Optogenetic tools enable the causal examination of how specific cell types contribute to brain circuit functions. A long-standing question is whether it is possible to independently activate two distinct neural populations in mammalian brain tissue. Such a capability would enable the examination of how different synapses or pathways interact to support computation. Here we report two new channelrhodopsins, Chronos and Chrimson, obtained through the de novo sequencing and physiological characterization of opsins from over 100 species of algae. Chrimson is 45 nm red-shifted relative to any previous channelrhodopsin, important for scenarios where red light would be preferred; we show minimal visual system mediated behavioral artifact in optogenetically stimulated Drosophila. Chronos has faster kinetics than any previous channelrhodopsin, yet is effectively more light-sensitive. Together, these two reagents enable crosstalk-free two-color activation of neural spiking and downstream synaptic transmission in independent neural populations in mouse brain slice.

Ring attractors are a class of recurrent networks hypothesized to underlie the representation of heading direction. Such network structures, schematized as a ring of neurons whose connectivity depends on their heading preferences, can sustain a bump-like activity pattern whose location can be updated by continuous shifts along either turn direction. We recently reported that a population of fly neurons represents the animal's heading via bump-like activity dynamics. We combined two-photon calcium imaging in head-fixed flying flies with optogenetics to overwrite the existing population representation with an artificial one, which was then maintained by the circuit with naturalistic dynamics. A network with local excitation and global inhibition enforces this unique and persistent heading representation. Ring attractor networks have long been invoked in theoretical work; our study provides physiological evidence of their existence and functional architecture.

Many animals rely on an internal heading representation when navigating in varied environments [1][2][3][4][5][6][7][8][9][10] . How this representation is linked to the sensory cues defining different surroundings is unclear. In the fly brain, heading is represented by 'compass neurons' that innervate a ring-shaped structure, the ellipsoid body 3,11,12 . Each compass neuron receives inputs from visual-feature-selective 'ring neurons' [13][14][15][16] , providing the ideal substrate for the extraction of directional information from a visual scene. We combine two-photon calcium imaging and optogenetics in tethered flying flies with circuit modeling to show how the correlated activity of compass and visual neurons drives plasticity [17][18][19][20][21][22] , that flexibly transforms two-dimensional visual cues into a stable heading representation. We also describe how this plasticity enables the fly to convert a partial heading representation established from orienting within part of a novel setting into a complete heading representation. Our results provide mechanistic insight into memoryrelated computations essential for flexible navigation in varied surroundings.Internal representations of an animal's spatial relationship to its surroundings are essential for flexible navigation 3,[8][9][10] . Although these representations must be stable to be useful for planning and goal-oriented behavior, they must also adapt to changes in environmental and behavioral contexts. Indeed, the representations provided by head direction cells, grid cells, Reprints and permissions information is available at www.nature.com/reprints.Users may view, print, copy, and download text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use:http:// www.nature.com/authors/editorial_policies/license.html#terms