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Summary

Abstract

Introduction

Protocol

Representative Results

Discussion

Acknowledgements

Materials

References

Neuroscience

Multichannel Extracellular Recording in Freely Moving Mice

Published: May 26th, 2023

DOI:

10.3791/65245

1School of Life Sciences, South China Normal University, 2Panyu Central Hospital Joint Laboratory of Translational Medical Research, Panyu Central Hospital, South China Normal University
* These authors contributed equally

The protocol describes the methodology of extracellular recording in the motor cortex (MC) to reveal extracellular electrophysiological properties in freely moving conscious mice, as well as the data analysis of local field potentials (LFPs) and spikes, which is useful for evaluating the network neural activity underlying behaviors of interest.

The protocol aims to uncover the properties of neuronal firing and network local field potentials (LFPs) in behaving mice carrying out specific tasks by correlating the electrophysiological signals with spontaneous and/or specific behavior. This technique represents a valuable tool in studying the neuronal network activity underlying these behaviors. The article provides a detailed and complete procedure for electrode implantation and consequent extracellular recording in free-moving conscious mice. The study includes a detailed method for implanting the microelectrode arrays, capturing the LFP and neuronal spiking signals in the motor cortex (MC) using a multichannel system, and the subsequent offline data analysis. The advantage of multichannel recording in conscious animals is that a greater number of spiking neurons and neuronal subtypes can be obtained and compared, which allows the evaluation of the relationship between a specific behavior and the associated electrophysiological signals. Notably, the multichannel extracellular recording technique and the data analysis procedure described in the present study can be applied to other brain areas when conducting experiments in behaving mice.

The local field potential (LFP), an important component of extracellular signals, reflects the synaptic activity of large populations of neurons, which form the neural code for multiple behaviors1. Spikes generated by neuronal activity are considered to contribute to the LFP and are important for neural coding2. Alterations in spikes and LFPs have been proven to mediate several brain diseases, such as Alzheimer's disease, as well as emotions such as fear, etc.3,4. It is worth noting that many studies have highlighted that spike activity significantly differs ....

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All the mice were obtained commercially and maintained in a 12 h light/12 h dark cycle (light on at 08:00 A.M. local time) at a room temperature of 22-25 °C and a relative humidity of 50%-60%. The mice had access to a continuous supply of food and water. All the experiments were carried out in accordance with the Guidelines for Care and Use of Laboratory Animals of South China Normal University and approved by the Institutional Animal Ethics Committee. Male C57BL/6J mice aged 3-5 months old were used for the experim.......

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A high-pass (250 Hz) filter was applied to extract the multi-unit spikes from the raw signals (Figure 6A). Further, the recorded units from the MC of a normal mouse sorted by PCA were verified (Figure 7A-D), and the valley width and waveform duration of the units in the MC of the mouse were recorded. The results showed that both the valley width and waveform duration of the MC putative pyramidal neurons (Pyn) in mice are higher .......

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Multichannel recording in free-moving mice has been deemed to be a useful technology in neuroscience studies, but it is still quite challenging for beginners to record and analyze the signals. In the present study, we provide simplified guidelines for making micro-drive systems and performing electrode implantation, as well as simplified procedures for capturing and analyzing the electrical signals via spike sorting software and software for neurophysiological data analysis.

Given tha.......

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This work was supported by grants from the National Natural Science Foundation of China (31871170, 32170950, and 31970915), the Natural Science Foundation of Guangdong Province (2021A1515010804 and 2023A1515010899), the Guangdong Natural Science Foundation for Major Cultivation Project (2018B030336001), and the Guangdong Grant: Key Technologies for Treatment of Brain Disorders(2018B030332001).

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Name Company Catalog Number Comments
2.54 mm pin header YOUXIN Electronic Co., Ltd. 1 x 5 Applying for the movable micro-drive which can slide on its stulls.
Adobe Illustrator CC 2017 Adobe N/A To optimize images from GraphPad.
BlackRock Microsystems Blackrock Neurotech Cerebus This systems includes headsatge, DA convert, amplifier and computer.
Brass nut Dongguan Gaosi Technology Co., Ltd. M0.8 brass nut The nut fixes the position of screw.
Brass screw Dongguan Gaosi Technology Co., Ltd. M0.8 x 11 mm brass screw A screw that hold the movable micro-drive.
C57BL/6J Guangdong Zhiyuan Biomedical Technology Co., LTD. N/A 12 weeks of age.
Centrifuge tube Biosharp 15 mL; BS-150-M To store mice brain with sucrose sulutions.
Conducting paint Structure Probe, Inc. 7440-22-4 To improve the lead-connecting quality between connector pins and Ni-wires.
Conductive copper foil tape 3M 1181 To reduce interferenc.
Connector YOUXIN Electronic Co., Ltd. 2 x 10P To connect the headtage to micro-drive system.
DC Power supply Maisheng MS-305D A power device for  electrolytic lesion.
Dental cement Shanghai New Century Dental Materials Co., Ltd. N/A To fix the electrode arrays on mouse's skull after finishing the implantation.
Digital analog converter Blackrock 128-Channel A device that converts digital data into analog signals.
Epoxy resin Alteco N/A To cover pins.
Excel Microsoft N/A To summarize data after analysis.
Eye scissors JiangXi YuYuan Medical Equipment Co.,Ltd. N/A For surgery or cutting the Ni-chrome wire.
Fine forceps JiangXi YuYuan Medical Equipment Co.,Ltd. N/A For surgery.
Forceps JiangXi YuYuan Medical Equipment Co.,Ltd. N/A For surgery or assembling the mirco-drive system.
Freezing microtome Leica CM3050 S  Cut the mouse’s brain into slices
Fused silica capillary tubing Zhengzhou INNOSEP Scientific Co., Ltd. TSP050125 To  serve as the guide tubes for Ni-chrome wires.
Glass microelectrode Sutter Instrument Company BF100-50-10 To mark the desired locations for implantation using the filled ink.
GraphPad Prism 7 GraphPad Software N/A To analyze and visualize the results.
Guide-tube Polymicro technologies 1068150020 To load Ni-chrome wires.
Headstage Blackrock N/A A tool of transmitting signals.
Helium balloon Yili Festive products Co., Ltd. 24 inch To offset the weight of headstage and micro-drive system.
Ink Sailor Pen Co.,LTD. 13-2001 To mark the desired locations for implantation.
Iodine tincture Guangdong Hengjian Pharmaceutical Co., Ltd. N/A To disinfect mouse's scalp.
Lincomycin in Hydrochloride and Lidocaine  hydrochloride gel Hubei kangzheng pharmaceutical co., ltd. 10g A drug used to reduce inflammation.
Meloxicam Vicki Biotechnology Co., Ltd. 71125-38-7 To reduce postoperative pain in mice.
Micromanipulators Scientifica Scientifica IVM Triple For electrode arrays implantation.
Microscope  Nikon ECLIPSE Ni-E  Capture the images of brain sections
nanoZ impedance tester Plexon nanoZ To measure impedance or performing electrode impedance spectroscopy (EIS) for multichannel microelectrode arrays.
NeuroExplorer Plexon NeuroExplorer A tool for analyzing the electrophysiological data.
NeuroExplorer  Plexon, USA N/A A software.
Ni-chrome wire California Fine Wire Co. M472490 35 μm Ni-chrome wire.
Offline Sorter Plexon Offline Sorter A tool for sorting the recorded multi-units.
PCB board Hangzhou Jiepei Information Technology Co., Ltd. N/A Computer designed board.
Pentobarbital Sigma P3761 To anesthetize mice.
Pentobarbital sodium Sigma 57-33-0 To anesthetize the mouse.
Peristaltic pump Longer BT100-1F A device used for perfusion
Polyformaldehyde  Sangon Biotech A500684-0500 The main component of fixative solution for fixation of mouse brains 
PtCl4 Tianjin Jinbolan Fine Chemical Co., Ltd. 13454-96-1 Preparation for gold plating liquid.
Saline Guangdong Hengjian Pharmaceutical Co., Ltd. N/A To clean the mouse's skull.
Silver wire Suzhou Xinye Electronics Co., Ltd. 2 mm diameter Applying for ground and reference electrodes.
Skull drill RWD Life Science 78001 To drill carefully two small holes on mouse's skull.
Stainless steel screws YOUXIN Electronic Co., Ltd. M0.8 x 2 To protect the micro-drive system and link the ground and reference electrodes.
Stereotaxic apparatus RWD Life Science 68513 To perform the stereotaxic coordinates of bilateral motor cortex.
Sucrose Damao 57-50-1 To dehydrate the mouse brains  after perfusion.
Super glue Henkel AG & Co. PSK5C To fix the guide tube and Ni-chrome wire.
Temperature controller Harvard Apparatus TCAT-2 To maintain mouse's rectal temperature at 37°C
Tetracycline eye ointment Guangdong Hengjian Pharmaceutical Co., Ltd. N/A To protect the mouse's eyes during surgery.
Thread Rapala N/A To link ballon and headstage.
Vaseline Unilever plc N/A To cover the gap between electrode arrays and mouse's skull.

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