ArticleNature methods2026
A versatile platform for two-photon neuronal population voltage imaging across cortical depths.
Article in Nature methods, 2026. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Not yet cited in PubMed.
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Abstract
Genetically encoded voltage indicators have emerged as a tool for resolving neuronal spiking activity with high spatiotemporal resolution within genetically specific populations; however, their fast temporal dynamics, low signal-to-noise ratio (SNR) and fast photobleaching have posed substantial challenges limiting their broader utility and, together with suboptimal optical acquisition schemes, preventing their efficient scale-up to larger neuronal populations. Here we introduce a versatile, scalable, spatiotemporally and energetically efficient two-photon optical imaging system scheme based on a flexible lateral-temporal multiplexing (FlatMux) platform. We demonstrate FlatMux's capability and its flexible reconfigurability for meeting different recording requirements. This includes a large field-of-view mode, a 2-kHz high-speed mode, a deep-tissue imaging mode allowing recordings of cortical spiking activity at up to 500-µm depth, a dual-plane imaging mode allowing for simultaneous recording of population spiking activity of neurons in cortical L2/3 and L4, and a high-SNR imaging mode for recording of subthreshold neuronal activity and high-SNR spiking activity, all while minimizing pixel crosstalk and bleaching. Thus, FlatMux meets the challenging demands of multiphoton voltage imaging across the mammalian cortex and can be expected to enable a range of studies of complex brain functions at single-spike and single-trial level for large neuronal populations.
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