Medical Neuroscience Department Theses and Dissertations

Permanent URI for this collection

https://hdl.handle.net/1805/1721

Browse

Now showing 1 - 10 of 89

Aberrant Neural Activity in Cortico-Striatal-Limbic Circuitry Underlies Behavioral Deficits in a Mouse Model of Neurofibromatosis Type 1
(2022-05) Drozd, Hayley Paulina; McKinzie, David L.; Clapp, D. Wade; Shekhar, Anantha; Lukkes, Jodi L.; Lapish, Christopher L.; Block, Michelle L.
Nearly 18% of children are diagnosed with developmental disabilities. Autism spectrum disorders (ASDs) and attention deficit hyperactivity disorder (ADHD) are increasingly common developmental disabilities, but neither is well understood. ADHD and ASD are both prevalent in the genetic disorder Neurofibromatosis type 1 (NF1) which impairs the Ras-MAPK/ERK pathway through mutation of the neurofibromin gene (NF1+/−). More broadly, syndromic forms of developmental disorders are often caused by mutations of proteins in pathways interconnected with Ras including TSC1/2, FMR1, and SynGAP. Of NF1 patients, around 30-50% are diagnosed with ASDs and more than 60% with ADHD. These studies are the first to show that male mice haploinsufficient for the Nf1 gene (Nf1+/−) exhibit deficits in behavioral inhibition in multiple contexts, a key feature of ADHD. They exhibit hyperactivity and impulsivity in an open field, delay discounting task, and cliff avoidance reaction test, rescuable through treatment with the clinically effective ADHD drug, guanfacine (α2A adrenergic receptor agonist). Previous experiments in our lab identified social deficits including deficits in consolidation of social memory. Using optogenetics and awake behaving electrode recordings, we explored the role of the cortico-striatal-limbic circuitry in impulsivity and in social deficits in male Nf1+/− mice. Manipulation of the prefrontal cortex, nucleus accumbens, or basolateral amygdala through optogenetics rescued social deficits. These studies are the first to record brain activity in a preclinical model of NF1 during impulsive behavior, finding broad spectrum changes across slow, delta, theta, and gamma oscillatory frequencies and decreased synchrony of the prefrontal cortex and nucleus accumbens during a delay discounting task. Overall, Nf1+/− male mice with deletion of a single NF1 gene recapitulate cognitive phenotypes of NF1 patients and are a useful model system to identify alterations in neural circuitry associated with ASD and ADHD.
Air Pollution Exposure and the Lung-Brain Axis: Implications for Alzheimer's Disease
(2022-03) Greve, Hendrik Jacob; Oblak, Adrian; Block, Michelle; Nass, Richard; Landreth, Gary
Alzheimer’s disease (AD) is a devastating neurodegenerative disease that is expected to affect approximately 6.2 million Americans. Despite its high prevalence, the mechanisms underlying AD remain poorly understood. In recent years, increasing reports indicate that exposure to urban air pollution is a risk factor for the development of AD. However, the mechanistic underpinnings of this association are not well studied. Rats exposed to diesel exhaust (DE) showed neuroinflammation and impaired expression of TREM2 and disease-associated microglia (DAM), a cell subtype hypothesized to play beneficial roles during neurodegeneration, markers. Microglia in the cortex of rats exposed to DE, also showed decreased association with the vasculature, providing a novel link between the microglia and the brain vasculature. Examining the functional role of TREM2 during DE exposures, Trem2-/- mice showed an altered pro-inflammatory profile in both the brain and lungs in response to DE particles as well as altered phagocytic oxidase related gene expression. Examining another prominent component of air pollution, ozone (O3), in a mouse model of AD, it was discovered that subchronic O3 exposure exacerbates amyloid pathology through impaired microglial-plaque association in 5xFAD mice. 5xFAD mice exposed to O3 also showed increased expression of pro-inflammatory cytokines, increased markers of dystrophic neurites, and decreased expression of key acetylcholinergic pathway components. Examining the peri-plaque microenvironment, it was discovered that O3 dysregulates key DAM proteins and amyloid processing proteins. In the lung, it was found that O3 exacerbated immune cell infiltration in 5xFAD mice compared to WT controls, suggesting that ongoing amyloid pathology regulates pulmonary immune response to air pollution. To examine how O3-induced pulmonary immune responses may be signaling to the CNS, we examined the serum of 5xFAD mice, where HMGB1, VEGF, and IL-9 were upregulated. Injection of rHMGB1 into mice showed similar gene changes to 5xFAD mice exposed to O3, along with impaired Trem2 expression. Using a peripheral myeloid specific knock-out model of HMGB1, we also show that HMGB1 regulates O3-induced Trem2 expression impairment. Taken together, these data support that air pollution exposure impairs TREM2, DAM cells, and the microglial plaque response through a bidirectional lung-brain axis to exacerbate AD-like pathology.
Assessment of Ethanol and Nicotine Interactions in the Rat Model: Pharmacotherapeutics, Adolescence, and the Mesolimbic System
(2019-09) Waeiss, Robert Aaron; Truitt, William A.; Hudmon, Andy; Johnson, Philip L.; McBride, William J.; Rodd, Zachary A.
Alcohol use disorder (AUD) and nicotine dependence often result in serious health problems and are top contributors to preventable deaths worldwide. Co-addiction to alcohol and nicotine is the most common form of polysubstance abuse. Epidemiological studies indicate that more than 80% of individuals diagnosed with AUD concurrently use nicotine. The prevalence of alcohol and nicotine comorbidity may stem from interconnected mechanisms underlying these disorders. A better understanding of how these drugs interact and the biological basis behind the high comorbidity rates could generate key targets for the development of more effective treatments for AUD and nicotine dependence. The following experiments utilized four similar overall groups consisting of vehicle, ethanol (EtOH), nicotine (NIC), and EtOH+NIC. Chapter Two investigated the efficacy of naltrexone and varenicline, the pharmacological ‘gold standards’ for treating AUD and nicotine dependence, on voluntary drug intake by rats selectively bred for high EtOH drinking. The results indicated that the standard treatments for AUD and nicotine dependence were effective at reducing consumption of the targeted reinforcer but neither reduced EtOH+NIC co-use/abuse. Chapter Three examined the effects of peri-adolescent EtOH drinking on the ability of NIC infused into the posterior ventral tegmental area (pVTA) to stimulate dopamine release within the nucleus accumbens (NAc) shell during adulthood. The results suggest a cross-sensitization to NIC produced by peri-adolescent EtOH consumption demonstrated by a leftward and upward shift in the dose response curve. Chapter Four investigated the effects of intra-pVTA infusions on NAc shell neurochemistry, EtOH reward within the NAc shell, and the role of brain-derived neurotrophic factor (BDNF) on EtOH reward within that region. The data indicated that only EtOH+NIC significantly increased glutamate, dopamine, and BDNF in the NAc shell. Repeated pretreatment with EtOH+NIC also enhanced EtOH reward in the NAc shell and BDNF infusions were sufficient to recapitulate these findings. Collectively, the data indicate that concurrent exposure to EtOH and NIC results in unique neuroadaptations that promote future drug use. The failure to develop effective pharmacotherapeutics for AUD or nicotine dependence could be associated with examining potential targets in models that fail to reflect the impact of polydrug exposure.
Assessment of the dopamine system in addiction using positron emission tomography
(2014) Albrecht, Daniel Strakis; Hutchins, Gary D.; Saykin, Andrew J.; Kareken, David A.; Yoder, Karmen K.; Grahame, Nicholas J.
Drug addiction is a behavioral disorder characterized by impulsive behavior and continued intake of drug in the face of adverse consequences. Millions of people suffer the financial and social consequences of addiction, and yet many of the current therapies for addiction treatment have limited efficacy. Therefore, there is a critical need to characterize the neurobiological substrates of addiction in order to formulate better treatment options. In the first chapter, the striatal dopamine system is interrogated with [11C]raclopride PET to assess differences between chronic cannabis users and healthy controls. The results of this chapter indicate that chronic cannabis use is not associated with a reduction in striatal D2/D3 receptor availability, unlike many other drugs of abuse. Additionally, recent cannabis consumption in chronic users was negatively correlated with D2/D3 receptor availability. Chapter 2 describes a retrospective analysis in which striatal D2/D3 receptor availability is compared between three groups of alcohol-drinking and tobacco-smoking subjects: nontreatment-seeking alcoholic smokers, social-drinking smokers, and social-drinking non-smokers. Results showed that smokers had reduced D2/D3 receptor availability throughout the striatum, independent of drinking status. The results of the first two chapters suggest that some combustion product of marijuana and tobacco smoke may have an effect on striatal dopamine concentration. Furthermore, they serve to highlight the effectiveness of using baseline PET imaging to characterize dopamine dysfunction in addictions. The final chapter explores the use of [18F]fallypride PET in a proof-of-concept study to determine whether changes in cortical dopamine can be detected during a response inhibition task. We were able to detect several cortical regions of significant dopamine changes in response to the task, and the amount of change in three regions was significantly associated with task performance. Overall, the results of Chapter 3 validate the use of [18F]fallypride PET to detect cortical dopamine changes during a impulse control task. In summary, the results reported in the current document demonstrate the effectiveness of PET imaging as a tool for probing resting and activated dopamine systems in addiction. Future studies will expand on these results, and incorporate additional methods to further elucidate the neurobiology of addiction.
The Behavioral Role of Mu Opioid Receptors in Glutamatergic Neurons
(2021-10) Reeves, Kaitlin C.; Sheets, Patrick; Baucum, Anthony II; Yamamoto, Bryan; McKinzie, David; Yoder, Karmen
Mu opioid receptors (MORs) mediate the analgesic and rewarding effects of opioids. Most research has focused on MORs in GABAergic neurons; however, MORs are also in glutamatergic neurons and their role in opioid-related behaviors was unclear. Our lab previously showed that MORs inhibit glutamate transmission from vesicular glutamate transporter 2 (vGluT2)-expressing thalamostriatal synapses. The behavioral relevance of MORs in vGluT2-expressing neurons was unknown; therefore, I utilized a conditional MOR knockout mouse with MORs deleted in vGluT2-expressing neurons (MORflox-vGluT2cre). MORflox-vGluT2cre mice have disrupted opioid reward, locomotor stimulation, and withdrawal, compared to cre-recombinase negative littermate controls. However, other MOR-mediated behaviors, including opioid-induced antinociception, alcohol reward, and palatable substance consumption are intact. MORs are expressed in vGluT2 neurons in several reward-related brain regions, including the thalamus and lateral habenula (LHb). To determine whether MORs in these brain regions modulate opioid-related behaviors, an adeno-associated viral (AAV) vector encoding cre-recombinase was stereotaxically injected into the thalamus or LHb of MORflox mice to specifically delete MORs in these brain regions. Opioid reward and locomotor stimulation remained intact in both thalamic and LHb MOR knockout mice; however, basal locomotor activity was increased in LHb MOR knockout mice. Sucrose consumption was also intact in LHb MOR knockout mice. Interestingly, in LHb MOR KO mice opioid withdrawal-induced paw shakes were increased, while withdrawal-induced jumping was completely ablated. Our lab previously showed that MORs inhibit glutamate transmission from the anterior insular cortex (AIC), which is disrupted by in vivo alcohol exposure. To determine the role of AIC MORs, AIC MORs were deleted with AAV vectors. AIC MOR knockout mice had intact opioid, sucrose, and alcohol reward, but had increased basal locomotor activity. MORs in glutamatergic neurons are critical mediators of opioid reward; however, the specific glutamatergic neurons mediating the rewarding effects of opioids remains to be determined.
Brain Responses to Sugar: Implications for Alcohol Use Disorder and Obesity
(2024-05) Alessi, Jonathan P.; Yoder, Karmen K.; Kareken, David A.; Džemidžić, Mario; Considine, Robert V.; Harezlak, Jaroslaw
Obesity and alcohol use may together account for 640,000 adult deaths each year in the United States. In both cases, overconsumption drives untoward effects. Alcohol use and obesity also both relate to sweet liking, as sugar consumption is consistently linked to weight gain and intense sweet liking has been linked to an inherited risk for alcohol use disorder (AUD). However, the neural underpinnings of these associations are largely unknown. Thus, we used sugar-sweetened water administration during functional magnetic resonance imaging (fMRI) to probe these relationships in two studies. In the first, we tested the relationship between a known AUD risk factor, subjective response to alcohol, and the brain response to both sucrose and monetary reward in 140 young adults. We found a significant positive correlation between the enjoyable component of subjective responses to a standardized intravenous alcohol exposure and activation to high-concentration sucrose (but not monetary reward) in the right dorsal anterior insula and the supplementary motor area, supporting a role for these regions in AUD risk. In the second study, we investigated the neural mechanisms of sweet liking decreases following bariatric surgery, the most effective obesity treatment. Here, we evaluated the change in brain activation to sucrose in 24 women before (BMI 47.0 + 6.9 kg/m2) and 21 women after (BMI 37.6 + 6.5 kg/m2) bariatric surgery and compared the pre- and post-surgical activation patterns to those of 21 normal to overweight (BMI 23.5 + 2.5 kg/m2) control participants. Brain activation did not differ between controls and surgery participants at either time point. However, activation to sucrose in reward, but not sensory, regions decreased significantly after surgery, consistent with reduced drive to consume sweet foods. Together, these studies highlight the utility of quantifying brain responses to sweet taste as a method to understand the mechanisms underlying overconsumptive behavior.
CaMKII Phosphorylation of the Voltage-Gated Sodium Channel Nav1.6 Regulates Channel Function and Neuronal Excitability
(2021-01) Zybura, Agnes Sara; Cummins, Theodore R.; Hudmon, Andy; Baucum II, Anthony J.; Sheets, Patrick L.
Voltage-gated sodium channels (Navs) undergo remarkably complex modes of modulation to fine tune membrane excitability and neuronal firing properties. In neurons, the isoform Nav1.6 is highly enriched at the axon initial segment and nodes, making it critical for the initiation and propagation of neuronal impulses. Thus, Nav1.6 modulation and dysfunction may profoundly impact the input-output properties of neurons in normal and pathological conditions. Phosphorylation is a powerful and reversible mechanism that exquisitely modulates ion channels. To this end, the multifunctional calcium/calmodulin-dependent protein kinase II (CaMKII) can transduce neuronal activity through phosphorylation of diverse substrates to serve as a master regulator of neuronal function. Because Nav1.6 and CaMKII are independently linked to excitability disorders, I sought to investigate modulation of Nav1.6 function by CaMKII signaling to reveal an important mechanism underlying neuronal excitability. Multiple biochemical approaches show Nav1.6 is a novel substrate for CaMKII and reveal multi-site phosphorylation within the L1 domain; a hotspot for post-translational regulation in other Nav isoforms. Consistent with these findings, pharmacological inhibition of CaMKII reduces transient and persistent sodium currents in Purkinje neurons. Because Nav1.6 is the predominant sodium current observed in Purkinje neurons, these data suggest that Nav1.6 may be modulated through CaMKII signaling. In support of this, my studies demonstrate that CaMKII inhibition significantly attenuates Nav1.6 transient and persistent sodium currents and shifts the voltage-dependence of activation to more depolarizing potentials in heterologous cells. Interestingly, I show that these functional effects are likely mediated by CaMKII phosphorylation of Nav1.6 at S561 and T642, and that each phosphorylation site regulates distinct biophysical characteristics of the channel. These findings are further extended to investigate CaMKII modulation of disease-linked mutant Nav1.6 channels. I show that different Nav1.6 mutants display distinct responses to CaMKII modulation and reveal that acute CaMKII inhibition attenuates gain-of-function effects produced by mutant channels. Importantly, computational simulations modeling the effects of CaMKII inhibition on WT and mutant Nav1.6 channels demonstrate dramatic reductions in neuronal excitability in Purkinje and cortical pyramidal cell models. Together, these findings suggest that CaMKII modulation of Nav1.6 may be a powerful mechanism to regulate physiological and pathological neuronal excitability.
CaMKII regulation of astrocytic glutamate uptake
(2016-05-19) Chawla, Aarti R.; Hudmon, Andy; Cummins, Theodore; Oxford, Gerry S.; Chen, Jinhui; Hoang, Quyen
Glutamate clearance by astrocytes is an essential part of physiological excitatory neurotransmission. Failure to adapt or maintain low levels of glutamate in the central nervous system is associated with multiple acute and chronic neurodegenerative diseases. The primary excitatory amino acid transporters (EAATs) in human astrocytes are EAAT1 and EAAT2 (GLAST and GLT-1 respectively in rodents). While the inhibition of a ubiquitously-expressed serine/threonine protein kinase, the calcium/calmodulindependent kinase (CaMKII) results in diminished glutamate uptake in cultured primary rodent astrocytes, the molecular mechanism underlying this regulation is unknown. In order to delineate this mechanism, we use a heterologous expression model to explore CaMKII regulation of EAAT1 and EAAT2. In transiently transfected HEK293T cells, pharmacological inhibition of CaMKII and overexpression of a dominant-negative version of CaMKII (Asp136Asn) reduces [3H]-glutamate uptake by EAAT1, without altering EAAT2 mediated glutamate uptake. Surprisingly, overexpression of a constitutively active autophosphorylation mutant (Thr287Asp) to increase autonomous CaMKII activity and a mutant incapable of autophosphorylation (Thr287Val) had no effect on either EAAT1 or EAAT2 mediated glutamate uptake. Pulldown of FLAGtagged glutamate transporters suggests CaMKII does not interact with EAAT1 or EAAT2. SPOTS peptide arrays and recombinant GST-fusion proteins of the intracellular N- and C-termini of EAAT1 identified two potential phosphorylation sites at residues Thr26 and Thr37 in the N-terminus. Introducing an Ala (a non-phospho mimetic) but not an Asp (phosphomimetic) at Thr37 diminished EAAT1-mediated glutamate uptake, suggesting that the phosphorylation state of this residue is important for constitutive EAAT1 function. In sum, this is the first report of a glutamate transporter being identified as a direct CaMKII substrate. These findings indicate that CaMKII signaling is a critical driver of homeostatic glutamate uptake by EAAT1. Aberrations in basal CaMKII activity disrupt glutamate uptake, which can perpetuate glutamate-mediated excitotoxicity and result in cellular death.
Cell surface proteoglycans control astrocyte migration and retinal angiogenesis by regulating basement membrane assembly
(2015-12-15) Tao, Chenqi; Zhang, Xin
Elaborate vascularization of the retina is crucial for the development and functioning of the eye. The proper patterning of astrocytes is a key event preceding retinal angiogenesis by providing guidance cues for endothelial cells, yet how this is regulated still remains obscure. The dual function of proteoglycans in both extracellular matrix (ECM) composition and cell signal transduction suggests their potential in the regulation of astrocyte migration. The current study demonstrated that non-cell-autonomous regulation by neuroretina cell surface proteoglycan is crucial for PDGF-A regulated astrocyte migration. Ablation of glycosaminoglycan side chains of proteoglycans in neuroretina led to impaired astrocyte migration, incomplete retinal angiogenesis, and hyaloid vessel persistence. This is followed by severe photoreceptor degeneration as a result of reactive gliosis, which cannot be rescued by constitutively activated Kras signaling. Notably, inner limiting membrane (ILM), the basement membrane of the retina, was breached in proteoglycan-deficient retinae prior to the formation of astrocytic network. Herein we propose that cell surface proteoglycans are essential for the initial assembly of ILM, and this cannot be compensated by secreted ECM proteoglycans. In support of this, after removal of ILM in retinal explant by Collagenase digestion, establishment of a new ILM can be achieved by incubation with exogenous laminin-supplemented Matrigel. This basement membrane reconstitution failed, however, in proteoglycan-deficient retinae or in wild type samples digested with a combination of Heparinase and ChABC in addition to Collagenase. Taken together, our study reveals a novel function of neuroretinal cell surface proteoglycans in the initial assembly of basement membrane which subsequently serves as a permissive substratum necessary for astrocyte migration.
Cell-Specific Spinophilin Function Underlying Striatal Motor Adaptations Associated with Amphetamine-Induced Behavioral Sensitization
(2022-07) Watkins, Darryl Shumon; Yamamoto, Bryan K.; Atwood, Brady K.; Baucum, Anthony J. II; Hudmon, Andy; Logrip, Marian L.
Striatal-mediated pathological disease-states such as Obsessive-Compulsive Disorder (OCD), Parkinson’s Disease (PD), and psychostimulant drug addiction/abuse are coupled with distinct motor movement abnormalities. In addition, these disorders are associated with perturbed synaptic transmission. Proper synaptic transmission is critical for maintaining neuronal communication. Furthermore, in many striatal-dependent disease-states, the principle striatal neurons, medium spiny neurons (MSNs), exhibit differential perturbations in downstream signaling. Signal transduction pathways that are localized to the glutamatergic post-synaptic density (PSD) of GABAergic MSNs regulate protein phosphorylation in a tightly controlled manner. Alterations in the control of this phosphorylation in striatal MSNs are observed in myriad striatal pathological diseasestates and can give rise to perturbations in synaptic transmission. While serine/threonine kinases obtain substrate specificity, in part, by phosphorylating specific consensus sites, serine/threonine phosphatases such as protein phosphatase 1 (PP1) are much more promiscuous. To obtain substrate selectivity, PP1 associates with targeting proteins. The major targeting protein for PP1 in the PSD of striatal dendritic spines is spinophilin. Spinophilin not only binds PP1, but also concurrently interacts with myriad synaptic proteins. Interestingly, dopamine depletion, an animal model of PD, modulates spinophilin protein-protein interactions in the striatum. However, spinophilin function on basal striatal-mediated motor behaviors such as the rotarod or under hyperdopaminergic states such as those observed following psychostimulant-induced behavioral sensitization are less well characterized. To elucidate spinophilin function more specifically, we have generated multiple transgenic animals that allow for cell type-specific loss of spinophilin as well as cell-specific interrogation of spinophilin protein interactions. Here, I report the functional role of spinophilin in regulating striatal mediated motor behaviors and functional changes associated with amphetamine-induced locomotor sensitization. In addition, we define changes in spinophilin protein-protein interactions that may mediate these behavioral changes. Furthermore, global loss of spinophilin abrogates amphetamine-induced sensitization and plays a critical role in striatal motor learning and performance. The data suggest that the striatal spinophilin protein interactome is upregulated in MSNs following psychostimulant administration. In addition, loss of spinophilin changes protein expression in myriad psychostimulant-mediated striatal adaptations. Taken together the data suggests that spinophilin’s protein-protein interactions in the striatum are obligate for appropriate striatal mediated motor function.

Browse

Browsing Medical Neuroscience Department Theses and Dissertations by Title

Results Per Page

Sort Options