Indiana University-Purdue University Indianapolis (IUPUI) / Social processing is the reception, interpretation, and reciprocation of social information and is critical for mental health. The neural structures, circuits, and substrates regulating these complex mechanisms are not well understood. Social processing in the form of social safety learning, as measured by a rat model of social familiarity-induced anxiolysis (SoFiA), was impaired following mild blast traumatic brain injury (mbTBI). Initial findings indicated that mbTBI altered resting state network activity in the orbitofrontal cortex (OFC) and was associated with accumulation of neurotoxin marker, acrolein, in lateral prefrontal cortex (PFC) (including OFC), indicating OFC as a brain region of interest that may contribute to social processing. Measuring GABA and Glutamate-related gene expression in OFC of mbTBI or sham-exposed rat brain revealed specific elevations of metabotropic glutamate receptor type 1 and 5 (mGluR1/5) expression in mbTBI but not sham OFC. Exposure-naïve rats intracranially injected with mGluR1/5 agonist demonstrated attenuated SoFiA, and this coincided with an impairment of social recognition (SR) behavior. Additionally, inactivation of OFC by local intracranial injection of GABAA agonist, muscimol, impaired two different measures of SR in which two conspecifics, or members of the same species, one novel and one familiar, were presented and required discrimination. Novelty seeking, decision-making, memory, and gregariousness were tested in isolation to determine OFC contributions to these specific behavioral contributions to SR test performance. OFC inactivation did not impair novelty seeking, non-social decision-making, or non-social memory as measured by novel object recognition (NOR) test, or gregariousness or social decision-making as measure by social preference (SP) test. When measuring SR behavior via consecutive presentation of two different conspecifics, OFC inactivation did not impact SR. Therefore, OFC is not directly responsible for social recognition, but rather the discrimination or ability to act upon discrimination of two simultaneously present conspecifics. These data suggest a novel role for OFC in high order processing or execution of action based on social information. / 2 years (2021-05-24)

Accurate and high throughput methods of measuring animal behavior are critical for many branches of neuroscience, allowing for mechanistic studies and preclinical drug testing. Methodological limitations contribute to narrow investigations, which may overlook the interplay between distinct but related behaviors, like affective behaviors and executive function (EF). To prevent such oversight, researchers can perform test batteries, or multiple assessments in one study. However, test batteries often exclude cognitive behaviors due to their lengthy testing period. This dissertation first reviews current evidence related to the investigation and relation of affective, pain-like, and operant behaviors in rodent models. Then, I demonstrate the use of traditional and novel test batteries to investigate these behavioral changes in multiple mouse models. First, I investigated affective and pain-like behavior in mice lacking Nape-pld, a key enzyme that synthesizes lipid mediators which activate receptors in the endocannabinoid system. I found that these mice displayed reduced sucrose preference, but otherwise normal anxiety- and depression-like behavior, and had baseline differences in thermal nociception and inflammation response. Then, I investigated the affective, pain-like, and operant effects of chronic vapor exposure (CVE) to vehicle or nicotine (NIC). Regardless of NIC content, acute abstinence from CVE increased mechanical sensitivity and self-grooming, while chronic abstinence from NIC CVE resulted in motor stimulation. Other traditional anxiety- and depression-like behaviors were unchanged by CVE. In an operant test battery, acute abstinence from NIC CVE impaired acquisition, decreased sucrose motivation, and impaired the response to aversive rewards. Finally, I developed a protocol for the high throughput analysis of six operant tests which can be completed in as few as nineteen sessions, significantly fewer sessions than traditional operant tests. This battery investigates multiple aspects of goal-directed behavior and EF including operant acquisition, cognitive flexibility, reward devaluation, motivation via response to increased instrumental effort, cue devaluation or the extinction of learned behavior, and reacquisition. I validated several of these tests by demonstrating that lesions to specific subregions of the orbitofrontal cortex impaired cognitive flexibility and altered response to instrumental effort as observed in traditional operant tests. I then used this battery to investigate the effects of the P129T mutation, which results in a mutated version of the Fatty Acid Amide Hydrolase (FAAH) enzyme that is associated with addiction, in male and female mice. Knock-in animals displayed reduced activity in response to increasing instrumental effort, and reduced activity on the first day of an extinction test. Then, to encourage others to use this new operant battery I outlined how to efficiently collect data, shared a database for customizable analysis, and described common issues and how to solve them. This protocol has potential implications for many aspects of neuroscience including the investigation of novel therapeutics and the neural circuitry underlying behaviors. Together, the information in this dissertation demonstrates the utility of multi-faceted behavioral assays and the combination of traditional and novel approaches to collect more comprehensive behavioral data, which will allow researchers to better investigate neural circuitry underlying behaviors or the behavioral changes associated with novel therapeutics. / Doctor of Philosophy / By measuring animal behavior researchers can gain insight into how specific brain regions interact to influence choice and action. Limitations in testing methods mean that researchers may fail to investigate the relationship between distinct aspects of behavior, like the influence of emotional state or pain on cognition. To prevent such oversight researchers can perform a test battery, a specific series of multiple tests that measures several different aspects of behavior. Traditional test batteries often overlook cognitive or operant (learning to perform an action for reward) behaviors due to time constraints, which limits their translational potential. This dissertation provides a brief overview of the ways that researchers investigate affective (emotional), pain-like (physical discomfort), and goal-directed behaviors. It further has a broad focus on mouse models related to addiction or the endocannabinoid system (ECS), which is shown to play a role in mood, pain (e.g., perception, relief, and inflammation), and cognition. Using a traditional test battery, we demonstrate that mice lacking a key enzyme in the ECS have altered responses to sugar, heat, and inflammation, but display otherwise normal performance in anxiety-, depression-, and pain-like tests. Next, we used a combined traditional and operant battery to investigate the effects of chronic vapor exposure (CVE) and nicotine in mice. We found that regardless of nicotine content, acute abstinence from CVE increased physical sensitivity and self-grooming but spared other anxiety- and depression-like behaviors. Acute abstinence from nicotine CVE resulted in motor stimulation, impaired operant learning, lower motivation for sucrose reward, and an impaired ability to withhold responding when presented with a bitter reward. Finally, I outline a novel operant test battery that addresses the limitations of current operant chamber- or place-based batteries. Using this battery, I first demonstrate that it captures similar behavioral changes to those seen in traditional operant chambers. Then, I demonstrate that mice containing an ECS mutation associated with problem drug use in humans display less motivation for food reward in response to increased effort, and more quickly inhibit a learned behavior when reward delivery is interrupted. I also found that in response to increased effort for reward or bitter rewards, male mice are more likely to alter their behavioral strategy. To encourage others to use this new operant battery I outlined how to efficiently collect data, shared a database for customizable analysis, and described common issues and how to solve them. This protocol has the potential to improve upon traditional tasks while opening cognitive research to more scientists. This has implications for many fields of neuroscience, especially the investigation of novel therapeutics and investigation of the neural circuitry underlying various disorders.

The DNA base excision repair enzyme, poly(ADP-ribose) polymerase-1 (PARP1), is a multi-functional enzyme and a member of a subfamily of three PARPs that covalently build PAR polymers onto proteins to regulate their function. Drug inhibitors of PARPs have anti-cancer, anti-inflammatory, and neuroprotective effects. Recently, we reported elevated gene expression levels of PARP1 in postmortem brain tissues from donors who had an active major depressive disorder at the time of death. Since PARP1 gene expression is positively correlated with PARP1 activity, these findings indicate that elevated PARP1 activity may contribute to brain pathology associated with depressive behavior. Therefore, we speculated that drug inhibitors of PARP1 may have antidepressant properties. To determine whether a rodent model could be used to evaluate the role of PARP1 in depressive-like behaviors, rats were exposed to repeated psychological stressors (social defeat and chronic unpredictable stress) for 10 days. Anhedonia (estimated by sucrose preference) and brain PARP1 gene expression levels were measured. After stress exposure, rats exhibited significantly reduced sucrose preference and significantly higher levels of brain PARP1 gene expression. To examine potential antidepressant activity of PARP inhibitors, rats were administered PARP inhibitors or saline vehicle and were exposed to the Porsolt swim test or repeated social defeat and chronic unpredictable stress. Two PARP inhibitors were investigated, 3-aminobenzamide (3-AB) and 5-aminoisoquinolinone (5-AIQ). PARP inhibitors produced antidepressant-like effects in the Porsolt swim test similar to the common antidepressant fluoxetine by significantly decreasing immobility time and increasing latency to immobility. PARP1 inhibitors did not significantly affect locomotor activity or swim speeds, suggesting that antidepressant-like actions of these drugs were not secondary to a stimulant effect. Treatment of rats with a combination of 3-AB and fluoxetine, at low doses of these drugs that individually did not have antidepressant-like effects, significantly decreased immobility time and increased latency to immobility in the swim test. Finally, treatment of rats with 3-AB significantly increased sucrose preference and social interaction times relative to vehicle-treated control rats following repeated exposure to combined social defeat and unpredictable stress, exhibiting effects similar to fluoxetine treatment. These findings uncover PARP1 as a unique molecular target for the development of a novel class of antidepressants that could be used alone or in combination with existing antidepressants.

Epilepsy is a complex, chronic neurological disorder with diverse underlying etiologies characterized by the spontaneous occurrence of seizures. Epilepsy affects all ages from neonates to elderly adults, with the most recent CDC estimates stating that ~3 million adults and over 400,000 children are currently suffering from active epilepsy in the U.S. alone. In adults, the leading cause of epilepsy worldwide in central nervous system (CNS) infection, while in neonates the most common cause of seizures is hypoxic/ischemic encephalopathy (HIE). However, in both adults and neonates, current antiepileptic drugs (AEDs) are ineffective in 30-50% of patients, despite the availability of over 20 FDA approved AEDs with diverse molecular targets. This disparity highlights a critical need for novel therapeutics in seizure-susceptibility and epilepsy. The microbes that inhabit gut mucosal surfaces, termed the gut microbiota, have been increasingly implicated in the pathology of neurological diseases including epilepsy. This gut-brain axis is an intriguing therapeutic target in epilepsy as gut microbes can affect the CNS through multiple mechanisms including vagus nerve signaling, immune-gut interactions, and through production of microbial-metabolites including neurotransmitters, short chain fatty acids (SCFAs), lactate, vitamins, and S-equol. Furthermore, the gut microbiota is crucial for neurodevelopment, indicating that the gut-brain axis may be involved in pediatric seizure-susceptibility. This dissertation reviews current evidence on the role of gut metabolites in seizure-susceptibility in epilepsy, highlighting the microbial-derived metabolite S-equol as a potential novel AED. We then evaluate gut microbiome alterations in the Theiler's murine encephalomyelitis virus (TMEV) adult mouse model of CNS infection-induced seizures and find decreases in S-equol-producing bacteria in the gut microbiomes of TMEV-infected mice with seizure phenotypes. We characterize the effect of exogenous S-equol on neuronal function in vitro, demonstrating a reduction in neuronal excitation following S-equol exposure. We additionally characterize entorhinal cortex (ECTX) pyramidal neuronal hyperexcitability, and demonstrate the ability of exogenous S-equol to ameliorate CNS-infection-induced ECTX neuronal hyperexcitability ex vivo. Finally, we demonstrate that perinatal and postnatal exposure to antibiotics alters the gut microbiome and increases seizure-susceptibility following HIE exposure in p9/p10 mice, potentially via sex-specific alterations in neuronal function. Together, this dissertation evaluates the gut-brain axis in pediatric and adult mouse models of seizure-susceptibility and identifies the gut metabolite S-equol as a potential target for the treatment of seizures. / Doctor of Philosophy / Epilepsy, a disease defined by the occurrence of two or more spontaneous seizures, affects over 50 million people worldwide. This makes epilepsy one of the most common chronic neurological disorders across the globe. People with epilepsy suffer increased mortality, lower quality of life, and increased social stigma. There is currently a crisis in the treatment and management of epilepsy, because although over 20 different anti-epileptic drugs (AEDs) are available to patients, these drugs only work in ~70% of individuals with epilepsy, leaving 30% of patients with uncontrolled seizures. Currently available AEDs are designed to target classical central nervous system (CNS) components. However, a growing body of evidence suggests that epilepsy is related to complex systems throughout the body. Therefore, in this manuscript we explore novel therapeutic targets outside of the CNS for the management of seizures. Over 1000 species of bacteria live in the in the human gut, and are termed the gut microbiota. Gut microbes produce a variety of chemicals that circulate through the body and can even reach the brain. Interaction of chemicals produced by the gut microbiota and brain chemistry have been shown to affect disease outcomes in Autism Spectrum Disorder, Parkinson Disease, and other brain disorders. However, very few studies have examined the possibility of a role for the gut microbiota in epilepsy. In this dissertation, we review chemicals produced by the gut microbiota that may alter epilepsy biology. We additionally examine gut microbiota alterations in a rodent model of epilepsy, and identify a novel chemical, S-equol, that is produced by the gut microbiota and impacts epilepsy biology in our rodent model. Lastly, we explore how altering the maternal gut microbiota in rodents can influence seizure-susceptibility in infants.

<I>Neospora hughesi</I> is a recently described cause of equine protozoal myeloencephalitis (EPM). In the present study, we examined the susceptibility of BALB/c gamma-interferon gene knockout (gamma-INFKO), BALB/c, CD-1, and C57BL/6 strains of mice and gerbils to infection with tachyzoites of the Nh-A1 strain of <I>N. hughesi</I>. Only the gamma-IFNKO mice developed severe clinical disease following infection with <I>N. hughesi</I>. The most severe lesions were in the hearts of these mice. Two dogs fed the brains of mice, shown to contain <I>N. hughesi</I> tissue stages by cell culture and g-IFNKO mouse bioassay, did not shed <I>N. hughesi</I> oocysts over a 23 day observation period. We report important differences between the nucleotide and deduced amino acid sequences of the dense granule proteins GRA6 and GRA7 of <I>N. hughesi and N. caninum</I>. The newly defined proteins of <I>N. hughesi</I> are referred to as NhGRA6 and NhGRA7. From analysis of the sequences we found that there is a 14.8% difference in deduced amino acid sequence between NhGRA7 and NcGRA7, and a 4% difference between NhGRA6 and NcGRA6 in areas that could be compared. This thesis supports the identification of <I>N. hughesi</I> as a separate species from <I>N. caninum</I> and describes novel methods of distinguishing between the two. / Master of Science

No description available.

Electroconvulsive therapy (ECT) is one of the oldest and most effective treatments for depression; however, its biological underpinnings are poorly understood. Brain-derived neurotrophic factor (BDNF) and the hypothalamic-pituitary-adrenal (HPA) axis are two chemical messenger systems implicated in the antidepressant action and cognitive side effects of ECT. The Wistar-Kyoto (WKY) strain is a genetic model of depression that shows biological, cognitive, behavioural, and treatment-response abnormalities, making it potentially a useful model in which to investigate the underpinnings of the action of electroconvulsive stimulus (ECS: the amimal model of ECT). In addition, the WKY presents a potentially useful model for translational research on depression. The WKY strain is particularly valuable for the measurement of serum BDNF protein, for which the association with antidepressant treatments is much less clear (mostly stemming from investigations in humans) than that between brain BDNF and antidepressant treatments in rodent studies. The three studies presented add insight into the biological and behavioural effects of ECS. The first study (chapter 2) found no evidence of increased (R)-[11C]rolipram binding (an indirect marker of cyclic-adenosine monophosphate, cAMP) in the brain, despite significant increases of brain BDNF protein expression after repeated ECS. The second study (chapter 3) demonstrated the validity of the WKY strain in the investigation of ECS. Relative to Wistar controls, WKY showed similar antidepressant and cognitive effects (despite some abnormal behavioural responses), immediate but not sustained increases in brain BDNF protein, and a novel finding of increased extra-hypothalamic CRF after 5 daily ECS. The final study (chapter 4) demonstrated baseline strain differences in serum (WKY < Wistar) but not brain BDNF and, in both strains, no change in serum BDNF despite significant changes in brain BDNF after repeated ECS treatment. Preliminary results from a human pilot study investigating similar measures in a small group of people receiving ECT for depression are also presented. The results of this body of work advance our understanding of the activation and role of brain and serum measures of BDNF and the HPA axis in ECS/ECT, and raise important issues in the translation of research from basic science to the human condition of depression.

Electroconvulsive therapy (ECT) is one of the oldest and most effective treatments for depression; however, its biological underpinnings are poorly understood. Brain-derived neurotrophic factor (BDNF) and the hypothalamic-pituitary-adrenal (HPA) axis are two chemical messenger systems implicated in the antidepressant action and cognitive side effects of ECT. The Wistar-Kyoto (WKY) strain is a genetic model of depression that shows biological, cognitive, behavioural, and treatment-response abnormalities, making it potentially a useful model in which to investigate the underpinnings of the action of electroconvulsive stimulus (ECS: the amimal model of ECT). In addition, the WKY presents a potentially useful model for translational research on depression. The WKY strain is particularly valuable for the measurement of serum BDNF protein, for which the association with antidepressant treatments is much less clear (mostly stemming from investigations in humans) than that between brain BDNF and antidepressant treatments in rodent studies. The three studies presented add insight into the biological and behavioural effects of ECS. The first study (chapter 2) found no evidence of increased (R)-[11C]rolipram binding (an indirect marker of cyclic-adenosine monophosphate, cAMP) in the brain, despite significant increases of brain BDNF protein expression after repeated ECS. The second study (chapter 3) demonstrated the validity of the WKY strain in the investigation of ECS. Relative to Wistar controls, WKY showed similar antidepressant and cognitive effects (despite some abnormal behavioural responses), immediate but not sustained increases in brain BDNF protein, and a novel finding of increased extra-hypothalamic CRF after 5 daily ECS. The final study (chapter 4) demonstrated baseline strain differences in serum (WKY < Wistar) but not brain BDNF and, in both strains, no change in serum BDNF despite significant changes in brain BDNF after repeated ECS treatment. Preliminary results from a human pilot study investigating similar measures in a small group of people receiving ECT for depression are also presented. The results of this body of work advance our understanding of the activation and role of brain and serum measures of BDNF and the HPA axis in ECS/ECT, and raise important issues in the translation of research from basic science to the human condition of depression.

Electroconvulsive therapy (ECT) is one of the oldest and most effective treatments for depression; however, its biological underpinnings are poorly understood. Brain-derived neurotrophic factor (BDNF) and the hypothalamic-pituitary-adrenal (HPA) axis are two chemical messenger systems implicated in the antidepressant action and cognitive side effects of ECT. The Wistar-Kyoto (WKY) strain is a genetic model of depression that shows biological, cognitive, behavioural, and treatment-response abnormalities, making it potentially a useful model in which to investigate the underpinnings of the action of electroconvulsive stimulus (ECS: the amimal model of ECT). In addition, the WKY presents a potentially useful model for translational research on depression. The WKY strain is particularly valuable for the measurement of serum BDNF protein, for which the association with antidepressant treatments is much less clear (mostly stemming from investigations in humans) than that between brain BDNF and antidepressant treatments in rodent studies. The three studies presented add insight into the biological and behavioural effects of ECS. The first study (chapter 2) found no evidence of increased (R)-[11C]rolipram binding (an indirect marker of cyclic-adenosine monophosphate, cAMP) in the brain, despite significant increases of brain BDNF protein expression after repeated ECS. The second study (chapter 3) demonstrated the validity of the WKY strain in the investigation of ECS. Relative to Wistar controls, WKY showed similar antidepressant and cognitive effects (despite some abnormal behavioural responses), immediate but not sustained increases in brain BDNF protein, and a novel finding of increased extra-hypothalamic CRF after 5 daily ECS. The final study (chapter 4) demonstrated baseline strain differences in serum (WKY < Wistar) but not brain BDNF and, in both strains, no change in serum BDNF despite significant changes in brain BDNF after repeated ECS treatment. Preliminary results from a human pilot study investigating similar measures in a small group of people receiving ECT for depression are also presented. The results of this body of work advance our understanding of the activation and role of brain and serum measures of BDNF and the HPA axis in ECS/ECT, and raise important issues in the translation of research from basic science to the human condition of depression.

Electroconvulsive therapy (ECT) is one of the oldest and most effective treatments for depression; however, its biological underpinnings are poorly understood. Brain-derived neurotrophic factor (BDNF) and the hypothalamic-pituitary-adrenal (HPA) axis are two chemical messenger systems implicated in the antidepressant action and cognitive side effects of ECT. The Wistar-Kyoto (WKY) strain is a genetic model of depression that shows biological, cognitive, behavioural, and treatment-response abnormalities, making it potentially a useful model in which to investigate the underpinnings of the action of electroconvulsive stimulus (ECS: the amimal model of ECT). In addition, the WKY presents a potentially useful model for translational research on depression. The WKY strain is particularly valuable for the measurement of serum BDNF protein, for which the association with antidepressant treatments is much less clear (mostly stemming from investigations in humans) than that between brain BDNF and antidepressant treatments in rodent studies. The three studies presented add insight into the biological and behavioural effects of ECS. The first study (chapter 2) found no evidence of increased (R)-[11C]rolipram binding (an indirect marker of cyclic-adenosine monophosphate, cAMP) in the brain, despite significant increases of brain BDNF protein expression after repeated ECS. The second study (chapter 3) demonstrated the validity of the WKY strain in the investigation of ECS. Relative to Wistar controls, WKY showed similar antidepressant and cognitive effects (despite some abnormal behavioural responses), immediate but not sustained increases in brain BDNF protein, and a novel finding of increased extra-hypothalamic CRF after 5 daily ECS. The final study (chapter 4) demonstrated baseline strain differences in serum (WKY < Wistar) but not brain BDNF and, in both strains, no change in serum BDNF despite significant changes in brain BDNF after repeated ECS treatment. Preliminary results from a human pilot study investigating similar measures in a small group of people receiving ECT for depression are also presented. The results of this body of work advance our understanding of the activation and role of brain and serum measures of BDNF and the HPA axis in ECS/ECT, and raise important issues in the translation of research from basic science to the human condition of depression.