Expanded alternative splice isoform profiling of the mouse Cav3.1/α1G T-type calcium channel

<p>Abstract</p> <p>Background</p> <p>Alternative splicing of low-voltage-activated T-type calcium channels contributes to the molecular and functional diversity mediating complex network oscillations in the normal brain. Transcript scanning of the human <it>CACNA1G </it>gene has revealed the presence of 11 regions within the coding sequence subjected to alternative splicing, some of which enhance T-type current. In mouse models of absence epilepsy, elevated T-type calcium currents without clear increases in channel expression are found in thalamic neurons that promote abnormal neuronal synchronization. To test whether enhanced T-type currents in these models reflect pathogenic alterations in channel splice isoforms, we determined the extent of alternative splicing of mouse <it>Cacna1g </it>transcripts and whether evidence of altered transcript splicing could be detected in mouse absence epilepsy models.</p> <p>Results</p> <p>Transcript scanning of the murine <it>Cacna1g </it>gene detected 12 regions encoding alternative splice isoforms of Ca_v3.1/α1G T-type calcium channels. Of the 12 splice sites, six displayed homology to the human <it>CACNA1G </it>splice sites, while six novel mouse-specific splicing events were identified, including one intron retention, three alternative acceptor sites, one alternative donor site, and one exon exclusion. In addition, two brain region-specific alternative splice patterns were observed in the cerebellum. Comparative analyses of brain regions from four monogenic absence epilepsy mouse models with altered thalamic T-type currents and wildtype controls failed to reveal differences in <it>Cacna1g </it>splicing patterns.</p> <p>Conclusion</p> <p>The determination of six novel alternative splice sites within the coding region of the mouse <it>Cacna1g </it>gene greatly expands the potential biophysical diversity of voltage-gated T-type channels in the mouse central nervous system. Although alternative splicing of Ca_v3.1/α1G channels does not explain the enhancement of T-type current identified in four mouse models of absence epilepsy, post-transcriptional modification of T-type channels through this mechanism may influence other developmental neurological phenotypes.</p

Hypomyelination alters K+ channel expression in mouse mutants shiverer and Trembler

AbstractVoltage-gated K+ channels are localized to juxtaparanodal regions of myelinated axons. To begin to understand the role of normal compact myelin in this localization, we examined mKv1.1 and mKv1.2 expression in the dysmyelinating mouse mutants shiverer and Trembler. In neonatal wild-type and shiverer mice, the focal localization of both proteins in axon fiber tracts is similar, suggesting that cues other than mature myelin can direct initial K+ channel localization in shiverer mutants. In contrast, K+ channel localization is altered in hypomyelinated axonal fiber tracts of adult mutants, suggesting that abnormal myelination leads to channel redistribution. In shiverer adult, K+ channel expression is up-regulated in both axons and glia, as revealed by immunocytochemistry, RNase protection, and in situ hybridization studies. This up-regulation of K+ channels in hypomyelinated axon tracts may reflect a compensatory reorganization of ionic currents, allowing impulse conduction to occur in these dysmyelinating mouse mutants

Disruption of the NF-κB/IκBα Autoinhibitory Loop Improves Cognitive Performance and Promotes Hyperexcitability of Hippocampal Neurons

<p>Abstract</p> <p>Background</p> <p>Though originally discovered in the immune system as an important mediator of inflammation, NF-κB has recently been shown to play key roles in the central nervous system, such as synaptogenesis, synaptic plasticity, and cognition. NF-κB activity is normally tightly regulated by its primary inhibitor, IκBα, through a unique autoinhibitory loop. In this study, we tested the hypothesis that the IκBα autoinhibitory loop ensures optimal levels of NF-κB activity to promote proper brain development and function. To do so, we utilized knock-in mice which possess mutations in the IκBα promoter to disrupt the autoinhibitory loop (IκBα^M/MKI mice).</p> <p>Results</p> <p>Here, we show that these mutations delay IκBα resynthesis and enhance NF-κB activation in neurons following acute activating stimuli. This leads to improved cognitive ability on tests of hippocampal-dependent learning and memory but no change in hippocampal synaptic plasticity. Instead, hippocampal neurons from IκBα^M/MKI mice form more excitatory and less inhibitory synapses in dissociated cultures and are hyperexcitable. This leads to increased burst firing of action potentials and the development of abnormal hypersynchronous discharges <it>in vivo</it>.</p> <p>Conclusions</p> <p>These results demonstrate that the IκBα autoinhibitory loop is critical for titrating appropriate levels of endogenous NF-κB activity to maintain proper neuronal function.</p

Sodium/Hydrogen Exchanger Gene Defect in Slow-Wave Epilepsy Mutant Mice

AbstractThe “housekeeping” sodium/hydrogen exchanger, NHE1, mediates the electroneutral 1:1 exchange of Na+ and H+ across the plasma membrane. NHE1 is ubiquitous and is studied extensively for regulation of pH i, cell volume, and response to growth factors. We describe a spontaneous mouse mutant, s low-w ave e pilepsy, (swe), with a neurological syndrome including ataxia and a unique epilepsy phenotype consisting of 3/sec absence and tonic-clonic seizures. swe was fine-mapped on Chromosome 4 and identified as a null allele of Nhe1. Mutants show selective neuronal death in the cerebellum and brainstem but otherwise are healthy. This first example of a disease-causing mutation in an Nhe gene provides a new tool for studying the delicate balance of neuroexcitability and cell survival within the CNS

WWOX P47T partial loss-of-function mutation induces epilepsy, progressive neuroinflammation, and cerebellar degeneration in mice phenocopying human SCAR12

WWOX gene loss-of-function (LoF) has been associated with neuropathologies resulting in developmental, epileptic, and ataxic phenotypes of varying severity based on the level of WWOX dysfunction. WWOX gene biallelic germline variant p.Pro47Thr (P47T) has been causally associated with a new form of autosomal recessive cerebellar ataxia with epilepsy and intellectual disability (SCAR12, MIM:614322). This mutation affecting the WW1 protein binding domain of WWOX, impairs its interaction with canonical proline-proline-X-tyrosine motifs in partner proteins. We generated a mutant knock-in mouse model of Wwox P47T mutation that phenocopies human SCAR12. WwoxP47T/P47T mice displayed epilepsy, profound social behavior and cognition deficits, and poor motor coordination, and unlike KO models that survive only for 1 month, live beyond 1 year of age. These deficits progressed with age and mice became practically immobile, suggesting severe cerebellar dysfunction. WwoxP47T/P47T mice brains revealed signs of progressive neuroinflammation with elevated astro-microgliosis that increased with age. Cerebellar cortex displayed significantly reduced molecular and granular layer thickness and a strikingly reduced number of Purkinje cells with degenerated dendrites. Transcriptome profiling from various brain regions of WW domain LoF mice highlighted widespread changes in neuronal and glial pathways, enrichment of bioprocesses related to neuroinflammation, and severe cerebellar dysfunction. Our results show significant pathobiological effects and potential mechanisms through which WWOX partial LoF leads to epilepsy, cerebellar neurodegeneration, neuroinflammation, and ataxia. Additionally, the mouse model described here will be a useful tool to understand the role of WWOX in common neurodegenerative conditions in which this gene has been identified as a novel risk factor.Fil: Hussain, Tabish. University of Texas; Estados UnidosFil: Sanchez, Kevin. University of Texas at Austin; Estados UnidosFil: Crayton, Jennifer. University of Texas; Estados UnidosFil: Saha, Dhurjhoti. University of Texas; Estados UnidosFil: Jeter, Collene. University of Texas; Estados UnidosFil: Lu, Yue. University of Texas; Estados UnidosFil: Abba, Martín Carlos. Universidad Nacional de La Plata. Facultad de Ciencias Médicas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata; ArgentinaFil: Seo, Ryan. Baylor College of Medicine; Estados UnidosFil: Noebels, Jeffrey L.. Baylor College of Medicine; Estados UnidosFil: Fonken, Laura. University of Texas at Austin; Estados UnidosFil: Aldaz, C. Marcelo. University of Texas; Estados Unido

Amyloid-beta/Fyn–Induced Synaptic, Network, and Cognitive Impairments Depend on Tau Levels in Multiple Mouse Models of Alzheimer’s Disease

Alzheimer\u27s disease (AD), the most common neurodegenerative disorder, is a growing public health problem and still lacks effective treatments. Recent evidence suggests that microtubule-associated protein tau may mediate amyloid-β peptide (Aβ) toxicity by modulating the tyrosine kinase Fyn.Weshowed previously that tau reduction prevents, and Fyn overexpression exacerbates, cognitive deficits in human amyloid precursor protein (hAPP) transgenic mice overexpressing Aβ. However, the mechanisms by which Aβ, tau, and Fyn cooperate in AD-related pathogenesis remain to be fully elucidated. Here we examined the synaptic and network effects of this pathogenic triad. Tau reduction prevented cognitive decline induced by synergistic effects of Aβ and Fyn. Tau reduction also prevented synaptic transmission and plasticity deficits in hAPP mice. Using electroencephalography to examine network effects, we found that tau reduction prevented spontaneous epileptiform activity in multiple lines of hAPP mice. Tau reduction also reduced the severity of spontaneous and chemically induced seizures in mice overexpressing both Aβ and Fyn. To better understand these protective effects, we recorded wholecell currents in acute hippocampal slices from hAPP mice with and without tau. hAPP mice with tau had increased spontaneous and evoked excitatory currents, reduced inhibitory currents, and NMDA receptor dysfunction. Tau reduction increased inhibitory currents and normalized excitation/inhibition balance and NMDA receptor-mediated currents in hAPP mice. Our results indicate that Aβ, tau, and Fyn jointly impair synaptic and network function and suggest that disrupting the copathogenic relationship between these factors could be of therapeutic benefit

Genetic suppression of transgenic APP rescues hypersynchronous network activity in a mouse model of alzeimer\u27s disease

Alzheimer's disease (AD) is associated with an elevated risk for seizures that may be fundamentally connected to cognitive dysfunction. Supporting this link, many mouse models for AD exhibit abnormal electroencephalogram (EEG) activity in addition to the expected neuropathology and cognitive deficits. Here, we used a controllable transgenic system to investigate how network changes develop and are maintained in a model characterized by amyloid β (Aβ) overproduction and progressive amyloid pathology. EEG recordings in tet-off mice overexpressing amyloid precursor protein (APP) from birth display frequent sharp wave discharges (SWDs). Unexpectedly, we found that withholding APP overexpression until adulthood substantially delayed the appearance of epileptiform activity. Together, these findings suggest that juvenile APP overexpression altered cortical development to favor synchronized firing. Regardless of the age at which EEG abnormalities appeared, the phenotype was dependent on continued APP overexpression and abated over several weeks once transgene expression was suppressed. Abnormal EEG discharges were independent of plaque load and could be extinguished without altering deposited amyloid. Selective reduction of Aβ with a γ-secretase inhibitor has no effect on the frequency of SWDs, indicating that another APP fragment or the full-length protein was likely responsible for maintaining EEG abnormalities. Moreover, transgene suppression normalized the ratio of excitatory to inhibitory innervation in the cortex, whereas secretase inhibition did not. Our results suggest that APP overexpression, and not Aβ overproduction, is responsible for EEG abnormalities in our transgenic mice and can be rescued independently of pathology