In-cell recording and stimulation by engulfment mechanisms

Abstract

Neuroscientists have long been using microelectrodes to record and stimulate neural activity—both in vitro and in vivo. On one end of the spectrum of electrode-based techniques are the sharp-glass and patch micropipette electrodes; on the other end are dense arrays of metal-based microelectrodes. Glass micropipette electrodes enable intracellular recording of action potentials and synaptic potentials with excellent signal-to-noise ratio, but because of their bulkiness, they allow for the recording and stimulation of only several neurons at a time. In addition, the injury inflicted on the cell plasma membrane during electrode entry and recording limits the duration of the recording session, usually to a small number of hours at most. By contrast, multielectrode devices are able to record and stimulate much larger populations of neurons for durations of weeks and even months. This is made possible due to fabrication technologies that allow for a scalable design of hundreds or even thousands of electrodes. These devices, however, have been able to provide only extracellular recording and stimulation with limited signal-to-noise ratio due to the extracellular positioning of the electrode in respect to the neuron’s plasma membrane. The inability to record intracellular signals from many neurons and for long periods of time has thus far prevented neuroscience from answering the most basic and interesting questions regarding learning and memory in large populations of neurons. This is because the vast majority of neurons in complex nervous systems are usually “silent” and will generate an action potential only when their complex synaptic inputs integrate appropriately. We are therefore blind to the rich milieu of synaptic interactions, synaptic plasticity, and subthreshold network oscillations that reflect the state of the studied nervous system. This chapter describes a recently developed technique termed in-cell recording. This technique yielded for the first time simultaneous, multisite, long-term recordings of action potentials and subthreshold synaptic potentials with matching quality and signal-to-noise ratio of conventional intracellular glass electrodes and the scalability of fabricated multielectrode devices. The in-cell recording and stimulation technique makes use of an array of cell-noninvasive micrometer-size protruding gold mushroom-shaped microelectrodes (gMμEs). The key to the multielectrode in-cell recording approach is the outcome of three converging cell-biological principles: (a) the activation of endocytotic-like mechanisms in which the cultured cells are induced to actively engulf gMμEs that protrude from the substrate, (b) the generation of high seal resistance between the cell’s membrane and the engulfed gMμE, and (c) the localization of ionic channels (ohmic conductance) in the plasma membrane that faces the gMμE. We will describe the electrical, ultrastructural, and cell-biological properties of the interface between the cells and the gMμEs and provide the reader with a digest of the published studies carried out for the development of this technique.