Activity-dependent redistribution of Kv2.1 ion channels on rat spinal motoneurons

Romer, Shannon H.; Deardorff, Adam S.; Fyffe, Robert E.W.

doi:10.14814/phy2.13039

Cited by 20 publications

(20 citation statements)

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“…As previously described (Romer et al . , ), micrographs of immunolabelled lumbar MN images were obtained with an Olympus Fluoview FV1000 (Center Valley, PA, USA) confocal microscope with a 60× oil immersion objective (N.A 1.35) at 1.0 µm z ‐steps. All control slices from the same rats were imaged and analysed with the same imaging parameters as experimental slices.…”

Section: Methodsmentioning

confidence: 99%

“…Cell body Kv2.1 immunoreactive (IR) macroclusters (diameter >1.0 µm), were measured in en face single optical sections on lumbar MNs (Romer et al . , ).…”

Section: Methodsmentioning

confidence: 99%

“…We have previously demonstrated that Kv2.1 channels rapidly decluster on MNs immediately following either peripheral nerve injury or in vivo stimulation of peripheral nerves, suggesting that Kv2.1 channels have the capacity to dynamically respond to pathological and/or prolonged excitatory stimuli (Romer et al . , ). Here we explore the impact of blocking Kv2 channel conductances using the tarantula toxin stromatoxin (STX; Escoubas et al .…”

Section: Introductionmentioning

confidence: 99%

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A molecular rheostat: Kv2.1 currents maintain or suppress repetitive firing in motoneurons

Romer

Deardorff

Fyffe

2019

The Journal of Physiology

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Key points Kv2 currents maintain and regulate motoneuron (MN) repetitive firing properties. Kv2.1 channel clustering properties are dynamic and respond to both high and low activity conditions. The enzyme calcineurin regulates Kv2.1 ion channel declustering. In patholophysiological conditions of high activity, Kv2.1 channels homeostatically reduce MN repetitive firing. Modulation of Kv2.1 channel kinetics and clustering allows these channels to act in a variable way across a spectrum of MN activity states. Abstract Kv2.1 channels are widely expressed in the central nervous system, including in spinal motoneurons (MNs) where they aggregate as distinct membrane clusters associated with highly regulated signalling ensembles at specific postsynaptic sites. Multiple roles for Kv2 channels have been proposed but the physiological role of Kv2.1 ion channels in mammalian spinal MNs is unknown. To determine the contribution of Kv2.1 channels to rat α‐motoneuron activity, the Kv2 inhibitor stromatoxin was used to block Kv2 currents in whole‐cell current clamp electrophysiological recordings in rat lumbar MNs. The results indicate that Kv2 currents permit shorter interspike intervals and higher repetitive firing rates, possibly by relieving Na+ channel inactivation, and thus contribute to maintenance of repetitive firing properties. We also demonstrate that Kv2.1 clustering properties in motoneurons are dynamic and respond to both high and low activity conditions. Furthermore, we show that the enzyme calcineurin regulates Kv2.1 ion channel clustering status. Finally, in a high activity state, Kv2.1 channels homeostatically reduce motoneuron repetitive firing. These results suggest that the activity‐dependent regulation of Kv2.1 channel kinetics allows these channels to modulate repetitive firing properties across a spectrum of motoneuron activity states.

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Section: Methodsmentioning

confidence: 99%

“…Cell body Kv2.1 immunoreactive (IR) macroclusters (diameter >1.0 µm), were measured in en face single optical sections on lumbar MNs (Romer et al . , ).…”

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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A molecular rheostat: Kv2.1 currents maintain or suppress repetitive firing in motoneurons

Romer

Deardorff

Fyffe

2019

The Journal of Physiology

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“…Both channels localize to micrometer-sized clusters on the neuronal surface of the soma, proximal dendrites, and axon initial segment (AIS) in vivo and in vitro (2). Clustered Kv2.1 channels disperse in response to ischemic or hypoxic conditions, neuronal activity, and glutamate-induced excitotoxity via calcineurin-dependent dephosphorylation of the channel C terminus (3,4). While Kv2.1 clustering was first proposed to regulate channel voltage dependence (5), several studies indicate little connection between channel clustering and regulation of conductance (6)(7)(8).…”

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confidence: 99%

Kv2 potassium channels form endoplasmic reticulum/plasma membrane junctions via interaction with VAPA and VAPB

Johnson

Leek

Solé

et al. 2018

Proc. Natl. Acad. Sci. U.S.A.

144

202

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Kv2.1 exhibits two distinct forms of localization patterns on the neuronal plasma membrane: One population is freely diffusive and regulates electrical activity via voltage-dependent K conductance while a second one localizes to micrometer-sized clusters that contain densely packed, but nonconducting, channels. We have previously established that these clusters represent endoplasmic reticulum/plasma membrane (ER/PM) junctions that function as membrane trafficking hubs and that Kv2.1 plays a structural role in forming these membrane contact sites in both primary neuronal cultures and transfected HEK cells. Clustering and the formation of ER/PM contacts are regulated by phosphorylation within the channel C terminus, offering cells fast, dynamic control over the physical relationship between the cortical ER and PM. The present study addresses the mechanisms by which Kv2.1 and the related Kv2.2 channel interact with the ER membrane. Using proximity-based biotinylation techniques in transfected HEK cells we identified ER VAMP-associated proteins (VAPs) as potential Kv2.1 interactors. Confirmation that Kv2.1 and -2.2 bind VAPA and VAPB employed colocalization/redistribution, siRNA knockdown, and Förster resonance energy transfer (FRET)-based assays. CD4 chimeras containing sequence from the Kv2.1 C terminus were used to identify a noncanonical VAP-binding motif. VAPs were first identified as proteins required for neurotransmitter release in and are now known to be abundant scaffolding proteins involved in membrane contact site formation throughout the ER. The VAP interactome includes AKAPs, kinases, membrane trafficking machinery, and proteins regulating nonvesicular lipid transport from the ER to the PM. Therefore, the Kv2-induced VAP concentration at ER/PM contact sites is predicted to have wide-ranging effects on neuronal cell biology.

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“…Kv2.1 exhibits conditional phosphorylation-dependent clustering that can be regulated by neuronal activity and other stimuli (52-54, 80-82). In certain mammalian cell lines such as COS- 1 cells, Kv2.1 exhibits reversible cell-cycle dependent clustering and recruitment/stabilization of ER-PM junctions, presumably due to increased phosphorylation of Kv2.1 observed at the onset of M-phase (47).…”

Section: Resultsmentioning

confidence: 99%

Organizing neuronal ER-PM junctions is a conserved nonconducting function of Kv2 plasma membrane ion channels

Kirmiz

Palacio

Thapa

et al. 2018

Preprint

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27Endoplasmic reticulum (ER) and plasma membrane (PM) form junctions crucial to ion and lipid 28 signaling and homeostasis. The Kv2.1 ion channel is unique among PM proteins in organizing 29 ER-PM junctions. Here, we show that this organizing function is conserved between Kv2 family 30 members that differ in their biophysical properties, modulation and cellular expression. 31Manipulation of actin cytoskeleton surrounding Kv2 ER-PM junctions affects their spatial 32 organization. Kv2-containing ER-PM junctions overlap with those formed by canonical ER-PM 33 tethers. ER-PM junction organization by Kv2 channels is unchanged by point mutations that 34 eliminate ion conduction, but abolished by those that eliminate PM clustering without impacting 35 ion channel function. Kv2.2 is distinct in lacking the reversible modulation of junction organization 36 present in Kv2.1. Brain neurons in Kv2 double knockout mice have altered ER-PM junctions, 37 demonstrating a conserved in vivo function for Kv2 family members distinct from their canonical 38 role as ion-conducting channels shaping neuronal excitability. 39 40. CC-BY 4.0 International license It is made available under a was not peer-reviewed) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity.The copyright holder for this preprint (which . http://dx.doi.org/10.1101/296731 doi: bioRxiv preprint first posted online Apr. 6, 2018; 3 Introduction 41Membrane contacts between the endoplasmic reticulum (ER) and plasma membrane (PM), or 42 ER-PM junctions, are a ubiquitous feature of eukaryotic cells (1-4). These specialized sites at 43 which ER is held in close apposition (10-30 nm) and other excitable and non-excitable cell types. In brain neurons, Kv2 channels are distinct from 63 other Kv channels (17) in being specifically localized to high-density micron sized clusters 64 prominent on the soma, proximal dendrites, and axon initial segment (34-42). Kv2 channels also 65 form such clusters when exogenously expressed in cultured neurons and in heterologous cells 66 . CC-BY 4.0 International license It is made available under a was not peer-reviewed) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity.The copyright holder for this preprint (which . http://dx.doi.org/10.1101/296731 doi: bioRxiv preprint first posted online Apr. 6, 2018; 4 (35, 37, 39, 41-47 at/near RyR clusters in specific neuron types, including hippocampal CA1 pyramidal neurons and 114 layer 6 neocortical neurons (Figure 1). A similar juxtaposition of Kv2.2 and RyR clusters was seen 115 in CHNs (Figure 1). In these classes of neurons, Kv2.2 was often found coclustered with Kv2.1 at 116 these ER-PM junctions (Figure 1). Neurons in each preparation also contained RyR clusters that 117 did not appear to colocalize with Kv2.2 or Kv2.1 (Figure 1). These findings demonstrate that Kv2.2 118 . CC-BY 4.0 International license It is made available under a was not peer-reviewed) is the author/funder, who has granted ...

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Activity-dependent redistribution of Kv2.1 ion channels on rat spinal motoneurons

Cited by 20 publications

References 64 publications

A molecular rheostat: Kv2.1 currents maintain or suppress repetitive firing in motoneurons

A molecular rheostat: Kv2.1 currents maintain or suppress repetitive firing in motoneurons

Kv2 potassium channels form endoplasmic reticulum/plasma membrane junctions via interaction with VAPA and VAPB

Organizing neuronal ER-PM junctions is a conserved nonconducting function of Kv2 plasma membrane ion channels

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