Integrating multi-unit electrophysiology and plastic culture dishes for network neuroscience

Abstract

The electrophysiological characterisation of cultured neurons is of paramount importance for drug discovery, safety pharmacology and basic research in the neurosciences. Technologies offering low cost, low technical complexity and potential for scalability towards high-throughput electrophysiology on in vitro neurons would be advantageous, in particular for screening purposes. Here we describe a plastic culture substrate supporting low-complexity multi-unit loose-patch recording and stimulation of developing networks while retaining manufacturability compatible with low-cost and large-scale production. Our hybrid polydimethylsilane (PDMS)-on-polystyrene structures include chambers (6 mm in diameter) and microchannels (25 microm x 3.7 microm x 1 mm) serving as substrate-embedded recording pipettes. Somas are plated and retained in the chambers due to geometrical constraints and their processes grow along the microchannels, effectively establishing a loose-patch configuration without human intervention. We demonstrate that off-the-shelf voltage-clamp, current-clamp and extracellular amplifiers can be used to record and stimulate multi-unit activity with the aid of our dishes. Spikes up to 50 pA in voltage-clamp and 300 microV in current-clamp modes are recorded in sparse and bursting activity patterns characteristic of 1 week-old hippocampal cultures. Moreover, spike sorting employing principal component analysis (PCA) confirms that single microchannels support the recording of multiple neurons. Overall, this work suggests a strategy to endow conventional culture plasticware with added functionality to enable cost-efficient network electrophysiology.

Large Extracellular Spikes Recordable From Axons in Microtunnels

“…In 2009, Dworak and Wheeler [8] analyzed the relationship between signal amplitudes and electrode impedances and suggested that signals in microtunnels could be as large as 2.4 mV as measured between an electrode placed within the tunnel and a distant reference electrode that is bathed in media electrically coupled to the wells on either side of the tunnels. The Claverol-Tinturé lab reported large signals (up to 8 mV for a 2 × 6 μm tunnel) recorded differentially from two otherwise isolated wells connected by a tunnel within which axons grew and propagated signals [14], [15]. This group further reported signals from electrodes within the tunnel (referenced to a well) of up to 1.5 mV.…”

“…Here we expand upon the range of potential tunnel dimensions where this effect can be seen with conventionally sized microtunnels (3 μm × 10 μm, most commonly reported and commercially available) and commercially available microelectrode arrays (most commonly with 30 um diameter electrodes), beyond the specially fabricated electrodes designed to intersect the wells reported previously [8], [14], [15]. Further, we report amplitudes (as large as 4.5 mV) that often exceed what has been previously reported for electrodes in tunnels, with correspondingly large signal to noise ratios (up to 450).…”

In vitro Verification of a 3-D Regenerative Neural Interface Design: Examination of Neurite Growth and Electrical Properties Within a Bifurcating Microchannel Structure

“…This builds on in vitro studies that have described neurite growth into enclosed microchannels, as well as on previous reports on the recording of neuroelectric activity from within microchannels [7]- [11]. Microchannels ranging from 2.5 to 20 m wide are examined to see if this has an influence on the ability to counter fasciculation and to see if supporting evidence can be found for hypothesized mechanisms of initiating separated growth.…”

Novel Aspects of Neuronal Differentiation In Vitro and Monitoring with Advanced Biosensor Tools

“…Moreover, fabrication costs can be reduced to a minimum by simplifying fabrication procedures [74], materials [75] and increasing quantities of the devices produced.…”