Abstract
Monitoring voltage dynamics in defined neurons deep in the brain is critical for unraveling the function of neuronal circuits but is challenging due to the limited performance of existing tools. In particular, while genetically encoded voltage indicators have shown promise for optical detection of voltage transients, many indicators exhibit low sensitivity when imaged under two-photon illumination. Previous studies thus fell short of visualizing voltage dynamics in individual neurons in single trials. Here, we report ASAP2s, a novel voltage indicator with improved sensitivity. By imaging ASAP2s using random-access multi-photon microscopy, we demonstrate robust single-trial detection of action potentials in organotypic slice cultures. We also show that ASAP2s enables two-photon imaging of graded potentials in organotypic slice cultures and in Drosophila. These results demonstrate that the combination of ASAP2s and fast two-photon imaging methods enables detection of neural electrical activity with subcellular spatial resolution and millisecond-timescale precision.
Highlights
Neurons represent, process, and propagate information by controlling the potential across their plasma membrane
Having evaluated ASAP2s in vitro and in vivo, we deploy this new indicator to image voltage dynamics using randomaccess multi-photon microscopy in organotypic hippocampal slice cultures, where we demonstrate the ability of ASAP2s to detect action potentials, subthreshold depolarizations, and hyperpolarizations in individual cells in single trials
The faster kinetics of the ASAP indicators compared to ArcLight enabled a shorter time to peak when reporting cardiac action potentials (Figure 2E–F, Figure 2—figure supplement 1). These results demonstrate that ASAP2s can image cardiac action potentials, as previously shown with other voltage indicators in vitro (Kaestner et al, 2015; Tian et al, 2011; Leyton-Mange et al, 2014; Chang Liao et al, 2015; Werley et al, 2017) and in vivo (Chang Liao et al, 2015; Tsutsui et al, 2010; Hou et al, 2014)
Summary
Process, and propagate information by controlling the potential across their plasma membrane. In contrast to electrode-based methods, these optophysiological indicators allow monitoring without placement of physical probes near or in the neurons of interest. They can enable easier and less invasive measurement of activity from individual neurons and from their subcellular compartments such as axons and dendrites. Encoded calcium indicators are commonly used to detect the forms of neuronal activity that trigger calcium flux into neurons (Grienberger and Konnerth, 2012; Tian et al, 2012; Lin and Schnitzer, 2016).
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