A novel epilepsy mutation in the sodium channel SCN1A identifies a cytoplasmic domain for {beta} subunit interaction

A mutation in the sodium channel SCN1A was identified in a small Italian family with dominantly inherited generalized epilepsy with febrile seizures plus (GEFS+). The mutation, D1866Y, alters an evolutionarily conserved aspartate residue in the C-terminal cytoplasmic domain of the sodium channel {alpha} subunit. The mutation decreased modulation of the {alpha} subunit by {beta}1, which normally causes a negative shift in the voltage dependence of inactivation in oocytes. There was less of a shift with the mutant channel, resulting in a 10 mV difference between the wild-type and mutant channels in the presence of {beta}1. This shift increased the magnitude of the window current, which resulted in more persistent current during a voltage ramp. Computational analysis suggests that neurons expressing the mutant channels will fire an action potential with a shorter onset delay in response to a threshold current injection, and that they will fire multiple action potentials with a shorter interspike interval at a higher input stimulus. These results suggest a causal relationship between a positive shift in the voltage dependence of sodium channel inactivation and spontaneous seizure activity. Direct interaction between the cytoplasmic C-terminal domain of the wild-type{alpha} subunit with the {beta}1or {beta}3 subunit was first demonstrated by yeast two-hybrid analysis. The SCN1A peptide K1846-R1886 is sufficient for {beta} subunit interaction. Coimmunoprecipitation from transfected mammalian cells confirmed the interaction between the C-terminal domains of the {alpha} and {beta}1 subunits. The D1866Y mutation weakens this interaction, demonstrating a novel molecular mechanism leading to seizure susceptibility

Neuropeptide S-Mediated Facilitation of Synaptic Transmission Enforces Subthreshold Theta Oscillations within the Lateral Amygdala

The neuropeptide S (NPS) receptor system modulates neuronal circuit activity in the amygdala in conjunction with fear, anxiety and the expression and extinction of previously acquired fear memories. Using in vitro brain slice preparations of transgenic GAD67-GFP (Δneo) mice, we investigated the effects of NPS on neural activity in the lateral amygdala as a key region for the formation and extinction of fear memories. We are able to demonstrate that NPS augments excitatory glutamatergic synaptic input onto both projection neurons and interneurons of the lateral amygdala, resulting in enhanced spike activity of both types of cells. These effects were at least in part mediated by presynaptic mechanisms. In turn, inhibition of projection neurons by local interneurons was augmented by NPS, and subthreshold oscillations were strengthened, leading to their shift into the theta frequency range. These data suggest that the multifaceted effects of NPS on amygdaloid circuitry may shape behavior-related network activity patterns in the amygdala and reflect the peptide's potent activity in various forms of affective behavior and emotional memory

Genomic Analysis of wig-1 Pathways

Background: Wig-1 is a transcription factor regulated by p53 that can interact with hnRNP A2/B1, RNA Helicase A, and dsRNAs, which plays an important role in RNA and protein stabilization. in vitro studies have shown that wig-1 binds p53 mRNA and stabilizes it by protecting it from deadenylation. Furthermore, p53 has been implicated as a causal factor in neurodegenerative diseases based in part on its selective regulatory function on gene expression, including genes which, in turn, also possess regulatory functions on gene expression. In this study we focused on the wig-1 transcription factor as a downstream p53 regulated gene and characterized the effects of wig-1 down regulation on gene expression in mouse liver and brain. Methods and Results: Antisense oligonucleotides (ASOs) were identified that specifically target mouse wig-1 mRNA and produce a dose-dependent reduction in wig-1 mRNA levels in cell culture. These wig-1 ASOs produced marked reductions in wig-1 levels in liver following intraperitoneal administration and in brain tissue following ASO administration through a single striatal bolus injection in FVB and BACHD mice. Wig-1 suppression was well tolerated and resulted in the reduction of mutant Htt protein levels in BACHD mouse brain but had no effect on normal Htt protein levels nor p53 mRNA or protein levels. Expression microarray analysis was employed to determine the effects of wig-1 suppression on genome-wide expression in mouse liver and brain. Reduction of wig-1 caused both down regulation and up regulation of several genes

The Lysosome and Intracellular Signalling.

In addition to being the terminal degradative compartment of the cell's endocytic and autophagic pathways, the lysosome is a multifunctional signalling hub integrating the cell's response to nutrient status and growth factor/hormone signalling. The cytosolic surface of the limiting membrane of the lysosome is the site of activation of the multiprotein complex mammalian target of rapamycin complex 1 (mTORC1), which phosphorylates numerous cell growth-related substrates, including transcription factor EB (TFEB). Under conditions in which mTORC1 is inhibited including starvation, TFEB becomes dephosphorylated and translocates to the nucleus where it functions as a master regulator of lysosome biogenesis. The signalling role of lysosomes is not limited to this pathway. They act as an intracellular Ca2+ store, which can release Ca2+ into the cytosol for both local effects on membrane fusion and pleiotropic effects within the cell. The relationship and crosstalk between the lysosomal and endoplasmic reticulum (ER) Ca2+ stores play a role in shaping intracellular Ca2+ signalling. Lysosomes also perform other signalling functions, which are discussed. Current views of the lysosomal compartment recognize its dynamic nature. It includes endolysosomes, autolysosome and storage lysosomes that are constantly engaged in fusion/fission events and lysosome regeneration. How signalling is affected by individual lysosomal organelles being at different stages of these processes and/or at different sites within the cell is poorly understood, but is discussed