Behavioral Neuroscience publications

Tracking head position and orientation in small mammals is crucial for many applications in the field of behavioral neurophysiology, from the study of spatial navigation to the investigation of active sensing and perceptual representations. Many approaches to head tracking exist, but most of them only estimate the 2D coordinates of the head over the plane where the animal navigates. Full reconstruction of the pose of the head in 3D is much more more challenging and has been achieved only in handful of studies, which employed headsets made of multiple LEDs or inertial units. However, these assemblies are rather bulky and need to be powered to operate, which prevents their application in wireless experiments and in the small enclosures often used in perceptual studies. Here we propose an alternative approach, based on passively imaging a lightweight, compact, 3D structure, painted with a pattern of black dots over a white background. By applying a cascade of feature extraction algorithms that progressively refine the detection of the dots and reconstruct their geometry, we developed a tracking method that is highly precise and accurate, as assessed through a battery of validation measurements. We show that this method can be used to study how a rat samples sensory stimuli during a perceptual discrimination task and how a hippocampal place cell represents head position over extremely small spatial scales. Given its minimal encumbrance and wireless nature, our method could be ideal for high-throughput applications, where tens of animals need to be simultaneously and continuously tracked. NEW & NOTEWORTHY Head tracking is crucial in many behavioral neurophysiology studies. Yet reconstruction of the head’s pose in 3D is challenging and typically requires implanting bulky, electrically powered headsets that prevent wireless experiments and are hard to employ in operant boxes. Here we propose an alternative approach, based on passively imaging a compact, 3D dot pattern that, once implanted over the head of a rodent, allows estimating the pose of its head with high precision and accuracy.

In auditory-conditioned fear learning, the freezing response is independent of the sound frequencies used, but the frequency of the conditioned sound is considered distinct from those of unrelated sounds based on electrophysiological responses in the auditory system. Whether an emergent discriminative learning underlies auditory fear conditioning and which nuclei and pathways are involved in it remain unclear. Using behavioral and electrophysiological assays, we found that the response of medial prefrontal cortex (mPFC) neurons to a conditioned auditory stimulus (CS) was enhanced relative to the response to unrelated frequencies (UFs) after auditory fear conditioning, and mice could distinguish the CS during multifrequency testing, a phenomenon called emergent discriminative learning. After silencing the mPFC with muscimol, emergent discriminative learning was blocked. In addition, the pure tone responses of mPFC neurons were inhibited after injection of lidocaine in the ipsilateral primary auditory cortex (A1), and the emergent discriminative learning was blocked by silencing both sides of A1 with muscimol. This study, therefore, provides evidence for an emergent discriminative learning mediated by mPFC and A1 neurons after auditory fear conditioning.

Canonical reinforcement learning models postulate that dopamine neurons encode reward prediction error (RPE) and provide a teaching signal to striatal spiny projection neurons (SPNs) in the form of dopamine (DA) release. DA is thought to guide learning via dynamic modulation of protein kinase A (PKA) in SPNs. However, this fundamental assumption remains untested in behaving animals. Here we utilized multi-channel fiber photometry and fluorescence lifetime photometry (FLiP) to monitor the activity of DA neurons, extracellular DA levels, and net PKA activity in SPNs in the nucleus accumbens during learning. We found dynamic encoding of RPE in the activity of DA neurons, which is both necessary and sufficient to explain striatal DA levels and SPN PKA activity. The modulation of PKA in SPNs that express type-1 (D1R-SPNs) and type-2 (D2R-SPNs) DA receptors was dichotomous such that in each cell class it is selectively sensitive to increases and decreases in DA, respectively, and occur at and support different phases of learning. Thus, PKA-dependent pathways in D1R- and D2R-SPNs are asynchronously engaged by RPE-encoding DA signals to promote different aspects of reinforcement learning: the former responsible for the initial association between action and outcome and the latter responsible for refining the learned association.

Animal behavioral paradigms, such as classical conditioning and operant conditioning, are an important tool to study the neural basis of cognition and behavior. These paradigms involve manipulating sensory stimuli in a way that learning processes are induced under controlled experimental conditions. However, the majority of the commercially available equipment did not offer flexibility to manipulate stimuli. Therefore, the development of most versatile devices would allow the study of more complex cognitive functions. The purpose of this work is to present a low-cost, customized and wireless-operated chamber for animal behavior conditioning, based on the joint operation of two microcontroller modules: Arduino Due and ESP8266-12E. Our results showed that the auditory stimulation system allows setting the carrier frequency in the range of 1 Hz up to more than 100 kHz and the sound stimulus can be modulated in amplitude, also over a wide range of frequencies. Likewise, foot-shock could be precisely manipulated regarding its amplitude (from ∼200 μA to ∼1500 μA) and frequency (up to 20 pulses per second). Finally, adult rats exposed to a protocol of cued fear conditioning in our device showed consistent behavioral response and electrophysiological evoked responses in the midbrain auditory pathway. Furthermore, the device developed in the current study represents an open source alternative to develop customized protocols to study fear memory under conditions of varied sensory stimuli.

The ability to recognize sounds in noise is a key part of hearing, and the mechanisms by which the brain identifies sounds in noise are of considerable interest to scientists, clinicians and engineers. Yet we know little about the necessity of regions such as auditory cortex for hearing in noise, or how cortical processing of sounds is adversely affected by noise. Here we used reversible cortical inactivation and extracellular electrophysiology in ferrets performing a vowel discrimination task to identify and understand the causal contribution of auditory cortex to hearing in noise. Cortical inactivation by cooling impaired task performance in noisy but not clean conditions, while responses of auditory cortical neurons were less informative about vowel identity in noise. Simulations mimicking cortical inactivation indicated that effects of inactivation were related to the loss of information about sounds represented across neural populations. The addition of noise to target sounds drove spiking activity in auditory cortex and recruitment of additional neural populations that were linked to degraded behavioral performance. To suppress noise-related activity, we used continuous exposure to background noise to adapt the auditory system and recover behavioral performance in both ferrets and humans. Inactivation by cooling revealed that the benefits of continuous exposure were not cortically dependent. Together our results highlight the importance of auditory cortex in sound discrimination in noise and the underlying mechanisms through which noise-related activity and adaptation shape hearing.

Evidence suggests that the pathophysiology associated with epileptic susceptibility may disturb the functional connectivity of neural circuits and compromise the brain functions, even when seizures are absent. Although memory impairment is a common comorbidity found in patients with epilepsy, it is still unclear whether more caudal structures may play a role in cognitive deficits, particularly in those cases where there is no evidence of hippocampal sclerosis. This work used a genetically selected rat strain for seizure susceptibility (Wistar audiogenic rat, WAR) and distinct behavioral (motor and memory-related tasks) and electrophysiological (inferior colliculus, IC) approaches to access acoustic primary integrative network properties. The IC neural assemblies' response was evaluated by auditory transient (focusing on bottom-up processing) and steady-state evoked response (ASSR, centering on feedforward and feedback forces over neural circuitry). The results show that WAR displayed no disturbance in motor performance or hippocampus-dependent memory tasks. Nonetheless, WAR animals exhibited significative impairment for auditory fear conditioning (AFC) along with no indicative of IC plastic changes between the pre-conditioning and test phases (ASSR coherence analysis). Furthermore, WAR's IC response to transient stimuli presented shorter latency and higher amplitude compared with Wistar; and the ASSR analysis showed similar results for WAR and Wistar animals under subthreshold dose of pentylenetetrazol (pro-convulsive drug) for seizure-induction. Our work demonstrated alterations at WAR IC neural network processing, which may explain the associated disturbance on AFC memory.

The brain faces the difficult task of maintaining a stable representation of key features of the outside world in noisy sensory surroundings. How does the sensory representation change with noise, and how does the brain make sense of it? We investigated the effect of background white noise (WN) on tuning properties of neurons in mouse A1 and its impact on discrimination performance in a go/no-go task. We find that WN suppresses the activity of A1 neurons, which surprisingly increases the discriminability of tones spectrally close to each other. To confirm the involvement of A1, we optogenetically excited parvalbumin-positive (PV+) neurons in A1, which have similar effects as WN on both tuning properties and frequency discrimination. A population model suggests that the suppression of A1 tuning curves increases frequency selectivity and thereby improves discrimination. Our findings demonstrate that the cortical representation of pure tones adapts during noise to improve sensory acuity.

Cue-evoked midbrain dopamine (DA) neuron activity reflects expected value, but its influence on reward assessment is unclear. In mice performing a trial-based operant task, we test if bidirectional manipulations of cue or operant-associated DA neuron activity drive learning as a result of under- or overexpectation of reward value. We target optogenetic manipulations to different components of forced trials, when only one lever is presented, and assess lever biases on choice trials in the absence of photomanipulation. Although lever biases are demonstrated to be flexible and sensitive to changes in expected value, augmentation of cue or operant-associated DA signaling does not significantly alter choice behavior, and blunting DA signaling during any component of the forced trials reduces choice trial responses on the associated lever. These data suggest cue-evoked DA helps maintain cue-value associations but does not encode expected value as to set the benchmark against which received reward is judged.

Energy homeostasis requires precise measurement of the quantity and quality of ingested food. The vagus nerve innervates the gut and can detect diverse interoceptive cues, but the identity of the key sensory neurons and corresponding signals that regulate food intake remains unknown. Here, we use an approach for target-specific, single-cell RNA sequencing to generate a map of the vagal cell types that innervate the gastrointestinal tract. We show that unique molecular markers identify vagal neurons with distinct innervation patterns, sensory endings, and function. Surprisingly, we find that food intake is most sensitive to stimulation of mechanoreceptors in the intestine, whereas nutrient-activated mucosal afferents have no effect. Peripheral manipulations combined with central recordings reveal that intestinal mechanoreceptors, but not other cell types, potently and durably inhibit hunger-promoting AgRP neurons in the hypothalamus. These findings identify a key role for intestinal mechanoreceptors in the regulation of feeding.

While substantial task-related neural activity has been observed during motor tasks in rodent primary motor cortex and premotor cortex, the long-term stability of these responses in healthy rats is uncertain, limiting the interpretability of longitudinal changes in the specific patterns of neural activity associated with learning or motor recovery following injury. This study examined the stability of task-related neural activity associated with execution of two distinct reaching tasks in healthy rodents. A novel automated rodent behavioral apparatus was constructed and rats were trained to perform a reaching task combining a 'gross' lever press and a 'fine' pellet retrieval. In each animal, two chronic microelectrode arrays were implanted in motor cortex spanning the caudal forelimb area (rodent primary motor cortex) and the rostral forelimb area (rodent premotor cortex). We recorded multiunit spiking and local field potential activity from 10 days to 7-10 weeks post-implantation to characterize the patterns of neural activity observed during each task component and analyzed the consistency of channel-specific task-related neural activity. Task-related changes in neural activity were observed on the majority of channels. While the task-related changes in multi-unit spiking and local field potential spectral power were consistent over several weeks, spectral power changes were more stable, despite the trade-off of decreased spatial and temporal resolution. These results show that neural activity in rodent primary and premotor cortex is associated with specific phases of reaching movements with stable patterns of task-related activity across time, establishing the relevance of the rodent for future studies designed to examine changes in task-related neural activity during recovery from focal cortical lesions.

Imprinted genes are highly expressed in the hypothalamus; however, whether specific imprinted genes affect hypothalamic neuromodulators and their functions is unknown. It has been suggested that Prader-Willi syndrome (PWS), a neurodevelopmental disorder caused by lack of paternal expression at chromosome 15q11-q13, is characterised by hypothalamic insufficiency. Here, we investigate the role of the paternally expressed Snord116 gene within the context of sleep and metabolic abnormalities of PWS, and we report a novel role of this imprinted gene in the function and organisation of the two main neuromodulatory systems of the lateral hypothalamus (LH), namely, the orexin (OX) and melanin concentrating hormone (MCH) systems. We observe that the dynamics between neuronal discharge in the LH and the sleep-wake states of mice with paternal deletion of Snord116 (PWScr m+/p− ) are compromised. This abnormal state-dependent neuronal activity is paralleled by a significant reduction in OX neurons in the LH of mutants. Therefore, we propose that an imbalance between OX- and MCH-expressing neurons in the LH of mutants reflects a series of deficits manifested in the PWS, such as dysregulation of rapid eye movement (REM) sleep, food intake and temperature control. Highlights Snord116 regulates neuronal activity in the lateral hypothalamus (LH), which is time-locked with cortical states of sleep. Loss of Snord116 reduces orexin neurons in the LH and affects sleep homeostasis and thermoregulation in mice. Snord116 and Peg3 independently control orexin expression in the LH. Paternally expressed alleles maximize the patrilineal effects in the control of REM sleep by the LH in mammals.

Manipulations that enhance neuroplasticity may inadvertently create opportunities for maladaptation. We have previously used passive exposures to non-traumatic white noise to open windows of plasticity in the adult rat auditory cortex and induce frequency-specific functional reorganizations of the tonotopic map. However, similar reorganizations in the central auditory pathway are thought to contribute to the generation of hearing disorders such as tinnitus and hyperacusis. Here, we investigate whether noise-induced reorganizations are accompanied by electrophysiological or behavioral evidence of tinnitus or hyperacusis in adult Long-Evans rats. We used a 2-week passive exposure to moderate-intensity (70 dB SPL) broadband white noise to reopen a critical period for spectral tuning such that a second 1-week exposure to 7 kHz tone pips produced an expansion of the 7 kHz frequency region in the primary auditory cortex (A1). We demonstrate for the first time that this expansion also takes place in the ventral auditory field (VAF). Sound exposure also led to spontaneous and sound-evoked hyperactivity in the anterior auditory field (AAF). Rats were assessed for behavioral evidence of tinnitus or hyperacusis using gap and tone prepulse inhibition of the acoustic startle response. We found that sound exposure did not affect gap-prepulse inhibition. However, sound exposure led to an improvement in prepulse inhibition when the prepulse was a 7 kHz tone, showing that exposed rats had enhanced sensorimotor gating for the exposure frequency. Together, our electrophysiological and behavioral results provide evidence of hyperacusis but not tinnitus in sound-exposed animals. Our findings demonstrate that periods of prolonged noise exposure may open windows of plasticity that can also be understood as windows of vulnerability, potentially increasing the likelihood for maladaptive plasticity to take place.

Social cues of threat are widely reported [1–3], whether actively produced to trigger responses in others, such as the emission of alarm calls, or by-products of an encounter with a predator, like the defensive behaviors themselves, such as an escape flight [4–14]. Although the recognition of social alarm cues is often innate [15–17], in some instances it requires experience to trigger defensive responses [4,7]. One mechanism proposed for how learning from self-experience contributes to social behavior is that of auto-conditioning, whereby subjects learn to associate their own behaviors with the relevant trigger events. Through this process the same behaviors, now displayed by others, gain meaning. [18,19 but see: 20]. Although it has been shown that only animals with prior experience with shock display observational freezing [21–25] suggesting that auto-conditioning could mediate this process, evidence for this hypothesis was lacking. Previously we found that when a rat freezes, the silence that results from immobility triggers observational freezing in its cage-mate, provided the cage-mate had experienced shocks before [24]. Hence, in our study auto-conditioning would correspond to rats learning to associate shock with their own response to it – freezing. Using a combination of behavioral and optogenetic manipulations, here we show that freezing becomes an alarm cue by a direct association with shock. Our work shows that auto-conditioning can indeed modulate social interactions, expanding the repertoire of cues that mediate social information exchange, providing a framework to study how the neural circuits involved in the self-experience of defensive behaviors overlap with the ones involved in socially triggered defensive behaviors.

The weakly electric gymnotiform fish produce a rhythmic electric organ discharge (EOD) used for communication and active electrolocation. The EOD frequency is entrained to a medullary pacemaker nucleus. During communication and exploration, this rate can be modulated by a pre-pacemaker network, resulting in specific patterns of rate modulation, including stereotyped communication signals and dynamic interactions with conspecifics known as a Jamming Avoidance Response (JAR). One well-known stereotyped signal is the chirp, a brief upward frequency sweep usually lasting less than 500 ms. The abrupt change in frequency has dramatic effects on phase precession between two signalers. We report here on chirping in Brachyhypopmus cf. sullivani, Microsternarchus cf. bilineatus Lineage C, and Steatogenys cf. elegans during conspecific playback experiments. Microsternarchus also exhibits two behaviors that include chirp-like extreme frequency modulations, EOD interruptions with hushing silence and tumultuous rises, and these are described in terms of receiver impact. These behaviors all have substantial impact on interference caused by conspecifics and may be a component of the JAR in some species. Chirps are widely used in electronic communications systems, sonar, and other man-made active sensing systems. The brevity of the chirp, and the phase disruption it causes, makes chirps effective as attention-grabbing or readiness signals. This conforms to the varied assigned functions across gymnotiforms, including pre-combat aggressive or submissive signals or during courtship and mating. The specific behavioral contexts of chirp expression vary across species, but the physical structure of the chirp makes it extremely salient to conspecifics. Chirps may be expected in a wide range of behavioral contexts where their function depends on being noticeable and salient. Further, in pulse gymnotiforms, the chirp is well structured to comprise a robust jamming signal to a conspecific receiver if specifically timed to the receiver's EOD cycle. Microsternarchus and Steatogenys exploit this feature and include chirps in dynamic jamming avoidance behaviors. This may be an evolutionary re-use of a circuitry for a specific signal in another context.

Humans and other animals effortlessly identify natural sounds and categorize them into behaviorally relevant categories. Yet, the acoustic features and neural transformations that enable sound recognition and the formation of perceptual categories are largely unknown. Here, using multichannel neural recordings in the auditory midbrain of unanesthetized female rabbits, we first demonstrate that neural ensemble activity in the auditory midbrain displays highly structured correlations that vary with distinct natural sound stimuli. These stimulus-driven correlations can be used to accurately identify individual sounds using single-response trials, even when the sounds do not differ in their spectral content. Combining neural recordings and an auditory model, we then show how correlations between frequency-organized auditory channels can contribute to discrimination of not just individual sounds but sound categories. For both the model and neural data, spectral and temporal correlations achieved similar categorization performance and appear to contribute equally. Moreover, both the neural and model classifiers achieve their best task performance when they accumulate evidence over a time frame of approximately 1-2 seconds, mirroring human perceptual trends. These results together suggest that time-frequency correlations in sounds may be reflected in the correlations between auditory midbrain ensembles and that these correlations may play an important role in the identification and categorization of natural sounds.

Noise-exposed rat pups provide a model of early deprivation of sensory input to the central auditory system, allowing the study of developmental neuroplasticity. Our previous results have demonstrated that a brief exposure of rats to broadband noise (125 dB SPL 8 min, 14th postnatal day) at the onset of hearing resulted in an altered intensity perception and frequency discrimination in adulthood despite normal hearing thresholds. In this study, we assessed the gap-detection ability and possible presence of tinnitus- and hyperacusis-like behavior in adult rats after the same neonatal acoustic trauma, using measurements of the acoustic startle response (ASR) in quiet and noisy environments and its prepulse inhibition by gaps in noise (gap-PPI). A significant deficit in the ability to detect gap was observed in the exposed rats when 55 dB SPL broadband noise was used as background. An increase of noise intensity to 65-75 dB SPL led to strengthening of the gap-PPI in exposed animals, which approached the gap-PPI values of control animals at these levels. Behavioral signs of tinnitus (gap detection deficits in 10 kHz narrow band noise) were found in 25% of exposed rats. An increased sensitivity to continuous noise was manifested in all exposed rats by suppression of the ASR at significantly lower background noise levels than in the controls. This effect was particularly pronounced in rats with tinnitus-like behavior. Our results indicate that neonatal acoustic trauma, producing only a transient threshold shift, may produce permanent abnormalities in suprathreshold auditory functions and the development of tinnitus and hyperacusis-like behavior.

Neurofibromatosis type 1 (NF1) is an autosomal dominant disorder whose neurodevelopmental symptoms include impaired executive function, attention, and spatial learning and could be due to perturbed mesolimbic dopaminergic circuitry. However, these circuits have never been directly assayed in vivo. We employed the genetically encoded optical dopamine sensor dLight1 to monitor dopaminergic neurotransmission in the ventral striatum of NF1 mice during motivated behavior. Additionally, we developed novel systemic AAV vectors to facilitate morphological reconstruction of dopaminergic populations in cleared tissue. We found that NF1 mice exhibit reduced spontaneous dopaminergic neurotransmission that was associated with excitation/inhibition imbalance in the ventral tegmental area and abnormal neuronal morphology. NF1 mice also had more robust dopaminergic and behavioral responses to salient visual stimuli, which were independent of learning, and rescued by optogenetic inhibition of non-dopaminergic neurons in the VTA. Overall, these studies provide a first in vivo characterization of dopaminergic circuit function in the context of NF1 and reveal novel pathophysiological mechanisms.

Dravet syndrome (DS) is a severe early-onset epilepsy associated with heterozygous loss-of-function mutations in SCN1A . Animal models of DS with global Scn1a haploinsufficiency recapitulate the DS phenotype, including seizures, premature death, and impaired spatial memory performance. Spatial memory requires hippocampal sharp-wave ripples (SPW-Rs), which consist of high-frequency field potential oscillations (ripples, 100–260 Hz) superimposed on a slower SPW. Published in vitro electrophysiologic recordings in DS mice demonstrate reduced firing of GABAergic inhibitory neurons, which are essential for the formation of SPW-R complexes. Here, in vivo electrophysiologic recordings of hippocampal local field potential in both male and female mice demonstrate that Scn1a haploinsufficiency slows intrinsic ripple frequency and reduces the rate of SPW-R occurrence. In DS mice, peak ripple-band power is shifted to lower frequencies, average intertrough intervals of individually detected ripples are slower, and the rate of SPW-R generation is reduced, while SPW amplitude remains unaffected. These alterations in SPW-R properties, in combination with published reductions in interneuron function in DS, suggest a direct link between reduced inhibitory neuron excitability and impaired SPW-R function. A simple interconnected, conductance-based in silico interneuron network model was used to determine whether reduced sodium conductance is sufficient to slow ripple frequency, and stimulation with a modeled SPW demonstrates that reduced sodium conductance alone is sufficient to slow oscillatory frequencies. These findings forge a potential mechanistic link between impaired SPW-R generation and Scn1a mutation in DS mice, expanding the set of disorders in which SPW-R dysfunction contributes to impaired memory. SIGNIFICANCE STATEMENT Disruption of sharp-wave ripples, a characteristic hippocampal rhythm coordinated by the precise timing of GABAergic interneurons, impairs spatial learning and memory. Prior in vitro patch-clamp recordings in brain slices from genetic mouse models of Dravet syndrome (DS) reveal reduced sodium current and excitability in GABAergic interneurons but not excitatory cells, suggesting a causal role for impaired interneuron activity in seizures and cognitive impairment. Here, heterozygous Scn1a mutation in DS mice reduces hippocampal sharp-wave ripple occurrence and slows internal ripple frequency in vivo and a simple in silico model demonstrates reduction in interneuron function alone is sufficient to slow model oscillations. Together, these findings provide a plausible pathophysiologic mechanism for Scn1a gene mutation to impair spatial memory.

To understand the cortical neuronal dynamics behind movement generation and control most studies focused on tasks where actions were planned and then executed, using different instances of visuomotor transformations. However, to fully understand the dynamics related to movement control one must also study how movements are actively inhibited. Inhibition, indeed, represents the first level of control both when different alternatives are available and only one solution could be adopted, and when it is necessary to maintain the current position. We recorded neuronal activity from a multielectrode array in the dorsal premotor (PMd) cortex of monkeys performing a countermanding reaching task that requires, in a subset of trials, to cancel a planned movement before its onset. In the analysis of the neuronal state-space of PMd we found a subspace in which activities conveying temporal information were confined during active inhibition and position holding. Movement execution required activities to escape from this subspace toward an orthogonal subspace and, furthermore, surpass a threshold associated with the maturation of the motor plan. These results revealed further details in the neuronal dynamics underlying movement control, extending the hypothesis that neuronal computation confined in an -output-null- subspace does not produce movements.

Cocaine-associated memories are critical drivers of relapse in cocaine-dependent individuals that can be evoked by exposure to cocaine or stress. Whether these environmental stimuli recruit similar molecular and circuit-level mechanisms to promote relapse remains largely unknown. Here, using cocaine- and stress-primed reinstatement of cocaine conditioned place preference to model drug-associated memories, we find that cocaine drives reinstatement by increasing the duration that mice spend in the previously cocaine-paired context whereas stress increases the number of entries into this context. Importantly, both forms of reinstatement require Cav1.2 L-type Ca2+ channels (LTCCs) in cells of the prelimbic cortex that project to the nucleus accumbens core (PrL→NAcC). Utilizing fiber photometry to measure circuit activity in vivo in conjunction with the LTCC blocker, isradipine, we find that LTCCs drive differential recruitment of the PrL→ NAcC pathway during cocaine- and stress-primed reinstatement. While cocaine selectively activates PrL→NAcC cells prior to entry into the cocaine-paired chamber, a measure that is predictive of duration in that chamber, stress increases persistent activity of this projection, which correlates with entries into the cocaine-paired chamber. Using projection-specific chemogenetic manipulations, we show that PrL→NAcC activity is required for both cocaine- and stress-primed reinstatement, and that activation of this projection in Cav1.2-deficient mice restores reinstatement. These data indicate that LTCCs are a common mediator of cocaine- and stress-primed reinstatement. However, they engage different patterns of behavior and PrL→NAcC projection activity depending on the environmental stimuli. These findings establish a framework to further study how different environmental experiences can drive relapse, and supports further exploration of isradipine, an FDA-approved LTCC blocker, as a potential therapeutic for the prevention of relapse in cocaine-dependent individuals.