DREADD receptors (Designer Receptor Exclusively Activated by Designer Drugs) are engineered G-protein receptors which are exclusively activated by the inert-molecule CNO (clozapine-N-oxide) and to a lesser degree clozapine. DREADD receptors are often used, to identify the circuitry and cellular signals that specify behavior, perceptions, emotions, motivation, and motor functions in species ranging from flies to nonhuman primates. Heterologous overexpression of non-native genes in biological systems, is often confounded by varying degrees of overexpression and result in non-specific effects. Thus, our purpose is to determine effects on cell signaling pathways following varying levels of DREADD receptor expression. Our hypothesis is that, increased levels of DREADD receptor expression increase non-canonical cellular signaling pathway activation. We used human embryonic kidney cells (HEK cells), as our model system, then expressed various levels of DREADD receptors using Lipofectamine transfection and DNA-plasmids expressing Gs- and Gi-coupled DREADD receptors (Canonically, Gs-increases cAMP and Gi-decreases cAMP). After transfection, we treated transfected cells with 1µM of CNO or VEH (a control) for 5-30min and then lysed the cells with RIPA Buffer, and prepared samples for western blotting to detect activation of proteins of interest in the GPCR non-canonical signaling pathways (example: kinases and transcription factors). We used anti-HA antibodies to ensure dosage of receptor expression, and used a panel of antibodies to detect activated proteins in well-established non-canonical GPCR signaling pathways. Western blot results were measured using Odyssey scanner and the Image studio software suite. We expect to see an increase in non-canonical signaling with increased expression of both Gi- and Gs- coupled, specifically in the MEK/ERK and CREB signaling pathways. In summary, our findings will highlight additional considerations regarding the level of receptor expression when using DREADD receptors in vitro and in vivo experiments, and specifically measure the off-target “non-canonical” signaling pathways induced by DREADD receptor overexpression.

The ability to quantify rat and mice ultrasonic vocalizations (USVs) can provide ethological validity to animal models of affective disorders. Vocalization frequencies are key indicators of a rodent’s subjective state and are closely tied to their behavior. For example, 50-kHz calls in rats are emitted during positive events such as sucrose drinking, and are attributed to a positive affective state. In contrast, 22-kHz calls are ascribed to a negative affective state, and are emitted during negative events such as shock. Current methods for manually scoring vocalizations are slow and labor intensive, while automated programs lack the accuracy to produce satisfactory scoring under non-ideal recording conditions. In order to create a more precise program for detecting these USVs, we applied deep learning, which has revolutionized the field of bioacoustics through image detection and recognition of audio files. By converting soundwaves produced by mice and rats into spectrograms, we can train convolutional neural networks to distinguish between vocalizations and noise. Using these methods, we created the software DeepSqueak©, a Deep Learning Based System for Quantifying USVs and Extended Analysis Kit, which outperforms current state-of-the-art, commercially available software. Hundreds of rat calls from the 18-75kHz were manually isolated from spectrograms and used as training data for our neural network, allowing it to differentiate calls from noise. As a result, USV detections using DeepSqueak are far more rapid and accurate in comparison to existing software. The detected calls produced from the software were then reviewed and re-entered into the neural network as another training set to consolidate its precision and efficiency. With the speed and reliability of DeepSqueak, quantification of USVs can become more widespread and lead to a wealth of new knowledge relating to affective disorders.

Research has shown that drinking motives (or reasons for consuming alcohol) are strong predictors of alcohol use; moreover, enhancement and social motives are among the strongest predictors of alcohol use. Given limited research to date, the purpose of the current study is to examine how drinking motives relate to drinking alone and also to drinking with friends. College student drinkers ages 18-24 were recruited for an ongoing study. To be eligible, participants had to report alcohol use two or more days a week and one or more occasions of heavy episodic drinking in the last two weeks. Baseline data on alcohol use, drinking motives, and drinking context were used for the current analyses (N = 292 to date; 52% female, 71% Caucasian, mean age = 20.17 years). Participants were asked to report how often they drank alone and with friends on a scale from “never” to “daily or almost daily.” In addition, they were asked to report motives for drinking on each of five subscales using response options from “almost never/never” to “almost always/always." The current study will use regression analyses to test associations with four of the subscales: coping-anxiety, coping-depression, enhancement, and social motives. We hypothesize that higher coping motives will be associated with being more likely to report drinking alone and with friends. Additionally, we hypothesize that higher social and enhancement motives will be associated with being less likely to report drinking alone and more likely to report drinking with friends. Results from this study will provide a better understanding of why people decide to drink and in which different social contexts, thus providing valuable information to guide interventions on how to better incorporate drinking motives and social context.

The prevalence of alcohol use disorder (AUD) and associated alcohol-related health conditions is disproportionately high among individuals experiencing homelessness. This disparity results in relatively high use of publicly funded services, particularly high-cost healthcare (e.g., emergency department and inpatient hospital services). A recent study of harm-reduction treatment for alcohol (HaRT-A) showed it was efficacious in reducing alcohol-related harm. The objective of this secondary study was to test whether HaRT-A had effects on inpatient and emergency department service utilization as well. Participants (N=167; 24% women) in this secondary study were recruited from community-based clinical and social services settings and were randomized to receive the HaRT-A intervention or services as usual. Outcomes were post-pre differences in number of inpatient and emergency department visits from 6 months prior and subsequent to study participation. There were no significant group effects on changes in emergency department and inpatient visits (ps > .78). HaRT-A did not precipitate significant changes in utilization of emergency and inpatient services by individuals experiencing homelessness and AUD. The study time frame might have been too short to document changes in such statistically rare outcomes. It is also possible that healthcare outcomes are too distal to show the impact of such a brief intervention. Future treatment trials should include longer data collection time frames to test treatment effects on more distal yet important health-related outcomes.

Stress is highly correlated with negative physical health impacts and an estimated 75 to 90% of primary care visits are attributed to stress related conditions. Currently available treatments for anxiety are not always effective and very limited, therefore identifying new targets for the development of medication is important. Recently, neuropeptides have drawn interest as potential targets for drug therapies. Pituitary adenylate cyclase-activating polypeptide, or PACAP, is a neuropeptide that has been linked to stress responses in a variety of brain regions. For example, the amygdala is a crucial brain region for mediating chronic stress, acute stress and fear and contains numerous PACAP-expressing neurons. My research focuses on the amygdala and the expression of PACAP following an acute stress. In my experiment, I quanitfied PACAP levels both with and without acute stress. After stressing rats using a forced swim protocol, I measured PACAP and mRNA expression in the amygdala using RNAscope in situ hybridization. Brain slices were imaged using fluorescence microscopy to quantify the RNA scope signal. To analyze these images quantitatively, I have developed a CellProfiler code to accurately and rapidly analyze the proportion of neurons expressing PACAP mRNA as well as the intensity of expression within the neurons. I hypothesize that PACAP mRNA expression will be increased in the rats subjected to the acute stressor, indicating that PACAP is an important mediator for the neural stress response. By determining this relationship between neuropeptide expression and stress response, PACAP antagonists could be potentially used in drug therapies to help patients experiencing stress related health conditions.

Roughly 20% of young adults in the United States experience some sort of mental health disorder. The majority of this research has focused on young adults in general or 4-year college students, however less is known about the mental health and coping strategies of young adults not currently attending school. The purpose of this study was to compare mental health, the use of alcohol and/or other substances as a coping mechanism for stress, and alcohol and marijuana use among young adults who attend 4-year colleges and those not attending school. A subsample (N=552; 56% female, 59.5% White, M age=20.58, SD=1.7) of young adults using baseline data from a larger longitudinal study on health behaviors was used for the current analyses. The subsample included 355 students currently attending 4-year college and 197 young adults not attending school (highest education status of these individuals included less than a high school diploma (1.5%), high school diploma (19.3%), some college (15.2%), 2-year degree (10.1%), 4-year college degree (52.8%), and graduate degree (1%). Findings indicated that young adults not in school reported significantly higher rates of depression, anxiety, and perceived stress in the past month than students in 4-year colleges. Similarly, findings indicated that non-students reported being high from marijuana for more hours, higher rates of alcohol use and coping with stress through alcohol and/or substance use in the past month than 4-year college students. Further research is needed, which focuses on young adults who are not currently attending 4-year colleges as they may face different life stressors and cope differently than 4-year college students.

Autism Spectrum Disorder (ASD) is a neurodevelopmental disorder involving impairments in social communication and repetitive and restrictive behaviors. Children with ASD are oftentimes diagnosed with cognitive impairment, specifically below average intelligence scores and atypical brain function in relation to early developmental regression. In order to analyze neural activity, Electroencephalogram (EEG) is used to monitor brain activation during certain tasks. EEG is a non-invasive tool that uses scalp electrodes to record neural activity frequency. EEG is divided into frequency bands, including the Alpha band frequency (neural activity between 6 and 12 Hz), which is present during the relaxed awake state of individuals and thought to be related to functional inhibition. Previous EEG studies consistently detected abnormalities in resting state alpha among individuals with ASD when compared to neurotypical counterparts. Findings suggested that reduced resting state alpha power was exhibited in brain regions involved in sensorimotor skills and higher-order cognitive functioning. Although many studies compare the difference between ASD group and neurotypical group, more research is needed to study the alpha power patterns within the ASD group. This study aims to compare EEG peak alpha power and average alpha power between high functioning (NVIQ>100) and low functioning children with ASD. 100 participants (50 males and 50 females) aged 6-18 participated in the study. All participants met criteria for autism diagnosis on a clinician administered standard interaction, and completed the DAS-II, which measures verbal and non-verbal cognitive abilities. Participants watched calm screensaver like videos while high density EEG was collected. We hypothesize that the children with ASD and lower functioning will have reduced average alpha power and reduced peak alpha frequency when compared to higher functioning children with ASD. The results of this study will provide insight into of differences within children with ASD.

The reduction of drug craving and subsequent relapse is central to the recovery process for individuals with substance use disorders. One of the goals of this project is to improve pharmacological treatment interventions for substance use disorders by better understanding how the neurotransmitter dopamine is involved in relapse. We investigated the relationship between drug craving and dopamine release in the nucleus accumbens core (NAC), an area of the brain that plays an integral role in reward-based learning and addiction, in male rats that have been trained to consume cocaine or alcohol. We tested the effects of L-DOPA, a chemical precursor to dopamine and a treatment that could reduce drug consumption, on craving and relapse-like behavior. Recent evidence from our lab suggests that decreased dopamine release in the NAC correlates with increased cocaine consumption in rats, and that L-DOPA administration attenuates drug intake. Preliminary data from our current project shows a similar effect of L-DOPA on alcohol or cocaine relapse. Our characterization of dopamine release in the NAC during both periods of drug abuse and withdrawal provides new insights for future studies and our use of innovative electrode implants for monitoring dopamine neurotransmission will pilot the technology for other drug self-administration studies. This will help to determine whether L-DOPA should be considered for treatment of cocaine and alcohol addiction in humans.

Autism spectrum disorder (ASD) is a developmental disorder in which children have difficulty with intercommunication, repetitive and restrictive behaviors, and atypical sensory processing. The CDC reports ASD is about 4.5 times more prevalent in males than in females. Less is known of gender differences in diagnosis and symptoms like atypical sensory processing. In a study of sensory behaviors in adults with ASD, females scored significantly higher in “unusual sensory response”. In past studies higher sensory processing sensitivities in ASD have been connected to more externalizing behaviors. This study’s aims are to: (1) compare sensory processing of typically developed (TD) children with children diagnosed with ASD, (2) determine if there is a difference in sensory sensitivity between females and males with ASD, (3) examine the relationship between sensory sensitivity and externalizing behaviors in adolescents with ASD, and (4) explore gender differences in the correlation between the two. 102 children with ASD (45 females, 57 males) and 105 TD children (57 males, 48 females) aged 11 to 17 completed the self-report adolescent Dunn Sensory Profile which asks about sensory related behaviors. Participants’ parents completed the Child Behavior Checklist, which provided scores of externalizing behaviors. We predict the children with ASD will have significantly greater sensory sensitivity than TD children, females with ASD will show more sensory sensitivity than males with ASD, and greater sensory sensitivity will relate to more externalizing behaviors. We will explore potential gender differences in correlation of externalizing behaviors and sensory sensitivity. Through this study, differences in the way males and females with ASD process sensory information and its relation to behavior may be found, furthering our understanding of gender differences and behaviors in ASD.

Autism spectrum disorder (ASD) is one of the fastest growing childhood disorders, becoming a major public health concern. Studies have shown that children with ASD are more likely to be exposed to perinatal complications than typically developing (TD) children. Sex differences have also been shown to affect ASD diagnoses. However, variables that affect severity of ASD are still understudied. Understanding factors that impact ASD severity may assist in establishing methods to identify ASD outcomes, and target children at high risk for worse outcomes with treatment. The study aims to investigate IQ differences in children with ASD on variables including birth weight, gestational age, sex, and birth order. 150 children (75 males and 75 females) aged 6 to 18 years with a confirmed diagnosis of Autism participated in the study. Autism diagnoses of the participants were confirmed via ADOS-2, a clinician-child measure that scores on child’s social, repetitive behaviors and communication skills. Parents completed a self-reported demographic questionnaire about family demographics, and the Autism Center of Excellence subject medical history form, a medical interview with information on pregnancy complications and child medical history. All subjects completed the DAS-II evaluation tool that measures the cognitive abilities (IQ) of children on verbal and non-verbal domains. We hypothesize that children with lower IQ levels (full scale IQ<80) will have lower average birth weight, average smaller gestational ages and a higher number of pregnancy related complications. We also expect that given the higher rate of ASD incidence in boys, within the ASD group, there will be a main effect of sex with more males (n=75) having higher rate of pregnancy complications than females (n=75). Results will help establish a better understanding of other prenatal variables that may be correlated to later severities of ASD, and provide support for treatment targeted at prenatal risk.

Autism Spectrum Disorder (ASD) is a collection of neurodevelopmental disorders characterized by social-communicative and behavioral impairments and affects approximately 1 in 68 children in the U.S. Children with ASD, on average, score lower on language measures than their typically developing peers and suffer from social impairments involving speech quality, social-communication, and interpersonal interaction. Children with ASD often report a desire for more peer social interaction, but have poor social support and more loneliness than their typically developing peers. In typical development, siblings can positively influence social skills and promote language, cognitive development, and understanding of emotions. This study (1) compared the social communication and language ability in children with ASD with siblings to those with ASD without siblings, (2) compared the severity level of autism spectrum-related symptoms between the two groups, and (3) investigated whether age differences of the siblings affect these measures. Participants included children with ASD with siblings (n=20) and children with ASD without siblings (n=50), between the ages of 8 and 17 years. Social communication was measured via clinical report using the Autism Diagnostic Observation Schedule (ADOS-2) and by parent report using the Social Responsiveness Scale (SRS-2). To assess language ability, participants completed the Differential Ability Scales, 2nd Edition (DAS-II) and the Clinical Evaluation of Language Fundamentals (CELF-4). To measure autism severity, we used ADOS-2. We predicted that (1) children with ASD with siblings would exhibit better social communication and higher language ability than those without siblings, (2) ASD children with siblings would experience less severe autism spectrum-related symptoms, and (3) ASD children who are closer in age with their siblings would show more benefit of having a sibling than those that are farther in age. The results of this study will provide insight into social and language benefits of siblings in children with ASD.

Diet, lifestyle, and environment can influence an individual’s behavior and brain function through epigenetic mechanisms. The Lattemann lab is investigating the effect that one factor, high fat diets, has on epigenetic regulation in hippocampus a key area for learning, memory and cognition. Previously, the lab determined that rats fed high fat diets exhibited increased sucrose motivation, and this behavioral effect was later linked to stearic and palmitic fatty acids explicitly. It was also shown that high fat, high sugar diets are associated with impaired place memory in rats. Most recently, the lab has analyzed hippocampal chromatin from rats fed high stearic and palmitic acid diets and is in the process of determining the effect of these diets on histone modifications. Histones regulate gene expression by binding to DNA: modifications to these histones influence how readily the DNA is transcribed, and how the gene expressed. These modifications are being screened for using Western blot analysis. In addition to screening for modifications in histones, the Lattemann lab also directly analyzed DNA for the expression of target genes. This screening procedure included measuring expression of candidate histone-target genes using PCR, and ChIP-Seq (Chromatin Immunoprecipitation and unbiased gene screening) assays. By locating histone modifications and quantitating the expression of hippocampal candidate genes, the lab has provided an epigenetic explanation for the behavioral effect of the diets, and a possible molecular mechanism by which the diets can induce this effect.

Autism Spectrum Disorder (ASD) is a neurodevelopmental disorder that impairs an individual's social, communicational, and behavioral skills. Males are 4.5 times more likely to be diagnosed with ASD compared to females. One possible reason for the difference in diagnostic rates is that females may be better able to mask their symptoms and therefore go undiagnosed more often than males. If females with autism are better at masking, then it is possible that standard diagnostic tools underestimate their impairment due to the clinician not seeing the full extent of individual's symptoms. The goal of this study is to understand the potential for sex bias in two measures of ASD by examining measurement consistency between parent and clinician report of ASD symptoms. 140 children (70 males, 70 females) aged 8-17 years with a confirmed diagnosis of autism participated in this study. Participants' diagnoses were confirmed via the ADOS-2, a clinician-child interaction measure that scores the child's communication, social skills, and restricted or repetitive behaviors. Parents completed the Social Responsiveness Scale (SRS-2), a survey about their child's social and autism-related impairments. To look at potential bias, we first compared males and females with ASD on both the ADOS and SRS to examine sex differences in autism severity. Second, we examined the relation between parent and clinician report for differences in consistency of ASD symptoms by sex. If females are better at masking their autistic traits, we expect a lower correlation between ADOS and SRS scores in females compared to males, indicating lower parent-clinician agreement. Third, we explore these analyses in children who had clinical diagnoses but who did not meet research criteria for ASD (n = 20). Understanding the relationship between sex and measurement tools should help inform whether traditional diagnostic measures are as effective in diagnosing both males and females.

Autism Spectrum Disorder (ASD) is a neurodevelopmental disorder characterized by marked impairments in social skills, communication, and behavioral aspects of typical development. Research has shown that individuals with ASD often present with psychiatric comorbidities such as Depression, Anxiety, Obsessive Compulsive Disorder, Attention Deficit Hyperactivity Disorder and Bipolar Disorder. Comorbidities can have a broad and significant impact on a child’s health and wellbeing and such conditions are significantly more prevalent in those with autism than in the general population. This study aims to explore the relationship between psychiatric comorbidities and autism severity in children with ASD. 100 children (males=50, females=50) aged 6-18 participated in the study. All participants met the Autism diagnostic criteria on the ADOS-2, a child-clinician interaction measure that measures child’s social, repetitive behaviors and communication skills. Parents completed the Autism Center of Excellence subject medical history form, with information on child medical history. Problem behaviors and low IQ scores in children with ASD may be the symptom of underlying psychiatric comorbidities instead of the autism itself. Due to this, treatment of comorbid medical conditions may result in dramatic improvements in the quality of life of both children with autism and their caregivers. In this study, we expect that children with a higher rate of psychiatric comorbidities will score higher ADOS calibrated severity scores with a higher rate of autism symptoms and will score lower on IQ measures. Results will help establish a better recognition of comorbid medical conditions as critically important issues to address in treatment approaches.
 

The mesolimbic dopamine pathway begins in the midbrain Ventral Tegmental Area (VTA) and projects to the nucleus accumbens (NAc) and is fundamental in regulating goal-directed behaviors. Abnormalities in this pathway are implicated in a number of neuropsychiatric disorders including drug addiction and depression. However, there is still a lot unknown about the specific circuitry and cell types that contribute to these behaviors and diseases. This project aims to provide a deeper understanding of how different dopamine populations in the VTA contribute to reward behavior. We use Cre-driver mouse lines to isolate genetically distinct VTA dopamine subpopulations. Using this strategy, we classified three different subpopulations: Corticotrophin releasing hormone receptor one (Crhr1-Cre), Cholecystokinin (Cck-Cre) and Tachykinin receptor three (Tacr3 Cre). Interestingly, these populations have inclusive and complementary projections to the NAc and it’s two subdivisions, the NAcCORE and NAcSHELL. We utilize optogenetic techniques to activate each subpopulation individually. To test learning acquisition, mice without any prior training were placed into an operant box containing two levers that when pressed would result in optical excitation of the targeted VTA subpopulation. We found that optical stimulation of the Crhr1 subpopulation, which projects to the NAcCORE, can drive lever pressing instrumental learning. Activation of Cck neurons, which project to the NAcSHELL, is unable to promote learning. However, if animals first learned an instrumental behavior to obtain food reward, subsequent activation of Cck neurons would maintain instrumental responding. Interestingly, activation of Tacr3 (projects to both NAcCORE and NAcSHELL), resulted in rapid instrumental learning and maintenance of high levels of lever pressing. These results establish independent roles for Crhr1 and Cck dopamine neurons in instrumental reward learning and motivation, as well as a requirement for co-activation of NAcCORE and NAcSHELL pathways (Tacr3) for maximal reinforcement responding, thus providing more specified targets for therapeutic interventions.

At one point or another, every student is faced with some sort of academic failure that can in turn have negative impacts on self-esteem or future academic performance. Being able to overcome such obstacles requires resiliency and the ability to view failure as a learning opportunity. Growth Mindset (GM) interventions aim to shift a student’s perception of intelligence as a quality that can be developed over time and enhance their approach to overcoming barriers. Originally, the GM was developed as an evidence-based intervention to be delivered by trained professionals. However, bringing this intervention into the real-world calls for a more feasible and accessible model. To bridge this gap, the current study utilizes the role of teachers as a valuable resource in building capacity within schools to attain sustainable, long-term goals regarding student success. The purpose of our research is to explore how a GM intervention delivered by teachers affects students’ beliefs about the brain’s ability to change (neuroplasticity). This project utilized a randomized control design with longitudinal follow-up to evaluate the efficacy of the GM intervention in comparison to curriculum as usual delivered in a rural district in the Pacific Northwest region. At two separate time points, middle and high school students completed an online self-report survey that assessed three constructs: theory of intelligence (whether intelligence can be changed), effort beliefs (the relationship between effort and outcomes), and learning goals (goals towards improving academic abilities). We hypothesize that students assigned to the intervention would show more positive implicit attitudes towards malleable intelligence, beliefs about effort, and learning goals. By understanding how students view themselves within an educational context, we can improve the measures taken to further engage and empower students. Future research should evaluate the quantitative outcomes of academic performance (i.e. grade point average) in relation to a GM intervention.

This project explores the relationship between anxiety and stimulant misuse. Both hormonal and social factors influence the prevalence rate of anxiety among females, which has consistently been higher than those of men. We hypothesize that anxious women are more likely than anxious men to take stimulant drugs in order to minimize the dysphoric feeling and relieve pressure. College students across 9 campuses who had previously reported at least one non-medical use of prescription stimulants in the last 12 months were invited to participate in a research study. Correlation analysis between anxiety and stimulant misuse and a moderation analysis were conducted. Consistent with our hypothesis, results revealed that women who experience symptoms of anxiety have a higher likelihood of misusing stimulant drugs compared to men who experience symptoms of anxiety. As gender moderated the relationship between anxiety and stimulant misuse, more attention needs to be paid to women with anxiety symptoms. Family members, friends and the society could provide more help and care for stressed females, preventing them from taking more stimulants.

The US homeless population is disproportionately affected by alcohol use disorder (AUD). This disparity leads to greater health and behavioral problems which precipitates greater utilization and associated costs of publicly funded services, such as the emergency healthcare and criminal justice systems. To address AUD in this population, researchers worked with homeless individuals with AUD and agencies that serve them to develop the Harm Reduction Treatment for Alcohol (HaRT-A). HaRT-A participants evinced improved alcohol outcomes compared to treatment-as-usual control participants. The present study’s aim was to test a secondary research question: Does HaRT-A significantly reduce the high costs from emergency healthcare and criminal justice system utilization in people experiencing homelessness and AUD? Participants (N=167; 24% women) were recruited for the parent HaRT-A study. Cost data were collected from participants’ use of the King County Correctional Facility and Harborview Medical Center. Jail costs included the costs of booking, days spent in jail, and use of supplementary jail services. Emergency healthcare system costs included the costs of emergency department visits and inpatient hospital admissions. All costs were added up for the 24 weeks prior and subsequent to study participation. Mann-Whitney U tests were conducted to assess group differences on post-pre cost change scores. Analyses indicated no significant group differences on post-pre changes in costs associated with utilization of emergency healthcare and jail services (ps > .58). There are several possible explanations for the lack of significant group differences. First, the time frame was likely too short to detect differences in statistically rare outcomes. Second, HaRT-A had a statistically significant impact on proximal outcomes; however, the brief nature of the treatment (4 sessions) might have limited its ability to impact the more distal variables measured in this study. Finally, homelessness creates multiple challenges that may require more intensive and multilevel interventions to fully address.

The lateral habenula (LHb) is part of the habenular complex in the dorsal diencephalon. Uniquely positioned anatomically to participate in reward-related brain circuits, this nucleus is an important regulator of the midbrain dopaminergic system that is known to be involved in the neuronal circuitry underlying various feeding behaviors. However, very little is known about the precise role of this nucleus in operant food self-administration and relapse to food seeking. Here, we utilized the DREADD (Designer Receptors Exclusively Activated by Designer Drugs) technology that we have successfully used to bidirectionally manipulate LHb neuronal activity associated with motivated behaviors. Male, Long Evans rats were infused with adeno-associated virus expressing Gi/o coupled human M4 (hM4Di) receptors into the LHb bilaterally. Approximately 10-12 days following viral infusion, rats were trained to self-administer food pellets for 10 days on a fixed ratio 1, 20 second timeout reinforcement schedule; food pellet delivery was paired with a compound tone and light cue. Activation of hM4Di by the pharmacologically inert synthetic ligand clozapine-N-oxide (CNO) activates G protein-activated inwardly rectifying K+ (GIRK) currents and results in hyperpolarization of neuronal activity. Our data indicate that activation of hM4Di in the LHb with CNO (1 and 3 mg/kg, i.p) had no influence on food reinforced operant responding, or the motivation to self-administer food under a progressive ratio schedule. Studies are currently underway to determine the effect of activation of Gi/o-coupled signaling in the LHb on the reinstatement of food seeking.

Dopamine neurotransmission is thought to be involved in economic decision making. One aspect of the decision making is the vigor by which a decision is made, and its concomitant action. Previous studies have found baseline/ambient dopamine levels to be connected to vigor, however much is still unknown about ambient dopamine. The lack of understanding of the role of ambient dopamine is due to technical limitations. The limitations of fast scan cyclic voltammetry (FSCV) lead us to fuse FSCV with fast-scan controlled adsorption voltammetry (FSCAV) which will allow us to take direct measurements of ambient dopamine. Here we seek to develop a new technical approach to better study ambient dopamine and its involvement in decision making. To test our hypothesis, we used fast-scan cyclic voltammetry to measure levels of dopamine by applying two triangle waveforms at 1Hz to measure electrodes. The electrodes were tested in vitro against different concentrations of dopamine (0.0625-1M), one time with a 50kOhm resistor and one time without. Subsequently, the ability to predict concentrations was tested on electrically evoked dopamine, in vivo. Here we demonstrate that the standard curves collected are significantly linear. We also tested the effect adding 50 kOhm resistor to the measuring circuit in order to mimic brain tissue impedance. As a result of adding this impedance, we found a significant drop in sensitivity. Moreover, preliminary data suggest that unknown concentrations can be predicted using this approach., however more work needs to be done both in vitro and in vivo. Through this analysis, we can extend our results to the investigation of dopamine transmission functions and assess decision making. This will help us determine the different alterations of dopamine transmission and behavior in hopes to further advance development of therapeutic strategies for mental illness.

Diagnostic tests in mental health can often lead to inconclusive diagnoses, or diagnoses that take months to come to. The focus of this study is to identify neuropsychiatric conditions earlier and more accurately using voice analysis. Our study tested the ability of a speech analysis software to determine whether a patient suffers from a given mental illness, since mental illness can affect speech patterns. For example, one standard tested against in depression is a slower speaking rate. This study examined the relationship between voice as an indicator of those illnesses by analyzing short, random speech samples from patients with a diagnosed psychological or neurological condition. Our large team of interns was divided into several partnerships that focused on collecting samples of one specific disease. These samples were then analyzed for speech markers related to that condition by initially extracting audio, linguistic, and prosodic features and then using various statistical tools, mainly in python, to build predictive models using these features. These samples, along with controls from unafflicted patients, were then cross-referenced against an objective scale rating of the given condition, such as the PHQ-9 questionnaire for depression. The degree of correlation revealed by this analysis was used to determine if specific voice features were predictive of illness. We hope to make this software accessible to individual practitioners in a mobile application, with the potential to allow an individual practitioner to identify mental and neurological conditions much sooner, more accurately, and less invasively, allowing for earlier and more effective treatment.

Serotonin (5-hydroxytryptamine, or 5-HT) signaling plays a key role in regulating a range of behaviors related to reward learning, motivation, and addiction. 5-HT6 receptors in particular are notable for their localization to primary cilia in neurons, and abundant expression in the striatum, a brain region critical for its role in mediating reward seeking behavior. In addition, 5-HT6 receptors signal through the cilia-specific type 3 adenylyl cyclase (AC3). Previous overexpression studies in my lab show that 5-HT6 receptors in the nucleus accumbens (NAc) modulate cocaine taking in a pathway-specific manner. To further characterize the role of endogenous 5-HT6 receptor signaling in cocaine taking, we are knocking out 5-HT6 receptor or AC3 expression in vivo using a pathway-specific CRISPR-Cas9 approach. We are injecting Cre-dependent AAV CRISPR-Cas9 into the NAc to target specific pathways, and we are validating successful knockout using immunohistochemistry (IHC). We are using an intersectional surgery approach, injecting CRISPR-Cas9 into the NAc, and Cre into either the VTA (direct pathway) or the Ventral Pallidum (indirect pathway) therefore only neurons with projections to both regions will express the CRISPR-Cas 9 enzyme. IHC staining is carried out on brain slices to confirm successful knockout of 5-HT6 or AC3 by staining for Cas9 and either AC3 or 5-HT6. The presence of Cas 9 positive neurons that lack 5-HT6 or AC3 staining will confirm the approach. We effectively stained for AC3 in rat brain slices, allowing us to validate AC3 knockout. Explicit staining of 5-HT6 and AC3 is crucial for continuing the study of 5-HT6 receptors signaling and their effects on cocaine reinforcements. This project aims to provide insight into the endogenous function of 5-HT6 receptor signaling in drug-seeking behavior.