Altered phosphorylation and localization of the A-type channel, Kv4.2 in status epilepticus

The Cain Foundation Laboratories, Department of Pediatrics, Houston, Texas, USA.
Journal of Neurochemistry (Impact Factor: 4.28). 06/2008; 106(4):1929-40. DOI: 10.1111/j.1471-4159.2008.05508.x
Source: PubMed


Extracelluar signal-regulated kinase (ERK) pathway activation has been demonstrated following convulsant stimulation; however, little is known about the molecular targets of ERK in seizure models. Recently, it has been shown that ERK phosphorylates Kv4.2 channels leading to down-regulation of channel function, and substantially alters dendritic excitability. In the kainate model of status epilepticus (SE), we investigated whether ERK phosphorylates Kv4.2 and whether the changes in Kv4.2 were evident at a synaptosomal level during SE. Western blotting was performed on rat hippocampal whole cell, membrane, synaptosomal, and surface biotinylated extracts following systemic kainate using an antibody generated against the Kv4.2 ERK sites and for Kv4.2, ERK, and phospho-ERK. ERK activation was associated with an increase in Kv4.2 phosphorylation during behavioral SE. During SE, ERK activation and Kv4.2 phosphorylation were evident at the whole cell and synaptosomal levels. In addition, while whole-cell preparations revealed no alterations in total Kv4.2 levels, a decrease in synaptosomal and surface expression of Kv4.2 was evident after prolonged SE. These results demonstrate ERK pathway coupling to Kv4.2 phosphorylation. The finding of decreased Kv4.2 levels in hippocampal synaptosomes and surface membranes suggest additional mechanisms for decreasing the dendritic A-current, which could lead to altered intrinsic membrane excitability during SE.

Download full-text


Available from: Yajun Ren, Oct 05, 2015
15 Reads
  • Source
    • "All samples were then stored at -80°C until used. Hippocampi were homogenized in ice-cold homogenization buffer (0.32 M sucrose, 1 mM EDTA, 5 mM Hepes) containing protease inhibitor cocktail (Sigma, USA) and processed for western blotting as previously described (Lugo et al., 2008). Through this procedure we produced total homogenate samples and crude synaptosomes. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Many genes have been implicated in the underlying cause of autism but each gene accounts for only a small fraction of those diagnosed with autism. There is increasing evidence that activity-dependent changes in neuronal signaling could act as a convergent mechanism for many of the changes in synaptic proteins. One candidate signaling pathway that may have a critical role in autism is the PI3K/AKT/mTOR pathway. A major regulator of this pathway is the negative repressor phosphatase and tensin homolog (PTEN). In the current study we examined the behavioral and molecular consequences in mice with neuron subset-specific deletion of PTEN. The knockout (KO) mice showed deficits in social chamber and social partition test. KO mice demonstrated alterations in repetitive behavior, as measured in the marble burying test and hole-board test. They showed no changes in ultrasonic vocalizations emitted on postnatal day 10 or 12 compared to wildtype (WT) mice. They exhibited less anxiety in the elevated-plus maze test and were more active in the open field test compared to WT mice. In addition to the behavioral alterations, KO mice had elevation of phosphorylated AKT, phosphorylated S6, and an increase in S6K. KO mice had a decrease in mGluR but an increase in total and phosphorylated fragile X mental retardation protein. The disruptions in intracellular signaling may be why the KO mice had a decrease in the dendritic potassium channel Kv4.2 and a decrease in the synaptic scaffolding proteins PSD-95 and SAP102. These findings demonstrate that deletion of PTEN results in long-term alterations in social behavior, repetitive behavior, activity, and anxiety. In addition, deletion of PTEN significantly alters mGluR signaling and many synaptic proteins in the hippocampus. Our data demonstrates that deletion of PTEN can result in many of the behavioral features of autism and may provide insights into the regulation of intracellular signaling on synaptic proteins.
    Frontiers in Molecular Neuroscience 04/2014; 7(1):27. DOI:10.3389/fnmol.2014.00027 · 4.08 Impact Factor
  • Source
    • "All samples were stored at −80°C until used. Hippocampi were homogenized in ice-cold homogenization buffer (100 mM Tris-HCl, pH 7.4, 0.32 M sucrose, 1 mM EDTA, 5 mM Hepes) containing protease inhibitor cocktail (Roche, Alameda, CA, USA) and processed for western blotting as previously described [31]. The protein concentration was determined using the Bradford Protein Assay (Bio Rad, Hercules, CA, USA). "
    [Show abstract] [Hide abstract]
    ABSTRACT: Cognitive impairments are prominent sequelae of prolonged continuous seizures (status epilepticus; SE) in humans and animal models. While often associated with dendritic injury, the underlying mechanisms remain elusive. The mammalian target of rapamycin complex 1 (mTORC1) pathway is hyperactivated following SE. This pathway modulates learning and memory and is associated with regulation of neuronal, dendritic, and glial properties. Thus, in the present study we tested the hypothesis that SE-induced mTORC1 hyperactivation is a candidate mechanism underlying cognitive deficits and dendritic pathology seen following SE. We examined the effects of rapamycin, an mTORC1 inhibitor, on the early hippocampal-dependent spatial learning and memory deficits associated with an episode of pilocarpine-induced SE. Rapamycin-treated SE rats performed significantly better than the vehicle-treated rats in two spatial memory tasks, the Morris water maze and the novel object recognition test. At the molecular level, we found that the SE-induced increase in mTORC1 signaling was localized in neurons and microglia. Rapamycin decreased the SE-induced mTOR activation and attenuated microgliosis which was mostly localized within the CA1 area. These findings paralleled a reversal of the SE-induced decreases in dendritic Map2 and ion channels levels as well as improved dendritic branching and spine density in area CA1 following rapamycin treatment. Taken together, these findings suggest that mTORC1 hyperactivity contributes to early hippocampal-dependent spatial learning and memory deficits and dendritic dysregulation associated with SE.
    PLoS ONE 03/2013; 8(3):e57808. DOI:10.1371/journal.pone.0057808 · 3.23 Impact Factor
  • Source
    • "This effectively reduces the number of functional dendritic ion channels, even though the total HCN1 channel protein content remains unchanged. Similar alterations in ion channel trafficking to the surface membrane post-SE have been described for A-type K + channels and GABA A -receptor subunits, and thus may be a common theme in acquired channelopathy (Goodkin et al., 2008; Lugo et al., 2008; Terunuma et al., 2008). These latter examples are phosphorylation dependent , although such a mechanism has not yet been established for altered trafficking of HCN1 channels. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Ion channel dysfunction or “channelopathy” is a proven cause of epilepsy in the relatively uncommon genetic epilepsies with Mendelian inheritance. But numerous examples of acquired channelopathy in experimental animal models of epilepsy following brain injury have also been demonstrated. Our understanding of channelopathy has grown due to advances in electrophysiology techniques that have allowed the study of ion channels in the dendrites of pyramidal neurons in cortex and hippocampus. The apical dendrites of pyramidal neurons comprise the vast majority of neuronal surface membrane area, and thus the majority of the neuronal ion channel population. Investigation of dendritic ion channels has demonstrated remarkable plasticity in ion channel localization and biophysical properties in epilepsy, many of which produce hyperexcitability and may contribute to the development and maintenance of the epileptic state. Herein we review recent advances in dendritic physiology and cell biology, and their relevance to epilepsy.
    Epilepsia 12/2012; 53(s9). DOI:10.1111/epi.12033 · 4.57 Impact Factor
Show more