KCC2 antagonism increases neuronal network excitability but disrupts ictogenesis in vitro

The potassium-chloride cotransporter 2 (KCC2) plays a role in epileptiform synchronization, but it remains unclear how it influences such a process. Here, we used tetrode recordings in the in vitro rat entorhinal cortex (EC) to analyze the effects of the KCC2 antagonist VU0463271 on 4-aminopyridine (4AP)-induced ictal and interictal activity. During 4AP application, ictal events were associated with significant increases in interneurons and principal cells activities. VU0463271 application transformed ictal discharges to shorter ictal-like events that were not accompanied by significant increases in interneuron or principal cell firing. Interictal events persisted during VU0463271 application at an accelerated frequency of occurrence with significant increases in interneuron and principal cell activity. Further analysis revealed that interneuron and principal cell firing rate during 4AP-induced interictal events were increased after VU0463271 application without changes in synchronicity. Overall, our results demonstrate that in the EC, KCC2 antagonism enhances both interneuron and principal cell excitability, while paradoxically decreasing the ability of neuronal networks to generate structured ictal events. NEW & NOTEWORTHY We are the first to use tetrode recordings in the entorhinal cortex to demonstrate that antagonizing potassium-chloride cotransporter 2 (KCC2) function abolishes ictal discharges and the associated, dynamic changes in single-unit firing in the in vitro 4-aminopyrine model of epileptiform synchronization. Interictal discharges were, however, shorter and more frequent during KCC2 antagonism, while the associated single-unit activity increased, suggesting augmented neuronal excitability. Our findings highlight the complex role of KCC2 in disease pathology.

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Single-unit Activity in the in vitro Entorhinal Cortex During Carbachol-induced Field Oscillations

Neuroscience ◽

10.1016/j.neuroscience.2018.03.003 ◽

2018 ◽

Vol 379 ◽

pp. 1-12 ◽

Cited By ~ 4

Author(s):

Li-Yuan Chen ◽

Maxime Lévesque ◽

Mauro Cataldi ◽

Massimo Avoli

Keyword(s):

Entorhinal Cortex ◽

Unit Activity ◽

Single Unit ◽

Single Unit Activity ◽

Induced Field

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Contribution of near-threshold currents to intrinsic oscillatory activity in rat medial entorhinal cortex layer II stellate cells

Journal of Neurophysiology ◽

10.1152/jn.00743.2011 ◽

2013 ◽

Vol 109 (2) ◽

pp. 445-463 ◽

Cited By ~ 24

Author(s):

Anne Boehlen ◽

Christian Henneberger ◽

Uwe Heinemann ◽

Irina Erchova

Keyword(s):

Entorhinal Cortex ◽

Neuronal Excitability ◽

Spatial Information ◽

Sodium Current ◽

Stellate Cells ◽

Frequency Component ◽

Oscillatory Activity ◽

Potential Oscillations ◽

Near Threshold

The temporal lobe is well known for its oscillatory activity associated with exploration, navigation, and learning. Intrinsic membrane potential oscillations (MPOs) and resonance of stellate cells (SCs) in layer II of the entorhinal cortex are thought to contribute to network oscillations and thereby to the encoding of spatial information. Generation of both MPOs and resonance relies on the expression of specific voltage-dependent ion currents such as the hyperpolarization-activated cation current ( IH), the persistent sodium current ( INaP), and the noninactivating muscarine-modulated potassium current ( IM). However, the differential contributions of these currents remain a matter of debate. We therefore examined how they modify neuronal excitability near threshold and generation of near-threshold MPOs and resonance in vitro. We found that resonance mainly relied on IH and was reduced by IH blockers and modulated by cAMP and an IM enhancer but that neither of the currents exhibited full control over MPOs in these cells. As previously reported, IH controlled a theta-frequency component of MPOs such that blockade of IH resulted in fewer regular oscillations that retained low-frequency components and high peak amplitude. However, pharmacological inhibition and augmentation of IM also affected MPO frequencies and amplitudes. In contrast to other cell types, inhibition of INaP did not result in suppression of MPOs but only in a moderation of their properties. We reproduced the experimentally observed effects in a single-compartment stochastic model of SCs, providing further insight into the interactions between different ionic conductances.

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How to build a grid cell

Philosophical Transactions of the Royal Society B Biological Sciences ◽

10.1098/rstb.2012.0520 ◽

2014 ◽

Vol 369 (1635) ◽

pp. 20120520 ◽

Cited By ~ 8

Author(s):

Christoph Schmidt-Hieber ◽

Michael Häusser

Keyword(s):

Entorhinal Cortex ◽

Action Potentials ◽

Path Integration ◽

Grid Cell ◽

Medial Entorhinal Cortex ◽

Synaptic Inputs ◽

New Findings ◽

Cell Firing

Neurons in the medial entorhinal cortex fire action potentials at regular spatial intervals, creating a striking grid-like pattern of spike rates spanning the whole environment of a navigating animal. This remarkable spatial code may represent a neural map for path integration. Recent advances using patch-clamp recordings from entorhinal cortex neurons in vitro and in vivo have revealed how the microcircuitry in the medial entorhinal cortex may contribute to grid cell firing patterns, and how grid cells may transform synaptic inputs into spike output during firing field crossings. These new findings provide key insights into the ingredients necessary to build a grid cell.

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Activity-dependent feedback modulation of spike patterning of supraoptic nucleus neurons by endogenous adenosine

AJP Regulatory Integrative and Comparative Physiology ◽

10.1152/ajpregu.00744.2005 ◽

2006 ◽

Vol 291 (1) ◽

pp. R83-R90 ◽

Cited By ~ 15

Author(s):

P. M. Bull ◽

C. H. Brown ◽

J. A. Russell ◽

M. Ludwig

Keyword(s):

Electrical Activity ◽

Hazard Function ◽

Supraoptic Nucleus ◽

Neuronal Excitability ◽

Cell Activity ◽

Male Rats ◽

Firing Rates ◽

Endogenous Adenosine

Neuropeptide secretion from the dendrites of hypothalamic magnocellular supraoptic nucleus (SON) neurons contributes to the regulation of neuronal activity patterning, which ultimately determines their peptide output from axon terminals in the posterior pituitary gland. SON dendrites also secrete a number of other neuromodulators, including ATP. ATP degrades to adenosine in the extracellular space to complement transported adenosine acting on pre- and postsynaptic SON A1 receptors to reduce neuronal excitability, measured in vitro. To assess adenosine control of electrical activity in vivo, we made extracellular single-unit recordings of the electrical activity of SON neurons in anesthetized male rats. Microdialysis application (retrodialysis) of the A1 receptor antagonist, 8-cyclopentyl-1,3-dimethylxanthine (CPT) increased phasic vasopressin cell intraburst firing rates progressively over the first 5 s by 4.5 ± 1.6 Hz ( P < 0.05), and increased burst duration by 293 ± 64% ( P < 0.05). Hazard function plots were generated from interval interspike histograms and revealed that these effects were associated with increased postspike excitability. In contrast, CPT had no effect on the firing rates and hazard function plot profiles of continuously active vasopressin and oxytocin cells. However, CPT significantly increased clustering of spikes, as quantified by the index of dispersion, in oxytocin cells and continuously active vasopressin cells (by 267 ± 113% and 462 ± 67%, respectively, P < 0.05). Indeed, in 4 of 5 continuously active vasopressin cells, CPT induced a pseudophasic activity pattern. Together, these results indicate that endogenous adenosine is involved in the local control of SON cell activity in vivo.

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Diverse antiepileptic drugs increase the ratio of background synaptic inhibition to excitation and decrease neuronal excitability in neurones of the rat entorhinal cortex in vitro

Neuroscience ◽

10.1016/j.neuroscience.2010.02.021 ◽

2010 ◽

Vol 167 (2) ◽

pp. 456-474 ◽

Cited By ~ 13

Author(s):

S.D. Greenhill ◽

R.S.G. Jones

Keyword(s):

Antiepileptic Drugs ◽

Entorhinal Cortex ◽

Neuronal Excitability ◽

Synaptic Inhibition

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An In vitro distal colon-inferior splanchnic nerve preparation: Single unit responses to colorectal distension and bradykinin

Gastroenterology ◽

10.1016/s0016-5085(01)81630-9 ◽

2001 ◽

Vol 120 (5) ◽

pp. A328-A328

Author(s):

J YUWEI ◽

Y WANG ◽

J MCROBERTS ◽

A MAYER

Keyword(s):

Single Unit ◽

Splanchnic Nerve ◽

Distal Colon ◽

Colorectal Distension

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Anti-tumor cell activity and in vitro profile of the next generation CXCR4 antagonist Balixafortide

Annals of Oncology ◽

10.1093/annonc/mdy272.312 ◽

2018 ◽

Vol 29 ◽

pp. viii103

Author(s):

J. Zimmermann ◽

T. Remus ◽

G. Lemercier ◽

D. Barker ◽

D. Obrecht ◽

...

Keyword(s):

Tumor Cell ◽

Cell Activity ◽

Next Generation ◽

Cxcr4 Antagonist

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Osseointegrating and phase-oriented micro-arc-oxidized titanium dioxide bone implants

Journal of Applied Biomaterials & Functional Materials ◽

10.1177/22808000211006878 ◽

2021 ◽

Vol 19 ◽

pp. 228080002110068

Author(s):

Hsien-Te Chen ◽

Hsin-I Lin ◽

Chi-Jen Chung ◽

Chih-Hsin Tang ◽

Ju-Liang He

Keyword(s):

Titanium Dioxide ◽

High Surface ◽

Cell Activity ◽

Active Surface ◽

Rutile Phase ◽

Hydroxyl Groups ◽

Bone Implant ◽

Implant System

Here, we present a bone implant system of phase-oriented titanium dioxide (TiO2) fabricated by the micro-arc oxidation method (MAO) on β-Ti to facilitate improved osseointegration. This (101) rutile-phase-dominant MAO TiO2 (R-TiO2) is biocompatible due to its high surface roughness, bone-mimetic structure, and preferential crystalline orientation. Furthermore, (101) R-TiO2 possesses active and abundant hydroxyl groups that play a significant role in enhancing hydroxyapatite formation and cell adhesion and promote cell activity leading to osseointegration. The implants had been elicited their favorable cellular behavior in vitro in the previous publications; in addition, they exhibit excellent shear strength and promote bone–implant contact, osteogenesis, and tissue formation in vivo. Hence, it can be concluded that this MAO R-TiO2 bone implant system provides a favorable active surface for efficient osseointegration and is suitable for clinical applications.

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184 Two types of anti-TIGIT antibodies with distinct binding epitope and functional activities

Journal for ImmunoTherapy of Cancer ◽

10.1136/jitc-2020-sitc2020.0184 ◽

2020 ◽

Vol 8 (Suppl 3) ◽

pp. A198-A198

Author(s):

Tingting Zhong ◽

Xinghua Pang ◽

Zhaoliang Huang ◽

Na Chen ◽

Xiaoping Jin ◽

...

Keyword(s):

T Cells ◽

Mixed Culture ◽

Cell Activity ◽

Cross Reactivity ◽

Binding Activity ◽

Jurkat Cells ◽

List Type ◽

Dependent Manner ◽

Vitro Assay

BackgroundTIGIT is an inhibitory receptor mainly expressed on natural killer (NK) cells, CD8+ T cells, CD4+ T cells and Treg cells. TIGIT competes with CD226 for binding with CD155. In cancers, CD155 has been reported to up-regulate on tumor cells, and TIGIT was found to increase on TILs.1 Activation of TIGIT/CD155 pathway would mediate immunosuppression in tumor; while blockade of TIGIT promotes anti-tumor immune response.MethodsAK126 and AK113 are two humanized anti-human TIGIT monoclonal antibodies developed by Akesobio. Binding activity of AK126 and AK113 to human TIGIT, and competitive binding activity with CD155 and CD112, were performed by using ELISA, Fortebio, and FACS assays. Cross-reactivity with cynomolgus monkey TIGIT and epitope binning were also tested by ELISA assay. In-vitro assay to investigate the activity to promote IL-2 secretion was performed in mixed-culture of Jurkat-TIGIT cells and THP-1 cells.ResultsAK126 and AK113 could specifically bind to human TIGIT with comparative affinity and effectively blocked the binding of human CD155 and CD112 to human TIGIT. X-ray crystal structure of TIGIT and PVR revealed the C’-C’’ loop and FG loop regions of TIGIT are the main PVR interaction regions.2 The only amino acid residue differences in these regions between human and monkey TIGIT are 70C and 73D. AK126 binds to both human and monkey TIGIT, AK113 binds only to monkey TIGIT. This suggests that these residues are required for AK113 binding to human TIGIT, but not required for AK126. Interestingly, results from cell-based assays indicated that AK126 and AK113 showed significantly different activity to induce IL-2 secretion in mixed-culture of Jurkat-TIGIT cells and THP-1 cells (figure 1A and B), in which AK126 had a comparable capacity of activity to 22G2, a leading TIGIT mAb developed by another company, to induce IL-2 secretion, while, AK113 showed a significantly higher capacity than 22G2 and AK126.Abstract 184 Figure 1Anti-TIGIT Antibodies Rescues IL-2 Production in Vitro T-Cell Activity Assay in a dose dependent manner. Jurkat-TIGIT cells (Jurkat cells engineered to over-express human TIGIT) were co-cultured with THP-1 cells, and stimulated with plate-bound anti-CD3 mAb in the presence of TIGIT ligand CD155 (A) or CD112 (B) with anti-TIGIT antibodies. After incubated for 48h at 37° C and 5.0% CO2, IL-2 levels were assessed in culture supernatants by ELISA. Data shown as mean with SEM for n = 2.ConclusionsWe discovered two distinct types of TIGIT antibodies with differences in both epitope binding and functional activity. The mechanism of action and clinical significance of these antibodies require further investigation.ReferencesSolomon BL, Garrido-Laguna I. TIGIT: a novel immunotherapy target moving from bench to bedside. Cancer Immunol Immunother 2018;67:1659–1667.Stengel KF, Harden-Bowles K, Yu X, et al. Structure of TIGIT immunoreceptor bound to poliovirus receptor reveals a cell-cell adhesion and signaling mechanism that requires cis-trans receptor clustering. Proc Natl Acad Sci USA 2012;109:5399–5404.

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