recording neural activity of unfolded hippocampal tissue using penetrating microelectrode array

Recording Neural Activity of Unfolded Hippocampal Tissue Using Penetrating Microelectrode Array

In this procedure, fill one bottle with normal aCSF and another bottle with 4-AP aCSF. Bubble the solutions with 95% oxygen, 5% carbon dioxide from the beginning of the experiments. Use a trivalve connector to control which solution will be selected during an experiment.
Next, connect a vacuum tube to the outlet of the chamber to remove the solution into a dust container. Heat the pipeline before delivering the solution into the recording chamber and keep it at 35 degrees Celsius controlled temperature. After closing the inlet and outlet of the recording chamber, use a custom-made glass pipette dropper to transfer the unfolded hippocampus to the recording chamber.
Under the microscope, position the unfolded hippocampus with its obvious side facing down, CA3 area pointing away, and CA1 field pointing towards the researcher. Carefully aspirate away the solution in the chamber using a vacuum pipette from the edge of the recording chamber until the chamber is dried and the tissue lies on the array.
Then, carefully place a custom-made tissue anchor on top of the tissue to hold the unfolded hippocampus onto the array. Refill the recording chamber with a few drops of solution. Gradually open the inlet and outlet to adjust the flow rate to about two drops per second in the IV drip chamber.
Incubate the tissue in the recording chamber with normal aCSF for about one minute. Then, switch the solution supply to 4-AP dissolved aCSF, and adjust the flow rate properly. Incubate the tissue in 4-AP dissolved aCSF for about 5 to 10 minutes before recording.

recording-neural-activity-unfolded-hippocampal-tissue-using

Universidad de Cartagena UDEC

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JoVE Encyclopedia of Experiments

Neuroscience

Neurophysiology

Electrophysiology Techniques

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All procedures involving animal samples have been reviewed and approved by the appropriate animal ethical review.
1. Solutions for Surgery and Experimental Recording
<ol>
	<li>Prepare a normal artificial cerebrospinal fluid (aCSF) buffer containing (mM) NaCl (sodium chloride) 124, KCl (potassium chloride) 3.75, KH2PO4 (potassium dihydrogen phosphate) 1.25, MgSO4 (magnesium sulfate) 2, NaHCO3 (sodium bicarbonate) 26, Dextrose 10, and CaCl2 (calcium chloride) 2. Use this normal aCSF for tissue recovery after dissection and for the washing system at the beginning of the experiment.</li>
	<li>Prepare sucrose aCSF that is used during the hippocampus dissection and contains (mM): Sucrose 220, KCl 2.95, NaH2PO4 (sodium dihydrogen phosphate) 1.3, MgSO4 2, NaHCO3 26, Dextrose 10, and CaCl2 2. In order to induce epileptiform activity in the intact hippocampal tissue preparation, add 4-Aminopyridine (4-AP) to normal aCSF at the concentration of 100 &#181;M.</li>
</ol>
2. Placing the Unfolded Hippocampus onto the penetrating microelectrode array (PMEA) to Record the Neural Activity
<ol>
	<li>To prepare the experimental setup, fill one bottle with normal aCSF and another bottle with 4-AP aCSF. In both bottles, bubble 95% O2/ 5% CO2 from the very beginning of each experiment. Use a tri-valve connector to control which solution will be selected during an experiment. Connect a vacuum tube at the outlet of the chamber to pump the solution into a dust container. Heat the pipeline before delivering it into the recording chamber and keep the solution at a controlled temperature level (35 &#176;C).</li>
	<li>When the recording chamber&#39;s inlet and outlet are closed, use a custom-made glass pipette dropper to transfer and place the unfolded hippocampus into the chamber. Under the microscope, position the unfolded hippocampus using a regular small paint brush while the tissue is floating in the solution. Place the unfolded hippocampus with its alveus side facing down, the CA3 area pointing away, and the CA1 field pointing towards the researcher.</li>
	<li>Carefully suck away the solution in the chamber using a vacuum pipette from the edge of the recording chamber to lower the solution level until the chamber is dried and the tissue is lowered onto the array. Then, carefully place a custom-made tissue anchor (Figure 1) (No. 7 in Specific Materials and Equipment) on top of the tissue to hold the unfolded hippocampus onto the array. Put a few drops of solution into the recording chamber to refill it, and gradually open the inlet and outlet to adjust the flow rate to about two drops per second in the IV drip chamber.</li>
	<li>Incubate the tissue in the recording chamber with normal aCSF for about 1 min to recover, then switch the solution supply to 4-AP dissolved aCSF and adjust the flow rate properly. Incubate the tissue in 4-AP dissolved aCSF for about 5 to 10 min, and then the researcher could start the software to record the signal when spontaneous activity appears.</li>
</ol>

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>small paint brush</td>
			<td>Lowe&#39;s</td>
			<td>tem #: 105657 Model #: 90219</td>
			<td>The one with the smallest size in a normal paint brush package</td>
		</tr>
		<tr>
			<td>Fire polished glass help tool</td>
			<td>N/A</td>
			<td>N/A</td>
			<td>This tool was fire polished and made from the regular Pasteur glass pipettes.</td>
		</tr>
		<tr>
			<td>Custom made glass needle</td>
			<td>N/A</td>
			<td>N/A</td>
			<td>This tool was fire polished and made from the regular Pasteur glass pipettes.</td>
		</tr>
		<tr>
			<td>Custom made glass tool with a metal wire loop</td>
			<td>N/A</td>
			<td>N/A</td>
			<td>This tool was fire polished and made from the regular Pasteur glass pipettes with a reshaped metal wire loop.</td>
		</tr>
		<tr>
			<td>Custom made glass solution dropper</td>
			<td>N/A</td>
			<td>N/A</td>
			<td>This tool was made from the regular Pasteur glass pipettes with its tips cut and a rubber head attached with the cut end.</td>
		</tr>
		<tr>
			<td>Custom made tissue anchor</td>
			<td>N/A</td>
			<td>N/A</td>
			<td>Nylon fiber mesh was glued on a insulated copper wire ring. The tissue anchor was hold by an micromanipulator.</td>
		</tr>
		<tr>
			<td>Custom fabricated microelectrode array</td>
			<td>N/A</td>
			<td>N/A</td>
			<td>More detail about the array please refer to Kibler, et al, 2011.</td>
		</tr>
		<tr>
			<td>Custom made filter and amplifiers circuits for the array</td>
			<td>N/A</td>
			<td>N/A</td>
			<td>More detail about the array please refer to Kibler, et al, 2011.</td>
		</tr>
		<tr>
			<td>Data acquisition processor 3400a</td>
			<td>Microstar Laboratories</td>
			<td>N/A</td>
			<td>This is a complete data acquisition system with A/D converter.</td>
		</tr>
	</tbody>
</table>

Source: Zhang, M., et al. <a href="https://appjove.unicartagenaproxy.elogim.com/v/52601">Neural Activity Propagation in an Unfolded Hippocampal Preparation with a Penetrating Micro-electrode Array.</a> J. Vis. Exp. (2015).
This video demonstrates the procedure to set up a recording chamber with a penetrating microelectrode array (PMEA) to capture neural signals from an unfolded hippocampus. This method allows us to study the speed and direction of propagation of neural activity within the unfolded hippocampus in vitro.

<img alt="Figure 1" src="/files/ftp_upload/22722/22722_Figure1.jpg" /> 
Figure 1. Experimental setup. (A) A plastic cover lid with screws is placed over the circuits to protect it against possible water damage. The recording chamber has both inlet and outlet tubes to carry the solution flow. A custom-made tissue anchor glued with a Nylon fiber mesh is used to secure the tissue during the experiments. (B) Picture taken from the bottom of the glass sub...

All procedures involving animal samples have been reviewed and approved by the appropriate animal ethical review.
1. Solutions for Surgery and ...

Take a recording chamber containing the penetrating microelectrode array or PMEA, which records electrical signals from neural tissue.
Connect the inlet to bottles containing oxygenated aCSF and oxygenated aCSF with 4-aminopyridine or 4-AP using a trivalve connector.
Attach the outlet to a vacuum tube.
Heat the pipeline between the inlet and trivalve to physiological temperature.
With the inlet and outlet closed, transfer the unfolded hippocampus into the chamber.
Under a microscope, position the hippocampus on the PMEA.
Remove the solution from the chamber.
Place a tissue anchor to hold the tissue onto the PMEA electrodes.
Open the inlet and outlet to start the aCSF flow.
Incubate the tissue with aCSF to allow neuronal acclimatization.
Then, flow in aCSF with 4-AP. The 4-AP blocks the neuronal potassium channels, extending the action potential duration.
The PMEA captures the electrical signals proportional to the enhanced neural firing from hippocampal neurons.

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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.

小鼠脑切片准备扣押样活性的直接电流刺激和多电极阵列记录

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

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JoVE Journal

神经科学

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.

鼠脑切片偶联微电极阵列癫痫活性的记录与调控

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

Recording Neural Activity of Unfolded Hippocampal Tissue Using Penetrating Microelectrode Array

成績單

Recording Neural Activity of Unfolded Hippocampal Tissue Using Penetrating Microelectrode Array