Optogenetics publications

Information is transmitted between brain regions through the release of neurotransmitters from long-range projecting axons. Understanding how the activity of such long-range connections contributes to behavior requires efficient methods for reversibly manipulating their function. Chemogenetic and optogenetic tools, acting through endogenous G-protein-coupled receptor pathways, can be used to modulate synaptic transmission, but existing tools are limited in sensitivity, spatiotemporal precision or spectral multiplexing capabilities. Here we systematically evaluated multiple bistable opsins for optogenetic applications and found that the Platynereis dumerilii ciliary opsin ( Pd CO) is an efficient, versatile, light-activated bistable G-protein-coupled receptor that can suppress synaptic transmission in mammalian neurons with high temporal precision in vivo. Pd CO has useful biophysical properties that enable spectral multiplexing with other optogenetic actuators and reporters. We demonstrate that Pd CO can be used to conduct reversible loss-of-function experiments in long-range projections of behaving animals, thereby enabling detailed synapse-specific functional circuit mapping.

Dopamine neurons in the ventral tegmental area support intracranial self-stimulation (ICSS), yet the cognitive representations underlying this phenomenon remain unclear. Here, 20-Hz stimulation of dopamine neurons, which approximates a physiologically relevant prediction error, was not sufficient to support ICSS beyond a continuously reinforced schedule and did not endow cues with a general or specific value. However, 50-Hz stimulation of dopamine neurons was sufficient to drive robust ICSS and was represented as a specific reward to motivate behavior. The frequency dependence of this effect is due to the rate (not the number) of action potentials produced by dopamine neurons, which differently modulates dopamine release downstream.

The feedback projections from cortical layer 6 (L6CT) to the sensory thalamus have long been implicated in playing a primary role in gating sensory signaling but remain poorly understood. To causally elucidate the full range of effects of these projections, we targeted silicon probe recordings to the whisker thalamocortical circuit of awake mice selectively expressing Channelrhodopsin-2 in L6CT neurons. Through optogenetic manipulation of L6CT neurons, multi-site electrophysiological recordings, and modeling of L6CT circuitry, we establish L6CT neurons as dynamic modulators of ongoing spiking in the ventral posteromedial nucleus of the thalamus (VPm), either suppressing or enhancing VPm spiking depending on L6CT neurons’ firing rate and synchrony. Differential effects across the cortical excitatory and inhibitory sub-populations point to an overall influence of L6CT feedback on cortical excitability that could have profound implications for regulating sensory signaling across a range of ethologically relevant conditions.

Temporally coordinated neural activity is central to nervous system function and purposeful behavior. Still, there is a paucity of evidence demonstrating how this coordinated activity within cortical and subcortical regions governs behavior. We investigated this between the primary motor (M1) and contralateral cerebellar cortex as rats learned a neuroprosthetic/brain-machine interface (BMI) task. In neuroprosthetic task, actuator movements are causally linked to M1 “direct” neurons that drive the decoder for successful task execution. However, it is unknown how task-related M1 activity interacts with the cerebellum. We observed a notable 3 to 6 hertz coherence that emerged between these regions’ local field potentials (LFPs) with learning that also modulated task-related spiking. We identified robust task-related indirect modulation in the cerebellum, which developed a preferential relationship with M1 task–related activity. Inhibiting cerebellar cortical and deep nuclei activity through optogenetics led to performance impairments in M1-driven neuroprosthetic control. Together, these results demonstrate that cerebellar influence is necessary for M1-driven neuroprosthetic control.

Stimulus-specific adaptation (SSA), the reduction of neural activity to a common stimulus that does not generalize to other, rare stimuli, is an essential property of our brain. Although well characterized in adults, it is still unknown how it develops during adolescence and what neuronal circuits are involved. Using in vivo electrophysiology and optogenetics in the lemniscal pathway of the mouse auditory system, we observed SSA to be stable from postnatal day 20 (P20) in the inferior colliculus, to develop until P30 in the auditory thalamus and even later in the primary auditory cortex (A1). We found this maturation process to be experience-dependent in A1 but not in thalamus and to be related to alterations in deep but not input layers of A1. We also identified corticothalamic projections to be implicated in thalamic SSA development. Together, our results reveal different circuits underlying the sequential SSA maturation and provide a unique perspective to understand predictive coding and surprise across sensory systems.

The auditory steady-state response (ASSR) is a cortical oscillation induced by trains of 40 Hz acoustic stimuli. While the ASSR has been widely used in clinic measurement, the underlying neural mechanism remains poorly understood. In this study, we investigated the contribution of different stages of auditory thalamocortical pathwaymedial geniculate body (MGB), thalamic reticular nucleus (TRN), and auditory cortex (AC)to the generation and regulation of 40 Hz ASSR in C57BL/6 mice of both sexes. We found that the neural response synchronizing to 40 Hz sound stimuli was most prominent in the GABAergic neurons in the granular layer of AC and the ventral division of MGB (MGBv), which were regulated by optogenetic manipulation of TRN neurons. Behavioral experiments confirmed that disrupting TRN activity has a detrimental effect on the ability of mice to discriminate 40 Hz sounds. These findings revealed a thalamocortical mechanism helpful to interpret the results of clinical ASSR examinations. Significance Statement Our study contributes to clarifying the thalamocortical mechanisms underlying the generation and regulation of the auditory steady-state response (ASSR), which is commonly used in both clinical and neuroscience research to assess the integrity of auditory function. Combining a series of electrophysiological and optogenetic experiments, we demonstrate that the generation of cortical ASSR is dependent on the lemniscal thalamocortical projections originating from the ventral division of medial geniculate body to the GABAergic interneurons in the granule layer of the auditory cortex. Furthermore, the thalamocortical process for ASSR is strictly regulated by the activity of thalamic reticular nucleus (TRN) neurons. Behavioral experiments confirmed that dysfunction of TRN would cause a disruption of mice's behavioral performance in the auditory discrimination task.

Cortical representations supporting many cognitive abilities emerge from underlying circuits comprised of several different cell types. However, cell type-specific contributions to rate and timing-based cortical coding are not well-understood. Here, we investigated the role of parvalbumin neurons in cortical complex scene analysis. Many complex scenes contain sensory stimuli which are highly dynamic in time and compete with stimuli at other spatial locations. Parvalbumin neurons play a fundamental role in balancing excitation and inhibition in cortex and sculpting cortical temporal dynamics; yet their specific role in encoding complex scenes via timing-based coding, and the robustness of temporal representations to spatial competition, has not been investigated. Here, we address these questions in auditory cortex of mice using a cocktail party-like paradigm, integrating electrophysiology, optogenetic manipulations, and a family of spike-distance metrics, to dissect parvalbumin neurons’ contributions towards rate and timing-based coding. We find that suppressing parvalbumin neurons degrades cortical discrimination of dynamic sounds in a cocktail party-like setting via changes in rapid temporal modulations in rate and spike timing, and over a wide range of time-scales. Our findings suggest that parvalbumin neurons play a critical role in enhancing cortical temporal coding and reducing cortical noise, thereby improving representations of dynamic stimuli in complex scenes.

Fear extinction is a form of inhibitory learning that suppresses the expression of aversive memories and plays a key role in the recovery of anxiety and trauma-related disorders. Here, using male mice, we identify a cerebello-thalamo-cortical pathway regulating fear extinction. The cerebellar fastigial nucleus (FN) projects to the lateral subregion of the mediodorsal thalamic nucleus (MD), which is reciprocally connected with the dorsomedial prefrontal cortex (dmPFC). The inhibition of FN inputs to MD in male mice impairs fear extinction in animals with high fear responses and increases the bursting of MD neurons, a firing pattern known to prevent extinction learning. Indeed, this MD bursting is followed by high levels of the dmPFC 4 Hz oscillations causally associated with fear responses during fear extinction, and the inhibition of FN-MD neurons increases the coherence of MD bursts and oscillations with dmPFC 4 Hz oscillations. Overall, these findings reveal a regulation of fear-related thalamo-cortical dynamics by the cerebellum and its contribution to fear extinction.

In humans, the level of consciousness is assessed by quantifying the spatiotemporal complexity of cortical responses using Perturbational Complexity Index (PCI) and related PCIst (st, state transitions). Here we validate PCIst in freely moving rats and mice by showing that it is lower in NREM sleep and slow wave anesthesia than in wake or REM sleep, as in humans. We then show that (1) low PCIst is associated with the occurrence of an OFF period of neuronal silence; (2) stimulation of deep, but not superficial, cortical layers leads to reliable PCIst changes across sleep/wake and anesthesia; (3) consistent PCIst changes are independent of which single area is being stimulated or recorded, except for recordings in mouse prefrontal cortex. These experiments show that PCIst can reliably measure vigilance states in unresponsive animals and support the hypothesis that it is low when an OFF period disrupts causal interactions in cortical networks.

The common marmoset has emerged as a key model in neuroscience. Marmosets are small in size, show great potential for genetic modification and exhibit complex behaviors. Thus, it is necessary to develop technology that enables monitoring and manipulation of the underlying neural circuits. Here, we describe a novel approach to record and optogenetically manipulate neural activity in awake, behaving marmosets. Our design utilizes a light-weight, 3D printed titanium chamber that can house several high-density silicon probes for semi-chronic recordings, while enabling simultaneous optogenetic stimulation. We demonstrate the application of our method in male marmosets by recording multi- and single-unit data from areas V1 and V6 with 192 channels simultaneously, and show that optogenetic activation of excitatory neurons in area V6 can influence behavior in a detection task. This method may enable future studies to investigate the neural basis of perception and behavior in the marmoset.

The cerebral cortex is spontaneously active during sleep, yet it is unclear how this global cortical activity is spatiotemporally organized, and whether such activity not only reflects sleep states but also contributes to sleep state switching. Here we report that cortex-wide calcium imaging in mice revealed distinct sleep stage-dependent spatiotemporal patterns of global cortical activity, and modulation of such patterns could regulate sleep state switching. In particular, elevated activation in the occipital cortical regions (including the retrosplenial cortex and visual areas) became dominant during rapid-eye-movement (REM) sleep. Furthermore, such pontogeniculooccipital (PGO) wave-like activity was associated with transitions to REM sleep, and optogenetic inhibition of occipital activity strongly promoted deep sleep by suppressing the NREM-to-REM transition. Thus, whereas subcortical networks are critical for initiating and maintaining sleep and wakefulness states, distinct global cortical activity also plays an active role in controlling sleep states.

Sensory responses typically vary depending on the recent history of sensory experience. This is essential for processes, including adaptation, efficient coding, and change detection. In the auditory cortex (AC), the short-term history dependence of sound-evoked (onset) responses has been well characterized. Yet many AC neurons also respond to sound terminations, and little is known about the history dependence of these "offset" responses, whether the short-term dynamics of onset and offset responses are correlated, or how these properties are distributed among cell types. Here we presented awake male and female mice with repeating noise burst stimuli while recording single-unit activity from primary AC. We identified parvalbumin and somatostatin interneurons through optotagging, and also separated narrow-spiking from broad-spiking units. We found that offset responses are typically less depressive than onset responses, and this result was robust to a variety of stimulus parameters, controls, measurement types, and selection criteria. Whether a cell's onset response facilitates or depresses does not predict whether its offset response facilitates or depresses. Cell types differed in the dynamics of their onset responses, and in the prevalence, but not the dynamics, of their offset responses. Finally, we clustered cells according to spiking responses and found that response clusters were associated with cell type. Each cluster contained cells of several types, but even within a cluster, cells often showed cell type-specific response dynamics. We conclude that onset and offset responses are differentially influenced by recent sound history, and discuss the implications of this for the encoding of ongoing sound stimuli.SIGNIFICANCE STATEMENT Sensory neuron responses depend on stimulus history. This history dependence is crucial for sensory processing, is precisely controlled at individual synapses and circuits, and is adaptive to the specific requirements of different sensory systems. In the auditory cortex, neurons respond to sound cessation as well as to sound itself, but how history dependence is used along this separate, "offset" information stream is unknown. We show that offset responses are more facilitatory than sound responses, even in neurons where sound responses depress. In contrast to sound onset responses, offset responses are absent in many cells, are relatively homogeneous, and show no cell type-specific differences in history dependence. Offset responses thus show unique response dynamics, suggesting their unique functions.

Neural activity exhibits oscillations, bursts, and resonance, enhancing responsiveness at preferential frequencies. For example, theta-frequency bursting and resonance in granule cells facilitate synaptic transmission and plasticity mechanisms at the input stage of the cerebellar cortex. However, whether theta-frequency bursting of Purkinje cells is involved in generating rhythmic behavior has remained neglected. We recorded and optogenetically modulated the simple and complex spike activity of Purkinje cells while monitoring whisker movements with a high-speed camera of awake, head-fixed mice. During spontaneous whisking, both simple spike activity and whisker movement exhibit peaks within the theta band. Eliciting either simple or complex spikes at frequencies ranging from 0.5 to 28 Hz, we found that 8 Hz is the preferred frequency around which the largest movement is induced. Interestingly, oscillatory whisker movements at 8 Hz were also generated when simple spike bursting was induced at 2 and 4 Hz, but never via climbing fiber stimulation. These results indicate that 8 Hz is the resonant frequency at which the cerebellar-whisker circuitry produces rhythmic whisking.

Chronic Levodopa therapy, the gold-standard treatment for Parkinson’s Disease (PD), leads to the emergence of involuntary movements, called levodopa-induced dyskinesia (LID). Cerebellar stimulation has been shown to decrease LID severity in PD patients. Here, in order to determine how cerebellar stimulation induces LID alleviation, we performed daily short trains of optogenetic stimulations of Purkinje cells (PC) in freely moving LID mice. We demonstrated that these stimulations are sufficient to suppress LID or even prevent their development. This symptomatic relief is accompanied by the normalization of aberrant neuronal discharge in the cerebellar nuclei, the motor cortex and the parafascicular thalamus. Inhibition of the cerebello-parafascicular pathway counteracted the beneficial effects of cerebellar stimulation. Moreover, cerebellar stimulation reversed plasticity in D1 striatal neurons and normalized the overexpression of FosB, a transcription factor causally linked to LID. These findings demonstrate LID alleviation and prevention by daily PC stimulations, which restore the function of a wide motor network, and may be valuable for LID treatment.

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.

Integration of information across the senses is critical for perception and is a common property of neurons in the cerebral cortex, where it is thought to arise primarily from corticocortical connections. Much less is known about the role of subcortical circuits in shaping the multisensory properties of cortical neurons. We show that stimulation of the whiskers causes widespread suppression of sound-evoked activity in mouse primary auditory cortex (A1). This suppression depends on the primary somatosensory cortex (S1), and is implemented through a descending circuit that links S1, via the auditory midbrain, with thalamic neurons that project to A1. Furthermore, a direct pathway from S1 has a facilitatory effect on auditory responses in higher-order thalamic nuclei that project to other brain areas. Crossmodal corticofugal projections to the auditory midbrain and thalamus therefore play a pivotal role in integrating multisensory signals and in enabling communication between different sensory cortical areas.

Fear conditioning is a form of associative learning that is known to involve different brain areas, notably the amygdala, the prefrontal cortex and the periaqueductal grey (PAG). Here, we describe the functional role of pathways that link the cerebellum with the fear network. We found that the cerebellar fastigial nucleus (FN) sends glutamatergic projections to vlPAG that synapse onto glutamatergic and GABAergic vlPAG neurons. Chemogenetic and optogenetic manipulations revealed that the FN-vlPAG pathway controls bi-directionally the strength of the fear memories, indicating an important role in the association of the conditioned and unconditioned stimuli, a function consistent with vlPAG encoding of fear prediction error. Moreover, FN-vlPAG projections also modulate extinction learning. We also found a FN-parafascicular thalamus pathway, which may relay cerebellar influence to the amygdala and modulates anxiety behaviors. Overall, our results reveal multiple contributions of the cerebellum to the emotional system.

The cerebellum is involved in the control of voluntary and autonomic rhythmic behaviors, yet it is unclear to what extent it coordinates these in concert. We studied Purkinje cell activity during unperturbed and perturbed respiration in lobules simplex, crus 1, and crus 2. During unperturbed (eupneic) respiration, complex spike and simple spike activity encode the phase of ongoing sensorimotor processing. In contrast, when the respiratory cycle is perturbed by whisker stimulation, mice concomitantly protract their whiskers and advance their inspiration in a phase-dependent manner, preceded by increased simple spike activity. This phase advancement of respiration in response to whisker stimulation can be mimicked by optogenetic stimulation of Purkinje cells and prevented by cell-specific genetic modification of their AMPA receptors, hampering increased simple spike firing. Thus, the impact of Purkinje cell activity on respiratory control is context and phase dependent, highlighting a coordinating role for the cerebellar hemispheres in aligning autonomic and sensorimotor behaviors.

Somatostatin improves visual perception by reducing excitatory inputs to the parvalbumin-positive interneurons in the cortex.

Sensory information–driven spikes are synchronized across cortical layers by distinct subtypes of interneurons.