scholarly journals Development and validation of a potent and specific inhibitor for the CLC-2 chloride channel

2020 ◽  
Vol 117 (51) ◽  
pp. 32711-32721
Author(s):  
Anna K. Koster ◽  
Austin L. Reese ◽  
Yuri Kuryshev ◽  
Xianlan Wen ◽  
Keri A. McKiernan ◽  
...  
Keyword(s):  
Central Nervous System ◽  
Nervous System ◽  
Chloride Channel ◽  
Pyramidal Neurons ◽  
Brain Slices ◽  
Specific Inhibitor ◽  
Kinetic Studies ◽  
Computational Docking ◽  
Hippocampal Pyramidal Neurons ◽  
The Central Nervous System

CLC-2 is a voltage-gated chloride channel that is widely expressed in mammalian tissues. In the central nervous system, CLC-2 appears in neurons and glia. Studies to define how this channel contributes to normal and pathophysiological function in the central nervous system raise questions that remain unresolved, in part due to the absence of precise pharmacological tools for modulating CLC-2 activity. Herein, we describe the development and optimization of AK-42, a specific small-molecule inhibitor of CLC-2 with nanomolar potency (IC50= 17 ± 1 nM). AK-42 displays unprecedented selectivity (>1,000-fold) over CLC-1, the closest CLC-2 homolog, and exhibits no off-target engagement against a panel of 61 common channels, receptors, and transporters expressed in brain tissue. Computational docking, validated by mutagenesis and kinetic studies, indicates that AK-42 binds to an extracellular vestibule above the channel pore. In electrophysiological recordings of mouse CA1 hippocampal pyramidal neurons, AK-42 acutely and reversibly inhibits CLC-2 currents; no effect on current is observed on brain slices taken from CLC-2 knockout mice. These results establish AK-42 as a powerful tool for investigating CLC-2 neurophysiology.

scholarly journals Development and validation of a potent and specific inhibitor for the CLC-2 chloride channel

2020 ◽  
Author(s):  
Anna K. Koster ◽  
Austin L. Reese ◽  
Yuri Kuryshev ◽  
Xianlan Wen ◽  
Keri A. McKiernan ◽  
...  
Keyword(s):  
Central Nervous System ◽  
Nervous System ◽  
Small Molecule ◽  
Chloride Channel ◽  
Brain Slices ◽  
Specific Inhibitor ◽  
Small Molecule Inhibitor ◽  
Causative Role ◽  
Molecule Inhibitor ◽  
The Central Nervous System

AbstractCLC-2 is a voltage-gated chloride channel that is widely expressed in many mammalian tissues. In the central nervous system (CNS), CLC-2 is expressed in neurons and glia. Studies to define how this channel contributes to normal and pathophysiological function in the CNS have been controversial, in part due to the absence of precise pharmacological tools for modulating CLC-2 activity. Herein, we describe the development and optimization of AK-42, a specific small-molecule inhibitor of CLC-2 with nanomolar potency (IC50 = 17 ± 1 nM). AK-42 displays unprecedented selectivity (>1000-fold) over CLC-1, the closest CLC-2 homolog, and exhibits no off-target engagement against a panel of 58 common channels, receptors, and transporters expressed in brain tissue. Computational docking, validated by mutagenesis and kinetic studies, indicates that AK-42 binds to an extracellular vestibule above the channel pore. In electrophysiological recordings of mouse CA1 hippocampal pyramidal neurons, AK-42 acutely and reversibly inhibits CLC-2 currents; no effect on current is observed on brain slices taken from CLC-2 knockout mice. These results establish AK-42 as a powerful new tool for investigating CLC-2 neurophysiology.Significance StatementThe CLC-2 ion channel facilitates selective passage of Cl− ions across cell membranes. In the central nervous system (CNS), CLC-2 is expressed in both neurons and glia and is proposed to regulate electrical excitability and ion homeostasis. CLC-2 has been implicated in various CNS disorders, including certain types of epilepsy and leukodystrophy. Establishing a causative role for CLC-2 in neuropathologies, however, has been limited by the absence of selective reagents that enable acute and specific channel modulation. Our studies have resulted in the identification of a highly potent, small-molecule inhibitor that enables specific block of CLC-2 Cl− currents in hippocampal brain slices. This precise molecular tool should enable future efforts to identify and treat CLC-2-related disease.

scholarly journals The antihelminthic moxidectin enhances tonic GABA currents in rodent hippocampal pyramidal neurons

2018 ◽  
Vol 119 (5) ◽  
pp. 1693-1698
Author(s):  
Jay Spampanato ◽  
Anne Gibson ◽  
F. Edward Dudek
Keyword(s):  
Central Nervous System ◽  
Nervous System ◽  
Gaba Receptors ◽  
Pyramidal Neurons ◽  
Ex Vivo ◽  
Companion Animals ◽  
Brain Slices ◽  
Dependent Manner ◽  
Hippocampal Pyramidal Neurons ◽  
Mammalian Central Nervous System

Macrocyclic lactones (MLs) are commonly used treatments for parasitic worm and insect infections in humans, livestock, and companion animals. MLs target the invertebrate glutamate-activated chloride channel that is not present in vertebrates. MLs are not entirely inert in vertebrates, though; they have been reported to have activity in heterologous expression systems consisting of ligand-gated ion channels that are present in the mammalian central nervous system (CNS). However, these compounds are typically not able to reach significant concentrations in the CNS because of the activity of the blood-brain barrier P-glycoprotein extrusion system. Despite this, these compounds are able to reach low levels in the CNS that may be useful in the design of novel “designer” ligand-receptor systems that can be used to directly investigate neuronal control of behavior in mammals and have potential for use in treating human neurological diseases. To determine whether MLs might affect neurons in intact brains, we investigated the activity of the ML moxidectin (MOX) at native GABA receptors. Specifically, we recorded tonic and phasic miniature inhibitory postsynaptic currents (mIPSCs) in ex vivo brain slices. Our data show that MOX potentiated tonic GABA currents in a dose-dependent manner but had no concomitant effects on phasic GABA currents (i.e., MOX had no effect on the amplitude, frequency, or decay kinetics of mIPSCs). These studies indicate that behavioral experiments that implement a ML-based novel ligand-receptor system should take care to control for potential effects of the ML on native tonic GABA receptors.NEW & NOTEWORTHY We have identified a novel mechanism of action in the mammalian central nervous system for the antihelminthic moxidectin, commonly prescribed to animals worldwide and currently being evaluated for use in humans. Specifically, moxidectin applied to rodent brain slices selectively enhanced the tonic GABA conductance of hippocampal pyramidal neurons.

scholarly journals 5-HT1c receptor is a prominent serotonin receptor subtype in the central nervous system

1989 ◽  
Vol 86 (17) ◽  
pp. 6793-6797 ◽  
Author(s):  
S M Molineaux ◽  
T M Jessell ◽  
R Axel ◽  
D Julius
Keyword(s):  
Central Nervous System ◽  
Nervous System ◽  
Serotonin Receptor ◽  
Pyramidal Neurons ◽  
Receptor Subtype ◽  
Hippocampal Pyramidal Neurons ◽  
Nociceptive Transmission ◽  
Central Processing ◽  
Cell Groups ◽  
The Central Nervous System

Neurons in rat central nervous system (CNS) that express 5-HT1c receptor mRNA have been localized by in situ hybridization histochemistry. The 5-HT1c receptor is expressed in a wide variety of cortical and subcortical neurons including hippocampal pyramidal neurons, neurons within most of the central monoaminergic cell groups, neurons in thalamic sensory relay nuclei, and neurons involved in the central processing and regulation of nociceptive transmission. Therefore, the 5-HT1c receptor is a prominent but poorly characterized central subclass of serotonin (5-HT) receptor. The distribution of the 5-HT1c receptor within the CNS is considerably more widespread than that of the structurally and functionally related 5-HT2 receptor.

7. The use of brain slices and dissociated neurons to explore the multiplicity of 5-HT's action in the central nervous system

1994 ◽  
Vol 52 (1) ◽  
pp. A3-A4
Author(s):  
J.S. Kelly ◽  
N.J. Penington ◽  
P.M. Larkman ◽  
H. Hecimovic ◽  
H. McAllister-Williams
Keyword(s):  
Central Nervous System ◽  
Nervous System ◽  
Brain Slices ◽  
The Central Nervous System

scholarly journals ENTÉRATE DE CÓMO CAMBIA EL CEREBRO CUANDO SE LESIONA UN NERVIO

2016 ◽  
Vol 21 (1Supl) ◽  
pp. 279-285 ◽  
Author(s):  
Julieta Troncoso
Keyword(s):  
Central Nervous System ◽  
Nervous System ◽  
Facial Nerve ◽  
Pyramidal Neurons ◽  
Dendritic Morphology ◽  
Nerve Lesion ◽  
Nerve Injuries ◽  
Somatosensory Stimulation ◽  
Facial Nerve Lesion ◽  
The Central Nervous System

<p>Desde hace algunos años el grupo de investigación de Neurofisiología Comportamental de la Universidad Nacional de Colombia ha venido evaluando los cambios que ocurren en el sistema nervioso central luego de la lesión de un nervio periférico. Específicamente trabajamos con el modelo de lesión del nervio facial en roedores para evaluar las modificaciones funcionales y estructurales que ocurren en la corteza sensoriomotora primaria luego de la lesión. Al lesionarse el nervio facial, el cerebro entra en un programa de reorganización que incluye cambios electrofisiológicos en las neuronas de la corteza motora que comandan los movimientos faciales (M1). En este sentido, las células de la corteza motora cerebral se vuelven más excitables y modifican su respuesta ante estímulos sensoriales. La reorganización tras la lesión también incluye cambios morfológicos en M1: las células piramidales de la corteza motora retraen su árbol dendrítico y disminuye la densidad de sus espinas dendríticas. En asociación con estos cambios, las células de M1 disminuyen transitoriamente su inmunorreactividad para NeuN (marcador específico de núcleos neuronales) y aumentan la expresión de GAP43 (proteína de crecimiento axonal). Esto indica, posiblemente, un cambio metabólico celular en asociación con la búsqueda de nuevas dianas sinápticas. Finalmente, hallamos que la glía circundante en M1 (tanto astrocitos como microglía) se activa de manera muy temprana luego de lesiones del nervio facial. Esto podría indicar que el remodelamiento estructural y funcional hallado en las neuronas corticales es el resultado de la interacción entre la activación de la glía circundante y las células piramidales de M1 (aunque se necesitan muchos experimentos adicionales que así lo demuestren).</p><p> </p><p>Abstract</p><p>Our research group (Neurofisiología Comportamental, Universidad Nacional de Colombia) has evaluated changes in the central nervous system induced by peripheral nerve injuries. We have characterized facial nerve lesion-induced structural and functional changes in primary motor cortex pyramidal neurons (M1) in rodents. Following the lesion, M1 neurons modified their spontaneous basal firing frequency: they become more excitable. Moreover, we found changes in evoked-activity with somatosensory stimulation after facial nerve lesion. Morphologically, it was found that facial nerve lesion induced long-lasting changes in the dendritic morphology of M1 pyramidal neurons. Dendritic branching of the pyramidal cells underwent overall shrinkage and dendrites suffered transient spine pruning. Additionally, we evaluated the reorganization processes in the central nervous system by using both neuronal and glial markers. Decreased NeuN (neuronal nuclei antigen) immunoreactivity and increased GAP-43 (growth-associated protein 43) immunoreactivity were found M1 after facial nerve lesion. In addition, we also observed astrogliosis and microglial activation sourrounding M1 early after facial nerve injury. Taken together these findings suggest that facial nerve lesions induce widespread reorganization in M1 including neuronal shrinkage, axon sprouting as well as astrocytic and microglia activation. These results suggest that facial nerve injuries elicit active remodeling due to pyramidal neuron and glia interaction (although additional experiments that demonstrate it are needed)</p>

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