Multichannel Extracellular Recording in Freely Moving Mice

We aim to uncover the properties of neuronal firing and network local field potentials in behaving mice, carrying out specific tasks by correlating the electrophysiological signal with behaviors. Multichannel extracellular recording by the microdrive system has been shown to be a suitable and efficient technology for medium neural activity during behavioral tests. Multichannel recording in free-moving mice has been deemed to be a useful technology in neuroscience studies, but it's still quite challenging for beginners to acquire and analyze those signals.

We introduced how to perform the multichannel extracellular recording in freely moving mice with a more steady and lightweight microdrive system and optimize the processes on recording and data analysis for the beginners. We are going to increase the number of channels and reduce the volume of microdriver system in a new version. Begin by assembling the microdrive system.

Connect two computer-designed boards using two skulls and a screw that holds the movable microdrive and attach the connector to one board. Ensure the microdrive can carry two sets of eight guide tubes for each side of the motor cortex, or MC region. Cut the guide tubes to the same length.

Cut 16 nichrome wires, each measuring approximately five centimeters long and 35 micrometers in diameter. Load the wires into the guide tubes and apply glue to fix them. Strip the wire insulation and twine each exposed wire to each pin from the connector following the channel map and the reference and ground electrodes.

Then, slowly coat conducting paint onto each pin. Cover the pins using epoxy resin, then perform gold plating via an impedance tester to reduce the impedance of the electrode tips to approximately 350 kiloohms. For electrode array implantation, fix the anesthetized mouse in a stereotaxic apparatus and use a temperature controller to maintain its rectal temperature at 37 degrees Celsius.

Then, make a small midline incision to expose the skull. Remove the residual tissue using scissors and clean the skull with sterile saline-soaked cotton buds. Using a glass microelectrode filled with ink, mark the desired locations of the bilateral MC for implantation.

Next, using a skull drill, carefully drill two small holes on both the left and right sides of the coordinated skull in the MC regions. Gently remove the dura mater from the holes with fine forceps. Then, insert the microdrive system into the center of the holes using a micromanipulator at 10 micrometers per second.

After finishing the insertion, fill the petroleum jelly into the dental cement walls and join the bottom plate of the microdrive system and the dental cement walls with the mixed dental cement. For the multichannel recording, move down the electrode arrays by twisting the screw on the movable part of the microdrive system one day in advance. Hold the head of an awake mouse lightly and carefully, then link the center of the head stage and a helium balloon with a thread to offset the weight of the head stage and the microdrive system.

Capture raw signals using the recording electrodes and multi-channel systems by sampling at 30 kilohertz in the recording software, then digitize using a digital analog converter from the multi-channel systems. Extract the local field potential, or LFP signals, from the raw data by resampling at 10 kilohertz, then using a notch filter, remove the 50 hertz line noise. For spike sorting and analysis, in the spike sorting software, click on file, then open and NEV files to open the spike data sampled at 30 kilohertz.

Click on info to select the unsorted channel, then choose sort, change sort method, and use K means. Press the button valley seeking sort, then K means sorting to obtain the sorted units. Next, in the neurophysiological data analysis software, open the sorted spike file by clicking on file, import data, and Blackrock file.

To obtain the autocorrelogram for the selected unit, click on analysis, then autocorrelograms and set the parameters. Load the sorted spike data, then click on analysis and interspike interval histograms to obtain the interspike interval histogram, and then set the desired parameters. Click on analysis, then cross corellograms to obtain the cross corellogram between two sorted unit events, and then set the reference events and parameters.

For LFP analysis, click on file, import data, and Blackrock file to open the continuous signal data sampled at 10 kilohertz. Then, by clicking analysis and spectrum for continuous, analyze the power spectrum for the LFP from the selected channel. Next, click on analysis, then coherence for continuous to analyze the coherence for two LFPs from the left and right sides of the MC.Then, click on analysis followed by correlation with continuous variables to analyze the correlation between two LFPs from the left and right sides of the MC.Once done, click on results, then numerical results to save the power spectrum density, coherence, and correlation with a dot XLS file name extension.

To analyze correlations between the spike and LFP, click on file, then import data and Blackrock file to open the continuous signal data and spike data. Then, click on analysis and coherence analysis to analyze the coherence between the spikes and LFP from the selected channel. Finally, click on results, then numerical results to save the results of the spike field coherence with a dot XLS file name extension.

The valley width and waveform duration of the units in the MC of the mouse showed that both the valley width and waveform duration of the MC putative pyramidal neurons in mice are higher than those of the putative inner neurons. The cross corellogram between putative pyramidal neurons and interneurons indicated that the putative pyramidal neurons spiking occurs before the putative inner neurons with a window of 18 milliseconds. In the LFP analysis, the LFPs of the left and right MC in normal mice were similar in the power spectrum, suggesting synchronized activities between the left and right MC.Further, the coherence and correlation between the left and right MC were calculated.

The curve of spike field coherence in the MC of a normal mouse showed a stronger low gamma coherence for the putative interneurons compared to the pyramidal neurons.