Optogenetics publications

Serotonergic neurons in the rostral ventral medulla (RVM) contribute to bidirectional control of pain through modulation of spinal and trigeminal nociceptive networks. Deficits in this pathway are believed to contribute to pathologic pain states, but whether changes in serotonergic mechanisms are pro- or antinociceptive is debated. We used a combination of optogenetics and fiber photometry to examine these mechanisms more closely. We find that optogenetic activation of RVM serotonergic afferents in the spinal cord of naive mice produces mechanical hypersensitivity and conditioned place aversion (CPA). Neuropathic pain, produced by chronic constriction injury of the infraorbital nerve (CCI-ION), evoked a tonic increase in serotonin (5HT) concentrations within the spinal trigeminal nucleus caudalis (SpVc), measured with liquid chromatography-tandem mass spectroscopy (LC-MS/MS). By contract, CCI-ION had no effect on the phasic serotonin transients in SpVc, evoked by noxious pinch, and measured with fiber photometry of a serotonin sensor. These findings suggest that serotonin release in the spinal cord is pronociceptive and that an increase in sustained serotonin signaling, rather than phasic or event driven increases, potentiate nociception in models of chronic pain.

Social behaviors are innate and supported by dedicated neural circuits, but the molecular identities of these circuits and how they are established developmentally and shaped by experience remain unclear. Here we show that medial amygdala (MeA) cells originating from two embryonically parcellated developmental lineages have distinct response patterns and functions in social behavior in male mice. MeA cells expressing the transcription factor Foxp2 (MeA Foxp2 ) are specialized for processing male conspecific cues and are essential for adult inter-male aggression. By contrast, MeA cells derived from the Dbx1 lineage (MeA Dbx1 ) respond broadly to social cues, respond strongly during ejaculation and are not essential for male aggression. Furthermore, MeA Foxp2 and MeA Dbx1 cells show differential anatomical and functional connectivity. Altogether, our results suggest a developmentally hardwired aggression circuit at the MeA level and a lineage-based circuit organization by which a cell’s embryonic transcription factor profile determines its social information representation and behavioral relevance during adulthood.

Adaptive behavior in a dynamic environment often requires rapid revaluation of stimuli that deviates from well-learned associations. The divergence between stable value-encoding and appropriate behavioral output remains a critical test to theories of dopamine’s function in learning, motivation, and motor control. Yet how dopamine neurons are involved in the revaluation of cues when the world changes to alter our behavior remains unclear. Here we make use of pharmacology, in vivo electrophysiology, fiber photometry, and optogenetics to resolve the contributions of the mesolimbic dopamine system to the dynamic reorganization of reward-seeking. Male and female rats were trained to discriminate when a conditioned stimulus would be followed by sucrose reward by exploiting the prior, non-overlapping presentation of a separate discrete cue - an occasion setter. Only when the occasion setter’s presentation preceded the conditioned stimulus did the conditioned stimulus predict sucrose delivery. As a result, in this task we were able to dissociate the average value of the conditioned stimulus from its immediate expected value on a trial-to-trial basis. Both the activity of ventral tegmental area dopamine neurons and dopamine signaling in the nucleus accumbens were essential for rats to successfully update behavioral responding in response to the occasion setter. Moreover, dopamine release in the nucleus accumbens following the conditioned stimulus only occurred when the occasion setter indicated it would predict reward. Downstream of dopamine release, we found that single neurons in the nucleus accumbens dynamically tracked the value of the conditioned stimulus. Together these results reveal a novel mechanism within the mesolimbic dopamine system for the rapid revaluation of motivation.

Objective: An environmental context, which reliably predicts food availability, can increase the appetitive food drive within the same environment context. However, hunger is required for the development of such a context-induced feeding (CIF) response, suggesting the neural circuits sensitive to hunger link an internal energy state with a particular environment context. Since Agouti related peptide (AgRP) neurons are activated by energy deficit, we hypothesised that AgRP neurons are both necessary and sufficient to drive CIF. Methods: To examine the role of AgRP neurons in the CIF process, we used fibre photometry with GCaMP7f, chemogenetic activation of AgRP neurons, as well as optogenetic control of AgRP neurons to facilitate acute temporal control not permitted with chemogenetics. Results: A CIF response at test was only observed when mice were fasted during context training and AgRP population activity at test showed an attenuated inhibitory response to food, suggesting increased food-seeking and/or decreased satiety signalling drives the increased feeding response at test. Intriguingly, chemogenetic activation of AgRP neurons during context training did not increase CIF, suggesting precise temporal firing properties may be required. Indeed, termination of AgRP neuronal photostimulation during context training (ON-OFF in context), in the presence or absence of food, increased CIF. Moreover, photoinhibition of AgRP neurons during context training in fasted mice was sufficient to drive a subsequent CIF in the absence of food. Conclusions: Our results suggest that AgRP neurons regulate the acquisition of CIF when the acute inhibition of AgRP activity is temporally matched to context exposure. These results establish acute AgRP inhibition as a salient neural event underscoring the effect of hunger on associative learning.

The kindling model has been used extensively by researchers to study the neurobiology of temporal lobe epilepsy (TLE) due to its capacity to induce intensification of seizures by the progressive recruitment of additional neuronal clusters into epileptogenic networks. We applied repetitive focal optogenetic activation of putative excitatory neurons in the dorsal CA1 area of the hippocampus of mice to investigate the role of inhibitory signaling during this process. This experimental protocol resulted in a kindling phenotype that was maintained for 2 weeks after the animals were fully kindled. As a result of the different phases of optogenetic kindling (OpK), key inhibitory signaling elements, such as KCC2 and NKCC1, exhibited distinct temporal and spatial dynamics of regulation. These alterations in protein expression were related to the distinct pattern of ictal activity propagation through the different hippocampal sublayers. Our results suggest the KCC2 disruption in the contralateral hippocampus of fully kindled animals progressively facilitated the creation of pathological pathways for seizure propagation through the hippocampal network. Upon completion of kindling, we observed animals that were restimulated after a rest period of 14-day showed, besides a persistent KCC2 downregulation, an NKCC1 upregulation in the bilateral dentate gyrus and hippocampus-wide loss of parvalbumin-positive interneurons. These alterations observed in the chronic phase of OpK suggest that the hippocampus of rekindled animals continued to undergo self-modifications during the rest period. The changes resulting from this period suggest the possibility of the development of a mirror focus on the hippocampus contralateral to the site of optical stimulations. Our results offer perspectives for preventing the recruitment and conversion of healthy neuronal networks into epileptogenic ones among patients with epilepsy.

In the classical model of the basal ganglia, direct pathway striatal projection neurons (dSPNs) send projections to the substantia nigra (SNr) and entopeduncular nucleus to regulate motor function. Recent studies have re-established that dSPNs also possess axon collaterals within the globus pallidus (GPe) (bridging collaterals), yet the significance of these collaterals for behavior is unknown. Here we use in vivo optical and chemogenetic tools combined with deep learning approaches in mice to dissect the roles of dSPN GPe collaterals in motor function. We find that dSPNs projecting to the SNr send synchronous motor-related information to the GPe via axon collaterals. Inhibition of native activity in dSPN GPe terminals impairs motor activity and function via regulation of Npas1 neurons. We propose a model by which dSPN GPe axon collaterals (striatopallidal Go pathway) act in concert with the canonical terminals in the SNr to support motor control by inhibiting Npas1 neurons.

We develop soft and stretchable fatigue-resistant hydrogel optical fibers that enable optogenetic modulation of peripheral nerves in naturally behaving animals during persistent locomotion. The formation of polymeric nanocrystalline domains within the hydrogels yields fibers with low optical losses of 1.07 dB cm-1, Young's modulus of 1.6 MPa, stretchability of 200% and fatigue strength of 1.4 MPa against 30,000 stretch cycles. The hydrogel fibers permitted light delivery to the sciatic nerve, optogenetically activating hindlimb muscles in Thy1::ChR2 mice during 6-week voluntary wheel running assays while experiencing repeated deformation. The fibers additionally enabled optical inhibition of pain hypersensitivity in an inflammatory model in TRPV1::NpHR mice over an 8-week period. Our hydrogel fibers offer a motion-adaptable and robust solution to peripheral nerve optogenetics, facilitating the investigation of somatosensation.

Rapid-eye-movement (REM) sleep is accompanied by intense cortical activity, underlying its wake-like electroencephalogram (EEG). The neural activity inducing REM sleep is thought to originate from subcortical circuits in brainstem and hypothalamus. However, whether cortical neurons can also trigger REM sleep has remained unknown. Here, we show in mice that the medial prefrontal cortex (mPFC) strongly promotes REM sleep. Bidirectional optogenetic manipulations demonstrate that excitatory mPFC neurons promote REM sleep through their projections to the lateral hypothalamus (LH) and regulate phasic events, reflected in accelerated EEG theta oscillations and increased eye-movement density during REM sleep. Calcium imaging reveals that the majority of LH-projecting mPFC neurons are maximally activated during REM sleep and a subpopulation is recruited during phasic theta accelerations. Our results delineate a cortico-hypothalamic circuit for the top-down control of REM sleep and identify a critical role of the mPFC in regulating phasic events during REM sleep.

The amygdala, cholinergic basal forebrain, and higher-order auditory cortex (HO-AC) regulate brain-wide plasticity underlying auditory threat learning. Here, we perform multi-regional extracellular recordings and optical measurements of acetylcholine (ACh) release to characterize the development of discriminative plasticity within and between these brain regions as mice acquire and recall auditory threat memories. Spiking responses are potentiated for sounds paired with shock (CS+) in the lateral amygdala (LA) and optogenetically identified corticoamygdalar projection neurons, although not in neighboring HO-AC units. Spike- or optogenetically triggered local field potentials reveal enhanced corticofugal-but not corticopetal-functional coupling between HO-AC and LA during threat memory recall that is correlated with pupil-indexed memory strength. We also note robust sound-evoked ACh release that rapidly potentiates for the CS+ in LA but habituates across sessions in HO-AC. These findings highlight a distributed and cooperative plasticity in LA inputs as mice learn to reappraise neutral stimuli as possible threats.

Priming, a change in the mental processing of a stimulus as a result of prior encounter with a related stimulus, has been observed repeatedly and studied extensively in humans. Yet currently, there is no behavioral model of short-term priming in lab animals, precluding research on the neurobiological basis of priming. Here, we describe an auditory discrimination paradigm for studying response priming in freely moving mice. We find a priming effect in success rate in all mice tested on the task. In contrast, we do not find a priming effect in response times. Compared to non-primed discrimination trials, the addition of incongruent prime stimuli reduces success rate more than congruent prime stimuli, suggesting a cognitive mechanism based on differential interference. The results establish the short-term priming phenomenon in rodents, and the paradigm opens the door to studying the cellular-network basis of priming.

Preclinical and clinical studies implicate endocannabinoids (eCBs) in fear extinction, but the underlying neural circuit basis of these actions is unclear. Here, we employed in vivo optogenetics, eCB biosensor imaging, ex vivo electrophysiology, and CRISPR-Cas9 gene editing in mice to examine whether basolateral amygdala (BLA)-projecting medial prefrontal cortex (mPFC) neurons represent a neural substrate for the effects of eCBs on extinction. We found that photoexcitation of mPFC axons in BLA during extinction mobilizes BLA eCBs. eCB biosensor imaging showed that eCBs exhibit a dynamic stimulus-specific pattern of activity at mPFC→BLA neurons that tracks extinction learning. Furthermore, using CRISPR-Cas9-mediated gene editing, we demonstrated that extinction memory formation involves eCB activity at cannabinoid CB1 receptors expressed at vmPFC→BLA synapses. Our findings reveal the temporal characteristics and a neural circuit basis of eCBs' effects on fear extinction and inform efforts to target the eCB system as a therapeutic approach in extinction-deficient neuropsychiatric disorders.

Background: Commonly encountered nontraumatic, moderate noise is increasingly implicated in anxiety; however, the neural substrates underlying this process remain unclear. Objectives: We investigated the neural circuit mechanism through which chronic exposure to moderate-level noise causes anxiety-like behaviors. Methods: Mice were exposed to chronic, moderate white noise [85 decibel (dB) sound pressure level (SPL)], 4 h/d for 4 wk to induce anxiety-like behaviors, which were assessed by open field, elevated plus maze, light-dark box, and social interaction tests. Viral tracing, immunofluorescence confocal imaging, and brain slice patch-clamp recordings were used to characterize projections from auditory brain regions to the lateral amygdala. Neuronal activities were characterized by in vivo multielectrode and fiber photometry recordings in awake mice. Optogenetics and chemogenetics were used to manipulate specific neural circuitry. Results: Mice chronically (4 wk) exposed to moderate noise (85 dB SPL, 4 h/d) demonstrated greater neuronal activity in the lateral amygdala (LA), and the LA played a critical role in noise-induced anxiety-like behavior in these model mice. Viral tracing showed that the LA received monosynaptic projections from the medial geniculate body (MG) and auditory cortex (ACx). Optogenetic excitation of the MG→LA or ACx→LA circuits acutely evoked anxiety-like behaviors, whereas their chemogenetic inactivation abolished noise-induced anxiety-like behavior. Moreover, mice chronically exposed to moderate noise were more susceptible to acute stress, with more neuronal firing in the LA, even after noise withdrawal. Discussion: Mice exposed to 4 wk of moderate noise (85 dB SPL, 4 h/d) demonstrated behavioral and physiological differences compared to controls. The neural circuit mechanisms involved greater excitation from glutamatergic neurons of the MG and ACx to LA neurons under chronic, moderate noise exposure, which ultimately promoted anxiety-like behaviors. Our findings support the hypothesis that nontraumatic noise pollution is a potentially serious but unrecognized public health concern. https://doi.org/10.1289/EHP12532.

Epilepsy is characterized by spontaneous non-provoked seizures, yet the mechanisms that trigger a seizure and allow its evolution remain underexplored. To dissect out phases of ictogenesis, we evoked hypersynchronous activity with optogenetic stimulation. Focal optogenetic activation of putative excitatory neurons in the mouse hippocampal CA1 reliably evoked convulsive seizures in awake mice. A time-vs-time pulsogram plot characterized the evolution of the EEG pulse response from a light evoked response to induced seizure activity. Our results depict ictogenesis as a stepwise process comprised of three distinctive phases demarcated by two transition points. The induction phase undergoes the first transition to reverberant phase activity, followed by the second transition into the paroxysmal phase or a seizure. Non-seizure responses are confined to either induction or reverberant phases. The pulsogram was then constructed in seizures recorded from a murine model of temporal lobe epilepsy and it depicted a similar reverberance preceding spontaneous seizures. The discovery of these distinct phases of ictogenesis may offer means to abort a seizure before it develops.

Aims Major depressive disorder is a severe psychiatric disorder that afflicts ~17% of the world population. Neuroimaging investigations of depressed patients have consistently reported the dysfunction of the basolateral amygdala in the pathophysiology of depression. However, how the BLA and related circuits are implicated in the pathogenesis of depression is poorly understood. Methods Here, we combined fiber photometry, immediate early gene expression (c‐fos), optogenetics, chemogenetics, behavioral analysis, and viral tracing techniques to provide multiple lines of evidence of how the BLA neurons mediate depressive‐like behavior. Results We demonstrated that the aversive stimuli elevated the neuronal activity of the excitatory BLA neurons (BLA CAMKII neurons). Optogenetic activation of CAMKII neurons facilitates the induction of depressive‐like behavior while inhibition of these neurons alleviates the depressive‐like behavior. Next, we found that the chemogenetic inhibition of GABAergic neurons in the BLA (BLA GABA ) increased the firing frequency of CAMKII neurons and mediates the depressive‐like phenotypes. Finally, through fiber photometry recording and chemogenetic manipulation, we proved that the activation of BLA GABA neurons inhibits BLA CAMKII neuronal activity and alleviates depressive‐like behavior in the mice. Conclusion Thus, through evaluating BLA GABA and BLA CAMKII neurons by distinct interaction, the BLA regulates depressive‐like behavior.

With concurrent global epidemics of chronic pain and opioid use disorders, there is a critical need to identify, target and manipulate specific cell populations expressing the mu-opioid receptor (MOR). However, available tools and transgenic models for gaining long-term genetic access to MOR+ neural cell types and circuits involved in modulating pain, analgesia and addiction across species are limited. To address this, we developed a catalog of MOR promoter ( MORp ) based constructs packaged into adeno-associated viral vectors that drive transgene expression in MOR+ cells. MORp constructs designed from promoter regions upstream of the mouse Oprm1 gene ( mMORp ) were validated for transduction efficiency and selectivity in endogenous MOR+ neurons in the brain, spinal cord, and periphery of mice, with additional studies revealing robust expression in rats, shrews, and human induced pluripotent stem cell (iPSC)-derived nociceptors. The use of mMORp for in vivo fiber photometry, behavioral chemogenetics, and intersectional genetic strategies is also demonstrated. Lastly, a human designed MORp ( hMORp ) efficiently transduced macaque cortical OPRM1 + cells. Together, our MORp toolkit provides researchers cell type specific genetic access to target and functionally manipulate mu-opioidergic neurons across a range of vertebrate species and translational models for pain, addiction, and neuropsychiatric disorders.

Motivated behaviors are often studied in isolation to assess labeled lines of neural connections underlying innate actions. However, in nature, multiple systems compete for expression of goal-directed behaviors via complex neural networks. Here, we examined flexible survival decisions in animals tasked with food seeking under predation threat. We found that predator exposure rapidly induced physiological, neuronal, and behavioral adaptations in mice highlighted by reduced food seeking and consumption contingent on current threat level. Diminishing conflict via internal state or external environment perturbations shifted feeding strategies. Predator introduction and/or selective manipulation of danger-responsive cholecystokinin (Cck) cells of the dorsal premammilary nucleus (PMd) suppressed hunger-sensitive Agouti-related peptide (AgRP) neurons, providing a mechanism for threat-evoked hypophagia. Increased caloric need enhanced food seeking under duress through AgRP pathways to the bed nucleus of the stria terminalis (BNST) and/or lateral hypothalamus (LH). Our results suggest oscillating interactions between systems underlying self-preservation and food seeking to promote optimal behavior.

Visual stimuli that deviate from the current context elicit augmented responses in the primary visual cortex (V1). These heightened responses, known as "deviance detection," require local inhibition in the V1 and top-down input from the anterior cingulate area (ACa). Here, we investigated the mechanisms by which the ACa and V1 interact to support deviance detection. Local field potential recordings in mice during an oddball paradigm showed that ACa-V1 synchrony peaks in the theta/alpha band (≈10 Hz). Two-photon imaging in the V1 revealed that mainly pyramidal neurons exhibited deviance detection, while contextually redundant stimuli increased vasoactive intestinal peptide (VIP)-positive interneuron (VIP) activity and decreased somatostatin-positive interneuron (SST) activity. Optogenetic drive of ACa-V1 inputs at 10 Hz activated V1-VIPs but inhibited V1-SSTs, mirroring the dynamics present during the oddball paradigm. Chemogenetic inhibition of V1-VIPs disrupted Aca-V1 synchrony and deviance detection in the V1. These results outline temporal and interneuron-specific mechanisms of top-down modulation that support visual context processing.

The endocannabinoid (eCB) system is a key modulator of glutamate release within limbic neurocircuitry and thus heavily modulates stress responsivity and adaptation. The ventral hippocampus (vHPC)-basolateral amygdala (BLA) circuit has been implicated in the expression of negative affective states following stress exposure and is modulated by retrograde eCB signaling. However, the mechanisms governing eCB release and the causal relationship between vHPC-BLA eCB signaling and stress-induced behavioral adaptations are not known. Here, we utilized in vivo optogenetic- and biosensor-based approaches to determine the temporal dynamics of activity-dependent and stress-induced eCB release at vHPC-BLA synapses. Furthermore, we demonstrate that genetic deletion of cannabinoid type-1 receptors selectively at vHPC-BLA synapses decreases active stress coping and exacerbates stress-induced avoidance and anhedonia phenotypes. These data establish the in vivo determinants of eCB release at limbic synapses and demonstrate that eCB signaling within vHPC-BLA circuitry serves to counteract adverse behavioral consequences of stress.

Hippocampal theta phase precession is involved in spatiotemporal coding and generating multineural spike sequences, but how precession originates remains unresolved. To determine whether precession can be generated directly in CA1 and disambiguate multiple competing mechanisms, we used optogenetic activation to impose artificial place fields in pyramidal cells of mice running on a linear track. More than a third of the CA1 artificial fields exhibited synthetic precession that persisted for a full cycle. In contrast, artificial fields in the parietal cortex did not exhibit synthetic precession. The findings are incompatible with precession models based on inheritance, spreading activation, dual-input, or inhibition-excitation summation. Thus, a precession generator resides locally within CA1.

Interneuron populations within the nucleus accumbens (NAc) orchestrate excitatory-inhibitory balance, undergo experience-dependent plasticity, and gate-motivated behavior, all biobehavioral processes heavily modulated by endogenous cannabinoid (eCB) signaling. While eCBs are well known to regulate synaptic plasticity onto NAc medium spiny neurons and modulate NAc function at the behavioral level, how eCBs regulate NAc interneuron function is less well understood. Here, we show that eCB signaling differentially regulates glutamatergic and feedforward GABAergic transmission onto NAc somatostatin–expressing interneurons (NAc SOM+ ) in an input-specific manner, while simultaneously increasing postsynaptic excitability of NAc SOM+ neurons, ultimately biasing toward vHPC (ventral hippocampal), and away from BLA (basolateral amygdalalar), activation of NAc SOM+ neurons. We further demonstrate that NAc SOM+ are activated by stress in vivo and undergo stress-dependent plasticity, evident as a global increase in intrinsic excitability and an increase in excitation–inhibition balance specifically at vHPC, but not BLA, inputs onto NAc SOM+ neurons. Importantly, both forms of stress-induced plasticity are dependent on eCB signaling at cannabinoid type 1 receptors. These findings reveal eCB-dependent mechanisms that sculpt afferent input and excitability of NAc SOM+ neurons and demonstrate a key role for eCB signaling in stress-induced plasticity of NAc SOM+ -associated circuits.