapses, loss of Ntn1 function in inhibitory neurons is significantly detrimental to mesolimbic system function.

Mass spectrometry imaging reveals glycine distribution in the developing and adult mouse brain

Cbln1 belongs to the C1q and tumour necrosis factor superfamily, and plays crucial roles as a cerebellar granule cell-derived transneuronal regulator for synapse integrity and plasticity in Purkinje cells. Although Cbln2-Cbln4 are also expressed in the brain and could form heteromeric complexes with Cbln1, their precise expressions remain unclear. Here, we investigated gene expression of the Cbln family in developing and adult C57BL mouse brains by reverse transcriptase-polymerase chain reaction (RT-PCR), Northern blot, and high-resolution in situ hybridization (ISH) analyses. In the adult brain, spatial patterns of mRNA expression were highly differential depending on Cbln subtypes. Notably, particularly high levels of Cbln mRNAs were expressed in some nuclei and neurons, whereas their postsynaptic targets often lacked or were low for any Cbln mRNAs, as seen for cerebellar granule cells/Purkinje cells, entorhinal cortex/hippocampus, intralaminar group of thalamic nuclei/caudate-putamen, and dorsal nucleus of the lateral lemniscus/central nucleus of the inferior colliculus. In the developing brain, Cbln1, 2, and 4 mRNAs appeared as early as embryonic day 10-13, and exhibited transient up-regulation during the late embryonic and neonatal periods. For example, Cbln2 mRNA was expressed in the cortical plate of the developing neocortex, displaying a high rostromedial to low caudolateral gradient. In contrast, Cbln3 mRNA was selective to cerebellar granule cells throughout development, and its onset was as late as postnatal day 7-10. These results will provide a molecular-anatomical basis for future studies that characterize roles played by the Cbln family.

For the most part, the production of new brain cells in mammals occurs only in immature individuals at the time the nervous system is undergoing development. One notable exception is the hippocampus, a part of the brain involved in memory and spatial perception. Hippocampal cells called dentate granule cells (DGCs) develop in adults as well as in young animals. How these “adult-born” cells build their connections with the rest of the brain, and the extent to which they resemble “pup-born” cells, is of great interest to those who would like to coax other parts of adult brains to make new cells as a strategy for reversing the loss of function from trauma or degenerative disorders. The development of DGCs is linked to hippocampus-related learning and behavioral changes. Developing, adult-born DGCs have different properties than mature neurons that arose when the brain was developing. Do the adult-born cells keep their distinct traits, or do they become functionally similar to the cells that developed early in life? To find out, Diego Laplagne, Alejandro Schinder, and colleagues compared the structure and function of these “new kids on the block” with DGCs that developed in the perinatal period in mice. The researchers’ first task was to figure out a way to distinguish between pup-born and adult-born DGCs in brain tissue that contained both. To accomplish that task, they used retroviruses to introduce one kind of fluorescent protein into the developing DGCs at 7 days after birth and a second protein into the adult mouse brain at 42 days after birth. As a result of this treatment, the pup-born cells fluoresced green and the adult-born cells fluoresced red, making them readily distinguishable in brain slices. Once they could tell the two types of cells apart, the researchers began testing a variety of electrophysiological traits of the connections between DGCs and neurons providing excitatory and inhibitory inputs. Using brain slices obtained from 19-week-old mice that had undergone the retrovirus labeling earlier in life, they looked at glutamatergic (excitatory) nerves connecting the hippocampus with the entorhinal cortex, another brain area associated with memory. When they stimulated the afferent excitatory neurons (which carry information from the neocortex to the hippocampus), the researchers evoked similar excitatory postsynaptic currents (EPSCs) in both pup-born and adult-born DGCs. When given paired-pulse stimulation, in which two pulses of electricity are given close together, the two types of cells showed a very similar reduction in EPSCs with the second pulse, suggesting that the short-term plasticity of the synapses is identical. And when the cells were given high-frequency stimulation, EPSC amplitude was depressed in both cell types in a similar manner. Thus, the researchers concluded that excitatory inputs to pup-born and adult-born DGCs are functionally similar. Next the researchers looked at GABAergic (inhibitory) inputs from interneurons that connect to the body and dendrites of the DGCs. Using brain slices from 14-week-old mice, they stimulated the incoming inhibitory neurons from the granule cell layer (GCL) and molecular layer (ML) of the hippocampus and found no significant difference in the amplitude and kinetics of inhibitory postsynaptic currents (IPSCs) that resulted. Spontaneous IPSCs, which add information about GABAergic inputs from areas other than the GCL and ML, exhibited similar frequency, amplitude, and kinetics in the pup-born and adult-born cells. Having shown that pup-born and adult-born DGCs respond to both excitatory and inhibitory inputs in the same way, the researchers next turned their attention to how the two types of cells integrate the signals from the various inputs to produce an action potential (which transmits the signal), or spike. Spiking probability varied among neurons but was not distinguishable between the two cell types, further supporting the earlier indications that adult-born and pup-born DGCs function in fundamentally the same way. With glutamatergic and GABAergic inputs handled similarly and the action-potential response indistinguishable, the researchers concluded that mature adult-born and pup-born DGCs are functionally similar. This means that at least some neurons that develop in adult brains can form connections that are indistinguishable from connections formed by neurons that develop early in life—a hopeful finding for those who have set their sights on one day being able to repair damaged or deteriorated brain tissue.

NFATc4 has recently been identified as playing an important role in variety of activity-dependent neuronal processes, including hippocampal plasticity, axonal growth, neuronal survival, and apoptosis. However, a systematic study examining the distribution of NFATc4 within the nervous system has yet to be conducted. With this in mind, we sought to determine the regional localization of NFATc4 within the adult mouse brain. Interestingly, NFATc4 was expressed broadly, but not uniformly, throughout various brain structures. The highest levels of NFATc4 expression were localized to the hippocampus, olfactory bulb, and various hypothalamic nuclei. Other brain regions that expressed NFATc4 included the cerebellum, striatum, globus pallidus, amygdala, neocortex, and brainstem nuclei. Given NFATc4's widespread expression, these results are consistent with the notion that NFATc4 may underlie activity-dependent neuronal plasticity throughout the adult brain.

Experience alters brain structure, but the underlying mechanism remained unknown. Structural plasticity reveals that brain function is encoded in generative changes to cells that compete with destructive processes driving neurodegeneration. At an adult critical period, experience increases fiber number and brain size in Drosophila. Here, we asked if Toll receptors are involved. Tolls demarcate a map of brain anatomical domains. Focusing on Toll-2, loss of function caused apoptosis, neurite atrophy and impaired behaviour. Toll-2 gain of function and neuronal activity at the critical period increased cell number. Toll-2 induced cycling of adult progenitor cells via a novel pathway, that antagonized MyD88-dependent quiescence, and engaged Weckle and Yorkie downstream. Constant knock-down of multiple Tolls synergistically reduced brain size. Conditional over-expression of Toll-2 and wek at the adult critical period increased brain size. Through their topographic distribution, Toll receptors regulate neuronal number and brain size, modulating structural plasticity in the adult brain.

Brain inflammation induces post-synaptic changes during early synapse formation in adult-born hippocampal neurons

Microglia, as the resident brain immune cells, can exhibit a broad range of activation phenotypes, which have been implicated in a multitude of central nervous system disorders. Current widely studied microglial cell lines are mainly derived from neonatal rodent brain that can limit their relevance to homeostatic function and disease-related neuroimmune responses in the adult brain. Recently, an adult mouse brain-derived microglial cell line has been established; however, a comprehensive proteome dataset remains lacking. Here, an optimization method for sensitive and rapid quantitative proteomic analysis of microglia is described that involves suspension trapping (S-Trap) for efficient and reproducible protein extraction from a limited number of microglial cells expected from an adult mouse brain (≈300000). Using a 2-h gradient on a 75-cm UPLC column with a modified data dependent acquisition method on a hybrid quadrupole-Orbitrap mass spectrometer, 4855 total proteins have been identified where 4698 of which are quantifiable by label-free quantitation with a median and average coefficient of variation (CV) of 6.7% and 10.6%, respectively. This dataset highlights the high depth of proteome coverage and related quantitation precision of the adult-derived microglial proteome including proteins associated with several key pathways related to immune response. Data are available via ProteomeXchange with identifier PXD012006.

MeCP2 is a critical epigenetic regulator in brain and its abnormal expression or compromised function leads to a spectrum of neurological disorders including Rett Syndrome and autism. Altered expression of the two MeCP2 isoforms, MeCP2E1 and MeCP2E2 has been implicated in neurological complications. However, expression, regulation and functions of the two isoforms are largely uncharacterized. Previously, we showed the role of MeCP2E1 in neuronal maturation and reported MeCP2E1 as the major protein isoform in the adult mouse brain, embryonic neurons and astrocytes. Recently, we showed that DNA methylation at the regulatory elements (REs) within the Mecp2 promoter and intron 1 impact the expression of Mecp2 isoforms in differentiating neural stem cells. This current study is aimed for a comparative analysis of temporal, regional and cell type-specific expression of MeCP2 isoforms in the developing and adult mouse brain. MeCP2E2 displayed a later expression onset than MeCP2E1 during mouse brain development. In the adult female and male brain hippocampus, both MeCP2 isoforms were detected in neurons, astrocytes and oligodendrocytes. Furthermore, MeCP2E1 expression was relatively uniform in different brain regions (olfactory bulb, striatum, cortex, hippocampus, thalamus, brainstem and cerebellum), whereas MeCP2E2 showed differential enrichment in these brain regions. Both MeCP2 isoforms showed relatively similar distribution in these brain regions, except for cerebellum. Lastly, a preferential correlation was observed between DNA methylation at specific CpG dinucleotides within the REs and Mecp2 isoform-specific expression in these brain regions. Taken together, we show that MeCP2 isoforms display differential expression patterns during brain development and in adult mouse brain regions. DNA methylation patterns at the Mecp2 REs may impact this differential expression of Mecp2/MeCP2 isoforms in brain regions. Our results significantly contribute towards characterizing the expression profiles of Mecp2/MeCP2 isoforms and thereby provide insights on the potential role of MeCP2 isoforms in the developing and adult brain.

Peroxisome proliferator-activated receptors (PPARs) are nuclear hormone receptors that act as ligand-activated transcription factors. PPAR agonists have well-documented anti-inflammatory and neuroprotective roles in the central nervous system. Recent evidence suggests that PPAR agonists are attractive therapeutic agents for treating neurodegenerative diseases as well as addiction. However, the distribution of PPAR mRNA and protein in brain regions associated with these conditions (i.e. prefrontal cortex, nucleus accumbens, amygdala, ventral tegmental area) is not well defined. Moreover, the cell type specificity of PPARs in mouse and human brain tissue has yet to be investigated. We utilized quantitative PCR and double immunofluorescence microscopy to determine that both PPAR mRNA and protein are expressed ubiquitously throughout the adult mouse brain. We found that PPARs have unique cell type specificities that are consistent between species. PPARα was the only isotype to colocalize with all cell types in both adult mouse and adult human brain tissue. Overall, we observed a strong neuronal signature, which raises the possibility that PPAR agonists may be targeting neurons rather than glia to produce neuroprotection. Our results fill critical gaps in PPAR distribution and define novel cell type specificity profiles in the adult mouse and human brain.

Zika Virus Infects Neural Progenitors in the Adult Mouse Brain and Alters Proliferation