Remote and precisely controlled activation of the brain is a fundamental challenge in the development of brain–machine interfaces for neurological treatments. Low-frequency ultrasound stimulation can be used to modulate neuronal activity deep in the brain, especially after expressing ultrasound-sensitive proteins. But so far, no study has described an ultrasound-mediated activation strategy whose spatiotemporal resolution and acoustic intensity are compatible with the mandatory needs of brain–machine interfaces, particularly for visual restoration. Here we combined the expression of large-conductance mechanosensitive ion channels with uncustomary high-frequency ultrasonic stimulation to activate retinal or cortical neurons over millisecond durations at a spatiotemporal resolution and acoustic energy deposit compatible with vision restoration. The in vivo sonogenetic activation of the visual cortex generated a behaviour associated with light perception. Our findings demonstrate that sonogenetics can deliver millisecond pattern presentations via an approach less invasive than current brain–machine interfaces for visual restoration.
Remote, precisely controlled activation of the brain is a fundamental challenge in the development of brain-machine interfaces providing feasible rehabilitation strategies for neurological disorders. Low-frequency ultrasound stimulation can be used to modulate neuronal activity deep in the brain1–7, but this approach lacks spatial resolution and cellular selectivity and loads the brain with high levels of acoustic energy. The combination of the expression of ultrasound-sensitive proteins with ultrasound stimulation (‘sonogenetic stimulation’) can provide cellular selectivity and higher sensitivity, but such strategies have been subject to severe limitations in terms of spatiotemporal resolution in vivo8–10, precluding their use for real-life applications. We used the expression of large-conductance mechanosensitive ion channels (MscL) with high-frequency ultrasonic stimulation for a duration of milliseconds to activate neurons selectively at a relatively high spatiotemporal resolution in the rat retina ex vivo and the primary visual cortex of rodents in vivo. This spatiotemporal resolution was achieved at low energy levels associated with negligible tissue heating and far below those leading to complications in ultrasound neuromodulation6,11. We showed, in an associative learning test, that sonogenetic stimulation of the visual cortex generated light perception. Our findings demonstrate that sonogenetic stimulation is compatible with millisecond pattern presentation for visual restoration at the cortical level. They represent a step towards the precise transfer of information over large distances to the cortical and subcortical regions of the brain via an approach less invasive than that associated with current brain-machine interfaces and with a wide range of applications in neurological disorders.
Optogenetics has revolutionized neurosciences by allowing fine control of neuronal activity. An important aspect for this control is assessing the activation and/or adjusting the stimulation, which requires imaging the entire volume of optogenetically-induced neuronal activity. An ideal technique for this aim is fUS imaging, which allows one to generate brain-wide activation maps with submesoscopic spatial resolution. However, optical stimulation of the brain with blue light might lead to non-specific activations at high irradiances. fUS imaging of optogenetic activations can be obtained at these wavelengths using lower light power (< 2mW) but it limits the depth of directly activatable neurons from the cortical surface. Our main goal was to report that we can detect specific optogenetic activations in V1 even in deep layers following stimulation at the cortical surface. Here, we show the possibility to detect deep optogenetic activations in anesthetized rats expressing the red-shifted opsin ChrimsonR in V1 using fUS imaging. We demonstrate the optogenetic specificity of these activations and their neuronal origin with electrophysiological recordings. Finally, we show that the optogenetic response initiated in V1 spreads to downstream (LGN) and upstream (V2) visual areas.
Over the last 15 years, optogenetics has changed fundamental research in neuroscience and is now reaching toward therapeutic applications. Vision restoration strategies using optogenetics are now at the forefront of these new clinical opportunities. But applications to human patients suffering from retinal diseases leading to blindness raise important concerns on the long-term functional expression of optogenes and the efficient signal transmission to higher visual centers. Here, we demonstrate in non-human primates continued expression and functionality at the retina level ∼20 months after delivery of our construct. We also performed in vivo recordings of visually evoked potentials in the primary visual cortex of anesthetized animals. Using synaptic blockers, we isolated the in vivo cortical activation resulting from the direct optogenetic stimulation of primate retina. In conclusion, our work indicates long-term transgene expression and transmission of the signal generated in the macaque retina to the visual cortex, two important features for future clinical applications.
After revolutionizing neuroscience, optogenetic therapy has entered successfully in clinical trials for restoring vision to blind people with degenerative eye diseases, such as retinitis pigmentosa. These clinical trials still have to evaluate the visual acuity achieved by patients and to determine if it reaches its theoretical limit extrapolated from ex vivo experiments. Different strategies are developed in parallel to reduce required light levels and improve information processing by targeting various cell types. For patients with vision loss due to optic atrophy, as in the case of glaucoma, optogenetic cortical stimulation is hampered by light absorption and scattering by the brain tissue. By contrast, ultrasound waves can diffuse widely through the dura mater and the brain tissue as indicated by ultrasound imaging. Based on our recent results in rodents, we propose the sonogenetic therapy relying on activation of the mechanosensitive channel as a very promising vision restoration strategy with a suitable spatiotemporal resolution. Genomic approaches may thus provide efficient brain machine interfaces for sight restoration.
To better understand how the brain allows primates to perform various set of tasks, the ability to simultaneously record the activity of the brain at multiple temporal and spatial scales is challenging but necessary. In non-human primates, combined fMRI and electrophysiological recordings have not disentangle the contributions of spiking activity to the neurovascular response. Here, we combined functional ultrasound imaging (fUS) of cerebral blood volume (CBV) and recording of single-unit activities (SUA) in visual and fronto-medial cortices of behaving macaques. We computed task-induced and SUA-induced CBV activation maps. We demonstrate that SUA provides a significant estimate of the neurovascular response below the typical fMRI voxel spatial resolution of 2mm3. Furthermore, our results also show that single unit and CBV activities are statistically uncorrelated during the resting states but correlate during behaving tasks. Conversely, during the resting states, CBV activities across known connected brain areas are correlated but decorrelate at the onset of the tasks as expected if participating in the default mode network (DMN). These results have important implications for interpreting functional imaging findings collected with fMRI or fUS while one constructs inferences of spiking activities during resting-state or while primates perform tasks.
Optogenetic stimulation of the primary visual cortex (V1) is a promising therapy for sight restoration, but it remains unclear what total cerebral volume is activated after surface stimulation. In this study, we expressed the red-shifted opsin ChrimsonR in excitatory neurons within V1 in rats, and used the fine spatial resolution provided by functional ultrasound imaging (fUS) over the whole depth of the brain to investigate the brain response to focal surface stimulation. We observed optogenetic activation of a high proportion of the volume of V1. Extracellular recordings confirmed the neuronal origin of this activation. Moreover, neuronal responses were even located in deep layers under conditions of low irradiance, spreading to the LGN and V2, consistent with a normal visual information process. This study paves the way for the use of optogenetics for cortical therapies, and highlights the value of coupling fUS with optogenetics.
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