Coordinated downregulation of KCC2 and GABAA receptor contributes to inhibitory dysfunction during seizure induction

Wan, Li; Chen, Lulan; Yu, Jiangning; Wang, Guoxiang; Wu, Zheng; Qian, Binbin; Liu, Xu; Wang, Yun

doi:10.1016/j.bbrc.2020.08.082

Cited by 9 publications

(8 citation statements)

References 30 publications

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“…Previous studies have focused on functional inhibition mediated by FUR, however, no prior research has explored whether FUR affects KCC2 expression. Here in our current study, similar as previous reported ( Wan et al, 2020 ), 100 µM FUR showed the depolarization shift of the E GABA which suggested an inhibition on KCC2 in mature neurons. But the effect of 100 µM FUR is suspected to result from inhibition of both KCC2 and NKCC1, since FUR also serves as an inhibitor for NKCC1, and the inhibition constant (Ki) for KCC2 and NKCC1 is 25 µM ( Payne, 1997 ) and 40 µM ( Gillen et al, 1996 ), respectively.…”

Section: Discussionsupporting

confidence: 93%

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Furosemide prevents membrane KCC2 downregulation during convulsant stimulation in the hippocampus

Chen

Wan

et al. 2022

IBRO Neuroscience Reports

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Section: Discussionsupporting

confidence: 93%

“…The epileptiform activity had long durations even after CTZ was washed out ( Qi et al, 2006 ). FUR seemed to prevent KCC2 downregulation by CTZ( Wan et al, 2020 ). In this study, we aimed to further investigate the effects of FUR on mKCC2 levels with and without CTZ-induced seizures using western blot (WB) and immunostaining.…”

Section: Introductionmentioning

confidence: 99%

Furosemide prevents membrane KCC2 downregulation during convulsant stimulation in the hippocampus

Chen

Wan

et al. 2022

IBRO Neuroscience Reports

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“…Our results revealed a novel mode of hUC-MSC transplantation that specifically targeting the reactivation of GABA A R subunits β3 and γ2 in combination with KCC2 in spinal neurons to facilitate the functional recovery of SCI (Figure 7 ). SCI downregulates the KCC2 expression, leading to an increase in [Cl⁻] i , which could restore an immature neuron state with the functional transition of GABA A R activation from hyperpolarization to depolarization 48 . Thereafter, this depolarization activates Ca 2+ influx through voltage-gated calcium channels (VGCC) and induces BDNF, which in turn increases the expression and cell surface presence of GABA A Rs via the transcriptional and posttranslational regulations 49 , 50 .…”

Section: Discussionmentioning

confidence: 99%

hUC-MSC-mediated recovery of subacute spinal cord injury through enhancing the pivotal subunits β3 and γ2 of the GABA_A receptor

Cao¹,

Chen²,

Huang³

et al. 2022

Theranostics

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Rationale: Spinal cord injury (SCI) remains an incurable neurological disorder leading to permanent and profound neurologic deficits and disabilities. Human umbilical cord-derived mesenchymal stem cells (hUC-MSCs) are particularly appealing in SCI treatment to curtail damage, restore homeostasis and possible neural relay. However, the detailed mechanisms underlying hUC-MSC-mediated functional recovery of SCI have not been fully elucidated. The purpose of our current study is to identify novel therapeutic targets and depict the molecular mechanisms underlying the hUC-MSC-mediated recovery of subacute SCI. Methods: Adult female rats suffering from subacute incomplete thoracic SCI were treated with intrathecal transplantation of hUC-MSCs. The beneficial effects of hUC-MSCs on SCI repair were evaluated by a series of behavioral analyses, motor evoked potentials (MEPs) recording of hindlimb and immunohistochemistry. We carried out extensive transcriptome comparative analyses of spinal cord tissues at the lesion site from the subacute phase of SCI (sub-SCI) either treated without (+PBS) or with hUC-MSCs (+MSC) at 0 (sub-SCI), 1, 2, and 4 weeks post-transplantation (wpt), as well as normal spinal cord segments of intact/sham rats (Intact). Adeno-associated virus (AAV)-mediated neuron-specific expression system was employed to functionally screen specific γ-aminobutyric acid type A receptor (GABA A R) subunits promoting the functional recovery of SCI in vivo . The mature cortical axon scrape assay and transplantation of genetically modified MSCs with either overexpression or knockdown of brain-derived neurotrophic factor (BDNF) were employed to demonstrate that hUC-MSCs ameliorated the reduction of GABA A R subunits in the injured spinal cord via BDNF secretion in vitro and in vivo , respectively. Results: Comparative transcriptome analysis revealed the GABAergic synapse pathway is significantly enriched as a main target of hUC-MSC-activated genes in the injured spinal cord. Functional screening of the primary GABA A R subunits uncovered that Gabrb3 and Garbg2 harbored the motor and electrophysiological recovery-promoting competence. Moreover, targeting either of the two pivotal subunits β3 or γ2 in combination with/without the K + /Cl - cotransporter 2 (KCC2) reinforced the therapeutic effects. Mechanistically, BDNF secreted by hUC-MSCs contributed to the upregulation of GABA A R subunits (β3 & γ2) and KCC2 in the injured neurons. Conclusions: Our study identifies a novel mode for hUC-MSC-mediated locomotor recovery of SCI through synergistic upregulation of GABA A R β3 and γ2 along with KCC2 by BDNF secretion, indicating the significance of restoring the excitation/inhibition balan...

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“…However, these manipulations do not affect KCC2 dwell time near GABAergic synapses, suggesting the existence of synapse-specific molecular interactions contributing to the formation of perisynaptic KCC2 clusters. Although KCC2 was shown to interact with both GABA A (Huang et al, 2012;Wan et al, 2020) and GABA B (Wright et al, 2017) receptors, the molecular mechanisms of KCC2 clustering near GABAergic synapses remain to be identified. Here, we addressed this issue using proteomic analysis of the KCC2 interactome in cortical principal neurons.…”

Section: Introductionmentioning

confidence: 99%

Gephyrin Interacts with the K-Cl Cotransporter KCC2 to Regulate Its Surface Expression and Function in Cortical Neurons

et al. 2021

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The K 1 -Clcotransporter KCC2, encoded by the Slc12a5 gene, is a neuron-specific chloride extruder that tunes the strength and polarity of GABA A receptor-mediated transmission. In addition to its canonical ion transport function, KCC2 also regulates spinogenesis and excitatory synaptic function through interaction with a variety of molecular partners. KCC2 is enriched in the vicinity of both glutamatergic and GABAergic synapses, the activity of which in turn regulates its membrane stability and function. KCC2 interaction with the submembrane actin cytoskeleton via 4.1N is known to control its anchoring near glutamatergic synapses on dendritic spines. However, the molecular determinants of KCC2 clustering near GABAergic synapses remain unknown. Here, we used proteomics to identify novel KCC2 interacting proteins in the adult rat neocortex. We identified both known and novel candidate KCC2 partners, including some involved in neuronal development and synaptic transmission. These include gephyrin, the main scaffolding molecule at GABAergic synapses. Gephyrin interaction with endogenous KCC2 was confirmed by immunoprecipitation from rat neocortical extracts. We showed that gephyrin stabilizes plasmalemmal KCC2 and promotes its clustering in hippocampal neurons, mostly but not exclusively near GABAergic synapses, thereby controlling KCC2-mediated chloride extrusion. This study identifies gephyrin as a novel KCC2 anchoring molecule that regulates its membrane expression and function in cortical neurons.

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Coordinated downregulation of KCC2 and GABAA receptor contributes to inhibitory dysfunction during seizure induction

Cited by 9 publications

References 30 publications

Furosemide prevents membrane KCC2 downregulation during convulsant stimulation in the hippocampus

Furosemide prevents membrane KCC2 downregulation during convulsant stimulation in the hippocampus

hUC-MSC-mediated recovery of subacute spinal cord injury through enhancing the pivotal subunits β3 and γ2 of the GABA_A receptor

Gephyrin Interacts with the K-Cl Cotransporter KCC2 to Regulate Its Surface Expression and Function in Cortical Neurons

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Coordinated downregulation of KCC2 and GABAA receptor contributes to inhibitory dysfunction during seizure induction

Cited by 9 publications

References 30 publications

Furosemide prevents membrane KCC2 downregulation during convulsant stimulation in the hippocampus

Furosemide prevents membrane KCC2 downregulation during convulsant stimulation in the hippocampus

hUC-MSC-mediated recovery of subacute spinal cord injury through enhancing the pivotal subunits β3 and γ2 of the GABAA receptor

Gephyrin Interacts with the K-Cl Cotransporter KCC2 to Regulate Its Surface Expression and Function in Cortical Neurons

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hUC-MSC-mediated recovery of subacute spinal cord injury through enhancing the pivotal subunits β3 and γ2 of the GABA_A receptor