In Vitro Testing of Voltage Indicators: Archon1, ArcLightD, ASAP1, ASAP2s, ASAP3b, Bongwoori-Pos6, BeRST1, FlicR1, and Chi-VSFP-Butterfly.

Genetically encoded voltage indicators (GEVIs) could potentially be used for mapping neural circuits at the plane of synaptic potentials and plateau potentials-two blind spots of GCaMP-based imaging. In the last year alone, several laboratories reported significant breakthroughs in the quality of GEVIs and the efficacy of the voltage imaging equipment. One major obstacle of using well performing GEVIs in the pursuit of interesting biological data is the process of transferring GEVIs between laboratories, as their reported qualities (e.g., membrane targeting, brightness, sensitivity, optical signal quality) are often difficult to reproduce outside of the laboratory of the GEVI origin. We have tested eight available GEVIs (Archon1, ArcLightD, ASAP1, ASAP2s, ASAP3b, Bongwoori-Pos6, FlicR1, and chi-VSFP-Butterfly) and two voltage-sensitive dyes (BeRST1 and di-4-ANEPPS). We used the same microscope, lens, and optical detector, while the light sources were interchanged. GEVI voltage imaging was attempted in the following three preparations: (1) cultured neurons, (2) HEK293 cells, and (3) mouse brain slices. Systematic measurements were successful only in HEK293 cells and brain slices. Despite the significant differences in brightness and dynamic response (ON rate), all tested indicators produced reasonable optical signals in brain slices and solid in vitro quality properties, in the range initially reported by the creator laboratories. Side-by-side comparisons between GEVIs and organic dyes obtained in HEK293 cells and brain slices by a "third party" (current data) will be useful for determining the right voltage indicator for a given research application.

Corrigendum: Single-Neuron Level One-Photon Voltage Imaging With Sparsely Targeted Genetically Encoded Voltage Indicators.

[This corrects the article DOI: 10.3389/fncel.2019.00039.].

Single-Neuron Level One-Photon Voltage Imaging With Sparsely Targeted Genetically Encoded Voltage Indicators.

Voltage imaging of many neurons simultaneously at single-cell resolution is hampered by the difficulty of detecting small voltage signals from overlapping neuronal processes in neural tissue. Recent advances in genetically encoded voltage indicator (GEVI) imaging have shown single-cell resolution optical voltage recordings in intact tissue through imaging naturally sparse cell classes, sparse viral expression, soma restricted expression, advanced optical systems, or a combination of these. Widespread sparse and strong transgenic GEVI expression would enable straightforward optical access to a densely occurring cell type, such as cortical pyramidal cells. Here we demonstrate that a recently described sparse transgenic expression strategy can enable single-cell resolution voltage imaging of cortical pyramidal cells in intact brain tissue without restricting expression to the soma. We also quantify the functional crosstalk in brain tissue and discuss optimal imaging rates to inform future GEVI experimental design.

Mechanisms of Spontaneous Electrical Activity in the Developing Cerebral Cortex-Mouse Subplate Zone.

Subplate (SP) neurons exhibit spontaneous plateau depolarizations mediated by connexin hemichannels. Postnatal (P1-P6) mice show identical voltage pattern and drug-sensitivity as observed in slices from human fetal cortex; indicating that the mouse is a useful model for studying the cellular physiology of the developing neocortex. In mouse SP neurons, spontaneous plateau depolarizations were insensitive to blockers of: synaptic transmission (glutamatergic, GABAergic, or glycinergic), pannexins (probenecid), or calcium channels (mibefradil, verapamil, diltiazem); while highly sensitive to blockers of gap junctions (octanol), hemichannels (La3+, lindane, Gd3+), or glial metabolism (DLFC). Application of La3+ (100 µM) does not exert its effect on electrical activity by blocking calcium channels. Intracellular application of Gd3+ determined that Gd3+-sensitive pores (putative connexin hemichannels) reside on the membrane of SP neurons. Immunostaining of cortical sections (P1-P6) detected connexins 26, and 45 in neurons, but not connexins 32 and 36. Vimentin-positive glial cells were detected in the SP zone suggesting a potential physiological interaction between SP neurons and radial glia. SP spontaneous activity was reduced by blocking glial metabolism with DFLC or by blocking purinergic receptors by PPADS. Connexin hemichannels and ATP release from vimentin-positive glial cells may underlie spontaneous plateau depolarizations in the developing mammalian cortex.