eoe sub articles

EoE Sub Articles

All procedures involving animal samples have been reviewed and approved by the appropriate animal ethical review committee. 
1. Recording Synaptic Responses in a Submersion Slice Chamber
NOTE: See the Table of Materials.
<ol>
	<li>Pre-warm the aCSF solution in a water bath to about 28-30 &#176;C (this will prevent bubble formation in the recording chamber).</li>
	<li>Start the recycling system (4-5 mL/min) and activate the inline heater to equilibrate the system for at least 1 h.</li>
	<li>Presoak a small piece of lens cleaning paper and a nylon mesh fixed on a U-shaped platinum wire in the submersion slice chamber for a few minutes (Figure 1).</li>
	<li>Remove the U-shaped platinum wire.</li>
	<li>Switch off the recycling and place a slice on the lens cleaning paper.</li>
	<li>Immediately place the slice-holding mesh on top of the slice, switch on the pump, and let the slice equilibrate for 30 min without dipping the objective into the aCSF.</li>
	<li>Fill the borosilicate micropipette (i.e., the recording electrode) with aCSF (tip resistance: 1-2 M&#937;, filled 1/3 with aCSF) and mount it into the pipette holder.</li>
	<li>Place the tested epoxy-insulated tungsten stimulation electrode in the manipulator holder and the reference wires in the slice chamber.</li>
	<li>Add a second reference electrode to the chamber and connect it to the reference socket of the headstage of the recording electrode (Figure 1A).</li>
	<li>In the amplifier control software, click on the &#34;zero clamp mode&#34; box and proceed by double-clicking the &#34;output gain list&#34; box. Choose a gain of &#34;100,&#34; double-click the &#34;high pass filter list box (Bessel),&#34; choose &#34;0.1 Hz,&#34; and double-click the &#34;low pass filter list box (AC)&#34; to choose &#34;3 kHz.&#34;</li>
	<li>In the recording software, click the on &#34;Acquire | Open Protocol.&#34; Choose a protocol that has settings allowing for episodic stimulations and the digitalization of amplified potentials at 10-20 kHz for 50-100 ms and that automatically triggers a stimulus isolator 10 ms after the start of an episodic recording.</li>
	<li>Place the stimulation and recording electrodes in line and parallel to the stratum pyramidale, for example (Figure 1B and Figure 2).</li>
	<li>Click &#34;Acquire | Edit Protocol&#34; and choose the &#34;trigger&#34; tab (in the pop-up window). Click on the &#34;trigger source&#34; box and choose &#34;space bar&#34; as the trigger source. Click the &#34;OK&#34; button. Click the &#34;record&#34; button to start the acquisition and note the pop-up window with a trigger button.</li>
	<li>In the stimulus isolator software, click on the &#34;voltage control&#34; box and enter &#34;0.&#34; Click the &#34;download&#34; button. Click on the &#34;recording software&#34; window and press the space bar. Repeat this cycle by sequentially entering values from 1 to 8, for example, at 1 mV steps in the &#34;voltage control&#34; box.</li>
	<li>Correlate the stimulation strength with fEPSP-slope values and determine the stimulation strength required to get 40% of the fEPSP-slope maximum. Click the &#34;voltage control box&#34; and enter the determined value. Click the &#34;download&#34; button. 
	NOTE: A stimulus isolator is used to apply brief voltage or current pulses to brain tissue. The biphasic pulses can have 100 &#181;s per phase.</li>
	<li>In the recording software, click the stop button and then the &#34;Acquire&#34; menu. Click on &#34;Edit Protocol&#34; and choose the &#34;trigger&#34; tab in the pop-up window. Click on the trigger source box and choose the &#34;internal timer&#34; as the trigger source. Click the &#34;OK&#34; button. Click the record button that starts the automatic recording of field potentials every 30 s, for example. Click the &#34;stop&#34; button after 30-60 min.</li>
	<li>Click on the &#34;Acquire&#34; menu, click on &#34;Open Protocol,&#34; choose the desired induction protocol of synaptic plasticity, and click the &#34;OK&#34; button. Click the &#34;record&#34; button to automatically activate the high-frequency stimulation and recording. Click the &#34;stop&#34; button. 
	NOTE: In case of studies relating to synaptic plasticity, apply one of the standard induction paradigms for long-term potentiation or long-term depression after prolonged baseline recording. Representative examples of the resulting modulation of the synaptic transmission are depicted in Figure 4 and Figure 5.</li>
	<li>In the recording software, click &#34;Acquire | Open Protocol&#34; and choose the same protocol as in step 4.12. Click the &#34;OK&#34; button and then click &#34;record.&#34; Keep automatically running the field potential recordings for 2-4 h, for example. Click the &#34;stop&#34; button to terminate the recording.</li>
	<li>In the case of pharmaceutical studies, apply the compound directly to the aCSF reservoir if permanent compound administration is desired (Figure 3).</li>
</ol>

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Electrophysiology equipment and materials</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Vertical Pipette Puller</td>
			<td>Narishige, Japan</td>
			<td>PC-10</td>
			<td></td>
		</tr>
		<tr>
			<td>Vibration isolation table</td>
			<td>Meirits, Japan</td>
			<td>ADZ-A0806</td>
			<td></td>
		</tr>
		<tr>
			<td>Submerged type recording chamber</td>
			<td>Warner Instruments</td>
			<td>RC-26GLP</td>
			<td></td>
		</tr>
		<tr>
			<td>Thermostatic water bath</td>
			<td>Zhongcheng Yiqi, China</td>
			<td>HH-1</td>
			<td></td>
		</tr>
		<tr>
			<td>4 Axis Micromanipulator</td>
			<td>Sutter, USA</td>
			<td>MP-285, MP-225</td>
			<td></td>
		</tr>
		<tr>
			<td>Platinum Wire</td>
			<td>World Precision Instruments</td>
			<td>PTP406</td>
			<td></td>
		</tr>
		<tr>
			<td>Amplifier</td>
			<td>Molecular Devices, USA</td>
			<td>Multiclamp 700B</td>
			<td></td>
		</tr>
		<tr>
			<td>Data Acquisition System</td>
			<td>Molecular Devices, USA</td>
			<td>Digidata 1440A</td>
			<td></td>
		</tr>
		<tr>
			<td>Anaysis software</td>
			<td>Molecular Devices, USA</td>
			<td>Clampex 10.2</td>
			<td></td>
		</tr>
		<tr>
			<td>Fluorescence Microscope</td>
			<td>Nikon, Japan</td>
			<td>FN1</td>
			<td></td>
		</tr>
		<tr>
			<td>LED light source</td>
			<td>Lumen Dynamics Group, Canada</td>
			<td>X-cite 120LED</td>
			<td></td>
		</tr>
		<tr>
			<td>Micropipettes</td>
			<td>Harvard apparatus</td>
			<td>GC150TF</td>
			<td>extracelluar recording</td>
		</tr>
		<tr>
			<td>Borosilicate micropipettes</td>
			<td>Sutter, USA</td>
			<td>BF150-86</td>
			<td>patch clamp</td>
		</tr>
		<tr>
			<td>Tungsten electrode</td>
			<td>A-M Systems, USA</td>
			<td>575500</td>
			<td></td>
		</tr>
		<tr>
			<td>Peristaltic pump</td>
			<td>Longer, China</td>
			<td>BT00-300T</td>
			<td></td>
		</tr>
		<tr>
			<td>Tubes for peristaltic pump</td>
			<td>ISMATEC, Wertheim, Germany</td>
			<td>SC0309</td>
			<td>1x inflow, ID: 1.02mm</td>
		</tr>
		<tr>
			<td>Tubes for peristaltic pump</td>
			<td>ISMATEC, Wertheim, Germany</td>
			<td>SC0319</td>
			<td>2x tubes for outflow, ID: 2.79 mm</td>
		</tr>
		<tr>
			<td>CCD camera</td>
			<td>PCO, Germany</td>
			<td>pco.edge sCMOS</td>
			<td></td>
		</tr>
		<tr>
			<td>Lens cleaning paper</td>
			<td>Kodak</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>50 ml conical centrifuge tube</td>
			<td>Thermo scientific</td>
			<td>339652</td>
			<td></td>
		</tr>
		<tr>
			<td>Prechamber</td>
			<td>Warner Instruments</td>
			<td>BSC-PC</td>
			<td></td>
		</tr>
		<tr>
			<td>Inline heater</td>
			<td>Warner Instruments</td>
			<td>SF-28</td>
			<td></td>
		</tr>
		<tr>
			<td>Temperature Controller</td>
			<td>Warner Instruments</td>
			<td>TC-324B</td>
			<td></td>
		</tr>
		<tr>
			<td>Reagents required</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>NaCl</td>
			<td>Sinopharm Chemical Reagent, 
			China</td>
			<td>10019318</td>
			<td></td>
		</tr>
		<tr>
			<td>KCl</td>
			<td>Sinopharm Chemical Reagent, 
			China</td>
			<td>10016318</td>
			<td></td>
		</tr>
		<tr>
			<td>KH2PO4</td>
			<td>Sinopharm Chemical Reagent, 
			China</td>
			<td>10017618</td>
			<td></td>
		</tr>
		<tr>
			<td>MgCl2&#183;6H2O</td>
			<td>Sinopharm Chemical Reagent, 
			China</td>
			<td>10012818</td>
			<td></td>
		</tr>
		<tr>
			<td>CaCl2</td>
			<td>Sinopharm Chemical Reagent, 
			China</td>
			<td>10005861</td>
			<td></td>
		</tr>
		<tr>
			<td>NaHCO3</td>
			<td>Sinopharm Chemical Reagent, 
			China</td>
			<td>10018960</td>
			<td></td>
		</tr>
		<tr>
			<td>Glucose</td>
			<td>Sinopharm Chemical Reagent, 
			China</td>
			<td>10010518</td>
			<td></td>
		</tr>
		<tr>
			<td>NaH2PO4</td>
			<td>Sinopharm Chemical Reagent, 
			China</td>
			<td>20040718</td>
			<td></td>
		</tr>
		<tr>
			<td>HEPES&#160;</td>
			<td>Sigma&#160;</td>
			<td>H3375</td>
			<td></td>
		</tr>
		<tr>
			<td>Sodium pyruvate&#160;</td>
			<td>Sigma&#160;</td>
			<td>A4043</td>
			<td></td>
		</tr>
		<tr>
			<td>MgSO4</td>
			<td>Sinopharm Chemical Reagent, 
			China</td>
			<td>20025118</td>
			<td></td>
		</tr>
		<tr>
			<td>NaOH</td>
			<td>Sinopharm Chemical Reagent, 
			China</td>
			<td>10019718</td>
			<td></td>
		</tr>
	</tbody>
</table>

recording neuronal field excitatory postsynaptic potentials in acute hippocampal slices

Source: Weng, W., et al. <a href="https://appjove.unicartagenaproxy.elogim.com/v/55936">Recording Synaptic Plasticity in Acute Hippocampal Slices Maintained in a Small-volume Recycling-, Perfusion-, and Submersion-type Chamber System.</a> J. Vis. Exp. (2018).
This video demonstrates the procedure for setting up a brain slice in a submersion chamber to record synaptic activity. The process involves securing the slice on lens paper, maintaining a continuous flow of artificial cerebrospinal fluid (aCSF) to ensure neuronal viability, and accurately positioning the stimulation and recording electrodes relative to a specific brain region of interest. This setup allows for precise monitoring of baseline synaptic activity and the effects of electrical stimulation.

<img alt="Figure 1" src="/files/ftp_upload/22862/22862_Figure1.jpg" /> 
Figure 1: Components of outflow-carbogenation for experiments in a submersion slice chamber. (A) Presentation of the submersion slice chamber. The U-shaped platinum wire with nylon fibers holding a hippocampal slice is depicted. The silver wires were wrapped around a small cannula, creating a coil to increase the area of the silver wire ending. The stimulation reference wire was ...

Recording Neuronal Field Excitatory Postsynaptic Potentials in Acute Hippocampal Slices

All procedures involving animal samples have been reviewed and approved by the appropriate animal ethical review committee. 
1. Recording Synaptic ...

Place a mouse hippocampal slice into a submersion chamber filled with artificial cerebrospinal fluid, or aCSF.
Secure the slice with a holding mesh to prevent movement.
Continuously circulate oxygenated aCSF to maintain neuronal viability.
Install reference electrodes to provide a stable voltage reference.
Position the stimulation and recording electrodes in line with and parallel to the hippocampal CA1 stratum pyramidale.
Apply electrical pulses.
These pulses stimulate the neurons, causing voltage-gated ion channels to open, allowing positive ions to flow in, and triggering action potentials.
These action potentials propagate along the neurons and induce the release of excitatory neurotransmitters into the synaptic cleft.
The neurotransmitters bind to specific receptors on the postsynaptic neurons, allowing positive ion influx and resulting in a change in membrane potential &#8212; the excitatory postsynaptic potential, or EPSP.
The recording electrode measures the EPSP from a population of neurons within the recording region &#8212; the field EPSP.

recording-neuronal-field-excitatory-postsynaptic-potentials-acute

Universidad de Cartagena UDEC

Research

JoVE Encyclopedia of Experiments

Neuroscience

Neurophysiology

Electrophysiology Techniques

853 vistas. Fuente: Weng, W., et al. Registro de la plasticidad sináptica en cortes agudos del hipocampo mantenidos en un sistema de cámara de tipo reciclaje, perfusión y inmersión de pequeño volumen. J. Vis. Exp. (2018). Este video muestra el procedimiento para configurar un corte de cerebro en una cámara de inmersión para registrar la actividad sináptica. El proceso consiste en asegurar el corte en papel de lente, mantener un flujo continuo de líquido cefalorraquídeo artificial (aCSF) para gara...

Video: Registro de potenciales postsinápticos excitatorios del campo neuronal en cortes agudos del hipocampo - Concepto

Direct-current Stimulation and Multi-electrode Array Recording of Seizure-like Activity in Mice Brain Slice Preparation

Cathodal transcranial direct-current stimulation (tDCS) induces suppressive effects on drug-resistant seizures. To perform effective actions, the stimulation parameters (e.g., orientation, field strength, and stimulation duration) need to be examined in mice brain slice preparations. Testing and arranging the orientation of the electrode relative to the position of the mice brain slice are feasible. The present method preserves the thalamocingulate pathway to evaluate the effect of DCS on anterior cingulate cortex seizure-like activities. The results of the multichannel array recordings indicated that cathodal DCS significantly decreased the amplitude of the stimulation-evoked responses and duration of 4-aminopyridine and bicuculline-induced seizure-like activity. This study also found that cathodal DCS applications at 15 min caused long-term depression in the thalamocingulate pathway. The present study investigates the effects of DCS on thalamocingulate synaptic plasticity and acute seizure-like activities. The current procedure can test the optimal stimulation parameters including orientation, field strength, and stimulation duration in an in vitro mouse model. Also, the method can evaluate the effects of DCS on cortical seizure-like activities at both the cellular and network levels.

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.

La estimulación de corriente directa y multi-electrodo matriz de registro de la actividad convulsiva en ratones cerebro rebanada preparación

Cathodal transcranial direct-current stimulation (tDCS) induces suppressive effects on drug-resistant seizures. To perform effective actions, the ...

Investigación

JoVE Journal

Neurociencias

Recording and Modulation of Epileptiform Activity in Rodent Brain Slices Coupled to Microelectrode Arrays

Temporal lobe epilepsy (TLE) is the most common partial complex epileptic syndrome and the least responsive to medications. Deep brain stimulation (DBS) is a promising approach when pharmacological treatment fails or neurosurgery is not recommended. Acute brain slices coupled to microelectrode arrays (MEAs) represent a valuable tool to study neuronal network interactions and their modulation by electrical stimulation. As compared to conventional extracellular recording techniques, they provide the added advantages of a greater number of observation points and a known inter-electrode distance, which allow studying the propagation path and speed of electrophysiological signals. However, tissue oxygenation may be greatly impaired during MEA recording, requiring a high perfusion rate, which comes at the cost of decreased signal-to-noise ratio and higher oscillations in the experimental temperature. Electrical stimulation further stresses the brain tissue, making it difficult to pursue prolonged recording/stimulation epochs. Moreover, electrical modulation of brain slice activity needs to target specific structures/pathways within the brain slice, requiring that electrode mapping be easily and quickly performed live during the experiment. Here, we illustrate how to perform the recording and electrical modulation of 4-aminopyridine (4AP)-induced epileptiform activity in rodent brain slices using planar MEAs. We show that the brain tissue obtained from mice outperforms rat brain tissue and is thus better suited for MEA experiments. This protocol guarantees the generation and maintenance of a stable epileptiform pattern that faithfully reproduces the electrophysiological features observed with conventional field potential recording, persists for several hours, and outlasts sustained electrical stimulation for prolonged epochs. Tissue viability throughout the experiment is achieved thanks to the use of a small-volume custom recording chamber allowing for laminar flow and quick solution exchange even at low (1 mL/min) perfusion rates. Quick MEA mapping for real-time monitoring and selection of stimulating electrodes is performed by a custom graphic user interface (GUI).

We illustrate how to perform recording and electrical modulation of 4-aminopyridine-induced epileptiform activity in rodent brain slices using microelectrode arrays. A custom recording chamber maintains tissue viability throughout prolonged experimental sessions. Live electrode mapping and selection of stimulating pairs are performed by a custom graphical user interface.

Grabación y modulación de la actividad epileptiforme en rebanadas de cerebro de roedor juntada a arreglos de microelectrodos

Temporal lobe epilepsy (TLE) is the most common partial complex epileptic syndrome and the least responsive to medications. Deep brain stimulation (DBS) ...

Investigating Long-term Synaptic Plasticity in Interlamellar Hippocampus CA1 by Electrophysiological Field Recording

The study of synaptic plasticity in the hippocampus has focused on the use of the CA3-CA1 lamellar network. Less attention has been given to the longitudinal interlamellar CA1-CA1 network. Recently however, an associational connection between CA1-CA1 pyramidal neurons has been shown. Therefore, there is the need to investigate whether the longitudinal interlamellar CA1-CA1 network of the hippocampus supports synaptic plasticity.
We designed a protocol to investigate the presence or absence of long-term synaptic plasticity in the interlamellar hippocampal CA1 network using electrophysiological field recordings both in vivo and in vitro. For in vivo extracellular field recordings, the recording and stimulation electrodes were placed in a septal-temporal axis of the dorsal hippocampus at a longitudinal angle, to evoke field excitatory postsynaptic potentials. For in vitro extracellular field recordings, hippocampal longitudinal slices were cut parallel to the septal-temporal plane. Recording and stimulation electrodes were placed in the stratum oriens (S.O) and the stratum radiatum (S.R) of the hippocampus along the longitudinal axis. This enabled us to investigate the directional and layer specificity of evoked excitatory postsynaptic potentials. Already established protocols were used to induce long-term potentiation (LTP) and long-term depression (LTD) both in vivo and in vitro. Our results demonstrated that the longitudinal interlamellar CA1 network supports N-methyl-D-aspartate (NMDA) receptor-dependent long-term potentiation (LTP) with no directional or layer specificity. The interlamellar network, however, in contrast to the transverse lamellar network, did not present with any significant long-term depression (LTD).

We used recording and stimulation electrodes in longitudinal hippocampal brain slices and longitudinally positioned recording and stimulation electrodes in the dorsal hippocampus in vivo to evoke extracellular postsynaptic potentials and demonstrate long-term synaptic plasticity along the longitudinal interlamellar CA1.

Recording Neuronal Field Excitatory Postsynaptic Potentials in Acute Hippocampal Slices

Transcripción

Recording Neuronal Field Excitatory Postsynaptic Potentials in Acute Hippocampal Slices