A noninvasive optical approach for assessing chloride extrusion activity of the K-Cl cotransporter KCC2 in neuronal cells

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http://hdl.handle.net/10138/176706

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Ludwig , A , Rivera Baeza , C & Uvarov , P 2017 , ' A noninvasive optical approach for assessing chloride extrusion activity of the K-Cl cotransporter KCC2 in neuronal cells ' , BMC Neuroscience , vol. 18 , 23 . https://doi.org/10.1186/s12868-017-0336-5

Title: A noninvasive optical approach for assessing chloride extrusion activity of the K-Cl cotransporter KCC2 in neuronal cells
Author: Ludwig, Anastasia; Rivera Baeza, Claudio; Uvarov, Pavel
Contributor: University of Helsinki, Neuroscience Center
University of Helsinki, Helsinki In Vivo Animal Imaging Platform (HAIP)
University of Helsinki, Medicum
Date: 2017-01-31
Language: eng
Number of pages: 15
Belongs to series: BMC Neuroscience
ISSN: 1471-2202
URI: http://hdl.handle.net/10138/176706
Abstract: Background: Cation-chloride cotransporters (CCCs) are indispensable for maintaining chloride homeostasis in multiple cell types, but K-Cl cotransporter KCC2 is the only CCC member with an exclusively neuronal expression in mammals. KCC2 is critical for rendering fast hyperpolarizing responses of ionotropic.-aminobutyric acid and glycine receptors in adult neurons, for neuronal migration in the developing central nervous system, and for the formation and maintenance of small dendritic protrusions-dendritic spines. Deficit in KCC2 expression and/or activity is associated with epilepsy and neuropathic pain, and effective strategies are required to search for novel drugs augmenting KCC2 function. Results: We revised current methods to develop a noninvasive optical approach for assessing KCC2 transport activity using a previously characterized genetically encoded chloride sensor. Our protocol directly assesses dynamics of KCC2-mediated chloride efflux and allows measuring genuine KCC2 activity with good spatial and temporal resolution. As a proof of concept, we used this approach to compare transport activities of the two known KCC2 splice isoforms, KCC2a and KCC2b, in mouse neuronal Neuro-2a cells. Conclusions: Our noninvasive optical protocol proved to be efficient for assessment of furosemide-sensitive chloride fluxes. Transport activities of the N-terminal splice isoforms KCC2a and KCC2b obtained by the novel approach matched to those reported previously using standard methods for measuring chloride fluxes.
Subject: Genetically encoded chloride sensor
Slc12a5 gene
KCC2
Inhibition
GABA
GREEN FLUORESCENT PROTEIN
GENETICALLY-ENCODED CHLORIDE
THROUGHPUT FUNCTIONAL ASSAY
PRESYNAPTIC NERVE-TERMINALS
INTRACELLULAR CHLORIDE
HIPPOCAMPAL-NEURONS
RAT-BRAIN
SYNAPTIC INHIBITION
POTASSIUM CHANNELS
NEUROPATHIC PAIN
3112 Neurosciences
3124 Neurology and psychiatry
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