scholarly journals Maternal experience-dependent cortical plasticity in mice is circuit- and stimulus-specific and requires MECP2

2019 ◽  
Author(s):  
Billy Y. B. Lau ◽  
Keerthi Krishnan ◽  
Z. Josh Huang ◽  
Stephen D. Shea
Keyword(s):  
Pyramidal Neurons ◽  
Cortical Plasticity ◽  
Genetic Manipulation ◽  
Deep Layer ◽  
Neurodevelopmental Disorder ◽  
Wild Type ◽  
Maternal Experience ◽  
Auditory Responses ◽  
Sensory Activity ◽  
Protein Mecp2

ABSTRACTThe neurodevelopmental disorder Rett syndrome is caused by mutations in the gene Mecp2. Misexpression of the protein MECP2 is thought to contribute to neuropathology by causing dysregulation of plasticity. Female heterozygous Mecp2 mutants (Mecp2het) failed to acquire a learned maternal retrieval behavior when exposed to pups, an effect linked to disruption of parvalbumin-expressing inhibitory interneurons (PV+) in the auditory cortex. However, the consequences of dysregulated PV+ networks during early maternal experience for auditory cortical sensory activity are unknown. Here we show that maternal experience in wild-type adult female mice (Mecp2wt) triggers suppression of PV+ auditory responses. We also observe concomitant disinhibition of auditory responses in deep-layer pyramidal neurons that is selective for behaviorally-relevant pup vocalizations. These neurons also exhibit sharpened tuning for pup vocalizations following maternal experience. All of these neuronal changes are abolished in Mecp2het, yet a genetic manipulation of GABAergic networks that restores accurate retrieval behavior in Mecp2het also restores maternal experience-dependent plasticity of PV+. Our data are consistent with a growing body of evidence that cortical networks are particularly vulnerable to mutations of Mecp2 in PV+ neurons.

scholarly journals The Chloride Homeostasis of CA3 Hippocampal Neurons Is Not Altered in Fully Symptomatic Mepc2-null Mice

2021 ◽  
Vol 15 ◽  
Author(s):  
Yasmine Belaïdouni ◽  
Diabe Diabira ◽  
Jinwei Zhang ◽  
Jean-Charles Graziano ◽  
Francesca Bader ◽  
...  
Keyword(s):  
Hippocampal Neurons ◽  
Pyramidal Neurons ◽  
Neurodevelopmental Disorder ◽  
Reversal Potential ◽  
Wild Type ◽  
Chloride Homeostasis ◽  
Inhibitory Strength ◽  
Extrusion Mechanism ◽  
Ca3 Pyramidal Neurons ◽  
Chloride Extrusion

Rett syndrome (RTT) is an X-linked neurodevelopmental disorder caused mainly by mutations in the MECP2 gene. Mouse models of RTT show reduced expression of the cation-chloride cotransporter KCC2 and altered chloride homeostasis at presymptomatic stages. However, whether these alterations persist to late symptomatic stages has not been studied. Here we assess KCC2 and NKCC1 expressions and chloride homeostasis in the hippocampus of early [postnatal (P) day 30–35] and late (P50–60) symptomatic male Mecp2-null (Mecp2–/y) mice. We found (i) no difference in the relative amount, but an over-phosphorylation, of KCC2 and NKCC1 between wild-type (WT) and Mecp2–/y hippocampi and (ii) no difference in the inhibitory strength, nor reversal potential, of GABAA-receptor-mediated responses in Mecp2–/y CA3 pyramidal neurons compared to WT at any stages studied. Altogether, these data indicate the presence of a functional chloride extrusion mechanism in Mecp2–/y CA3 pyramidal neurons at symptomatic stages.

scholarly journals Network hyperexcitability in hippocampal slices from Mecp2 mutant mice revealed by voltage-sensitive dye imaging

2011 ◽  
Vol 105 (4) ◽  
pp. 1768-1784 ◽  
Author(s):  
Gaston Calfa ◽  
John J. Hablitz ◽  
Lucas Pozzo-Miller
Keyword(s):  
Pyramidal Neurons ◽  
Neurodevelopmental Disorder ◽  
Hippocampal Slices ◽  
Clinical Manifestations ◽  
Mutant Mice ◽  
Loss Of Function ◽  
Wild Type ◽  
Limbic Seizures ◽  
Ca3 Pyramidal Neurons ◽  
And Function

Dysfunctions of neuronal and network excitability have emerged as common features in disorders associated with intellectual disabilities, autism, and seizure activity, all common clinical manifestations of Rett syndrome (RTT), a neurodevelopmental disorder caused by loss-of-function mutations in the transcriptional regulator methyl-CpG-binding protein 2 (MeCP2). Here, we evaluated the consequences of Mecp2 mutation on hippocampal network excitability, as well as synapse structure and function using a combination of imaging and electrophysiological approaches in acute slices. Imaging the amplitude and spatiotemporal spread of neuronal depolarizations with voltage-sensitive dyes (VSD) revealed that the CA1 and CA3 regions of hippocampal slices from symptomatic male Mecp2 mutant mice are highly hyperexcitable. However, only the density of docked synaptic vesicles and the rate of release from the readily releasable pool are impaired in Mecp2 mutant mice, while synapse density and morphology are unaffected. The differences in network excitability were not observed in surgically isolated CA1 minislices, and blockade of GABAergic inhibition enhanced VSD signals to the same extent in Mecp2 mutant and wild-type mice, suggesting that network excitability originates in area CA3. Indeed, extracellular multiunit recordings revealed a higher level of spontaneous firing of CA3 pyramidal neurons in slices from symptomatic Mecp2 mutant mice. The neuromodulator adenosine reduced the amplitude and spatiotemporal spread of VSD signals evoked in CA1 of Mecp2 mutant slices to wild-type levels, suggesting its potential use as an anticonvulsant in RTT individuals. The present results suggest that hyperactive CA3 pyramidal neurons contribute to hippocampal dysfunction and possibly to limbic seizures observed in Mecp2 mutant mice and RTT individuals.

scholarly journals SEX-SPECIFIC TRAJECTORIES OF REELIN-DEPENDENT MATURATION OF DEEP LAYER PREFRONTAL NEURONS

2021 ◽  
Author(s):  
Thenzing J Silva Hurtado ◽  
Olivier Lassalle ◽  
Antoine Ameloot ◽  
Pascale Chavis
Keyword(s):  
Matrix Protein ◽  
Pyramidal Neurons ◽  
Deep Layer ◽  
Long Term Potentiation ◽  
Developmental Trajectory ◽  
Extracellular Matrix Protein ◽  
Wild Type ◽  
Functional Maturation

Throughout early adulthood, the anatomical and functional maturation of PFC circuitry continues under the influence of multiple extrinsic and intrinsic factors, most notably electrical activity, and molecular cues. We previously showed that the extracellular matrix protein reelin orchestrates the structural and functional maturation of deep layers medial PFC (mPFC) pyramidal neurons. Additionally, we reported that reelin haploinsufficiency is associated to prefrontal disruptions of long-term memory retention thereby illustrating the eminent role of reelin in cognitive maturation of the PFC. Prefrontal maturation follows a sex-specific developmental pattern, supporting the existence of sexual differences in the morphology-functional properties PFC. Here, we interrogated the role of reelin in the functional maturation of excitatory networks in the mPFC. The developmental trajectory of reelin's expression and deep layer pyramidal neurons synaptic plasticity was tracked in the mPFC of male and female mice, from the juvenile period to adulthood. To assess the role of reelin in both sexes, wild-type and heterozygous reeler mice (HRM) were compared. The results show that the maturational profile of reelin expression in the mPFC is sex-dependent and that the developmental trajectory of long-term potentiation is different between wild-type males and females. These data demonstrate reelin influence on prefrontal synapses is sex and period specific.

scholarly journals Lactose-Inducible System for Metabolic Engineering of Clostridium ljungdahlii

2014 ◽  
Vol 80 (8) ◽  
pp. 2410-2416 ◽  
Author(s):  
Areen Banerjee ◽  
Ching Leang ◽  
Toshiyuki Ueki ◽  
Kelly P. Nevin ◽  
Derek R. Lovley
Keyword(s):  
Ethanol Production ◽  
Genetic Manipulation ◽  
Electron Flow ◽  
Model Organism ◽  
Inducible Expression ◽  
Carbon Flow ◽  
Wild Type ◽  
Content Type ◽  
Microbial Electrosynthesis ◽  
Clostridium Ljungdahlii

ABSTRACTThe development of tools for genetic manipulation ofClostridium ljungdahliihas increased its attractiveness as a chassis for autotrophic production of organic commodities and biofuels from syngas and microbial electrosynthesis and established it as a model organism for the study of the basic physiology of acetogenesis. In an attempt to expand the genetic toolbox forC. ljungdahlii, the possibility of adapting a lactose-inducible system for gene expression, previously reported forClostridium perfringens, was investigated. The plasmid pAH2, originally developed forC. perfringenswith agusAreporter gene, functioned as an effective lactose-inducible system inC. ljungdahlii. Lactose induction ofC. ljungdahliicontaining pB1, in which the gene for the aldehyde/alcohol dehydrogenase AdhE1 was downstream of the lactose-inducible promoter, increased expression ofadhE130-fold over the wild-type level, increasing ethanol production 1.5-fold, with a corresponding decrease in acetate production. Lactose-inducible expression ofadhE1in a strain in whichadhE1and theadhE1homologadhE2had been deleted from the chromosome restored ethanol production to levels comparable to those in the wild-type strain. Inducing expression ofadhE2similarly failed to restore ethanol production, suggesting thatadhE1is the homolog responsible for ethanol production. Lactose-inducible expression of the four heterologous genes necessary to convert acetyl coenzyme A (acetyl-CoA) to acetone diverted ca. 60% of carbon flow to acetone production during growth on fructose, and 25% of carbon flow went to acetone when carbon monoxide was the electron donor. These studies demonstrate that the lactose-inducible system described here will be useful for redirecting carbon and electron flow for the biosynthesis of products more valuable than acetate. Furthermore, this tool should aid in optimizing microbial electrosynthesis and for basic studies on the physiology of acetogenesis.

scholarly journals Application of CRISPR/Cas9 Tools for Genome Editing in the White-Rot Fungus Dichomitus squalens

Biomolecules ◽  
2021 ◽  
Vol 11 (10) ◽  
pp. 1526
Author(s):  
Joanna E. Kowalczyk ◽  
Shreya Saha ◽  
Miia R. Mäkelä
Keyword(s):  
Genome Editing ◽  
Plant Biomass ◽  
Genetic Manipulation ◽  
White Rot ◽  
White Rot Fungus ◽  
Detailed Knowledge ◽  
Wild Type ◽  
Low Frequencies ◽  
Dichomitus Squalens

Dichomitus squalens is an emerging reference species that can be used to investigate white-rot fungal plant biomass degradation, as it has flexible physiology to utilize different types of biomass as sources of carbon and energy. Recent comparative (post-) genomic studies on D. squalens resulted in an increasingly detailed knowledge of the genes and enzymes involved in the lignocellulose breakdown in this fungus and showed a complex transcriptional response in the presence of lignocellulose-derived compounds. To fully utilize this increasing amount of data, efficient and reliable genetic manipulation tools are needed, e.g., to characterize the function of certain proteins in vivo and facilitate the construction of strains with enhanced lignocellulolytic capabilities. However, precise genome alterations are often very difficult in wild-type basidiomycetes partially due to extremely low frequencies of homology directed recombination (HDR) and limited availability of selectable markers. To overcome these obstacles, we assessed various Cas9-single guide RNA (sgRNA) ribonucleoprotein (RNP) -based strategies for selectable homology and non-homologous end joining (NHEJ) -based gene editing in D. squalens. We also showed an induction of HDR-based genetic modifications by using single-stranded oligodeoxynucleotides (ssODNs) in a basidiomycete fungus for the first time. This paper provides directions for the application of targeted CRISPR/Cas9-based genome editing in D. squalens and other wild-type (basidiomycete) fungi.

scholarly journals Daily oscillation of the excitation/inhibition ratio is disrupted in two mouse models of autism

2021 ◽  
Author(s):  
Michelle Bridi ◽  
Nancy Luo ◽  
Grace Kim ◽  
Caroline O'Ferrall ◽  
Ruchit Oatel ◽  
...  
Keyword(s):  
Mouse Models ◽  
Sensory Processing ◽  
Neurodevelopmental Disorder ◽  
Autism Spectrum ◽  
Behavioral State ◽  
Wild Type ◽  
Inhibitory Synaptic Transmission ◽  
Sleep Timing ◽  
Per Se ◽  
Daily Oscillation

Autism spectrum disorder (ASD) is a prevalent neurodevelopmental disorder involving sensory processing abnormalities. Alterations to the balance between excitation and inhibition (E/I ratio) are postulated to underlie behavioral phenotypes in ASD patients and mouse models. However, in primary visual cortex (V1) of wild type mice, the E/I ratio is not a fixed value, but rather oscillates across the 24h day. Therefore, we hypothesized that the E/I oscillation, rather than the overall E/I ratio, may be disrupted in ASD mouse models. To this end, we measured the E/I ratio in Fmr1 KO and BTBR mice, models of syndromic and idiopathic ASD, respectively. We found that the E/I ratio is dysregulated in both models, but in different ways: the oscillation is flattened in Fmr1 KO and phase-shifted in BTBR mice. These phenotypes cannot be explained by altered sleep timing, which was largely normal in both lines. Furthermore, we found that E/I dysregulation occurs due to alterations in both excitatory and inhibitory synaptic transmission in both models. These findings provide a crucial perspective on the E/I ratio in ASD, suggesting that ASD phenotypes may be produced by a mismatch of E/I to the appropriate behavioral state, rather than alterations to overall E/I levels per se.

Genetic Manipulation of Key Photosynthetic Enzymes in the C4 Plant Flaveria bidentis

1997 ◽  
Vol 24 (4) ◽  
pp. 477 ◽  
Author(s):  
Robert T. Furbank ◽  
Julie A. Chitty ◽  
Colin L.D. Jenkins ◽  
William C. Taylor ◽  
Stephen J. Trevanion ◽  
...  
Keyword(s):  
Enzyme Activity ◽  
Malate Dehydrogenase ◽  
Genetic Manipulation ◽  
Photosynthetic Performance ◽  
Covalent Modification ◽  
Wild Type ◽  
Bisphosphate Carboxylase ◽  
Photosynthetic Enzymes ◽  
Flaveria Bidentis

The NADP-malic enzyme type C4 dicot Flaveria bidentis (L.) Kuntze was transformed with antisense and cosense gene constructs that resulted in specific decreases in single photosynthetic enzymes. The enzymes targeted were ribulose-1,5-bisphosphate carboxylase/oxygenase [EC 4.1.1.39] (Rubisco), pyruvate, Pi dikinase [EC 2.7.9.1] (PPDK) and NADP malate dehydrogenase [EC 1.1.1.82] (NADP-MDH). These enzymes were chosen as targets because they have low activity compared to photosynthetic rates (Rubisco), are subject to complex covalent regulation (NADP-MDH), or both (PPDK). T1 progeny of a number of lines of these transformants were examined for the effects of these gene constructs on enzyme levels and photosynthetic performance. Rubisco antisense transformants expressing between 15 and 100% of wild-type enzyme activity were obtained. Pyruvate, Pi dikinase antisense lines were obtained with 40–100% wild-type levels. NADP malate dehydrogenase sense constructs caused a co-suppression of enzyme activity with some lines containing less than 2% of wild- type activity. Under saturating illumination, the control coefficients for these enzymes were determined to be up to 0.7 for Rubisco, 0.2–0.3 for PPDK and effectively zero for NADP-MDH. The implications of these observations for the regulation of photosynthetic flux and metabolism in C4 plants and the role of regulation by covalent modification are discussed.

scholarly journals 11 Study of Polynucleotide Kinase/Phosphatase (PNKP) Mutations Found in a Patient with Microcephaly, Seizures, and Developmental Delay (MCSZ) and Glioblastoma

2018 ◽  
Vol 45 (S3) ◽  
pp. S2-S2
Author(s):  
Bingcheng Jiang ◽  
Chibawanye I. Ene ◽  
Bonnie Cole ◽  
Jeff Ojemann ◽  
Sarah Leary ◽  
...  
Keyword(s):  
Human Cancer ◽  
Neurodevelopmental Disorder ◽  
Site Directed Mutagenesis ◽  
Expression Vectors ◽  
Wild Type ◽  
Double Mutation ◽  
Mutant Proteins ◽  
Human Cancer Cell Lines ◽  
Mammalian Expression ◽  
Polynucleotide Kinase

The enzyme polynucleotide kinase/phosphatase (PNKP) plays a key role in DNA repair by resolving the chemistry at DNA strand breaks. Mutations in PNKP (chromosome 19q13.4) are known to cause MCSZ, a serious neurodevelopmental disorder, but to date there has been no link to cancer initiation or progression. However, a child with MCSZ recently presented at Seattle Children's Hospital with a 3-cm glioblastoma. The child was shown to have two germline mutations in PNKP. To study the effects of the PNKP mutations found in this patient, we generated mutant PNKP cDNAs carrying either the individual mutations or the double mutation using site directed mutagenesis. These cDNAs were incorporated into bacterial and mammalian expression vectors. The bacterially expressed mutant proteins as well as the wild type have been purified and are undergoing testing for PNKP DNA kinase and phosphatase activity. The PNKP cDNAs, fused to GFP, were expressed in Hela and HCT116 human cancer cell lines. High-content analysis and micro-irradiation techniques are being used to determine PNKP localization within the cells and recruitment to damaged DNA. Our preliminary results indicate that the mutations alter the ratio of nuclear to cytoplasmic PNKP compared to the wild-type protein.

scholarly journals BRAFV600E expression in neural progenitors results in a hyperexcitable phenotype in neocortical pyramidal neurons

2020 ◽  
Vol 123 (6) ◽  
pp. 2449-2464
Author(s):  
Roman U. Goz ◽  
Gülcan Akgül ◽  
Joseph J. LoTurco
Keyword(s):  
Neuronal Excitability ◽  
Pyramidal Neurons ◽  
Mammalian Target Of Rapamycin ◽  
Neural Progenitors ◽  
Wild Type ◽  
Neuronal Phenotype ◽  
Neuronal Progenitors ◽  
Neocortical Neurons ◽  
Distinct Cell ◽  
Phosphoinositide 3 Kinase

This study is the first to report the cell autonomous effects of BRAFV600E mutations on the intrinsic neuronal excitability. We show that BRAFV600E alters multiple electrophysiological parameters in neocortical neurons. Similar excitability changes did not occur in cells neighboring BRAFV600E-expressing neurons, after overexpression of wild-type BRAF transgenes, or after introduction of mutations affecting the mammalian target of rapamycin (mTOR) or the catalytic subunit of phosphoinositide 3-kinase (PIK3CA). We conclude that BRAFV600E causes a distinct, cell autonomous, highly excitable neuronal phenotype when introduced somatically into neocortical neuronal progenitors.

scholarly journals Abundance and localization of human UBE3A protein isoforms

2020 ◽  
Vol 29 (18) ◽  
pp. 3021-3031 ◽  
Author(s):  
Carissa L Sirois ◽  
Judy E Bloom ◽  
James J Fink ◽  
Dea Gorka ◽  
Steffen Keller ◽  
...  
Keyword(s):  
Neurodevelopmental Disorder ◽  
Embryonic Stem ◽  
Protein Isoforms ◽  
Wild Type ◽  
Neuronal Function ◽  
Ube3a Gene ◽  
Recent Case Study ◽  
The Individual ◽  
Human Isoforms

Abstract Loss of UBE3A expression, a gene regulated by genomic imprinting, causes Angelman syndrome (AS), a rare neurodevelopmental disorder. The UBE3A gene encodes an E3 ubiquitin ligase with three known protein isoforms in humans. Studies in mouse suggest that the human isoforms may have differences in localization and neuronal function. A recent case study reported mild AS phenotypes in individuals lacking one specific isoform. Here we have used CRISPR/Cas9 to generate isogenic human embryonic stem cells (hESCs) that lack the individual protein isoforms. We demonstrate that isoform 1 accounts for the majority of UBE3A protein in hESCs and neurons. We also show that UBE3A predominantly localizes to the cytoplasm in both wild type and isoform-null cells. Finally, we show that neurons lacking isoform 1 display a less severe electrophysiological AS phenotype.

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