Minibox: Custom solo or semi-group housing chambers for long term housing of rats with miniscopes.

In this detailed procedure, we include open-source methodologies using 'solidworks' designs for creating solo or semi-group housing units for rats wearing miniscopes for long periods of time. Builds are optimized to preserve rat health and prevent hardware destruction. We include all prices and suggestions for purchasing strategies to reduce overall build-costs.•Chambers are optimized for long-term housing to protect rats wearing delicate headstages (e.g., miniscopes).•Designed to be low-cost, efficient supplement to operant chambers and provides numerous benefits to long-term miniscope imaging. The housing chambers can be augmented by installing cameras, commutators, or different types of floor grids depending on experimental conditions.•The chambers can also be secured to one another to create "rat-duplexes", allowing experimenters to control the degree of social isolation.

A modular, cost-effective, versatile, open-source operant box solution for long-term miniscope imaging, 3D tracking, and deep learning behavioral analysis.

In this procedure we have included an open-source method for a customized operant chamber optimized for long-term miniature microscope (miniscope) recordings. •The miniscope box is designed to function with custom or typical med-associates style accessories (e.g., houselights, levers, etc.).•The majority of parts can be directly purchased which minimizes the need for skilled and time-consuming labor.•We include designs and estimated pricing for a single box but it is recommended to build these in larger batches to efficiently utilize bulk ordering of certain components.

Examining a punishment-related brain circuit with miniature fluorescence microscopes and deep learning.

In humans experiencing substance use disorder (SUD), abstinence from drug use is often motivated by a desire to avoid some undesirable consequence of further use: health effects, legal ramifications, etc. This process can be experimentally modeled in rodents by training and subsequently punishing an operant response in a context-induced reinstatement procedure. Understanding the biobehavioral mechanisms underlying punishment learning is critical to understanding both abstinence and relapse in individuals with SUD. To date, most investigations into the neural mechanisms of context-induced reinstatement following punishment have utilized discrete loss-of-function manipulations that do not capture ongoing changes in neural circuitry related to punishment-induced behavior change. Here, we describe a two-pronged approach to analyzing the biobehavioral mechanisms of punishment learning using miniature fluorescence microscopes and deep learning algorithms. We review recent advancements in both techniques and consider a target neural circuit.

Acquired Alterations in Nucleus Accumbens Responsiveness to a Cocaine-Paired Discriminative Stimulus Preceding Rats' Daily Cocaine Consumption.

Resumption of drug taking is a primary focus for substance use disorder research and can be triggered by drug-associated environmental stimuli. The Nucleus Accumbens (NAc) is a key brain region which guides motivated behavior and is implicated in resumption. There remains a pressing need to characterize NAc neurons' responsiveness to drug associated stimuli during withdrawal and abstinence. We recorded discriminative stimulus (DS) induced NAc activity via in vivo single-unit electrophysiology in rats that self-administered cocaine. Male and female rats implanted with a jugular catheter and a microwire array in NAc Core and Shell self-administered cocaine under control of a 30s auditory DS for 6 hours per session across 14 consecutive days. Rats acquired tone discrimination within 4 sessions. To exclude pharmacological effects of circulating cocaine from all neural analyses, we studied changes in DS-induced firing only for trials preceding the first infusion of cocaine in each of the 14 sessions, which were defined as "pre-drug trials." NAc neuron responses were assessed prior to tone-evoked movement onset. Responsiveness to the DS tone was exhibited throughout all sessions by the NAc Core population, but only during Early sessions by the NAc Shell population. Both Core and Shell responded selectively to the DS, i.e., more strongly on drug taking trials, or Hits, than on Missed opportunities. These findings suggest that NAc Core and Shell play distinct roles in initiating cocaine seeking prior to daily cocaine consumption, and align with reports suggesting that as drug use becomes chronic, cue-evoked activity shifts from NAc Shell to NAc Core.

Using deep learning to study emotional behavior in rodent models.

Quantifying emotional aspects of animal behavior (e.g., anxiety, social interactions, reward, and stress responses) is a major focus of neuroscience research. Because manual scoring of emotion-related behaviors is time-consuming and subjective, classical methods rely on easily quantified measures such as lever pressing or time spent in different zones of an apparatus (e.g., open vs. closed arms of an elevated plus maze). Recent advancements have made it easier to extract pose information from videos, and multiple approaches for extracting nuanced information about behavioral states from pose estimation data have been proposed. These include supervised, unsupervised, and self-supervised approaches, employing a variety of different model types. Representations of behavioral states derived from these methods can be correlated with recordings of neural activity to increase the scope of connections that can be drawn between the brain and behavior. In this mini review, we will discuss how deep learning techniques can be used in behavioral experiments and how different model architectures and training paradigms influence the type of representation that can be obtained.

Dysbindin-1, BDNF, and GABAergic Transmission in Schizophrenia.

Schizophrenia is a psychiatric disorder characterized by hallucinations, anhedonia, disordered thinking, and cognitive impairments. Both genetic and environmental factors contribute to schizophrenia. Dysbindin-1 (DTNBP1) and brain-derived neurotrophic factor (BDNF) are both genetic factors associated with schizophrenia. Mice lacking Dtnbp1 showed behavioral deficits similar to human patients suffering from schizophrenia. DTNBP1 plays important functions in synapse formation and maintenance, receptor trafficking, and neurotransmitter release. DTNBP1 is co-assembled with 7 other proteins into a large protein complex, known as the biogenesis of lysosome-related organelles complex-1 (BLOC-1). Large dense-core vesicles (LDCVs) are involved in the secretion of hormones and neuropeptides, including BDNF. BDNF plays important roles in neuronal development, survival, and synaptic plasticity. BDNF is also critical in maintaining GABAergic inhibitory transmission in the brain. Two studies independently showed that DTNBP1 mediated activity-dependent BDNF secretion to maintain inhibitory transmission. Imbalance of excitatory and inhibitory neural activities is thought to contribute to schizophrenia. In this mini-review, we will discuss a potential pathogenetic mechanism for schizophrenia involving DTNBP1, BDNF, and inhibitory transmission. We will also discuss how these processes are interrelated and associated with a higher risk of schizophrenia development.

Striatal direct pathway neurons play leading roles in accelerating rotarod motor skill learning.

Dorsal striatum is important for movement control and motor skill learning. However, it remains unclear how the spatially and temporally distributed striatal medium spiny neuron (MSN) activity in the direct and indirect pathways (D1 and D2 MSNs, respectively) encodes motor skill learning. Combining miniature fluorescence microscopy with an accelerating rotarod procedure, we identified two distinct MSN subpopulations involved in accelerating rotarod learning. In both D1 and D2 MSNs, we observed neurons that displayed activity tuned to acceleration during early stages of trials, as well as movement speed during late stages of trials. We found a distinct evolution trajectory for early-stage neurons during motor skill learning, with the evolution of D1 MSNs correlating strongly with performance improvement. Importantly, optogenetic inhibition of the early-stage neural activity in D1 MSNs, but not D2 MSNs, impaired accelerating rotarod learning. Together, this study provides insight into striatal D1 and D2 MSNs encoding motor skill learning.

Characterization of operant social interaction in rats: effects of access duration, effort, peer familiarity, housing conditions, and choice between social interaction vs. food or remifentanil.

RATIONALE AND OBJECTIVE: Social factors play a critical role in drug addiction. We recently showed that rats will abstain from methamphetamine, cocaine, heroin, and remifentanil self-administration when given a choice between the addictive drug and operant social interaction. Here, we further characterized operant social interaction by determining the effects of access duration, effort, peer familiarity, and housing conditions. We also determined choice between social interaction vs. palatable food or remifentanil. METHODS: We first trained single-housed male and female rats to lever-press for social interaction with a sex- and age-matched peer. Next, we determined effects of access duration (3.75 to 240 s), effort (increasing fixed-ratio schedule requirements or progressive ratio schedule), peer familiarity (familiar vs. unfamiliar), and housing conditions (single vs. paired housing) on social self-administration. We also determined choice between social interaction vs. palatable food pellets or intravenous remifentanil (0, 1, 10 µg/kg/infusion). RESULTS: Increasing access duration to a peer decreased social self-administration under fixed ratio but not progressive ratio schedule; the rats showed similar preference for short vs. long access duration. Social self-administration under different fixed ratio requirements was higher in single-housed than in paired-housed rats and higher for a familiar vs. unfamiliar partner in single-housed but not paired-housed rats. Response rates of food-sated rats under increasing fixed-ratio requirements were higher for palatable food than for social interaction. The rats strongly preferred palatable food over social interaction and showed dose-dependent preference for social interaction vs. remifentanil. CONCLUSIONS: We identified parameters influencing the reinforcing effects of operant social interaction and introduce a choice procedure sensitive to remifentanil self-administration dose.

Detailed mapping of behavior reveals the formation of prelimbic neural ensembles across operant learning.

The prelimbic cortex (PrL) is involved in the organization of operant behaviors, but the relationship between longitudinal PrL neural activity and operant learning and performance is unknown. Here, we developed deep behavior mapping (DBM) to identify behavioral microstates in video recordings. We combined DBM with longitudinal calcium imaging to quantify behavioral tuning in PrL neurons as mice learned an operant task. We found that a subset of PrL neurons were strongly tuned to highly specific behavioral microstates, both task and non-task related. Overlapping neural ensembles were tiled across consecutive microstates in the response-reinforcer sequence, forming a continuous map. As mice learned the operant task, weakly tuned neurons were recruited into new ensembles, with a bias toward behaviors similar to their initial tuning. In summary, our data suggest that the PrL contains neural ensembles that jointly encode a map of behavioral states that is fine grained, is continuous, and grows during operant learning.

Circuit Investigation of Social Interaction and Substance Use Disorder Using Miniscopes.

Substance use disorder (SUD) is comorbid with devastating health issues, social withdrawal, and isolation. Successful clinical treatments for SUD have used social interventions. Neurons can encode drug cues, and drug cues can trigger relapse. It is important to study how the activity in circuits and embedded cell types that encode drug cues develop in SUD. Exploring shared neurobiology between social interaction (SI) and SUD may explain why humans with access to social treatments still experience relapse. However, circuitry remains poorly characterized due to technical challenges in studying the complicated nature of SI and SUD. To understand the neural correlates of SI and SUD, it is important to: (1) identify cell types and circuits associated with SI and SUD, (2) record and manipulate neural activity encoding drug and social rewards over time, (3) monitor unrestrained animal behavior that allows reliable drug self-administration (SA) and SI. Miniaturized fluorescence microscopes (miniscopes) are ideally suited to meet these requirements. They can be used with gradient index (GRIN) lenses to image from deep brain structures implicated in SUD. Miniscopes can be combined with genetically encoded reporters to extract cell-type specific information. In this mini-review, we explore how miniscopes can be leveraged to uncover neural components of SI and SUD and advance potential therapeutic interventions.

Chronic Fentanyl Self-Administration Generates a Shift toward Negative Affect in Rats during Drug Use.

Drug addiction is thought to be driven by negative reinforcement, and it is thought that a shift from positive affect upon initial exposure to negative affect after chronic exposure to a drug is responsible for maintaining self-administration (SA) in addicted individuals. This can be modeled in rats by analyzing ultrasonic vocalizations (USVs), a type of intraspecies communication indicative of affective state based on the frequency of the emission: calls in the 22 kHz range indicate negative affect, whereas calls in the 50 kHz range indicate positive affect. We employed a voluntary chronic, long-access model of fentanyl SA to analyze affective changes in the response to chronic fentanyl exposure. Male Sprague-Dawley rats self-administered either fentanyl (N = 7) or saline (N = 6) for 30 consecutive days and USVs were recorded at four different time points: the day before the first SA session (PRE), the first day of SA (T01), the last day of SA (T30), and the first day of abstinence (ABS). At T01, the ratio of 50 to 22 kHz calls was similar between the fentanyl and saline groups, but at T30, the ratio differed between groups, with the fentanyl group showing significantly fewer 50 kHz calls and more 22 kHz calls relative to saline animals. These results indicate a shift toward a negative affect during drug use after chronic exposure to fentanyl and support negative reinforcement as a main driving factor of opioid addiction.

Emergence of negative affect as motivation for drug taking in rats chronically self-administering cocaine.

RATIONALE: The role of negative affect as a motivational factor in animal models of drug addiction has been underexplored in the context of cocaine self-administration. OBJECTIVES: The present investigation studied the relationship between magnitude of affective response and quantity of cocaine consumed in order to clarify the affective components that drive drug use in a preclinical model. METHODS: Rats self-administered (SA) cocaine 6 h/day for 14 consecutive days while their ultrasonic vocalizations (USVs) were recorded. RESULTS: Animals displayed an increase in 50-kHz call rates (indicating positive affect) when their drug levels were rapidly rising and an increase in 22-kHz call rates (indicating negative affect) when forced to abstain. The rate of 50-kHz calls predicted drug consumption during the 1st week of SA, but not week two. Contrarily, there was a strongly predictive positive association between rate of 22-kHz calls and amount of drug consumed during the 2nd week of SA. CONCLUSIONS: Experimental results indicate that after chronic cocaine self-administration, negative affect emerges when animals are deprived of expected drug during withdrawal. Moreover, the increase in USVs indicating negative affect when deprived of drug was directly related to drug intake, concurrent with a decay in the direct relationship between USVs indicating positive affect and drug intake. The present preclinical support for the widely hypothesized shift from positive to negative affect as a salient motivational factor in human drug abuse adds to growing evidence of the unique value of rat USVs for understanding the role of emotion in drug addiction.