Seizures and Reduced Life Span in Mice Lacking the Potassium Channel Subunit Kv1.2, but Hypoexcitability and Enlarged Kv1 Currents in Auditory Neurons

295

392

Epilepsy is a devastating and poorly understood disease. Mutations in a secreted neuronal protein, leucine-rich glioma inactivated 1 (LGI1), were reported in patients with an inherited form of human epilepsy, autosomal dominant partial epilepsy with auditory features (ADPEAF). Here, we report an essential role of LGI1 as an antiepileptogenic ligand. We find that loss of LGI1 in mice (LGI1 −/− ) causes lethal epilepsy, which is specifically rescued by the neuronal expression of LGI1 transgene, but not LGI3. Moreover, heterozygous mice for the LGI1 mutation (LGI1 +/− ) show lowered seizure thresholds. Extracellularly secreted LGI1 links two epilepsy-related receptors, ADAM22 and ADAM23, in the brain and organizes a transsynaptic protein complex that includes presynaptic potassium channels and postsynaptic AMPA receptor scaffolds. A lack of LGI1 disrupts this synaptic protein connection and selectively reduces AMPA receptor-mediated synaptic transmission in the hippocampus. Thus, LGI1 may serve as a major determinant of brain excitation, and the LGI1 gene-targeted mouse provides a good model for human epilepsy.A ffecting 1-2% of the population, epilepsy is one of the most common neurological disorders. Epilepsy is characterized by recurrent unprovoked seizures and is caused by disturbances in the delicate balance between excitation and inhibition in neural circuits (1, 2). Recent human genetic studies have established the channelopathy concept for idiopathic (inherited) epilepsies: Many of the genes whose mutations cause human epilepsy encode ion channel subunits (1, 2). Examples include voltagegated ion channels (K + , Na + , Ca 2+ , and Cl -channels) and ligandgated ion channels (nicotinic acetylcholine and GABA A receptors), which regulate neuronal excitability.Leucine-rich glioma inactivated 1 (LGI1) is a unique human epilepsy-related gene in that it does not encode an ion channel subunit (3-5), but is a neuronal secreted protein (6). Mutations in LGI1 are linked to autosomal dominant partial epilepsy with auditory features (ADPEAF, also known as autosomal dominant lateral temporal lobe epilepsy [ADLTE]) (3-5), which is an inherited epileptic syndrome characterized by partial seizures with acoustic or visual hallucinations. So far, 25 LGI1 mutations have been described in familial ADPEAF patients and sporadic cases (7). Interestingly, at least six tested ADPEAF mutations all abolish LGI1 secretion (6,8).Recent proteomic analysis identified LGI1 as a subunit of presynaptic Kv 1 (shaker type)-voltage gated potassium channels (9). It was shown that LGI1 selectively prevents inactivation of the Kv 1 channel mediated by a cytoplasmic regulatory protein, Kvβ. Because LGI1 is a secreted protein, it remains unclear how LGI1 modulates a cytosolic potassium channel mechanism. LGI1 was also isolated from the brain as a component of a protein complex mediated by PSD-95, a representative postsynaptic scaffolding protein.LGI1 functions as a ligand for the epilepsy-related ADAM22 transmembrane protein, which is anchored by PS...

Section: Resultsmentioning

confidence: 99%

Disruption of LGI1–linked synaptic complex causes abnormal synaptic transmission and epilepsy

Fukata¹,

Lovero

Iwanaga³

et al. 2010

295

392

“…For instance, genome-wide analysis of developing neurons (26) identified MEF2 target genes that (i) decrease surface AMPAR expression [arc, JNK, SynGAP (27-31)], (ii) uncouple mGluR5 from its effector targets in the postsynaptic density and reduce the amplitude of AMPAR-and NMDARmediated excitatory postsynaptic currents [Homer1a (32-34)], (iii) internalize N-cadherin, a synaptic adhesion molecule implicated in synaptic transmission [PCDH (protocadherin) family members, including protocadherins 9, 10, and 17 (35)], and (iv) decrease neuronal excitability [voltage-gated K + channels (including kcna1 and kcna4) (36)(37)(38)]. Therefore, increasing MEF2-dependent transcription may orchestrate a coordinated increase in expression of genes that suppress excitatory synapse function.…”

Section: Discussionmentioning

confidence: 99%

Spine growth in the anterior cingulate cortex is necessary for the consolidation of contextual fear memory

Vetere

Restivo

Cole

et al. 2011

160

135

Remodeling of cortical connectivity is thought to allow initially hippocampus-dependent memories to be expressed independently of the hippocampus at remote time points. Consistent with this, consolidation of a contextual fear memory is associated with dendritic spine growth in neurons of the anterior cingulate cortex (aCC). To directly test whether such cortical structural remodeling is necessary for memory consolidation, we disrupted spine growth in the aCC at different times following contextual fear conditioning in mice. We took advantage of previous studies showing that the transcription factor myocyte enhancer factor 2 (MEF2) negatively regulates spinogenesis both in vitro and in vivo. We found that increasing MEF2-dependent transcription in the aCC during a critical posttraining window (but not at later time points) blocked both the consolidation-associated dendritic spine growth and subsequent memory expression. Together, these data strengthen the causal link between cortical structural remodeling and memory consolidation and, further, identify MEF2 as a key regulator of these processes.structural plasticity | remote memory | viral vector I n experimental animals, damage to the hippocampus disproportionately impacts recently acquired memories, with relative sparing of remote memories (1-8). Such observations have led to the idea that the hippocampus is transiently required for memory expression, with remote memory expression being exclusively dependent on the cortex (9). According to one model (10), posttraining hippocampal activity coordinates reactivation of memory traces in the cortex. This reactivation leads to the remodeling of cortical connections, allowing the memory to eventually be expressed independently of the hippocampus. A recent study in mice (5) provided correlative evidence for posttraining remodeling of neurons in the anterior cingulate cortex (aCC), a subregion of the prefrontal cortex that plays an essential role in remote memory expression (11). Increases in dendritic spine density on layer 2/3 pyramidal aCC neurons were observed 1 mo, but not 1 d, following contextual fear conditioning (5). As layer 2/3 pyramidal neurons send and receive long-range cortical connections (12), such changes may contribute to increased functional connectivity between the aCC and other cortical areas important for remote memory expression (13,14). However, whether this increase in aCC spine density is necessary for memory consolidation is not known. To test this, it would be necessary to evaluate whether preventing posttraining spinogenesis, specifically in this region, disrupts memory consolidation.The transcription factor myocyte enhancer factor 2 (MEF2) negatively regulates spinogenesis in an activity-dependent manner and therefore provides a tool to address this question. For example, increasing MEF2 function decreases the number of dendritic spines and excitatory synapses in vitro (15) and blocks increases in spine density normally observed following repeated cocaine administration in rat medium spiny...

“…Our data suggest that signaling pathways that impact levels of Kv1.2 phosphorylation may also regulate biosynthetic trafficking of Kv1.2-containing channels. Dramatic effects of lowering functional levels of Kv1.2-containing channels by pharmacological blockade leads to enhanced depolarization-induced neurotransmitter release (1) and seizures (28), and Kv1.2 knockout mice exhibit enhanced seizure susceptibility and die on average by postnatal day 17 (8). As such, regulation of Kv1.2 activity in mammalian neurons is crucial to normal brain function.…”

Section: Discussionmentioning

confidence: 99%

“…In mammalian brain, Kv1.2 is found prominently along unmyelinated axons, in nerve terminals and/or in preterminal axon segments, at axon initial segments, and at juxtaparanodes of myelinated axons, but it is also present in the dendrites of some neurons (7). The physiological importance of Kv1.2 is underscored by the recent finding that Kv1.2 knockout mice have enhanced seizure susceptibility and die in the third postnatal week (8).…”

mentioning

confidence: 99%

Trafficking-dependent phosphorylation of Kv1.2 regulates voltage-gated potassium channel cell surface expression

Yang¹,

Vacher²,

Park³

et al. 2007

Kv1.2 ␣-subunits are components of low-threshold, rapidly activating voltage-gated potassium (Kv) channels in mammalian neurons. Expression and localization of Kv channels is regulated by trafficking signals encoded in their primary structure. Kv1.2 is unique in lacking strong trafficking signals and in exhibiting dramatic cell-specific differences in trafficking, which is suggestive of conditional trafficking signals. Here we show that a cluster of cytoplasmic C-terminal phosphorylation sites regulates Kv1.2 trafficking. Using tandem MS to analyze Kv1.2 purified from rat, human, and mouse brain, we identified in each sample in vivo phosphoserine (pS) phosphorylation sites at pS434, pS440, and pS441, as well as doubly phosphorylated pS440/pS441. We also found these sites, as well as pS449, on recombinant Kv1.2 expressed in heterologous cells. We found that phosphorylation at pS440/pS441 is present only on the post-endoplasmic reticulum (ER)/cell surface pool of Kv1.2 and is not detectable on newly synthesized and ER-localized Kv1.2, on which we did observe pS449 phosphorylation. Elimination of PS440/PS441 phosphorylation by mutation reduces cell-surface expression efficiency and functional expression of homomeric Kv1.2 channels. Interestingly, mutation of S449 reduces phosphorylation at pS440/pS441 and also decreases Kv1.2 cell-surface expression efficiency and functional expression. These mutations also suppress trafficking of Kv1.2/ Kv1.4 heteromeric channels, suggesting that incorporation of Kv1.2 into heteromeric complexes confers conditional phosphorylation-dependent trafficking to diverse Kv channel complexes. These data support Kv1.2 phosphorylation at these clustered C-terminal sites as playing an important role in regulating trafficking of Kv1.2-containing Kv channels.ion channel ͉ mass spectrometry ͉ proteomics ͉ brain ͉ phosphospecific V oltage-gated potassium, or Kv, channels that form as tetramers of Kv1 ␣-subunits play important and diverse roles in regulating excitability of axons, nerve terminals, and dendrites of mammalian neurons (1). The major Kv1 ␣-subunits in mammalian brain, Kv1.1, Kv1.2, and Kv1.4, are found predominantly in heterotetrameric channel complexes (2-4). Homotetrameric Kv1.2 channels form low-threshold, sustained-or delayed-rectifier Kv channels (5) and, when coassembled with other Kv1 ␣-subunits, can generate a diverse array of channel subtypes (6). In mammalian brain, Kv1.2 is found prominently along unmyelinated axons, in nerve terminals and/or in preterminal axon segments, at axon initial segments, and at juxtaparanodes of myelinated axons, but it is also present in the dendrites of some neurons (7). The physiological importance of Kv1.2 is underscored by the recent finding that Kv1.2 knockout mice have enhanced seizure susceptibility and die in the third postnatal week (8).Biosynthetic trafficking of Kv1 channels from their site of translation in the rough endoplasmic reticulum (ER) to the cell surface is governed by a set of intrinsic targeting motifs encoded in primary s...