Dataset Information

PatcherBot: a single-cell electrophysiology robot for adherent cells and brain slices.

ABSTRACT: OBJECTIVE:Intracellular patch-clamp electrophysiology, one of the most ubiquitous, high-fidelity techniques in biophysics, remains laborious and low-throughput. While previous efforts have succeeded at automating some steps of the technique, here we demonstrate a robotic 'PatcherBot' system that can perform many patch-clamp recordings sequentially, fully unattended. APPROACH:Comprehensive automation is accomplished by outfitting the robot with machine vision, and cleaning pipettes instead of manually exchanging them. MAIN RESULTS:the PatcherBot can obtain data at a rate of 16 cells per hour and work with no human intervention for up to 3?h. We demonstrate the broad applicability and scalability of this system by performing hundreds of recordings in tissue culture cells and mouse brain slices with no human supervision. Using the PatcherBot, we also discovered that pipette cleaning can be improved by a factor of three. SIGNIFICANCE:The system is potentially transformative for applications that depend on many high-quality measurements of single cells, such as drug screening, protein functional characterization, and multimodal cell type investigations.

SUBMITTER: Kolb I

PROVIDER: S-EPMC7144284 | biostudies-literature | 2019 Aug

REPOSITORIES: biostudies-literature

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PatcherBot: a single-cell electrophysiology robot for adherent cells and brain slices.

Kolb Ilya I Landry Corey R CR Yip Mighten C MC Lewallen Colby F CF Stoy William A WA Lee John J Felouzis Amanda A Yang Bo B Boyden Edward S ES Rozell Christopher J CJ Forest Craig R CR

Journal of neural engineering 20190410 4

<h4>Objective</h4>Intracellular patch-clamp electrophysiology, one of the most ubiquitous, high-fidelity techniques in biophysics, remains laborious and low-throughput. While previous efforts have succeeded at automating some steps of the technique, here we demonstrate a robotic 'PatcherBot' system that can perform many patch-clamp recordings sequentially, fully unattended.<h4>Approach</h4>Comprehensive automation is accomplished by outfitting the robot with machine vision, and cleaning pipettes ...[more]

PMID: 30970335

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Project description:Although patch pipettes were initially designed to record extracellularly the elementary current events from muscle and neuron membranes, the whole-cell and loose cell-attached recording configurations proved to be useful tools for examination of signalling within and between nerve cells. In this Paton Prize Lecture, I will initially summarize work on electrical signalling within single neurons, describing communication between the dendritic compartments, soma and nerve terminals via forward- and backward-propagating action potentials. The newly discovered dendritic excitability endows neurons with the capacity for coincidence detection of spatially separated subthreshold inputs. When these are occurring during a time window of tens of milliseconds, this information is broadcast to other cells by the initiation of bursts of action potentials (AP bursts). The occurrence of AP bursts critically impacts signalling between neurons that are controlled by target-cell-specific transmitter release mechanisms at downstream synapses even in different terminals of the same neuron. This can, in turn, induce mechanisms that underly synaptic plasticity when AP bursts occur within a short time window, both presynaptically in terminals and postsynaptically in dendrites. A fundamental question that arises from these findings is: 'what are the possible functions of active dendritic excitability with respect to network dynamics in the intact cortex of behaving animals?' To answer this question, I highlight in this review the functional and anatomical architectures of an average cortical column in the vibrissal (whisker) field of the somatosensory cortex (vS1), with an emphasis on the functions of layer 5 thick-tufted cells (L5tt) embedded in this structure. Sensory-evoked synaptic and action potential responses of these major cortical output neurons are compared with responses in the afferent pathway, viz. the neurons in primary somatosensory thalamus and in one of their efferent targets, the secondary somatosensory thalamus. Coincidence-detection mechanisms appear to be implemented in vivo as judged from the occurrence of AP bursts. Three-dimensional reconstructions of anatomical projections suggest that inputs of several combinations of thalamocortical projections and intra- and transcolumnar connections, specifically those from infragranular layers, could trigger active dendritic mechanisms that generate AP bursts. Finally, recordings from target cells of a column reveal the importance of AP bursts for signal transfer to these cells. The observations lead to the hypothesis that in vS1 cortex, the sensory afferent sensory code is transformed, at least in part, from a rate to an interval (burst) code that broadcasts the occurrence of whisker touch to different targets of L5tt cells. In addition, the occurrence of pre- and postsynaptic AP bursts may, in the long run, alter touch representation in cortex.

Dataset Information

PatcherBot: a single-cell electrophysiology robot for adherent cells and brain slices.

Publications

PatcherBot: a single-cell electrophysiology robot for adherent cells and brain slices.

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