Robotic navigation to subcortical neural tissue for intracellular electrophysiology in vivo

Abstract

In vivo studies of neurophysiology using the whole cell patch-clamp technique enable exquisite access to both intracellular dynamics and cytosol of cells in the living brain but are underrepresented in deep subcortical nuclei because of fouling of the sensitive electrode tip. We have developed an autonomous method to navigate electrodes around obstacles such as blood vessels after identifying them as a source of contamination during regional pipette localization (RPL) in vivo. In mice, robotic navigation prevented fouling of the electrode tip, increasing RPL success probability 3 mm below the pial surface to 82% ( = 72/88) over traditional, linear localization (25%, = 24/95), and resulted in high-quality thalamic whole cell recordings with average access resistance (32.0 MΩ) and resting membrane potential (-62.9 mV) similar to cortical recordings in isoflurane-anesthetized mice. Whole cell yield improved from 1% ( = 1/95) to 10% ( = 9/88) when robotic navigation was used during RPL. This method opens the door to whole cell studies in deep subcortical nuclei, including multimodal cell typing and studies of long-range circuits. This work represents an automated method for accessing subcortical neural tissue for intracellular electrophysiology in vivo. We have implemented a novel algorithm to detect obstructions during regional pipette localization and move around them while minimizing lateral displacement within brain tissue. This approach leverages computer control of pressure, manipulator position, and impedance measurements to create a closed-loop platform for pipette navigation in vivo. This technique enables whole cell patching studies to be performed throughout the living brain.

Compensation of physiological motion enables high-yield whole-cell recordingin vivo

“…This suggests that the pipettes were slightly 393 indenting the surface of the cell's membranes in this study, immediately prior to gigasealing. 394 However, this is the same decrease in current that we have used in all previous studies with the 395 Autopatcher, both in the cortex and the thalamus (Kodandaramaiah et al, 2012;Stoy et al, 2017). 396…”

“…The motion compensation hardware and software described here is part of an automated suite of 410 tools for neuroscientists that enables the recording of whole-cell electrophysiology in previously 411 difficult-to-access tissue (Stoy et al, 2017). Additionally, the combination of this motion 412 compensation procedure and robotic localization during regional pipette localization with pipette The results of this optimization can be seen in Supplementary Figure 2.…”

“…To evaluate the success of the feedforward system at approximating the finite impulse response of 288 physiological sources of motion in the brain, whole-cell patch clamp trials were performed in the 289 cortex (n=7 mice, 70 trials) and in the thalamus (n=8 mice, 76 trials). Neuron hunting was 290 performed as we published previously (Kodandaramaiah et al, 2012;Stoy et al, 2017). The 291 forward progress of the pipette was halted when the pipette resistance increased above threshold 292 and the peak-to-peak amplitude of the resistance fluctuations was above a user-determined 293 threshold, indicating that the pipette was near a cell.…”

“…As in our previous work (Stoy et al, 2017) where we demonstrated an algorithm for navigation 157 around blood vessels during regional pipette localization (RPL), resistance measurements were 158 performed throughout RPL, rather than only before and after localization. Briefly, during 159 localization to the region of interest (cortex or thalamus for this study), the pipette resistance was 160 recorded in the aCSF on the surface of the tissue by applying a 20 mV amplitude, 128 Hz square 161 wave at 50% duty cycle and stored as a moving average of four periods.…”

Engineering microscale systems for fully autonomous intracellular neural interfaces

“…Recently, automated systems have been developed to reduce the "art" in the process of intracellular recording in vivo [10][11][12] . Kodandaramaiah and colleagues 10 developed a closedloop control system that used a temporal sequence of electrode impedance changes as a feedback signal to automate movement of electrode and whole-cell patching of neurons in the cortex and hippocampus of anesthetized, head-fixed mice. They recently improved the algorithm to automate localization of a pipette to deep cortical nuclei through autonomous detection and lateral navigation around blood vessels and obtained high-yield (10%) thalamic whole-cell recordings 13 . Desai et al 12 and Ota et al 11 developed similar algorithms to automate cortical whole-cell patching in awake, head-fixed, behaving mice and sharp micropipette recording in anesthetized, headfixed mice, respectively.…”