Summary

Direct-current stimulatie en multi-elektrode Array Registratie van Inbeslagneming-achtige activiteit in de muizen Brain Slice Voorbereiding

Published: June 07, 2016
doi:

Summary

Studies have shown that cathodal transcranial direct-current stimulation can produce suppressive effects on drug-resistant seizures. In this study, an in vitro experimental setup was devised in which the direct-current stimulation and multielectrode array recording of seizure-like activity were evaluated in mice brain slice preparation. The direct-current stimulation parameters were evaluated.

Abstract

Kathodische gepulste elektromagnetische velden (tDCS) induceert onderdrukkende effecten op de resistente aanvallen. Om effectief acties uit te voeren, de stimulatie parameters (bv, oriëntatie, veldsterkte, en duur stimulatie) nodig hebben bij muizen hersenen slice voorbereidingen worden onderzocht. Testen en regelen van de oriëntatie van de elektrode ten opzichte van de positie van de muizen hersenen slice haalbaar. De onderhavige werkwijze behoudt de thalamocingulate weg naar het effect van DCS op de anterior cingulate cortex inbeslagneming-achtige activiteiten te evalueren. De resultaten van de multikanaals matrix recordings aangegeven kathodische DCS significant af van de amplitude van de stimulatie opgewekte responsen en duur van 4-aminopyridine en bicuculline geïnduceerde aanvallen-achtige activiteit. Deze studie is ook gebleken dat de kathodische DCS toepassingen op 15 min veroorzaakt langdurige depressie in de thalamocingulate route. De huidige studie onderzoekt de effecten van DCS op thalamocingulate synaptische plasticiteit en acute inbeslagneming-achtige activiteiten. De huidige procedure kan testen de optimale stimulatie parameters zoals oriëntatie, veldsterkte en de duur stimuleren in een in vitro muismodel. Ook kan de werkwijze de effecten van DCS corticale aanvallen-achtige activiteiten zowel op cellulaire als netwerkniveau evalueren.

Introduction

Epilepsy is a common neurological disorder. Thirty percent of patients with epilepsy suffer from drug-resistant seizures1. Transcranial direct-current stimulation (tDCS) provides a noninvasive approach to control or alter network activities across large brain areas, such as seizures. Clinical studies have shown that tDCS effectively treats intractable seizures2 and can produce both short- and long-term suppressive effects on seizures3-5. However, the therapeutic mechanism of tDCS actions is still unclear. The brain slice model presented is an in vitro method to investigate how the therapeutic mechanism of tDCS actions alters the symptoms of seizure-like brain activities. Accordingly, to achieve its optimal effects, specific stimulation parameters including orientation, field strength, and stimulation duration need to be tested in an experimental model. Previous studies have shown that the orientation of the electric field is important to obtain therapeutic effects6. Thus, testing and arranging the orientation of electrodes relative to the position of the tested brain slice are feasible.

Frontal lobe epilepsy and anterior cingulate cortex (ACC) seizures are often drug-resistant7,8. Some studies have reported the application of tDCS in the cingulate cortex9-11. tDCS is shown to affect vigilance, decision making and emotion through alteration of ACC activities, and can modulate neuronal excitability and seizure activity in this brain region12. Therefore, suppressive effects of tDCS on ACC seizures might be helpful for clinical treatment and the evaluation of alternative treatments.

The present protocol describes the preparation of an electrode in the recording chamber for DCS of a brain slice and its effect on seizure-like activity recording with a multielectrode array (MEA).

Protocol

Procedures die proefdieren te betrekken werden goedgekeurd door de Institutional Animal Care en Ingebruikname Comité, Academia Sinica, Taipei, Taiwan. 1. Voorbereiding Experimentele Solution en Apparatuur voor multielectrode Array Recording Bereid kunstmatige cerebrospinale vloeistof (aCSF; 124 mM NaCl, 4,4 mM KCI, 1 mM NaH 2 PO 3, 2 mM MgSO4, 2 mM CaCl2, 25 mM NaHCO 3 en 10 mM glucose, geborreld met 95% O2 en 5% C…

Representative Results

Voorbereiding van de Thalamocingulate Slice en MEA Recording System Setup De MT-ACC segment van muizen is een speciaal segment preparaat dat verkenning van de elektrofysiologische eigenschappen van het thalamocingulate route toelaat. Figuur 1A toont de wijze waarop de MT-ACC slice bereid. De hersenen van de muis werd snel verwijderd en bewaard in koud geoxygeneerd aCSF (Figuur 1A, a, b). Om su…

Discussion

In deze studie werden de effecten van de duur en oriëntatie van DCS ACC aanvallen-achtige activiteit getest. Stabiele data in muizenhersenen plakjes verkrijgen, hoe de integriteit van de MT-ACC pathway houden en om schade te voorkomen is het belangrijk, met name de stappen waarin twee schuine sneden ventrale en dorsale verlaging van de cortex gemaakt. Bovendien kan de tijd om de hersenen slice bereiden beïnvloeden ook de activiteit van de hersenen deel, dat zo snel mogelijk naar de hersenen vers en sterk te houden moe…

Disclosures

The authors have nothing to disclose.

Acknowledgements

We are grateful for the technical support from the Neural Circuit Electrophysiology Core at Academia Sinica. This work was supported by the National Science Council (102-2320-B-001-026-MY3 and 100-2311-B-001-003-MY3) and Neuroscience Program of Academia Sinica.

Materials

Anesthetic:
Isoflurane Halocarbon Products Corporation  NDC 12164-002-25 4%
Name Company Catalog Number Comments
aCSF (total:1L):
D(+)-Glucose MERCK 1.08337.1000 10 mM
Sodium hydrogen carbonate MERCK 1.06329.0500 25 mM
Sodium chloride MERCK 1.06404.1000 124 mM
(+)-Sodium L-ascorbate, >=98% SIGMA A4034-100G 0.15 g / 2 c.c
Magnesium sulfate, anhydrous,ReagentPlus SIGMA M7506-500G 2 mM
Calcium chloride dihydrate MERCK 1.02382.1000 2 mM
Sodium dihydrogen phosphate monohydrate MERCK 1.06346.1000 1 mM
Potassium chloride May & Baker LTD Dagenham England MS 7616 4.4 mM
Name Company Catalog Number Comments
Drugs:
(+)-Bicuculline TOCRIS 0130 5 µM in aCSF
4-Aminopyridine TOCRIS 0940 250 µM in aCSF
Name Company Catalog Number Comments
Brain slice Preparation:
Vibratome Vibratome Series 1000 Block slicing into 500 µm thick slices
Name Company Catalog Number Comments
MEA system:
Multielectrode array (MEA) probes: 6 x 10 planar MEA Multi Channel Systems 60MEA500/30iR-Ti-pr MEAS 6×10 electrode diameter, 30 µm; electrode spacing, 500 µm; impedance, 50 kΩ at 200 Hz
Multielectrode array (MEA) probes: 8 x 8 MEA  Ayanda Biosystems 60MEA200/10iR-Ti-pr MEAS 8×8 pyramidal-shaped electrode; diameter, 40 µm; tip height, 50 µm; electrode spacing, 200 µm; impedance, 1000 kΩ at 200 Hz
A 60-channel amplifier was used with a band-pass filter set between 0.1 Hz and 3 KHz at 1200X amplification Multi-Channel Systems MEA-1060-BC
MC Rack software at a 10 KHz sampling rate Multi-Channel Systems Software for data collect and recordings
control of a pulse generator Multi-Channel Systems STG 1002
slice anchor kits and hold-downs Warner Instruments SHD-26H/10; WI64-0250
Peristaltic Pump-minipuls3 Gilsom MINIPULS3 perfusion rate : 8 ml/min
Name Company Catalog Number Comments
Stimulation system:
Isolated stimulator A-M Systems Model 2100 intensity of ±350 μA , duration of 200 μs
Tungsten electrode A-M Systems 575300 placed in thalamus

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Cite This Article
Lu, H., Chang, W., Chang, W., Shyu, B. Direct-current Stimulation and Multi-electrode Array Recording of Seizure-like Activity in Mice Brain Slice Preparation. J. Vis. Exp. (112), e53709, doi:10.3791/53709 (2016).

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