Michigan State University

MSU Libraries

Digital Repository

Search results

(1 - 11 of 11)
Title
DEORPHANIZATION AND CHARACTERIZATION OF SEA LAMPREY OLFACTORY TRACE AMINE-ASSOCIATED RECEPTORS
Creator
JIA, LIANG
Date
2022
Collection
Electronic Theses & Dissertations
Description
The sense of smell plays an important role in mediating diverse behaviors in the animal kingdom. Odor detection in the sea lamprey is mediated by a limited number of odorant receptors (ORs) and trace amine-associated receptors (TAARs). Upon binding with odorants, the receptors are activated and subsequently activate the downstream neuronal signaling cascade that transforms the chemical information into electrophysiological signals. Odorous biogenic amines, when enriched in biological...
Show moreThe sense of smell plays an important role in mediating diverse behaviors in the animal kingdom. Odor detection in the sea lamprey is mediated by a limited number of odorant receptors (ORs) and trace amine-associated receptors (TAARs). Upon binding with odorants, the receptors are activated and subsequently activate the downstream neuronal signaling cascade that transforms the chemical information into electrophysiological signals. Odorous biogenic amines, when enriched in biological excretions, stimulate TAARs of the main olfactory epithelium and evoke innate behaviors in animals. I hypothesized that these biogenic amines are potent ligands for lamprey TAARs, and characterized the structural basis for amine recognition in these receptors. Chapter 1 describes discovery that spermine, an odorous polyamine in semen, serves as a sex pheromone in sea lamprey. Spermine potently stimulates the lamprey olfactory system, activates TAAR348 receptor, and attracts ovulated females. A novel antagonist to this receptor inhibits olfactory and female behavioral responses to spermine. This discovery elucidates a mechanism that male animals recruit mates through the release of chemical cues in ejaculates. In chapter 2, I demonstrated that two clades of independently evolved TAARs, represented by sea lamprey TAAR365 (sTAAR365) and mouse TAAR9 (mTAAR9), share a similar response profile. The results suggest a conserved mechanism whereby independently evolved TAAR receptors utilize convergent structural bases to detect various biogenic polyamines. In chapter 3, I found that a cadaverine-responsive sea lamprey TAAR receptor, TAAR346a, exhibits high basal activity when heterologously expressed in HEK293T cells. Triethylamine serves as an inverse agonist for TAAR346a that can specifically attenuate its high basal activity. These data support a model in which the inverse agonist recognizes only one of the two orthosteric sites used by the agonist as it elicits its inhibitory effect on the basal activity of the receptor. Further evidence was provided to highlight the importance of interhelical interactions in modulating ligand-independent activation of TAAR346a. Thus, this thesis contributes to a better understanding of sea lamprey olfaction and the structural basis of TAARs for amine recognition in vertebrate animals.
Show less
Title
Circuit-Specific Inhibition of Dopaminergic Signaling Associated with Phantom Gustatory Sensations in Disrupted-in-Schizophrenia-1 Mice
Creator
Fry, Benjamin R.
Date
2022
Collection
Electronic Theses & Dissertations
Description
Schizophrenia is a severe neuropsychiatric disorder characterized by a suite of symptoms occurring across cognitive (delayed processing, paraphasia, attentional deficits), negative (anhedonia, blunted affect, catatonia), and positive (hallucinations, delusions) domains. Antipsychotics are the most commonly prescribed medication to treat positive symptoms, however their use is complicated by substantial side-effects and inadequate efficacy. This reflects a lack of progress in understanding the...
Show moreSchizophrenia is a severe neuropsychiatric disorder characterized by a suite of symptoms occurring across cognitive (delayed processing, paraphasia, attentional deficits), negative (anhedonia, blunted affect, catatonia), and positive (hallucinations, delusions) domains. Antipsychotics are the most commonly prescribed medication to treat positive symptoms, however their use is complicated by substantial side-effects and inadequate efficacy. This reflects a lack of progress in understanding the precise neurobiological mechanisms underlying these symptoms, due in part to a lack of appropriate preclinical animal models. Here, I used an animal model of genetic vulnerability for neuropsychiatric illness known as Disrupted-in-schizophrenia-1 (DISC-1) to examine impaired reality testing, which reflects an aberrant internal representation of an absent event. In mice, this can be observed by an associatively evoked perception of an absent sweet taste. This effect is dopaminergically-dependent and associated with elevated activity in the insular cortex (IC). By combining sophisticated Pavlovian behavioral procedures with chemogenetic inhibition of dopamine neurons projecting from the ventral tegmental area (VTA) to the IC, I show that inactivation of the VTA --> IC dopaminergic circuitry leads to impaired reality testing in wild-type mice, and that DISC-1 mice have significantly less dopamine neurons which send projections to the IC, specifically. These data yield new insights with regard to the neurobiology underlying reality testing and the functional anatomical outcomes following perturbations of the DISC-1 genetic locus. My studies also suggest potential targets for the development of novel pharmacological treatments in humans with schizophrenia.
Show less
Title
An investigation of cerebellar morphology in childhood stuttering
Creator
Johnson, Chelsea Anna
Date
2022
Collection
Electronic Theses & Dissertations
Description
While many studies have connected structural and functional cerebellar differences to developmental stuttering, there are limited studies of cerebellar gray matter morphology in young children who stutter. These examinations include small sample sizes of children and use morphometry methods that might not be best suited for examining the cerebellum (e.g., Chang et al., 2008). This dissertation examines how the structure of specific cerebellar lobules differs in a larger cohort of children who...
Show moreWhile many studies have connected structural and functional cerebellar differences to developmental stuttering, there are limited studies of cerebellar gray matter morphology in young children who stutter. These examinations include small sample sizes of children and use morphometry methods that might not be best suited for examining the cerebellum (e.g., Chang et al., 2008). This dissertation examines how the structure of specific cerebellar lobules differs in a larger cohort of children who stutter and children who do not stutter as well as in persistent and recovered children who stutter. These data will provide evidence to better inform predictions of how the morphology of cerebellar areas are likely involved in aspects of speech motor control in developmental stuttering. In this study, gray matter morphology of the cerebellum was examined in children who do and do not stutter using voxel-based morphometry and a specialized toolbox and atlas for the cerebellum (Diedrichsen, 2006). Here we examined cerebellar gray matter volume (GMV) based on structural MRI data from children who stutter and children who do not stutter, 116 preschool-age children (stuttering N= 57) between the ages of 3-5 years, and a school-age cohort of 72 children (stuttering N=37) six years of age and up. This dissertation is the first study to examine cerebellar GMV in a large group of children who stutter using a specialized toolbox and atlas for the cerebellum. Results from this study showed that there were no overall significant group differences of lobular GMV between the stuttering and non-stuttering groups in any of the groups of children. There were significant age-related associations, however, that differentiated children who do and do not stutter in specific age ranges. In particular, the following cerebellar lobules differed significantly in GMV between children who do and do not stutter with age: 1) cerebellar lobule VII, which may correspond with cerebellar functions that support speech planning, 2) lobule VIII, which has been linked to various functions including corrections during perturbation studies, and 3) lobule IV which has been reported to be involved in feedforward control speech motor control processes. Notably, GMV of cerebellar lobule VI was associated negatively with Stuttering Severity Instrument (SSI) score in preschool-age persistent children who stutter. Associations between SSI score and GMV in cerebellar lobule VI may mean that feedforward control mechanisms are associated with the frequency of stuttering in children who stutter. In summary, significant findings of this investigation indicate that 1) children who do and do not stutter do not show an overall difference in cerebellar GMV, 2) GMV of the cerebellum is associated with SSI score, 3) age-related differences in GMV in the cerebellum differentiate children who do and do not stutter. The results from this study indicate that feedforward control is associated with disfluencies while age-related variations of cerebellar areas that may support both the feedback and feedforward control pathways are connected to aspects of stuttering, such as age.
Show less
Title
Structural connectivity of an interoception network in schizophrenia
Creator
Yao, Beier
Date
2022
Collection
Electronic Theses & Dissertations
Description
Interoception refers to the processing, integration, and interpretation of bodily signals by the brain. Interoception is key to not only basic survival, but also many cognitive processes, especially motivational and affective functioning. There is emerging evidence suggesting altered interoception in schizophrenia, but its neural underpinning has not been examined. The current study aims to investigate the structural connectivity of a putative interoception network in schizophrenia, and its...
Show moreInteroception refers to the processing, integration, and interpretation of bodily signals by the brain. Interoception is key to not only basic survival, but also many cognitive processes, especially motivational and affective functioning. There is emerging evidence suggesting altered interoception in schizophrenia, but its neural underpinning has not been examined. The current study aims to investigate the structural connectivity of a putative interoception network in schizophrenia, and its relationship with affective functioning and clinical symptoms. Thirty-five participants with schizophrenia (SZ) and 36 healthy control participants (HC) underwent diffusion tensor imaging (DTI) and performed tasks measuring emotional functioning. Probabilistic tractography was used to identify white matter tracts connecting the key hubs forming the interoception network (i.e., rostral and caudal anterior cingulate cortex, ventral anterior insula, dorsal mid and posterior insula, and amygdala). Microstructural integrity of these tracts was compared across groups and correlated with measures of emotional functioning and symptom severity. I found that SZ exhibited altered structural connectivity in the putative interoception network, compared to HC. The structural connectivity of the network was correlated with emotion recognition in HC, supporting a link between the interoception network and emotional functioning. However, this correlation was much weaker in SZ, suggesting less reliance on this network. I did not find a correlation between the structural connectivity and clinical symptoms in SZ. These findings suggest that altered interoception may play a role in illness mechanisms of schizophrenia, especially in relation to emotional deficits.
Show less
Title
NEUROTECHNOLOGY DESIGN FEATURES’ IMPACT ON THE FUNCTION AND IDENTITY OF REACTIVE ASTROCYTES
Creator
Riggins, Ti'Air
Date
2021
Collection
Electronic Theses & Dissertations
Description
Implantable neurotechnology offers substantial promise to improve the condition of many neurodegenerative diseases. Microelectrode arrays implanted in the brain have the capability to stimulate or record electrical activity from neighboring cells. However, shortly after implantation, a foreign body response occurs, which is what researchers believe decreases the electrical recording stability and longevity of signal detection of these devices. Established biomarkers such as astrogliosis, and...
Show moreImplantable neurotechnology offers substantial promise to improve the condition of many neurodegenerative diseases. Microelectrode arrays implanted in the brain have the capability to stimulate or record electrical activity from neighboring cells. However, shortly after implantation, a foreign body response occurs, which is what researchers believe decreases the electrical recording stability and longevity of signal detection of these devices. Established biomarkers such as astrogliosis, and stimuli such as the mechanical mismatch at the device-tissue interface, have been studied to understand the tissue response to the devices. However, the relationship of these factors with device performance is not well understood. Astrocytes play an important role in the brain’s immune system and recently, RNA analysis has confirmed transcriptional profiles of reactive astrocytes which are associated with specific injury states and neurodegenerative diseases. In this dissertation, I have investigated new biomarkers of astroglial reactivity at the electrode interface and characterized the surface topography and bending stiffness of devices. I induced two types of inflammatory astrocytic cell culture models, and I characterize each model’s reactivity in comparison to gene expression surrounding electrodes implanted in rat tissue. Atomic microscope microscopy (AFM) techniques were also used to measure surface roughness and bending stiffness as it may predict cellular adhesion and device performance. I aim to elucidate pathways in the neurological foreign body response which will give researchers new potential biomarkers to target to improve recording performance, motivating improved designs for implantable neurotechnology. The research presented in this dissertation investigates how design features influence the tissue interface and asks questions about possible ways to mitigate tissue response: (1) by exploring and summarizing the design space as a whole, suggesting ways to characterize designs and evaluating each designs’ successes and limitations (2) using a cutting edge imaging technique to image and measure material properties of three commonly used materials, (3) and creating a reactive tissue culture model, comparing its proteomic and genetic expression to the established rat model. Chapter 2 describes surface characterization techniques that could be used to better classify device features to predict performance and explores next generation probes from a design and performance standpoint. Chapter 3 uses atomic force microscopy to image and measure surface roughness on device surfaces while also measuring the bending stiffness to help determine possible micromotion in the brain. Here, we speculate what these findings mean for the performance and longevity of current probe design. Chapter 4 develops an astroglial culture model to mimic foreign body response in the brain and compare the genomic results to tissue culture near and far from the implanted device. Here, we report the transcriptomic results of the model in comparison to brain transcriptomic results, and what these biomarkers may implicate regarding tissue response and neurodegenerative signaling. This body of work uncovers knowledge recapitulating important factors of device features that affects tissue signaling at the tissue device interface, and biomarkers that play a role and cell signaling. Future directions aim at developing a more physiologically relevant tissue culture model that can predict clinical outcomes, and use high throughput screening techniques to help researchers address the challenge of long term suboptimal device performance.
Show less
Title
TOWARD PRECISION MEDICINE : EFFECTS OF THE COMMON VAL66MET BDNF VARIANT IN THE AGING BRAIN AND IMPLICATIONS FOR THE FUTURE OF PARKINSON’S DISEASE THERAPEUTICS
Creator
Mercado-Idziak, Natosha Marie
Date
2021
Collection
Electronic Theses & Dissertations
Description
The rs6265 (Val66Met) single nucleotide polymorphism in the gene for brain-derived neurotrophic factor (BDNF) is a common genetic variant that has been shown to alter therapeutic responses in patients with Parkinson’s disease (PD). Possession of the variant Met allele results in decreased activity-dependent release of BDNF by disrupting BDNF transport and sorting into synaptic vesicles. In the experiments detailed in this thesis, I examine the effects of the Val66Met SNP, and its interaction...
Show moreThe rs6265 (Val66Met) single nucleotide polymorphism in the gene for brain-derived neurotrophic factor (BDNF) is a common genetic variant that has been shown to alter therapeutic responses in patients with Parkinson’s disease (PD). Possession of the variant Met allele results in decreased activity-dependent release of BDNF by disrupting BDNF transport and sorting into synaptic vesicles. In the experiments detailed in this thesis, I examine the effects of the Val66Met SNP, and its interaction with aging, on therapeutic efficacy and the development of aberrant side-effects following primary dopamine (DA) neuron transplantation, a restorative experimental therapeutic approach for PD that is currently experiencing a robust revitalization following a decade-long worldwide moratorium. In particular, I hypothesized that rs6265-mediated dysfunctional BDNF signaling is an unrecognized contributor to the limited clinical benefit observed in a subpopulation of individuals with PD despite robust survival of grafted DA neurons and extensive integration into the host brain. I also hypothesized that this genetic variant contributes to the development of graft-induced dyskinesias (GID). To test these hypotheses, we generated a novel CRISPR knock-in rat model of the rs6265 BDNF SNP to investigate for the first time the influence of a common genetic polymorphism on graft survival, functional efficacy, and side-effect burden in subjects grafted with embryonic ventral mesencephalic DA neurons. In two sister studies, I compared these primary endpoints between wild-type (Val/Val) rats and those homozygous for the variant Met allele (Met/Met), in both young adult (8 m.o. at grafting) and middle-aged (15 m.o. at grafting) cohorts. In each study, rats were rendered unilaterally parkinsonian with intranigral 6-hydroxydopamine and primed with levodopa (12 mg/kg M-Fr) to induce stable expression of levodopa-induced dyskinesias (LID), the primary behavioral endpoint for assessing graft function. After levodopa priming, rats received an intrastriatal graft of embryonic ventral mesencephalic neurons (200,000 cells in young adult rats, 400,000 cells in middle-aged rats; E14 wild-type donors) or a sham graft. LID were evaluated for 9-10 weeks post-engraftment, and GID were assessed 24-48 hr prior to sacrifice. In young adult graft recipients, this research demonstrates that: 1) Met/Met rats display enhanced graft efficacy and paradoxically enriched graft-derived neurite outgrowth compared to Val/Val rats, and 2) the Met allele is strongly linked to GID development and this behavioral phenotype is correlated with neurochemical signatures of glutamatergic neurotransmission by grafted DA neurons. In middle-aged graft recipients, this research indicates that: 1) behavioral enhancement associated with the Met allele is maintained with advancing age, and 2) advanced age is associated with the induction of GID in rats of both genotypes despite the presence of widespread intrastriatal grafts. In this rapidly evolving era of precision medicine, understanding mechanisms underlying the beneficial versus detrimental impact of the Val66Met polymorphism, and/or its interaction with aging, will aid in the development of safe and optimized therapeutic approaches for remodeling the parkinsonian striatum.
Show less
Title
PERTURBATION OF ASTROCYTIC KEAP1-NRF2-ARE PATHWAY AND GLUTAMATE TRANSPORTER EXPRESSION IN SPINAL ASTROCYTIC DEGENERATION
Creator
Wiwatratana, Duanghathai
Date
2021
Collection
Electronic Theses & Dissertations
Description
Several cellular mechanisms are known to be involved in methylmercury (MeHg) induced central nervous system (CNS) toxicity, including the dysregulation of intracellular Ca2+, redox, and glutamate homeostasis. However, the factors that make particular neurons susceptible to MeHg toxicity, and the latency period of neurological signs and symptoms, have not yet been clearly delineated. For example, the spinal dorsal root ganglia (DRG) is the primary target of MeHg. Mercury (Hg) granules are...
Show moreSeveral cellular mechanisms are known to be involved in methylmercury (MeHg) induced central nervous system (CNS) toxicity, including the dysregulation of intracellular Ca2+, redox, and glutamate homeostasis. However, the factors that make particular neurons susceptible to MeHg toxicity, and the latency period of neurological signs and symptoms, have not yet been clearly delineated. For example, the spinal dorsal root ganglia (DRG) is the primary target of MeHg. Mercury (Hg) granules are first detected in spinal cord motor neurons (SMNs) in the non-symptomatic phase, whereas Hg granules are detected in glia later, following with neurological symptoms (Møller-Madsen, 1991). This finding suggested that the latent period (non-symptomatic phase) is associated with Hg accumulation in neurons, while the symptomatic phase occurs following Hg accumulation in glia, and the susceptibility is not associated with Hg granule accumulation in cells (Møller-Madsen, 1991). Astrocytes generally provide glutathione (GSH) for neurons to detoxify toxic insult. In the spinal cord, MeHg might perturb the antioxidant pathway, Keap1-Nrf2-ARE pathway in the spinal cord astrocytes (SCAs) consequently contribute to DRG or SMN susceptibility to MeHg toxicity. In this study, the comparative responses of different SCAs maturity to a non-toxic MeHg concentration (0.1 μM) suggested that the fully mature SCAs (Day in vitro 30; DIV30), were more susceptible to MeHg than SCAs on DIV14. The perturbation of the Keap1-Nrf2-ARE pathway in SCAs (DIV 30) during exposure to sub-toxic MeHg concentration (0.50 μM) caused a biphasic increase in antioxidant genes such as Keap1, Nrf2, Gclc, Abcc1 mRNAs expression. The concomitant increase of glutamate transporter Slc7a11 encoded for the system Xc-, and Slc1a3 encoded for EAAT1, and Slc1a2 encoded for EAAT2 expression during MeHg exposure might suggest the cooperative expression or function of these glutamate transporters. This concomitant expression was further demonstrated in studies using Nrf2-knockout (Nrf2-KO) derived SCAs. The increase of basal Slc7a11 mRNA, was concurrent to the increase of basal Slc1a3 and Slc1a2 mRNA expressions in Nrf2-KO derived SCA. The function of time of MeHg exposure indicated that Nrf2-KO derived SCAs were more susceptible to MeHg than the wild-type (WT)-derived SCAs. The pronounced susceptibility of Nrf2-KO derived SCAs was mainly due to the loss of GSH) metabolism and transport genes Gclc, GPx1, GPx4, and Abcc1 mRNAs in this genotype. MeHg significantly reduced these mRNA expressions in both genotypes. However, not all Nrf2-ARE regulated genes were affected by MeHg in similar ways in these genotypes. For example, MeHg induced the increase of Slc7a11 mRNA expression in WT-derived SCAs, but it appears to cause the reduction of this mRNA expression in Nrf2 KO-derived SCAs. Administration of antioxidant N-acetyl-L-cystine (NAC) in pretreatment (NP), co-treatment (CO), and post-treatment of MeHg (MP) prevented the reduction of SCAs metabolic functions for over 160h. The mechanism of NAC action in preventing MeHg induced SCAs degeneration is primarily due to its thiol antioxidant property.In conclusion, this study suggests that age and genetic predisposition contribute to SCAs susceptibility to MeHg toxicity. The dysregulation of the antioxidant pathways and glutamate homeostasis in SCAs potentially contributes to SMNs or DRG susceptible to MeHg.
Show less
Title
Heterogeneous Thalamic Reticular Nucleus Neurons and Their Functional Role in Thalamocortical Processing
Creator
Harding-Jackson, Laura
Date
2021
Collection
Electronic Theses & Dissertations
Description
The thalamic reticular nucleus (TRN) is an integral regulator of information flow between the thalamus and cortex. The TRN receives synaptic inputs from both cortical and thalamic regions and based upon this information it selectively inhibits thalamic activity. TRN neurons produce action potentials in two distinct modes: a fast, transient burst discharge from a hyperpolarized state, and a prolonged, tonic discharge from a relatively depolarized state. While previous studies have...
Show moreThe thalamic reticular nucleus (TRN) is an integral regulator of information flow between the thalamus and cortex. The TRN receives synaptic inputs from both cortical and thalamic regions and based upon this information it selectively inhibits thalamic activity. TRN neurons produce action potentials in two distinct modes: a fast, transient burst discharge from a hyperpolarized state, and a prolonged, tonic discharge from a relatively depolarized state. While previous studies have characterized burst discharge as a transient high frequency discharge (> 250 Hz), these electrophysiological studies reveal a highly variable range of burst frequencies (4- 342 Hz). In these studies, I aim to discover the mechanisms underlying these highly variable burst frequencies, as well as their functional role in thalamocortical processing.In chapter two, I found that bursts from TRN neurons with relatively higher frequency discharge (>100 Hz) contain more action potentials per burst. These neurons also have higher input resistances, broader action potentials, higher action potential thresholds, and larger somas. The amplitude of the T-type calcium channel-mediated low-threshold spike, which underlies the burst discharge, is positively correlated with both the burst discharge frequency and the number of action potentials per burst. I next investigated whether small conductance calcium-activated potassium channels (SK channels) could mediate the differences in burst firing rate and action potential number. Blocking SK channels increased the frequency and duration of the burst but did not increase the amplitude of the underlying T-type calcium current. Prior studies suggest that T-type calcium channels are distributed along the dendrites in TRN neurons with high frequency burst discharge. In chapter three, I examine the distribution of dendritic calcium activity within the lower frequency bursting neurons. While the calcium signal was lower in these neurons all along the dendrites, the calcium signal was evenly distributed across proximal, intermediate, and distal dendritic regions. Investigation of SK channel activity revealed significant location-specific effects. In lower frequency bursting neurons, SK channels had the greatest influence at proximal and distal locations. In higher frequency bursting neurons, SK channels had the greatest influence at proximal and intermediate dendritic locations. Heterogeneous TRN burst discharge frequencies may represent a diverse cell population with unique dendritic ion channel composition and distribution. These results may improve our understanding of the mechanisms of TRN neuron afferent synaptic integration as well as modulation of thalamocortical inhibition. In chapter four I investigate whether intrinsic properties of TRN neurons are altered in the Fmr1-KO mouse model of Fragile X Syndrome (FXS). Individuals with FXS experience a variety of comorbidities that could involve TRN function, such as altered sensory perceptions, sleep disorders, and epilepsy. Analysis of intrinsic cellular properties revealed no differences in TRN neuron properties. Further investigation of synaptic plasticity, which is an abnormal finding in several other brain regions in FXS, also revealed no pathology. These findings suggest that TRN dysfunction does not contribute to FXS pathology.
Show less
Title
NOVEL IMPACTS OF HOST-ENVIRONMENT INTERACTIONS IN ENTERIC GLIA THROUGH SEQUENCING AND IN-SITU EXPRESSION
Creator
Ponnampalam, Christine Dharshika
Date
2021
Collection
Electronic Theses & Dissertations
Description
The enteric nervous system (ENS) is comprised of enteric neurons and glia that facilitate essential gastrointestinal (GI) function including motility, visceral sensation, absorption, and gut permeability. Enteric neurons and glia are responsive to environmental cues and stressors ranging from the local gut microenvironment to the host’s psychosocial state and understanding how the ENS integrates these cues to modulate local and systemic function is critical. Novel roles for enteric neurons in...
Show moreThe enteric nervous system (ENS) is comprised of enteric neurons and glia that facilitate essential gastrointestinal (GI) function including motility, visceral sensation, absorption, and gut permeability. Enteric neurons and glia are responsive to environmental cues and stressors ranging from the local gut microenvironment to the host’s psychosocial state and understanding how the ENS integrates these cues to modulate local and systemic function is critical. Novel roles for enteric neurons in host-environmental interactions have been discovered using specialized sequencing technologies but these tools have not yet readily investigated enteric glia. The goal of this dissertation was to develop and utilize genetic technologies to characterize enteric glial responses to environmental mediators. First we adapted existing genetic tools to study molecular changes in the ENS and specifically enteric glia. We developed effective means of characterizing enteric glial expression within complex in vivo models using the RiboTag model with RNA-sequencing and subsequently visualized changes in gene expression within enteric ganglia in situ. We then utilized these techniques to investigate sex-specific responses to early life stress in enteric glia. Enteric glia from male and female mice have contrasting expression profiles including differences in GPCR signaling that could contribute to sex-specific ENS signaling mechanisms and ultimately GI disease outcomes. This supports recent findings of sexual dimorphism in glial functional connectivity and may highlight a critical difference in the way enteric glia communicate with other cell types between males and females. Additionally enteric glia from male mice ‘feminize’ following iiiearly life stress through altered expression of GI and neurological disease genes including mechanisms of glial-immune communication like type I interferon signaling. Together these data highlight striking differences in the physiologic molecular patterns and nature of stress response in enteric glia between males and females that likely contribute to sexually dimorphic GI disease patterns and symptom presentation. Next we investigated ENS type I interferon responses through the stimulator of interferon genes (STING) pathway. STING responds to both microbial and host mediators to contribute to GI inflammation. However the role of STING signaling in the gut is complex and can either exacerbate or ameliorate inflammation likely dependent on complex microenvironmental factors. We provide the first known investigation of STING expression and signaling within the ENS. STING is expressed in both enteric neurons and glia but IFNB is only expressed in enteric neurons. ENS STING is activated by its canonical ligands toproduce type I interferons. However this is likely primarily mediated through canonical activation of enteric neuronal STING and the contribution of enteric glial STING to type I IFN response is minor. Additionally enteric glial STING does not alter gastrointestinal outcomes during acute colitis within the DSS colitis model. Taken together these findings suggest enteric glia do not utilize STING for canonical type I IFN signaling or contribute to disease pathology in acute DSS colitis. Enteric glial STING may instead utilize primordial and specialized signaling pathways that more selectively alter local function. Together our data provide novel genetic tools and data to further uncover molecular functions in enteric glia and their role in GI and systemic health. Using these we discovered entirely novel molecular interaction effects between sex and early life stress that shift the framework of these risk factors in GI disease. Furthermore we highlight a novel potential mediator of ENS-microbe communication with STING. Our findings further characterize the molecular patterns used by glia in response to complex environmental factors and highlight unique heterogeneity in glial intercellular communication.
Show less
Title
Biomaterial and genetic tools to influence neuronal network formation, excitability, and maturity at the electrode interface
Creator
Setien-Grafals, Monica B.
Date
2020
Collection
Electronic Theses & Dissertations
Description
Understanding brain function remains a grand challenge of our time. Likewise, when neurodegeneration occurs, repair efforts are limited due to the highly heterogeneous and interconnected nature of the cerebral cortex. The drive to better understand normal brain function and pathological states has intensified demand for new technologies which can interrogate the nervous system with enhanced spatiotemporal resolution. Implanted brain electrodes are being used and developed to provide a deeper...
Show moreUnderstanding brain function remains a grand challenge of our time. Likewise, when neurodegeneration occurs, repair efforts are limited due to the highly heterogeneous and interconnected nature of the cerebral cortex. The drive to better understand normal brain function and pathological states has intensified demand for new technologies which can interrogate the nervous system with enhanced spatiotemporal resolution. Implanted brain electrodes are being used and developed to provide a deeper understanding for neurological injury and neurodegeneration. However, issues with biological integration come into play and potentially interfere with signal stability over time. Here, this work provides innovative tools that can be used to interface and control the tissue-electrode interface. In particular, we are interested in exploring surface chemistries, genetic tools, and electrode materials which favor neural regeneration around implanted electrodes. The research presented in this dissertation describes the exploration of biomaterial and genetic tools for interfacing the tissue-electrode interface: (1) characterization of surface chemistries presented to differentiating neural progenitors, and an understanding of the conditions which promote neurite outgrowth and electrophysiological maturation, (2) a blue-light inducible gene expression system, which could potentially be used to control gene expression at the implanted electrode interface, and (3) testing the impacts of "next-generation" electrode materials, such as diamond, as candidates for neural interfacing. Chapter 2 uncovers the study of various common substrates and their effects on rat neural progenitor cells, which can be used to create unique morphologies. Chapter 3 explores the use of an optogenetic system from a bacterial transcription factor (EL222) that allows for blue light-dependent transcriptional activation. Here, we validated the use of EL222 for spatial patterning of fluorescent reporter genes and developed stable expression in HEK293 cells, which can be used long-term for developing approaches for light-driven regeneration of neural circuitry. Chapter 4 reveals material and genetic factors that can affect cell structure and function. Here, we report the results of an initial characterization of the biocompatibility of the novel diamond-based materials, including conductive boron-doped polycrystalline diamond (BDD) and insulating polycrystalline diamond (PCD). The results presented will inform the transfer of the novel diamond substrate materials to sensing applications in the in vivo environment, where we expect to leverage the positive performance characteristics of the diamond materials displayed in vitro. Taken together, these chapters offer significant development of material and biological tools and that will help manage and mitigate challenges presented at the tissue-electrode interface. Future directions aim at exploring synergistic effects of electrode material and optogenetic control for controlling excitability and identity of cells at the interface, effectively bridging the divide between electronics and tissue.
Show less
Title
Enteric glial modulation of immune activation during inflammatory stress
Creator
Chow, Aaron Kin Yeung
Date
2020
Collection
Electronic Theses & Dissertations
Description
Gastrointestinal (GI) disorders such as inflammatory bowel disease (IBD), irritable bowel syndrome, and other functional GI disorders are major health concerns and account for about $26.4 billion in yearly costs in the United States alone. Many of these GI disorders manifest symptoms such as GI dysmotility, intestinal secretion and absorption dysfunction, and abdominal pain. Inflammation plays a major role in the pathogenesis of these diseases, and current therapies for many GI disorders aim...
Show moreGastrointestinal (GI) disorders such as inflammatory bowel disease (IBD), irritable bowel syndrome, and other functional GI disorders are major health concerns and account for about $26.4 billion in yearly costs in the United States alone. Many of these GI disorders manifest symptoms such as GI dysmotility, intestinal secretion and absorption dysfunction, and abdominal pain. Inflammation plays a major role in the pathogenesis of these diseases, and current therapies for many GI disorders aim to regulate the activation and progression of the inflammatory cascade. However, due to a gap in knowledge in how the immune system is regulated within the GI tract, there remains a lack of effective treatments for these common GI disorders. Interactions between the nervous system and immune system point to neurons having important roles in immune modulation, but the mechanisms of neuro-immune regulation in the gut is not completely understood.The enteric nervous system (ENS) consists of enteric neurons and enteric glia arranged in plexuses embedded in the gut wall. This neural network is responsible for the normal secretomotor functioning of the GI tract, and the disruption of the ENS network alters GI functioning and underlies pathological GI symptoms. As part of the ENS, enteric glia work in tandem with enteric neurons to coordinate GI functions. In addition to their contributions to maintain normal secretomotor functioning of the GI tract, enteric glia are activated by immunomodulatory signals, they can secrete and respond to cytokines, can exert immunosuppressive effects, and share characteristics with antigen presenting cells. Therefore, we hypothesize that enteric glia play an active role in immune regulation in the ENS.In this dissertation, we specifically examine the role enteric glial cells play as an antigen presenting cell to regulate immune activation. Our results show that enteric glia have the machinery necessary to act as an antigen presenting cell and can express major histocompatibility complex (MHC) type II molecules during inflammatory stress to interact with T-lymphocytes. Enteric glial MHC II expression has functional relevance, as it modulates the activation in Th17 and Treg subtypes, but not Th1 or Th2 T-lymphocyte subtypes. Although MHC II molecules are typically associated with the expression of phagocytosed extracellular antigens, our results show that enteric glia do not readily phagocytose extracellular antigens. Instead, MHC II expression in enteric glia is mediated by autophagy. The activation of autophagic pathways is necessary, but not sufficient in eliciting enteric glial MHC II expression. Finally, although enteric glia regulate T-lymphocyte activation, cytokine levels at the whole organism or regional tissue levels remain unchanged, suggesting that enteric glial cytokine effects primarily operate at the local microenvironment level.Our findings provide support for enteric glial cells having an active role as an immunomodulator. Specifically, we show that enteric glia modulate T-lymphocyte activation via autophagy-mediated MHC II expression and propose a novel mechanism of neuro-immune modulation in the gut.
Show less