Optogenetics seminars

Nuo Li (Baylor College of Medicine, USA) shares novel insights into coordinated interhemispheric large-scale neural network activity underpinning short-term memory in mice. Relevant techniques covered include: simultaneous multi-regional recordings using multiple 64-channel H probes during head-fixed behavior in mice. simultaneous optogenetics and population recording. analysis of population recordings to infer interactions between brain regions. Reference: Chen G, Kang B, Lindsey J, Druckmann S, Li N, (2021). Modularity and robustness of frontal cortex networks. Cell, 184(14):3717-3730.

How does our brain rapidly determine if something is good or bad? How do we know our place within a social group? How do we know how to behave appropriately in dynamic environments with ever-changing conditions? The Tye Lab is interested in understanding how neural circuits important for driving positive and negative motivational valence (seeking pleasure or avoiding punishment) are anatomically, genetically and functionally arranged. We study the neural mechanisms that underlie a wide range of behaviors ranging from learned to innate, including social, feeding, reward-seeking and anxiety-related behaviors. We have also become interested in “social homeostasis” -- how our brains establish a preferred set-point for social contact, and how this maintains stability within a social group. How are these circuits interconnected with one another, and how are competing mechanisms orchestrated on a neural population level? We employ optogenetic, electrophysiological, electrochemical, pharmacological and imaging approaches to probe these circuits during behavior.

In understanding circuit operations, a key issue is the extent to which neuronal spiking reflects local computation or responses to upstream inputs. Because pyramidal cells in CA1 do not have local recurrent projections, it is currently assumed that firing in CA1 is inherited from its inputs – thus, entorhinal inputs provide communication with the rest of the neocortex and the outside world, whereas CA3 inputs provide internal and past memory representations. Several studies have attempted to prove this hypothesis, by lesioning or silencing either area CA3 or the entorhinal cortex and examining the effect of firing on CA1 pyramidal cells. Despite the intense and careful work in this research area, the magnitudes and types of the reported physiological impairments vary widely across experiments. At least part of the existing variability and conflicts is due to the different behavioral paradigms, designs and evaluation methods used by different investigators. Simultaneous manipulations in the same animal or even separate manipulations of the different inputs to the hippocampal circuits in the same experiment are rare. To address these issues, I used optogenetic silencing of unilateral and bilateral mEC, of the local CA1 region, and performed bilateral pharmacogenetic silencing of the entire CA3 region. I combined this with high spatial resolution recording of local field potentials (LFP) in the CA1-dentate axis and simultaneously collected firing pattern data from thousands of single neurons. Each experimental animal had up to two of these manipulations being performed simultaneously. Silencing the medial entorhinal (mEC) largely abolished extracellular theta and gamma currents in CA1, without affecting firing rates. In contrast, CA3 and local CA1 silencing strongly decreased firing of CA1 neurons without affecting theta currents. Each perturbation reconfigured the CA1 spatial map. Yet, the ability of the CA1 circuit to support place field activity persisted, maintaining the same fraction of spatially tuned place fields, and reliable assembly expression as in the intact mouse. Thus, the CA1 network can maintain autonomous computation to support coordinated place cell assemblies without reliance on its inputs, yet these inputs can effectively reconfigure and assist in maintaining stability of the CA1 map.

The neural correlates of decision-making have been investigated extensively, and recent work aims to identify under what conditions cortex is actually necessary for making accurate decisions. We discovered that mice with distinct cognitive experiences, beyond sensory and motor learning, use different cortical areas and neural activity patterns to solve the same task, revealing past learning as a critical determinant of whether cortex is necessary for decision tasks. We used optogenetics and calcium imaging to study the necessity and neural activity of multiple cortical areas in mice with different training histories. Posterior parietal cortex and retrosplenial cortex were mostly dispensable for accurate performance of a simple navigation-based visual discrimination task. In contrast, these areas were essential for the same simple task when mice were previously trained on complex tasks with delay periods or association switches. Multi-area calcium imaging showed that, in mice with complex-task experience, single-neuron activity had higher selectivity and neuron-neuron correlations were weaker, leading to codes with higher task information. Therefore, past experience is a key factor in determining whether cortical areas have a causal role in decision tasks.

Brendan Ito (Cornell University, USA) and Teja Bollu (Salk Institute, USA) share unique insights into rapid online motor corrections during mouse licking, analogous to primate goal-oriented reaching. Techniques covered include large-scale single unit recording during behaviour with optogenetics, and a deep-learning-based neural network to resolve 3D tongue kinematics during licking.

Visual restoration is certainly the greatest challenge for brain-machine interfaces with the high pixel number and high refreshing rate. In the recent year, we brought retinal prostheses and optogenetic therapy up to successful clinical trials. Concerning visual restoration at the cortical level, prostheses have shown efficacy for limited periods of time and limited pixel numbers. We are investigating the potential of sonogenetics to develop a non-contact brain machine interface allowing long-lasting activation of the visual cortex. The presentation will introduce our genetic-based brain machine interfaces for visual restoration at the retinal and cortical levels.

Human movement disorders and pharmacological studies have long suggested molecular dopamine modulates the pace of the internal clock. But how does the endogenous dopaminergic system influence the timing of our movements? We examined the relationship between dopaminergic signaling and the timing of reward-related, self-timed movements in mice. Animals were trained to initiate licking after a self-timed interval following a start cue; reward was delivered if the animal’s first lick fell within a rewarded window (3.3-7 s). The first-lick timing distributions exhibited the scalar property, and we leveraged the considerable variability in these distributions to determine how the activity of the dopaminergic system related to the animals’ timing. Surprisingly, dopaminergic signals ramped-up over seconds between the start-timing cue and the self-timed movement, with variable dynamics that predicted the movement/reward time, even on single trials. Steeply rising signals preceded early initiation, whereas slowly rising signals preceded later initiation. Higher baseline signals also predicted earlier self-timed movement. Optogenetic activation of dopamine neurons during self-timing did not trigger immediate movements, but rather caused systematic early-shifting of the timing distribution, whereas inhibition caused late-shifting, as if dopaminergic manipulation modulated the moment-to-moment probability of unleashing the planned movement. Consistent with this view, the dynamics of the endogenous dopaminergic signals quantitatively predicted the moment-by-moment probability of movement initiation. We conclude that ramping dopaminergic signals, potentially encoding dynamic reward expectation, probabilistically modulate the moment-by-moment decision of when to move. (Based on work from Hamilos et al., eLife, 2021).

Dr. Looger will present updates on a variety of molecular tools for studying & manipulating neural circuits & other preparations. Topics include genetically encoded calcium indicators (including the new ultra-fast jGCaMP8 variants), neurotransmitter sensors (improved versions for following glutamate, GABA, acetylcholine, serotonin), optogenetic effectors including the new “enhanced Magnets” dimerizers, AAV serotypes for retrograde labeling & altered tropism, probes for correlative light-electron microscopy, chemical gene switches, etc. He will make all his slides freely available - so don’t worry about hurriedly taking notes; instead focus on questions and ideas for collaboration. Please bring your suggestions for molecular tools that would be transformative for the field.

Coping with threatening situations requires both identifying stimuli predicting danger and selecting adaptive behavioral responses in order to survive. The dorso medial prefrontal cortex (dmPFC) is a critical structure involved in the regulation of threat-related behaviour, yet it is still largely unclear how threat-predicting stimuli and defensive behaviours are associated within prefrontal networks in order to successfully drive adaptive responses. Over the past years, we used a combination we used a combination of extracellular recordings, neuronal decoding approaches, and state of the art optogenetic manipulations to identify key neuronal elements and mechanisms controlling defensive fear responses. I will present an overview of our recent work ranging from analyses of dedicated neuronal types and oscillatory and synchronization mechanisms to artificial intelligence approaches used to decode the activity or large population of neurons. Ultimately these analyses allowed the identification of high dimensional representations of defensive behavior unfolding within prefrontal networks.

Nearly all motivated behaviors require the ability to associate outcomes with specific actions and make adaptive decisions about future behavior. The nucleus accumbens (NAc) is integrally involved in these processes. The NAc is a heterogeneous population primarily composed of D1 and D2 medium spiny projection (MSN) neurons that are thought to have opposed roles in behavior, with D1 MSNs promoting reward and D2 MSNs promoting aversion. Here we examined what types of information are encoded by the D1 and D2 MSNs using optogenetics, fiber photometry, and cellular resolution calcium imaging. First, we showed that mice responded for optical self-stimulation of both cell types, suggesting D2-MSN activation is not inherently aversive. Next, we recorded population and single cell activity patterns of D1 and D2 MSNs during reinforcement as well as Pavlovian learning paradigms that allow dissociation of stimulus value, outcome, cue learning, and action. We demonstrated that D1 MSNs respond to the presence and intensity of unconditioned stimuli – regardless of value. Conversely, D2 MSNs responded to the prediction of these outcomes during specific cues. Overall, these results provide foundational evidence for the discrete aspects of information that are encoded within the NAc D1 and D2 MSN populations. These results will significantly enhance our understanding of the involvement of the NAc MSNs in learning and memory as well as how these neurons contribute to the development and maintenance of substance use disorders.

Optogenetics is a powerful tool that allows experimentalists to perturb neural circuits. What can we learn about a network from observing its response to perturbations? I will first describe the results of optogenetic activation of inhibitory neurons in mice cortex, and show that the results are consistent with inhibition stabilization. I will then move to experiments in which excitatory neurons are activated optogenetically, with or without visual inputs, in mice and monkeys. In some conditions, these experiments show a surprising result that the distribution of firing rates is not significantly changed by stimulation, even though firing rates of individual neurons are strongly modified. I will show in which conditions a network model of excitatory and inhibitory neurons can reproduce this feature.

Neurons in the suprachiasmatic nucleus (SCN, the brain’s master circadian clock) display a 24 hour cycle in the their rate of action potential discharge whereby firing rates are high during the light phase and lower during the dark phase. Although it is generally agreed that this cycle of activity is a key mediator of the clock’s neural and humoral output, surprisingly little is known about how changes in clock electrical activity can mediate scheduled physiological changes at different times of day. Using opto- and chemogenetic approaches in mice we have shown that the onset of electrical activity in vasopressin releasing SCN neurons near Zeitgeber time 22 (ZT22) activates glutamatergic thirst-promoting neurons in the OVLT (organum vasculosum lamina terminalis) to promote water intake prior to sleep. This effect is mediated by activity-dependent release of vasopressin from the axon terminals of SCN neurons which acts as a neurotransmitter on OVLT neurons. More recently we found that the clock receives excitatory input from a different subset of sodium sensing neurons in the OVLT. Activation of these neurons by a systemic salt load delivered at ZT19 stimulated the electrical activity of SCN neurons which are normally silent at this time. Remarkably, this effect induced an acute reduction in non-shivering thermogenesis and body temperature, which is an adaptive response to the salt load. These findings provide information regarding the mechanisms by which the SCN promotes scheduled physiological rhythms and indicates that the clock’s output circuitry can also be recruited to mediate an unscheduled homeostatic response.

The hippocampus is crucial for spatial navigation and episodic memory formation. Hippocampal place cells exhibit spatially selective activity within an environment and have been proposed to form the neural basis of a cognitive map of space that supports these mnemonic functions. However, the direct influence of place cell activity on spatial navigation behaviour has not yet been demonstrated. Using an ‘all-optical’ combination of simultaneous two-photon calcium imaging and two-photon holographically targeted optogenetics, we identified and selectively activated place cells that encoded behaviourally relevant locations in a virtual reality environment. Targeted stimulation of a small number of place cells was sufficient to bias the behaviour of animals during a spatial memory task, providing causal evidence that hippocampal place cells actively support spatial navigation and memory. Time permitting, I will also describe new experiments aimed at understanding the fundamental encoding mechanism that supports episodic memory, focussing on the role of hippocampal sequences across multiple timescales and behaviours.

Recent advances in optogenetics allow for stimulation of neurons with sub-millisecond spike jitter and single neuron selectivity. Already this precision has revealed new levels of cortical sensitivity: stimulating tens of neurons can yield changes in the mean firing rate of thousands of similarly tuned neurons. This extreme sensitivity suggests that cortical dynamics are near criticality. Criticality is often studied in neural systems as a non-equilibrium thermodynamic process in which scale-free patterns of activity, called avalanches, emerge between distinct states of spontaneous activity. While criticality is well studied, it is still unclear what these distinct states of spontaneous activity are and what responses we expect from stimulation of this activity. By answering these questions, optogenetic stimulation will become a new avenue for approaching criticality and understanding cortical dynamics. Here, for the first time, we study the effects of optogenetic-like stimulation on a model near criticality. We study a model of Inhibitory/Excitatory (I/E) Leaky Integrate and Fire (LIF) spiking neurons which display a region of high sensitivity as seen in experiments. We find that this region of sensitivity is, indeed, near criticality. We derive the Dynamic Mean Field Theory of this model and find that the distinct states of activity are asynchrony and synchrony. We use our theory to characterize response to various types and strengths of optogenetic stimulation. Our model and theory predict that asynchronous, near-critical dynamics can have two qualitatively different responses to stimulation: one characterized by high sensitivity, discrete event responses, and high trial-to-trial variability, and another characterized by low sensitivity, continuous responses with characteristic frequencies, and low trial-to-trial variability. While both response types may be considered near-critical in model space, networks which are closest to criticality show a hybrid of these response effects.

Inter-areal coherence between cortical field-potentials is a widespread phenomenon and depends on numerous behavioral and cognitive factors. It has been hypothesized that inter-areal coherence reflects phase-synchronization between local oscillations and flexibly gates communication. We reveal an alternative mechanism, where coherence results from and is not the cause of communication, and naturally emerges as a consequence of the fact that spiking activity in a sending area causes post-synaptic inputs both in the same area and in other areas. Consequently, coherence depends in a lawful manner on oscillatory power and phase-locking in a sending area and inter-areal connectivity. We show that changes in oscillatory power explain prominent changes in fronto-parietal beta-coherence with movement and memory, and LGN-V1 gamma-coherence with arousal and visual stimulation. Optogenetic silencing of a receiving area and E/I network simulations demonstrate that afferent synaptic inputs rather than spiking entrainment are the main determinant of inter-areal coherence. These findings suggest that the unique spectral profiles of different brain areas automatically give rise to large-scale inter-areal coherence patterns that follow anatomical connectivity and continuously reconfigure as a function of behavior and cognition.

Ever since Bliss and Lømo discovered the phenomenon of long-term potentiation (LTP) in rabbit dentate gyrus in the 1960s, Hebb’s rule—neurons that fire together wire together—gained popularity to explain learning and memory. Accumulating evidence, however, suggests that neural activity is homeostatically regulated. Homeostatic mechanisms are mostly interpreted to stabilize network dynamics. However, recent theoretical work has shown that linking the activity of a neuron to its connectivity within the network provides a robust alternative implementation of Hebb’s rule, although entirely based on negative feedback. In this setting, both natural and artificial stimulation of neurons can robustly trigger network rewiring. We used computational models of plastic networks to simulate the complex temporal dynamics of network rewiring in response to external stimuli. In parallel, we performed optogenetic stimulation experiments in the mouse anterior cingulate cortex (ACC) and subsequently analyzed the temporal profile of morphological changes in the stimulated tissue. Our results suggest that the new theoretical framework combining neural activity homeostasis and structural plasticity provides a consistent explanation of our experimental observations.

In our everyday lives, we form connections and build up social networks that allow us to function successfully as individuals and as a society. Our social networks tend to include well-connected individuals who link us to other groups of people that we might otherwise have limited access to. In addition, we are more likely to befriend individuals who a) live nearby and b) have mutual friends. Interestingly, neurons tend to do the same…until development is perturbed. Just like social networks, neuronal networks require highly connected hubs to elicit efficient communication at minimal cost (you can’t befriend everybody you meet, nor can every neuron wire with every other!). This talk will cover some of Alex’s work showing that microscopic (cellular scale) brain networks inferred from spontaneous activity show similar complex topology to that previously described in macroscopic human brain scans. The talk will also discuss what happens when neurodevelopment is disrupted in the case of a monogenic disorder called Rett Syndrome. This will include simulations of neuronal activity and the effects of manipulation of model parameters as well as what happens when we manipulate real developing networks using optogenetics. If functional development can be restored in atypical networks, this may have implications for treatment of neurodevelopmental disorders like Rett Syndrome.

Reflex circuits in the nervous system integrate changes in the environment with physiology. Compact clusters of brain neuron cell bodies, termed nuclei, are essential for receiving sensory input and for transmitting motor outputs to the body. These nucelii are critical relay stations which process incoming information and convert these signals to outgoing action potentials which regulate immune system functions. Thus, reflex neural circuits maintain parameters of immunological physiology within a narrow range optimal for health. Advances in neuroscience and immunology using optogenetics, pharmacogenetics, and functional mapping offer a new understanding of the importance of neural circuitry underlying immunity, and offer direct paths to new therapies.