Publications by authors named "Eric Delpire"

156 Publications

Large-Scale Proteomic Assessment of Urinary Extracellular Vesicles Highlights Their Reliability in Reflecting Protein Changes in the Kidney.

J Am Soc Nephrol 2021 09 6;32(9):2195-2209. Epub 2021 Jul 6.

Department of Biomedicine, Aarhus University, Aarhus, Denmark

Background: Urinary extracellular vesicles (uEVs) are secreted into urine by cells from the kidneys and urinary tract. Although changes in uEV proteins are used for quantitative assessment of protein levels in the kidney or biomarker discovery, whether they faithfully reflect (patho)physiologic changes in the kidney is a matter of debate.

Methods: Mass spectrometry was used to compare in an unbiased manner the correlations between protein levels in uEVs and kidney tissue from the same animal. Studies were performed on rats fed a normal or high K diet.

Results: Absolute quantification determined a positive correlation (Pearson =0.46 or 0.45, control or high K respectively, <0.0001) between the approximately 1000 proteins identified in uEVs and corresponding kidney tissue. Transmembrane proteins had greater positive correlations relative to cytoplasmic proteins. Proteins with high correlations (>0.9), included exosome markers Tsg101 and Alix. Relative quantification highlighted a monotonic relationship between altered transporter/channel abundances in uEVs and the kidney after dietary K manipulation. Analysis of genetic mouse models also revealed correlations between uEVs and kidney.

Conclusion: This large-scale unbiased analysis identifies uEV proteins that track the abundance of the parent proteins in the kidney. The data form a novel resource for the kidney community and support the reliability of using uEV protein changes to monitor specific physiologic responses and disease mechanisms.
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http://dx.doi.org/10.1681/ASN.2020071035DOI Listing
September 2021

Agmatine requires GluN2B-containing NMDA receptors to inhibit the development of neuropathic pain.

Mol Pain 2021 Jan-Dec;17:17448069211029171

Department of Neuroscience, University of Minnesota, Minneapolis, MN, USA.

A decarboxylated form of L-arginine, agmatine, preferentially antagonizes NMDArs containing Glun2B subunits within the spinal cord and lacks motor side effects commonly associated with non-subunit-selective NMDAr antagonism, namely sedation and motor impairment. Spinally delivered agmatine has been previously shown to reduce the development of tactile hypersensitivity arising from spinal nerve ligation. The present study interrogated the dependence of agmatine's alleviation of neuropathic pain (spared nerve injury (SNI) model) on GluN2B-containing NMDArs. SNI-induced hypersensitivity was induced in mice with significant reduction of levels of spinal GluN2B subunit of the NMDAr and their floxed controls. Agmatine reduced development of SNI-induced tactile hypersensitivity in controls but had no effect in subjects with reduced levels of GluN2B subunits. Ifenprodil, a known GluN2B-subunit-selective antagonist, similarly reduced tactile hypersensitivity in controls but not in the GluN2B-deficient mice. In contrast, MK-801, an NMDA receptor channel blocker, reduced hypersensitivity in both control and GluN2B-deficient mice, consistent with a pharmacological pattern expected from a NMDAr antagonist that does not have preference for GluN2B subtypes. Additionally, we observed that spinally delivered agmatine, ifenprodil and MK-801 inhibited nociceptive behaviors following intrathecal delivery of NMDA in control mice. By contrast, in GluN2B-deficient mice, MK-801 reduced NMDA-evoked nociceptive behaviors, but agmatine had a blunted effect and ifenprodil had no effect. These results demonstrate that agmatine requires the GluN2B subunit of the NMDA receptor for inhibitory pharmacological actions in pre-clinical models of NMDA receptor-dependent hypersensitivity.
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http://dx.doi.org/10.1177/17448069211029171DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8255568PMC
July 2021

Protein kinase D1 variant associated with human epilepsy and peripheral nerve hypermyelination.

Clin Genet 2021 08 2;100(2):176-186. Epub 2021 Jun 2.

Department of Anesthesiology, Vanderbilt University School of Medicine, Nashville, Tennessee, USA.

We report the case of a patient with severe progressive epilepsy and peripheral neuropathy and a novel de novo inactivating variant (p.E79X) in Protein Kinase D1 (PKD1). Using CRISPR/Cas9, we engineered the homologous variant in mice and showed that in the homozygote mouse, it recapitulated the patient peripheral nerve hypermyelination pathology. The lethality of the homozygote mouse prevented us from performing an assessment of locomotor behavior. The mutant heterozygote mouse; however, exhibited a significant increase in kainate-induced seizure activity over wild-type mice, supporting the hypothesis that the PKD1 variant is a candidate for the cause of the patient epilepsy. Because PKD1 was previously identified in a kinomic screen as an interacting partner of the K-Cl cotransporter 3 (KCC3), and since KCC3 is involved in peripheral nerve disease and brain hyperexcitability, one possible mechanism of action of PKD1 in disease is through KCC3. We show that catalytically inactive PKD1 stimulates KCC3 activity, consistent with tonic relief of inhibitory phosphorylation. Our findings implicate a novel role for PKD1 in the human nervous system, and uncover a mechanism that could serve as a potential target to promote nervous system myelination.
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http://dx.doi.org/10.1111/cge.13973DOI Listing
August 2021

Collagen IV dysfunction in glomerular basement membrane diseases. I. Discovery of a COL4A3 variant in familial Goodpasture's and Alport diseases.

J Biol Chem 2021 Jan-Jun;296:100590. Epub 2021 Mar 26.

Division of Nephrology and Hypertension, Department of Medicine, Vanderbilt University Medical Center, Nashville, Tennessee, USA; Center for Matrix Biology, Vanderbilt University Medical Center, Nashville, Tennessee, USA; Aspirnaut Program, Vanderbilt University Medical Center, Nashville, Tennessee, USA; Department of Pathology, Microbiology and Immunology, Vanderbilt University Medical Center, Nashville, Tennessee, USA; Department of Biochemistry, Vanderbilt University, Nashville, Tennessee, USA; Center for Structural Biology, Vanderbilt University, Nashville, Tennessee, USA; Department of Cell and Developmental Biology, Vanderbilt University, Nashville, Tennessee, USA; Vanderbilt-Ingram Cancer Center, Vanderbilt University, Nashville, Tennessee, USA. Electronic address:

Diseases of the glomerular basement membrane (GBM), such as Goodpasture's disease (GP) and Alport syndrome (AS), are a major cause of chronic kidney failure and an unmet medical need. Collagen IV is an important architectural element of the GBM that was discovered in previous research on GP and AS. How this collagen enables GBM to function as a permselective filter and how structural defects cause renal failure remain an enigma. We found a distinctive genetic variant of collagen IV in both a familial GP case and four AS kindreds that provided insights into these mechanisms. The variant is an 8-residue appendage at the C-terminus of the α3 subunit of the α345 hexamer. A knock-in mouse harboring the variant displayed GBM abnormalities and proteinuria. This pathology phenocopied AS, which pinpointed the α345 hexamer as a focal point in GBM function and dysfunction. Crystallography and assembly studies revealed underlying hexamer mechanisms, as described in Boudko et al. and Pedchenko et al. Bioactive sites on the hexamer surface were identified where pathogenic pathways of GP and AS converge and, potentially, that of diabetic nephropathy (DN). We conclude that the hexamer functions include signaling and organizing macromolecular complexes, which enable GBM assembly and function. Therapeutic modulation or replacement of α345 hexamer could therefore be a potential treatment for GBM diseases, and this knock-in mouse model is suitable for developing gene therapies.
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http://dx.doi.org/10.1016/j.jbc.2021.100590DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8100070PMC
August 2021

Sodium Transporters in Human Health and Disease.

Front Physiol 2020 25;11:588664. Epub 2021 Feb 25.

Department of Anesthesiology, School of Medicine, Vanderbilt University, Nashville, TN, United States.

Sodium (Na) electrochemical gradients established by Na/K ATPase activity drives the transport of ions, minerals, and sugars in both excitable and non-excitable cells. Na-dependent transporters can move these solutes in the same direction (cotransport) or in opposite directions (exchanger) across both the apical and basolateral plasma membranes of polarized epithelia. In addition to maintaining physiological homeostasis of these solutes, increases and decreases in sodium may also initiate, directly or indirectly, signaling cascades that regulate a variety of intracellular post-translational events. In this review, we will describe how the Na/K ATPase maintains a Na gradient utilized by multiple sodium-dependent transport mechanisms to regulate glucose uptake, excitatory neurotransmitters, calcium signaling, acid-base balance, salt-wasting disorders, fluid volume, and magnesium transport. We will discuss how several Na-dependent cotransporters and Na-dependent exchangers have significant roles in human health and disease. Finally, we will discuss how each of these Na-dependent transport mechanisms have either been shown or have the potential to use Na in a secondary role as a signaling molecule.
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http://dx.doi.org/10.3389/fphys.2020.588664DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7947867PMC
February 2021

Temporal manipulation of KCC3 expression in juvenile or adult mice suggests irreversible developmental deficit in hereditary motor sensory neuropathy with agenesis of the corpus callosum.

Am J Physiol Cell Physiol 2021 05 17;320(5):C722-C730. Epub 2021 Feb 17.

Department of Anesthesiology, Vanderbilt University School of Medicine, Nashville, Tennessee.

Hereditary motor sensory neuropathy (HMSN/ACC) with agenesis of the corpus callosum (ACC) has been documented in the French-derived populations of Charlevoix and Saguenay/Lac St. Jean in Quebec, Canada, as well as a few sporadic families throughout the world. HMSN/ACC occurs because of loss-of-function mutations in the potassium-chloride cotransporter 3 (KCC3). In HMSN/ACC, motor deficits occur early in infancy with rapid and continual deterioration of motor and sensory fibers into juvenile and adulthood. Genetic work in mice has demonstrated that the disease is caused by loss of KCC3 function in neurons and particularly parvalbumin (PV)-expressing neurons. Currently, there are no treatments or cures for HMSN/ACC other than pain management. As genetic counseling in Quebec has increased as a preventative strategy, most individuals with HSMN/ACC are now adults. The onset of the disease is unknown. In particular, it is unknown if the disease starts early during development and whether it can be reversed by restoring KCC3 function. In this study, we used two separate mouse models that when combined to the PV-Cre tamoxifen-inducible system allowed us to ) disrupt KCC3 expression in adulthood or juvenile periods; and ) reintroduce KCC3 expression in mice that first develop with a nonfunctional cotransporter. We show that disrupting or reintroducing KCC3 in the adult mouse has no effect on locomotor behavior, indicating that expression of KCC3 is critical during embryonic development and/or the perinatal period and that once the disease has started, reexpressing a functional cotransporter fails to change the course of HMSN/ACC.
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http://dx.doi.org/10.1152/ajpcell.00594.2020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8163575PMC
May 2021

Phenobarbital, midazolam, bumetanide, and neonatal seizures: The devil is in the details.

Epilepsia 2021 04 3;62(4):935-940. Epub 2021 Feb 3.

Department of Anesthesiology, Vanderbilt University Medical School, Nashville, Tennessee, USA.

Kaila, Löscher, and colleagues report that phenobarbital (PHB) and midazolam (MDZ) attenuate neonatal seizures following birth asphyxia, but the former only when applied before asphyxia and the latter before or after the triggering insult. In contrast, the NKCC1 chloride importer antagonist bumetanide (BUM) had no effect whether applied alone or with PHB. The observations are compelling and in accord with earlier studies. However, there are several general issues that deserve discussion. What is the clinical relevance of these data and the validity of animal models of encephalopathic seizures? Why is it that although they act on similar targets, these agents have different efficacy? Are both PHB and MDZ actions restricted to γ-aminobutyric acidergic (GABAergic) mechanisms? Why is BUM inefficient in attenuating seizures but capable of reducing the severity of other brain disorders? We suggest that the relative failure of antiepileptic drugs (AEDs) to treat this severe life-threatening condition is in part explicable by the recurrent seizures that shift the polarity of GABA, thereby counteracting their effects on their target. AEDs might be efficient after a few seizures but not recurrent ones. In addition, PHB and MDZ actions are not limited to GABA signals. BUM efficiently attenuates autism symptomatology notably in patients with tuberous sclerosis but does not reduce the recurrent seizures, illustrating the uniqueness of epilepsies. Therefore, the efficacy of AEDs to treat babies with encephalopathic seizures will depend on the history and severity of the seizures prior to their administration, challenging a universal common underlying mechanism.
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http://dx.doi.org/10.1111/epi.16830DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8035263PMC
April 2021

Locus coeruleus anchors a trisynaptic circuit controlling fear-induced suppression of feeding.

Neuron 2021 03 20;109(5):823-838.e6. Epub 2021 Jan 20.

Department of Physiology, Feinberg School of Medicine, Northwestern University, Chicago, IL, USA. Electronic address:

The circuit mechanisms underlying fear-induced suppression of feeding are poorly understood. To help fill this gap, mice were fear conditioned, and the resulting changes in synaptic connectivity among the locus coeruleus (LC), the parabrachial nucleus (PBN), and the central nucleus of amygdala (CeA)-all of which are implicated in fear and feeding-were studied. LC neurons co-released noradrenaline and glutamate to excite PBN neurons and suppress feeding. LC neurons also suppressed inhibitory input to PBN neurons by inducing heterosynaptic, endocannabinoid-dependent, long-term depression of CeA synapses. Blocking or knocking down endocannabinoid receptors in CeA neurons prevented fear-induced depression of CeA synaptic transmission and fear-induced suppression of feeding. Altogether, these studies demonstrate that LC neurons play a pivotal role in modulating the circuitry that underlies fear-induced suppression of feeding, pointing to new ways of alleviating stress-induced eating disorders.
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http://dx.doi.org/10.1016/j.neuron.2020.12.023DOI Listing
March 2021

Cannabinoid type 1 receptors in A2a neurons contribute to cocaine-environment association.

Psychopharmacology (Berl) 2021 Apr 16;238(4):1121-1131. Epub 2021 Jan 16.

Vanderbilt Brain Institute, Vanderbilt University, Nashville, TN, 37232, USA.

Rationale: Cannabinoid type 1 receptors (CB1Rs) are widely expressed within the brain's reward circuits and are implicated in regulating drug induced behavioral adaptations. Understanding how CB1R signaling in discrete circuits and cell types contributes to drug-related behavior provides further insight into the pathology of substance use disorders.

Objective And Methods: We sought to determine how cell type-specific expression of CB1Rs within striatal circuits contributes to cocaine-induced behavioral plasticity, hypothesizing that CB1R function in distinct striatal neuron populations would differentially impact behavioral outcomes. We crossed conditional Cnr1 mice and striatal output pathway cre lines (Drd1a -cre; D1, Adora2a -cre; A2a) to generate cell type-specific CB1R knockout mice and assessed their performance in cocaine locomotor and associative behavioral assays.

Results: Both knockout lines retained typical locomotor activity at baseline. D1-Cre x Cnr1 mice did not display hyperlocomotion in response to acute cocaine dosing, and both knockout lines exhibited blunted locomotor activity across repeated cocaine doses. A2a-cre Cnr1, mice did not express a preference for cocaine paired environments in a two-choice place preference task.

Conclusions: This study aids in mapping CB1R-dependent cocaine-induced behavioral adaptations onto distinct striatal neuron subtypes. A reduction of cocaine-induced locomotor activation in the D1- and A2a-Cnr1 knockout mice supports a role for CB1R function in the motor circuit. Furthermore, a lack of preference for cocaine-associated context in A2a-Cnr1 mice suggests that CB1Rs on A2a-neuron inhibitory terminals are necessary for either reward perception, memory consolidation, or recall. These results direct future investigations into CB1R-dependent adaptations underlying the development and persistence of substance use disorders.
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http://dx.doi.org/10.1007/s00213-021-05759-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8386588PMC
April 2021

Advances in the development of novel compounds targeting cation-chloride cotransporter physiology.

Authors:
Eric Delpire

Am J Physiol Cell Physiol 2021 03 23;320(3):C324-C340. Epub 2020 Dec 23.

Department of Anesthesiology, Vanderbilt University School of Medicine, Nashville, Tennessee.

For about half a century, the pharmacology of electroneutral cation-chloride cotransporters has been dominated by a few drugs that are widely used in clinical medicine. Because these diuretic drugs are so good at what they do, there has been little incentive in expanding their pharmacology. The increasing realization that cation-chloride cotransporters are involved in many other key physiological processes and the knowledge that different tissues express homologous proteins with matching transport functions have rekindled interest in drug discovery. This review summarizes the methods available to assess the function of these transporters and describe the multiple efforts that have made to identify new compounds. We describe multiple screens targeting KCC2 function and one screen designed to find compounds that discriminate between NKCC1 and NKCC2. Two of the KCC2 screens identified new inhibitors that are 3-4 orders of magnitude more potent than furosemide. Additional screens identified compounds that purportedly increase cell surface expression of the cotransporter, as well as several FDA-approved drugs that increase KCC2 transcription and expression. The technical details of each screen biased them toward specific processes in the life cycle of the transporter, making these efforts independent and complementary. In addition, each drug discovery effort contributes to our understanding of the biology of the cotransporters.
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http://dx.doi.org/10.1152/ajpcell.00566.2020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8294628PMC
March 2021

NKCC1: Newly Found as a Human Disease-Causing Ion Transporter.

Function (Oxf) 2021 3;2(1):zqaa028. Epub 2020 Nov 3.

Department of Anesthesiology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA.

Among the electroneutral Na-dependent chloride transporters, NKCC1 had until now evaded identification as a protein causing human diseases. The closely related transporters, NKCC2 and NCC have been identified some 25 years ago as responsible for Bartter and Gitelman syndromes: two renal-dependent salt wasting disorders. Absence of disease was most surprising since the NKCC1 knockout mouse was shown in 1999 to be viable, albeit with a wide range of deleterious phenotypes. Here we summarize the work of the past 5 years that introduced us to clinical cases involving NKCC1. The most striking cases are of 3 children with inherited mutations, who have complete absence of NKCC1 expression. These cases establish that lack of NKCC1 causes deafness; CFTR-like secretory defects with mucus accumulation in lung and intestine; severe xerostomia, hypotonia, dysmorphic facial features, and severe neurodevelopmental disorder. Another intriguing case is of a patient with a dominant deleterious allele. This mutation introduced a premature stop codon leading to a truncated protein. This mutant transporter seems to exert dominant-negative effect on wild-type transporter only in epithelial cells. The patient who suffers from lung, bladder, intestine, pancreas, and multiple endocrine abnormalities has, however, normal hearing and cognition. Finally, new reports substantiate the haploinsufficiency prediction of the gene. Cases with single allele mutations in have been linked to hearing loss and neurodevelopmental disorders.
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http://dx.doi.org/10.1093/function/zqaa028DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7727275PMC
November 2020

Mitochondria: an elegant means of partitioning and harnessing energy.

Am J Physiol Cell Physiol 2021 04 16;320(4):C463-C464. Epub 2020 Dec 16.

Department of Anesthesiology, Vanderbilt University School of Medicine, Nashville, Tennessee.

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http://dx.doi.org/10.1152/ajpcell.00592.2020DOI Listing
April 2021

Structures and an activation mechanism of human potassium-chloride cotransporters.

Sci Adv 2020 Dec 11;6(50). Epub 2020 Dec 11.

Department of Biophysics, and Department of Pathology of Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou 310058, China.

Potassium-chloride cotransporters KCC1 to KCC4 mediate the coupled export of potassium and chloride across the plasma membrane and play important roles in cell volume regulation, auditory system function, and γ-aminobutyric acid (GABA) and glycine-mediated inhibitory neurotransmission. Here, we present 2.9- to 3.6-Å resolution structures of full-length human KCC2, KCC3, and KCC4. All three KCCs adopt a similar overall architecture, a domain-swap dimeric assembly, and an inward-facing conformation. The structural and functional studies reveal that one unexpected N-terminal peptide binds at the cytosolic facing cavity and locks KCC2 and KCC4 at an autoinhibition state. The C-terminal domain (CTD) directly interacts with the N-terminal inhibitory peptide, and the relative motions between the CTD and the transmembrane domain (TMD) suggest that CTD regulates KCCs' activities by adjusting the autoinhibitory effect. These structures provide the first glimpse of full-length structures of KCCs and an autoinhibition mechanism among the amino acid-polyamine-organocation transporter superfamily.
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http://dx.doi.org/10.1126/sciadv.abc5883DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7732191PMC
December 2020

The Kainate Receptor Subunit GluK2 Interacts With KCC2 to Promote Maturation of Dendritic Spines.

Front Cell Neurosci 2020 26;14:252. Epub 2020 Aug 26.

HiLIFE Neuroscience Center, University of Helsinki, Helsinki, Finland.

Kainate receptors (KAR) play a crucial role in the plasticity and functional maturation of glutamatergic synapses. However, how they regulate structural plasticity of dendritic spines is not known. The GluK2 subunit was recently shown to coexist in a functional complex with the neuronal K-Cl cotransporter KCC2. Apart from having a crucial role in the maturation of GABAergic transmission, KCC2 has a morphogenic role in the maturation of dendritic spines. Here, we show that local inactivation of GluK2 expression in CA3 hippocampal neurons induces altered morphology of dendritic spines and reduction in mEPSC frequency. GluK2 deficiency also resulted in a strong change in the subcellular distribution of KCC2 as well as a smaller somatodendritic gradient in the reversal potential of GABA. Strikingly, the aberrant morphology of dendritic spines in GluK2-deficient CA3 pyramidal neurons was restored by overexpression of KCC2. GluK2 silencing in hippocampal neurons significantly reduced the expression of 4.1N and functional form of the actin filament severing protein cofilin. Consistently, assessment of actin dynamics using fluorescence recovery after photobleaching (FRAP) of β-actin showed a significant increase in the stability of F-actin filaments in dendritic spines. In conclusion, our results demonstrate that GluK2-KCC2 interaction plays an important role in the structural maturation of dendritic spines. This also provides novel insights into the connection between KAR dysfunction, structural plasticity, and developmental disorders.
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http://dx.doi.org/10.3389/fncel.2020.00252DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7479265PMC
August 2020

Stimulatory Role of SPAK Signaling in the Regulation of Large Conductance Ca-Activated Potassium (BK) Channel Protein Expression in Kidney.

Front Physiol 2020 2;11:638. Epub 2020 Jul 2.

Renal Division, Department of Medicine, Emory University School of Medicine, Atlanta, GA, United States.

SPS1-related proline/alanine-rich kinase (SPAK) plays important roles in regulating the function of numerous ion channels and transporters. With-no-lysine (WNK) kinase phosphorylates SPAK kinase to active the SPAK signaling pathway. Our previous studies indicated that WNK kinases regulate the activity of the large-conductance Ca-activated K (BK) channel and its protein expression via the ERK1/2 signaling pathway. It remains largely unknown whether SPAK kinase directly modulates the BK protein expression in kidney. In this study, we investigated the effect of SPAK on renal BK protein expression in both HEK293 cells and mouse kidney. In HEK293 cells, siRNA-mediated knockdown of SPAK expression significantly reduced BK protein expression and increased ERK1/2 phosphorylation, whereas overexpression of SPAK significantly enhanced BK expression and decreased ERK1/2 phosphorylation in a dose-dependent manner. Knockdown of ERK1/2 prevented SPAK siRNA-mediated inhibition of BK expression. Similarly, pretreatment of HEK293 cells with either the lysosomal inhibitor bafilomycin A1 or the proteasomal inhibitor MG132 reversed the inhibitory effects of SPAK knockdown on BK expression. We also found that there is no BK channel activity in PCs of CCD in SPAK KO mice using the isolated split-open tubule single-cell patching. In addition, we found that BK protein abundance in the kidney of SPAK knockout mice was significantly decreased and ERK1/2 phosphorylation was significantly enhanced. A high-potassium diet significantly increased BK protein abundance and SPAK phosphorylation levels, while reducing ERK1/2 phosphorylation levels. These findings suggest that SPAK enhances BK protein expression by reducing ERK1/2 signaling-mediated lysosomal and proteasomal degradations of the BK channel.
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http://dx.doi.org/10.3389/fphys.2020.00638DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7343913PMC
July 2020

AMPA Receptor Auxiliary Subunit GSG1L Suppresses Short-Term Facilitation in Corticothalamic Synapses and Determines Seizure Susceptibility.

Cell Rep 2020 07;32(3):107921

Department of Molecular Physiology and Biophysics, Vanderbilt University, School of Medicine, Nashville, TN 37232, USA; Vanderbilt Brain Institute, Vanderbilt University, School of Medicine, Nashville, TN 37232, USA; Center for Structural Biology, Vanderbilt University, School of Medicine, Nashville, TN 37232, USA. Electronic address:

The anterior thalamus (AT) is critical for memory formation, processing navigational information, and seizure initiation. However, the molecular mechanisms that regulate synaptic function of AT neurons remain largely unexplored. We report that AMPA receptor auxiliary subunit GSG1L controls short-term plasticity in AT synapses that receive inputs from the cortex, but not in those receiving inputs from other pathways. A canonical auxiliary subunit stargazin co-exists in these neurons but is functionally absent from corticothalamic synapses. In GSG1L knockout mice, AT neurons exhibit hyperexcitability and the animals have increased susceptibility to seizures, consistent with a negative regulatory role of GSG1L. We hypothesize that negative regulation of synaptic function by GSG1L plays a critical role in maintaining optimal excitation in the AT.
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http://dx.doi.org/10.1016/j.celrep.2020.107921DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7425083PMC
July 2020

SLC12A2 variants cause a neurodevelopmental disorder or cochleovestibular defect.

Brain 2020 08;143(8):2380-2387

Department of Anesthesiology, Vanderbilt University School of Medicine, Nashville, TN, USA.

The SLC12 gene family consists of SLC12A1-SLC12A9, encoding electroneutral cation-coupled chloride co-transporters. SCL12A2 has been shown to play a role in corticogenesis and therefore represents a strong candidate neurodevelopmental disorder gene. Through trio exome sequencing we identified de novo mutations in SLC12A2 in six children with neurodevelopmental disorders. All had developmental delay or intellectual disability ranging from mild to severe. Two had sensorineural deafness. We also identified SLC12A2 variants in three individuals with non-syndromic bilateral sensorineural hearing loss and vestibular areflexia. The SLC12A2 de novo mutation rate was demonstrated to be significantly elevated in the deciphering developmental disorders cohort. All tested variants were shown to reduce co-transporter function in Xenopus laevis oocytes. Analysis of SLC12A2 expression in foetal brain at 16-18 weeks post-conception revealed high expression in radial glial cells, compatible with a role in neurogenesis. Gene co-expression analysis in cells robustly expressing SLC12A2 at 16-18 weeks post-conception identified a transcriptomic programme associated with active neurogenesis. We identify SLC12A2 de novo mutations as the cause of a novel neurodevelopmental disorder and bilateral non-syndromic sensorineural hearing loss and provide further data supporting a role for this gene in human neurodevelopment.
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http://dx.doi.org/10.1093/brain/awaa176DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7447514PMC
August 2020

Osmotic Response of Dorsal Root Ganglion Neurons Expressing Wild-Type and Mutant KCC3 Transporters.

Cell Physiol Biochem 2020 Jun;54(4):577-590

Department of Anesthesiology and Neuroscience Graduate Program, Vanderbilt University School of Medicine, Nashville, TN, USA,

Background/aims: Loss-of-Function (LOF) of the potassium chloride cotransporter 3 (KCC3) results in hereditary sensorimotor neuropathy with Agenesis of the Corpus Callosum (HSMN/ACC). Our KCC3 knockout mouse recapitulated axonal swelling and tissue vacuolization observed in autopsies of individuals with HSMN/ACC. We previously documented the first human case of a KCC3 gain-of-function (GOF) in which the patient also exhibited severe peripheral neuropathy. Furthermore, the GOF mouse model exhibited shrunken axons implicating the cotransporter in cell volume homeostasis. It is unclear how both KCC3 LOF and GOF lead to peripheral neuropathy. Thus, we sought to study differences in cell volume regulation of dorsal root ganglion neurons isolated from different mouse lines.

Methods: Using wide-field microscopy, we measured calcein fluorescence intensity through pinhole measurements at the center of cells and compared cell swelling and cell volume regulation/recovery of wild-type, LOF, and GOF dorsal root ganglia neurons, as well as wild-type neurons treated with a KCC-specific inhibitor.

Results: In contrast to control neurons that swell and volume regulate under a hypotonic challenge, neurons lacking KCC3 swell but fail to volume regulate. Similar data were observed in wild-type neurons treated with the KCC inhibitor. We also show that sensory neurons expressing a constitutively active KCC3 exhibited a blunted swelling phase compared to wild-type neurons, questioning the purely osmotic nature of the swelling phase.

Conclusion: These findings demonstrate the integral role of KCC3 in cell volume homeostasis and support the idea that cell volume homeostasis is critical to the health of peripheral nerves.
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http://dx.doi.org/10.33594/000000241DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7393635PMC
June 2020

Endoplasmic reticulum retention and degradation of a mutation in SLC6A1 associated with epilepsy and autism.

Mol Brain 2020 05 12;13(1):76. Epub 2020 May 12.

Department of Neurology, Vanderbilt University Medical Center, Nashville, USA.

Mutations in SLC6A1, encoding γ-aminobutyric acid (GABA) transporter 1 (GAT-1), have been recently associated with a spectrum of epilepsy syndromes, intellectual disability and autism in clinic. However, the pathophysiology of the gene mutations is far from clear. Here we report a novel SLC6A1 missense mutation in a patient with epilepsy and autism spectrum disorder and characterized the molecular defects of the mutant GAT-1, from transporter protein trafficking to GABA uptake function in heterologous cells and neurons. The heterozygous missense mutation (c1081C to A (P361T)) in SLC6A1 was identified by exome sequencing. We have thoroughly characterized the molecular pathophysiology underlying the clinical phenotypes. We performed EEG recordings and autism diagnostic interview. The patient had neurodevelopmental delay, absence epilepsy, generalized epilepsy, and 2.5-3 Hz generalized spike and slow waves on EEG recordings. The impact of the mutation on GAT-1 function and trafficking was evaluated by H GABA uptake, structural simulation with machine learning tools, live cell confocal microscopy and protein expression in mouse neurons and nonneuronal cells. We demonstrated that the GAT-1(P361T) mutation destabilizes the global protein conformation and reduces total protein expression. The mutant transporter protein was localized intracellularly inside the endoplasmic reticulum (ER) with a pattern of expression very similar to the cells treated with tunicamycin, an ER stress inducer. Radioactive H-labeled GABA uptake assay indicated the mutation reduced the function of the mutant GAT-1(P361T), to a level that is similar to the cells treated with GAT-1 inhibitors. In summary, this mutation destabilizes the mutant transporter protein, which results in retention of the mutant protein inside cells and reduction of total transporter expression, likely via excessive endoplasmic reticulum associated degradation. This thus likely causes reduced functional transporter number on the cell surface, which then could cause the observed reduced GABA uptake function. Consequently, malfunctioning GABA signaling may cause altered neurodevelopment and neurotransmission, such as enhanced tonic inhibition and altered cell proliferation in vivo. The pathophysiology due to severely impaired GAT-1 function may give rise to a wide spectrum of neurodevelopmental phenotypes including autism and epilepsy.
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http://dx.doi.org/10.1186/s13041-020-00612-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7218610PMC
May 2020

Loss of non-canonical KCC2 functions promotes developmental apoptosis of cortical projection neurons.

EMBO Rep 2020 04 17;21(4):e48880. Epub 2020 Feb 17.

Molecular and Integrative Biosciences, Faculty of Biological and Environmental Sciences, University of Helsinki, Helsinki, Finland.

KCC2, encoded in humans by the SLC12A5 gene, is a multifunctional neuron-specific protein initially identified as the chloride (Cl ) extruder critical for hyperpolarizing GABA receptor currents. Independently of its canonical function as a K-Cl cotransporter, KCC2 regulates the actin cytoskeleton via molecular interactions mediated through its large intracellular C-terminal domain (CTD). Contrary to the common assumption that embryonic neocortical projection neurons express KCC2 at non-significant levels, here we show that loss of KCC2 enhances apoptosis of late-born upper-layer cortical projection neurons in the embryonic brain. In utero electroporation of plasmids encoding truncated, transport-dead KCC2 constructs retaining the CTD was as efficient as of that encoding full-length KCC2 in preventing elimination of migrating projection neurons upon conditional deletion of KCC2. This was in contrast to the effect of a full-length KCC2 construct bearing a CTD missense mutation (KCC2 ), which disrupts cytoskeletal interactions and has been found in patients with neurological and psychiatric disorders, notably seizures and epilepsy. Together, our findings indicate ion transport-independent, CTD-mediated regulation of developmental apoptosis by KCC2 in migrating cortical projection neurons.
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http://dx.doi.org/10.15252/embr.201948880DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7132201PMC
April 2020

A mutation in the Na-K-2Cl cotransporter-1 leads to changes in cellular metabolism.

J Cell Physiol 2020 10 10;235(10):7239-7250. Epub 2020 Feb 10.

Department of Anesthesiology, Vanderbilt University School of Medicine, Nashville, Tennessee.

The Na-K-Cl cotransporter-1 (NKCC1), by mediating the electroneutral transport of Na , K , and Cl plays an important role in cell volume regulation, epithelial transport, and the control of neuronal excitability. Recently, we reported the first known human mutation in SLC12A2, the gene encoding NKCC1. The 17-year old patient suffers from multiorgan failure. Laboratory tests conducted on muscle and liver biopsies of the patient showed abnormal increase in mitochondrial DNA copy number and increased glycogen levels, indicating the possibility that the transporter may play a role in energy metabolism. Here, we show that fibroblasts isolated from the patient demonstrate a significant increase in mitochondrial respiration, compared to fibroblasts isolated from healthy individuals. Similarly, Madin Darby canine kidney (MDCK) cells transfected with enhanced green fluorescent protein (EGFP)-tagged mutant NKCC1 DNA demonstrated increased mitochondrial respiration when compared to MDCK cells expressing EGFP-tagged wild-type (WT) cotransporter. Direct inhibition of the cotransporter through addition of bumetanide did not change the rate of basal respiration, but led to increased maximal mitochondrial respiration. Fibroblasts extracted from NKCC1 and NKCC1 mice also demonstrated a significant elevation in mitochondrial respiration, compared to fibroblasts isolated from their WT littermates. Expression of the mutant protein was associated with an increase in hydrogen peroxide and peroxidase activity and a decrease in messenger RNA transcript levels for protein involved in the unfolded protein response. These data reveal that cells expressing the mutant cotransporter demonstrate increased mitochondrial respiration and behave like they are experiencing a state of starvation.
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http://dx.doi.org/10.1002/jcp.29623DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7416433PMC
October 2020

Endocannabinoid Signaling Collapse Mediates Stress-Induced Amygdalo-Cortical Strengthening.

Neuron 2020 03 13;105(6):1062-1076.e6. Epub 2020 Jan 13.

Department of Psychiatry and Behavioral Sciences, Vanderbilt University Medical Center, Nashville, TN 37232, USA; The Vanderbilt Brain Institute, Vanderbilt University, Nashville, TN 37232, USA; Departments of Pharmacology and Molecular Physiology & Biophysics, Vanderbilt University School of Medicine, Nashville, TN 37232, USA; Vanderbilt Center for Addiction Research, Nashville, TN 37232, USA. Electronic address:

Functional coupling between the amygdala and the dorsomedial prefrontal cortex (dmPFC) has been implicated in the generation of negative affective states; however, the mechanisms by which stress increases amygdala-dmPFC synaptic strength and generates anxiety-like behaviors are not well understood. Here, we show that the mouse basolateral amygdala (BLA)-prelimbic prefrontal cortex (plPFC) circuit is engaged by stress and activation of this pathway in anxiogenic. Furthermore, we demonstrate that acute stress exposure leads to a lasting increase in synaptic strength within a reciprocal BLA-plPFC-BLA subcircuit. Importantly, we identify 2-arachidonoylglycerol (2-AG)-mediated endocannabinoid signaling as a key mechanism limiting glutamate release at BLA-plPFC synapses and the functional collapse of multimodal 2-AG signaling as a molecular mechanism leading to persistent circuit-specific synaptic strengthening and anxiety-like behaviors after stress exposure. These data suggest that circuit-specific impairment in 2-AG signaling could facilitate functional coupling between the BLA and plPFC and the translation of environmental stress to affective pathology.
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http://dx.doi.org/10.1016/j.neuron.2019.12.024DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7992313PMC
March 2020

Interpreting an apoptotic corpse as anti-inflammatory involves a chloride sensing pathway.

Nat Cell Biol 2019 12 2;21(12):1532-1543. Epub 2019 Dec 2.

The Center for Cell Clearance, University of Virginia, Charlottesville, VA, USA.

Apoptotic cell clearance (efferocytosis) elicits an anti-inflammatory response by phagocytes, but the mechanisms that underlie this response are still being defined. Here, we uncover a chloride-sensing signalling pathway that controls both the phagocyte 'appetite' and its anti-inflammatory response. Efferocytosis transcriptionally altered the genes that encode the solute carrier (SLC) proteins SLC12A2 and SLC12A4. Interfering with SLC12A2 expression or function resulted in a significant increase in apoptotic corpse uptake per phagocyte, whereas the loss of SLC12A4 inhibited corpse uptake. In SLC12A2-deficient phagocytes, the canonical anti-inflammatory program was replaced by pro-inflammatory and oxidative-stress-associated gene programs. This 'switch' to pro-inflammatory sensing of apoptotic cells resulted from the disruption of the chloride-sensing pathway (and not due to corpse overload or poor degradation), including the chloride-sensing kinases WNK1, OSR1 and SPAK-which function upstream of SLC12A2-had a similar effect on efferocytosis. Collectively, the WNK1-OSR1-SPAK-SLC12A2/SLC12A4 chloride-sensing pathway and chloride flux in phagocytes are key modifiers of the manner in which phagocytes interpret the engulfed apoptotic corpse.
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http://dx.doi.org/10.1038/s41556-019-0431-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7140761PMC
December 2019

Cryo-EM structures of NKCC1 and hKCC1: a new milestone in the physiology of cation-chloride cotransporters.

Am J Physiol Cell Physiol 2020 02 20;318(2):C225-C237. Epub 2019 Nov 20.

Department of Biophysics, Department of Pathology of Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou, China.

New milestones have been reached in the field of cation-Cl cotransporters with the recently released cryo-electron microscopy (EM) structures of the (zebrafish) Na-K-2Cl cotransporter (NKCC1) and the human K-Cl cotransporter (hKCC1). In this review we provide a brief timeline that identifies the multiple breakthroughs in the field of solute carrier 12 transporters that led to the structure resolution of two of its key members. While cation-Cl cotransporters share the overall architecture of carriers belonging to the amino acid-polyamine-organocation (APC) superfamily and some of their substrate binding sites, several new insights are gained from the two individual structures. A first major feature relates to the largest extracellular domain between transmembrane domain (TMD) 5 and TMD6 of KCC1, which stabilizes the dimer and forms a cap that likely participates in extracellular gating. A second feature is the conservation of the K and Cl binding sites in both structures and evidence of an unexpected second Cl coordination site in the KCC1 structure. Structural data are discussed in the context of previously published studies that examined the basic and kinetics properties of these cotransport mechanisms. A third characteristic is the evidence of an extracellular gate formed by conserved salt bridges between charged residues located toward the end of TMD3 and TMD4 in both transporters and the existence of an additional neighboring bridge in the hKCC1 structure. A fourth feature of these newly solved structures relates to the multiple points of contacts between the monomer forming the cotransporter homodimer units. These involve the TMDs, the COOH-terminal domains, and the large extracellular loop for hKCC1.
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http://dx.doi.org/10.1152/ajpcell.00465.2019DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7052613PMC
February 2020

GABA interneurons are the cellular trigger for ketamine's rapid antidepressant actions.

J Clin Invest 2020 03;130(3):1336-1349

Department of Psychiatry, Yale School of Medicine, New Haven, Connecticut, USA.

A single subanesthetic dose of ketamine, an NMDA receptor (NMDAR) antagonist, produces rapid and sustained antidepressant actions in depressed patients, addressing a major unmet need for the treatment of mood disorders. Ketamine produces a rapid increase in extracellular glutamate and synaptic formation in the prefrontal cortex, but the initial cellular trigger that initiates this increase and ketamine's behavioral actions has not been identified. To address this question, we used a combination of viral shRNA and conditional mutation to produce cell-specific knockdown or deletion of a key NMDAR subunit, GluN2B, implicated in the actions of ketamine. The results demonstrated that the antidepressant actions of ketamine were blocked by GluN2B-NMDAR knockdown on GABA (Gad1) interneurons, as well as subtypes expressing somatostatin (Sst) or parvalbumin (Pvalb), but not glutamate principle neurons in the medial prefrontal cortex (mPFC). Further analysis of GABA subtypes showed that cell-specific knockdown or deletion of GluN2B in Sst interneurons blocked or occluded the antidepressant actions of ketamine and revealed sex-specific differences that are associated with excitatory postsynaptic currents on mPFC principle neurons. These findings demonstrate that GluN2B-NMDARs on GABA interneurons are the initial cellular trigger for the rapid antidepressant actions of ketamine and show sex-specific adaptive mechanisms to GluN2B modulation.
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http://dx.doi.org/10.1172/JCI130808DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7269589PMC
March 2020

Novel Human NKCC1 Mutations Cause Defects in Goblet Cell Mucus Secretion and Chronic Inflammation.

Cell Mol Gastroenterol Hepatol 2020 23;9(2):239-255. Epub 2019 Oct 23.

Department of Anesthesiology, Vanderbilt University School of Medicine, Nashville, Tennessee. Electronic address:

Background & Aims: Infections resulting from intestinal yeast and bacteria affect a large number of patients with deficits in absorptive or secretory epithelial transport mechanisms. The basolateral Na-K-2Cl cotransporter (NKCC1) has been implicated in intestinal epithelial fluid secretion. Two patients with deleterious heterozygous (NKCC1-DFX, DFX for Asp-Phe-stop codon) or homozygous (Kilquist) mutations in SLC12A2 (NKCC1) suffered from gastrointestinal deficits. Because of chronic infections, the colon and the small intestine of the NKCC1-DFX patient were resected surgically.

Methods: To investigate how NKCC1 affects the integrity and function of the gut epithelia, we used a mouse model recapitulating the NKCC1-DFX patient mutation. Electron microscopy and immunostaining were used to analyze the integrity of the colonic mucus layers and immune cell infiltration. Fluorescence in situ hybridization was performed on the distal colon sections to measure bacteria translocation to the mucosa and submucosa. Citrobacter rodentium was used to measure mouse ability to clear enteric infection. A multiplex cytokine assay was used to analyze mouse inflammatory response to infection.

Results: We show that NKCC1-DFX expression causes defective goblet cell mucus granule exocytosis, leading to secretion of intact granules into the lumen of the large intestine. In addition, NKCC1-DFX colon submucosal glands secrete mucus that remained attached to the epithelium. Importantly, expression of the mutant NKCC1 or complete loss of NKCC1 function leads to aggravated inflammatory response to C rodentium infection. Compared with wild-type, NKCC1-DFX mice showed decreased expression of claudin-2, a tight junction protein involved in paracellular Na and water transport and enteric infection clearance.

Conclusions: Our data indicate that NKCC1-DFX impairs gut barrier function by affecting mucus secretion and immune properties.
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http://dx.doi.org/10.1016/j.jcmgh.2019.10.006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6957845PMC
May 2021

Cryo-EM structures of the human cation-chloride cotransporter KCC1.

Science 2019 10;366(6464):505-508

Department of Biophysics, Department of Pathology of Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou 310058, China.

Cation-chloride cotransporters (CCCs) mediate the coupled, electroneutral symport of cations with chloride across the plasma membrane and are vital for cell volume regulation, salt reabsorption in the kidney, and γ-aminobutyric acid (GABA)-mediated modulation in neurons. Here we present cryo-electron microscopy (cryo-EM) structures of human potassium-chloride cotransporter KCC1 in potassium chloride or sodium chloride at 2.9- to 3.5-angstrom resolution. KCC1 exists as a dimer, with both extracellular and transmembrane domains involved in dimerization. The structural and functional analyses, along with computational studies, reveal one potassium site and two chloride sites in KCC1, which are all required for the ion transport activity. KCC1 adopts an inward-facing conformation, with the extracellular gate occluded. The KCC1 structures allow us to model a potential ion transport mechanism in KCCs and provide a blueprint for drug design.
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http://dx.doi.org/10.1126/science.aay3129DOI Listing
October 2019

Developmentally regulated KCC2 phosphorylation is essential for dynamic GABA-mediated inhibition and survival.

Sci Signal 2019 10 15;12(603). Epub 2019 Oct 15.

Departments of Neurosurgery, Pediatrics, and Cellular and Molecular Physiology, Centers for Mendelian Genomics, Yale School of Medicine, New Haven, CT 06510, USA.

Despite its importance for γ-aminobutyric acid (GABA) inhibition and involvement in neurodevelopmental disease, the regulatory mechanisms of the K/Cl cotransporter KCC2 (encoded by ) during maturation of the central nervous system (CNS) are not entirely understood. Here, we applied quantitative phosphoproteomics to systematically map sites of KCC2 phosphorylation during CNS development in the mouse. KCC2 phosphorylation at Thr and Thr, which inhibits KCC2 activity, underwent dephosphorylation in parallel with the GABA excitatory-inhibitory sequence in vivo. Knockin mice expressing the homozygous phosphomimetic KCC2 mutations T906E/T1007E ( ), which prevented the normal developmentally regulated dephosphorylation of these sites, exhibited early postnatal death from respiratory arrest and a marked absence of cervical spinal neuron respiratory discharges. mice also displayed disrupted lumbar spinal neuron locomotor rhythmogenesis and touch-evoked status epilepticus associated with markedly impaired KCC2-dependent Cl extrusion. These data identify a previously unknown phosphorylation-dependent KCC2 regulatory mechanism during CNS development that is essential for dynamic GABA-mediated inhibition and survival.
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http://dx.doi.org/10.1126/scisignal.aaw9315DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7219477PMC
October 2019

variants in cause sporadic early-onset progressive sensorimotor neuropathy.

J Med Genet 2020 04 22;57(4):283-288. Epub 2019 Aug 22.

Institute of Medical Genetics and Applied Genomics, University of Tübingen, Tübingen, Germany

Background: Charcot-Marie-Tooth disease (CMT) is a clinically and genetically heterogeneous disorder of the peripheral nervous system. Biallelic variants in have been associated with autosomal-recessive hereditary motor and sensory neuropathy with agenesis of the corpus callosum (HMSN/ACC). We identified heterozygous de novo variants in in three unrelated patients with intermediate CMT.

Methods: We evaluated the clinical reports and electrophysiological data of three patients carrying de novo variants in identified by diagnostic trio exome sequencing. For functional characterisation of the identified variants, potassium influx of mutated KCC3 cotransporters was measured in oocytes.

Results: We identified two different de novo missense changes (p.Arg207His and p.Tyr679Cys) in in three unrelated individuals with early-onset progressive CMT. All presented with axonal/demyelinating sensorimotor neuropathy accompanied by spasticity in one patient. Cognition and brain MRI were normal. Modelling of the mutant KCC3 cotransporter in oocytes showed a significant reduction in potassium influx for both changes.

Conclusion: Our findings expand the genotypic and phenotypic spectrum associated with variants from autosomal-recessive HMSN/ACC to dominant-acting de novo variants causing a milder clinical presentation with early-onset neuropathy.
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http://dx.doi.org/10.1136/jmedgenet-2019-106273DOI Listing
April 2020
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