Browsing by Author "Williams, Christina L"

The cholinergic system is intricately linked with hippocampal memory. As well, choline is anti-inflammatory in the brain and periphery (Terrando et al., 2011; Rivera et al., 1998). However, few have analyzed the anti-inflammatory properties of choline as an alternate means by which cholinergic manipulations affect hippocampal memory throughout the lifespan. The first aim of this dissertation work sought to determine if dietary choline supplementation protects against the deleterious effects of air pollution in the developing brain. Pregnant mice were given a high-choline diet (approximately 4.5x the choline chloride in the control diet) or a synthetic control diet. As well, dams were exposed to a series of diesel particulate (DEP) or saline vehicle sessions throughout pregnancy. Mice were sacrificed and tissues were collected on embryonic day 18. The activation state of microglia, identified by quantifying morphology using Iba1+ immunohistochemical staining, was examined in the dentate gyrus of the hippocampus (DG), the paraventricular nucleus (PVN) of the hypothalamus, the basolateral amygdala (AMY), and the parietal cortex (PCX). As expected, we found that DEP led to increased microglial activation in the fetal DG in males. Choline supplementation partially prevented this increase in activation. Interestingly, these effects were region-specific: the opposite pattern is seen in the PVN, and no significant diesel effect was seen in the AMY and PCX. These findings suggest that prenatal choline supplementation throughout pregnancy may protect the fetal hippocampus against the neuroinflammation associated with air pollution. To analyze whether the acute effects of dietary choline seen prenatally also occur in adulthood, adult dietary choline supplementation alongside the tibial fracture model of post-operative cognitive dysfunction (POCD) was used. POCD occurs when increased neuroinflammation due to peripheral surgery leads to impairments in cognition. Differences were found in almost hippocampal-dependent behavior, astrocytic activation, and cell proliferation. Differences were time point-specific. In the hippocampus, astrocytic activation, cell proliferation, and hilar granule cells all increased 1 day after surgery, and these increases were blunted by dietary choline. An increase in hippocampal young neurons was found 2 weeks after surgery. However, both were blunted by choline supplementation. At both time points assessed, tibial fracture impaired novel object recognition performance, and dietary choline rescued these impairments. As well, dietary choline supplementation did not mitigate the increase in anxiety-related behavior – specifically implicating hippocampal actions of the nutrient. Because the hippocampal-dependent memory impairment and rescue is not time point-specific, but the neural effects of tibial fracture are each specific to a certain timepoint, the mechanisms of behavior are likely different at each time point. Building upon aim 2, aim 3 explores if perinatal choline supplementation can act via “programming” of the neuroimmune system in development to prevent POCD in adulthood. Perinatal choline supplementation prevented POCD and neuroinflammation due to peripheral surgery, but did not protect against increases in young neurons or hilar neurons. Perinatal choline nutrition, in addition to its already-known neuroprotection, is additionally protective against POCD and its associated neuroinflammation in adulthood. Taken together, this body of work concludes that dietary choline supplementation at various administration dates is protective in neuroinflammatory models in behavior and brain.

Altered dietary availability of the vital nutrient choline during early development leads to persistent changes in brain and behavior throughout adulthood. Prenatal choline supplementation during embryonic days (ED) 12-17 of the rodent gestation period enhances memory capacity and precision and hippocampal plasticity in adulthood, and protects against spatial learning and memory deficits shortly after excitotoxic seizures, whereas prenatal choline deficiency can compromise hippocampal memory and plasticity in adulthood. Recent evidence from our laboratory has determined that lifelong proliferation of newborn neurons in the adult hippocampus, a feature of adult hippocampal plasticity that has been implicated in some aspects of learning and memory, is modulated by early choline availability. Prenatal choline's effects on adult neurogenesis may be one mechanism for diet-induced cognitive changes throughout life and in response to injury, although little is known about the mechanisms underlying how prenatal choline alters adult neurogenesis or the neural mechanisms underlying prenatal choline supplementation's protection against cognitive deficits after seizures. To address these issues, the present set of experiments investigated how prenatal choline availability modulates specific properties of neurogenesis in the adult brain (in the intact brain and in response to injury), as well as hippocampal markers known to change in response to excitotoxin-induced seizures, and sought to relate changes in neurogenesis and in neuropathological markers following injury to changes in performance on spatial learning and memory tasks. Subjects in each experiment were adult offspring from rat dams that received either a control diet or diet supplemented with choline chloride or deficient of choline on ED 12-17. To measure neurogenesis, rats were given injections of the mitotic marker bromodeoxyurdine to label dividing cells in the hippocampus. Prenatal choline supplementation enhanced several properties of basal adult hippocampal neurogenesis (cell division and survival, neural stem/progenitor cell phenotype and proliferative capacity, trophic support), and this increase was associated with improvements in spatial working memory retention in a delayed-matching-to-place water maze task. In contrast, prenatal choline deficiency had little effect on basal adult hippocampal neurogenesis, and no effect on spatial memory performance. Prenatal choline supplementation also enhanced olfactory bulb neurogenesis without altering cell proliferation in the subventricular zone, while prenatal choline deficiency had no effect on either measure, showing for the first time that prenatal choline's effects on adult neurogenesis is similarly expressed in another distinct neurogenic region of the adult brain. Altered prenatal choline availability also modulated the hippocampal response to kainic acid-induced seizures where supplementation attenuated while deficiency had no effect on the injury-induced proliferative response of the dentate gyrus shortly after injury. Prenatal choline supplementation also attenuated other markers of hippocampal neuropathology shortly after seizures and promoted the long-term hippocampal recovery from seizures months after injury, including rescuing declines in adult hippocampal neurogenesis and in spatial memory performance in a standard water maze task. Taken together, these findings demonstrate a robust neuroprotective effect of prenatal choline supplementation that may be driven by enhanced adult hippocampal plasticity and trophic support prior to injury, and shed light on the mechanisms underlying how prenatal choline availability alters adult hippocampal neurogenesis, which may contribute to changes in memory capacity and precision both throughout life and following neural assault.

Opioid drug abuse represents a serious public health concern with few effective therapeutic strategies. A primary goal for researchers modeling substance abuse disorders has been the delineation of the biological and environmental factors that shape an individual’s susceptibility or resistance to the reinforcing properties of abused substances. Early-life environmental conditions are frequently implicated as critical mediators for later-life health outcomes, although the cellular and molecular mechanisms that underlie these effects have historically been challenging to identify. Previous work has shown that a neonatal handling procedure in rats (which promotes enriched maternal care) attenuates morphine conditioning, reduces morphine-induced glial activation in the nucleus accumbens (NAc), and increases microglial expression of the anti-inflammatory cytokine interleukin-10 (IL-10). The experiments described in this dissertation were thus designed to address if inflammatory signaling in the NAc may underlie the effects of early-life experience on later-life opioid drug-taking. The results demonstrate that neonatal handling attenuates intravenous self-administration of the opioid remifentanil in a drug concentration-dependent manner. Transcriptional profiling of the NAc reveals a suppression of pro-inflammatory cytokine and chemokine signaling molecules and an increase in anti-inflammatory IL-10 in handled rats following repeated exposure to remifentanil. To directly test the hypothesis that anti-inflammatory signaling can alter drug-taking behavior, bilateral intracranial injections of plasmid DNA encoding IL-10 (pDNA-IL-10) or control pDNA were delivered into the NAc of naïve rats. pDNA-IL-10 treatment reduces remifentanil self-administration in a drug concentration-dependent manner, similar to the previous observations in handled rats. Additional experiments confirmed that neither handling nor pDNA-IL-10 treatment alters operant responding for food or sucrose rewards. These results help define the conditions under which ventral striatal neuroimmune signaling may influence motivated behaviors for highly reinforcing opioid drugs.

It is commonly accepted that aerobic exercise increases hippocampal neurogenesis, learning and memory, as well as stress resiliency. However, human populations are widely variable in their inherent aerobic fitness as well as their capacity to show increased aerobic fitness following a period of regimented exercise. It is unclear whether these inherent or acquired components of aerobic fitness play a role in neurocognition. To isolate the potential role of inherent aerobic fitness, we exploited a rat model of high (HCR) and low (LCR) inherent aerobic capacity for running. At a baseline, HCR rats have two- to three-fold higher aerobic capacity than LCR rats. We found that HCR rats also had two- to three- fold more young neurons in the hippocampus than LCR rats as well as rats from the heterogeneous founder population. We then asked whether this enhanced neurogenesis translates to enhanced hippocampal cognition, as is typically seen in exercise-trained animals. Compared to LCR rats, HCR rats performed with high accuracy on tasks designed to test neurogenesis-dependent pattern separation ability by examining investigatory behavior between very similar objects or locations. To investigate whether an aerobic response to exercise is required for exercise-induced changes in neurogenesis and cognition, we utilized a rat model of high (HRT) and low (LRT) aerobic response to treadmill training. At a baseline, HRT and LRT rats have comparable aerobic capacity as measured by a standard treadmill fit test, yet after a standardized training regimen, HRT but not LRT rats robustly increase their aerobic capacity for running. We found that sedentary LRT and HRT rats had equivalent levels of hippocampal neurogenesis, but only HRT rats had an elevation in the number of young neurons in the hippocampus following training, which was positively correlated with accuracy on pattern separation tasks. Taken together, these data suggest that a significant elevation in aerobic capacity is necessary for exercise-induced hippocampal neurogenesis and hippocampal neurogenesis-dependent learning and memory. To investigate the potential for high aerobic capacity to be neuroprotective, doxorubicin chemotherapy was administered to LCR and HCR rats. While doxorubicin induces a progressive decrease in aerobic capacity as well as neurogenesis, HCR rats remain at higher levels on those measures compared to even saline-treated LCR rats. HCR and LCR rats that received exercise training throughout doxorubicin treatment demonstrated positive effects of exercise on aerobic capacity and neurogenesis, regardless of inherent aerobic capacity. Overall, these findings demonstrate that inherent and acquired components of aerobic fitness play a crucial role not only in the cardiorespiratory system but also the fitness of the brain.

Estradiol modulates the use of spatial navigation strategies in female rats. The presence of circulating estradiol enhances learning on tasks that require the use of a hippocampus-dependent place strategy and impairs learning on tasks that require the use of a dorsal striatum-dependent response strategy. When either strategy may be used successfully, estradiol biases females to use a place strategy. While this behavioral effect has been well-described in the young adult female rat, little is known about the mechanisms in the brain that underlie it or how it changes across age. The experiments in this dissertation examined how age, previous experience, and hormonal condition affect the ability of estradiol to modulate learning during explicit training of place and response tasks, as well as navigation strategy use during ambiguous navigation tasks. Age highly influenced the ability of estradiol to influence strategy use. While female rats could use place and response strategies to navigate by postnatal day (PD) 21, estradiol did not bias them to use a response strategy until PD26, just before puberty. In adulthood, previous navigation experience and estradiol interacted to influence navigation strategy use on a series of experiences to an ambiguous navigation task. And, estradiol impaired learning during explicit response training but did not affect place learning. In middle age, estradiol further impaired response learning but still did not affect place learning. Long-term hormone deprivation, however, was detrimental to acquisition of a place task but did not affect response learning. These experiments also examined the effects of estradiol on activity, plasticity, and reliability of neuronal ensembles in several subregions of the hippocampus and striatum during spatial navigation using cellular and molecular techniques that take advantage of the kinetics of the immediate-early genes c-fos and Arc. Increased activation and plasticity during active exploration across several subregions of the hippocampus and striatum reflected similar inputs to these neural systems and similar effects of exploration. However, estradiol modulated the plasticity and reliability of neuronal ensembles in the hippocampus and striatum specifically during goal-directed spatial navigation. Estradiol increased plasticity in CA1 of all behaviorally-trained rats, but only place strategy users displayed high reliability in this plasticity across training and probe trials on a navigation task. Estradiol prevented increase in plasticity and reliability in the dorsolateral striatum displayed by low estradiol response strategy users. These experiments reveal how several factors, including age, influence estradiol's modulation of spatial navigation strategy use and suggest functional mechanisms by which this modulation occurs.

Alzheimer’s Disease (AD) disproportionately impacts women; and the loss of ovarian hormones during the perimenopausal transition has been identified as a sex-specific risk factor. Previous studies have shown that the ovarian hormone, estrogen, utilizes its neuroprotective effects on tissues in the brain by aiding in cognitive function, exerting anti-inflammatory effects, promoting neuronal synaptic activity, and regulating energy biosynthesis. These effects are lost when circulating ovarian hormones are decreased. Additionally, during the perimenopausal transition women are experiencing similar neurological deficits found in AD patients such as reduced verbal acuity, memory deficits, delayed speech, etc. making early diagnosis of AD, if present, difficult. As a result, the window for therapeutic intervention is limited. Studies have shown that long-term physical exercise has been associated with a reduction in the rates of cognitive decline, dementia, and other related-neurodegenerative diseases. However, despite the strong evidence for greater female vulnerability, studies aiming to unravel the mechanisms that influence female susceptibility and the potential beneficial effects of exercise on cognitive function, menopause, and AD, are lacking.Here we sought to identify how hormonal changes during the perimenopausal transition influences the susceptibility of females to age-related cognitive decline and the effectiveness of physical exercise during this period in mouse models of AD. Based on a well-characterized neuropathological progression of the CVN-AD (APPSwDI/mNos2-/-) mouse model, we assessed mice at 24 weeks of age (WoA; mid AD-neuropathology) and at 36 WoA (late AD-neuropathology). Mice were treated with either the oil vehicle or 4-vinylcyclohexene diepoxide (VCD) to induce gradual ovarian failure. All mice were given pre- and post-cardiovascular tests at two timepoints, to assess the effects of exercise or being sedentary for 12 weeks. All 24 WoA mice remained sedentary throughout the study. Half of the 36 WoA CVN-AD mice remained sedentary while the other half were exercised with both voluntary wheel running and treadmill training for 12 weeks beginning at 24 WOA. Additionally, all mice were given a novel object recognition test (NOR) 1 week prior to sacrifice to assess short-term episodic memory. After sacrifice, uterine weights, body weights and total follicular counts were assessed. We found that VCD-treatment was effective in reducing uterine weights in all CVN-AD mouse models. Additionally, we found that at 36 WoA CVN-AD have a natural gradual loss in ovarian function. Exercise prior to the exacerbation of AD neuropathology and during the perimenopausal transition increased cardiovascular fitness, improved memory function, and increased the number of healthy ovarian follicles in comparison to sedentary 36 WoA CVN-AD mice. We then investigated the changes in forebrain metabolite levels in 36 WoA CVN-AD mice to identify whether metabolic changes in menopause-like ovarian failure were linked to AD progression and if exercise intervention could modify these effects. As a control, we used forebrain homogenates of sedentary 36 WoA NOS-/- (mNos2-/-) mice that were subjected to the same timelines of VCD- or oil-treatment. Forebrain samples were analyzed using the Biocrates MxP Quant 500 kit, 3 Flow-Injection-Analysis (FIA-MS) and 2 Ultra-High-Pressure Liquid Chromatography (UPLC). The CVN-AD genotype had significantly lower metabolite levels in comparison to NOS-/- mice and this effect was exacerbated by VCD-treatment. When evaluating the effects of exercise on the CVN-AD genotype we found that exercise significantly shifted the brain metabolome and increased metabolite levels in comparison to sedentary CVN-AD and NOS-/- mice. We then compared the interaction between exercise and VCD-treatment and found that exercise was able to reduce some of the negative effects associated with VCD. Within the sedentary CVN-AD treatment group, we found that VCD-treatment significantly increased the number of metabolite changes in comparison to Oil-treated mice. Whereas in the exercise CVN-AD treatment and NOS-/- control groups, VCD’s effects were dampened. These findings indicate that exercise was effective in reducing the effects of VCD-treatment, so much so, there was no difference in the number of metabolite changes between exercised CVN-AD and NOS-/- mice. We then sought to examine the potential role of menopause-like ovarian failure on the neuroinflammatory response and β-amyloid plaque deposition through the evaluation of overall microglial expression, homeostatic microglial expression, and β-amyloid plaque deposition in subregions of the hippocampus. We performed a triple immuno-fluorescence stain (Iba1, Tmem119 and β-amyloid) on brain slices through CA1, CA3 and dentate gyrus (DG) regions of sedentary and exercised 36 WoA CVN-AD mice and 36 WoA NOS-/- control mice. We used confocal microscopy to image the subregions of the hippocampus. Images were then analyzed in the ilastik software program to output cell and area counts of Iba1, Tmem119 and β-amyloid. VCD-treatment in sedentary CVN-AD mice significantly increased microglial proliferation in all subregions of the hippocampus. In comparison, exercise significantly reduced this effect. When evaluating Tmem119 expression, we found that in the CA1 region the CVN-AD genotype has significantly fewer healthy microglia in comparison to sedentary Oil-treated NOS-/- mice. When evaluating Tmem119 expression in the exercised CVN-AD mice we found that in the CA1 region, exercise was able to stave off some of the effects of the genotype and VCD-treatment, however, these effects did not occur in the CA3 and DG. Lastly, when evaluating how microglial expression coupled with exercise intervention and VCD-treatment affected β-amyloid plaque deposition in the CVN-AD mice we found no significant differences within any of the subregions. Taken together, these findings indicate that Aβ plaque deposition may occur independently from microglial expression and that regardless of exercise intervention and VCD-treatment, once Aβ plaques in the CVN-AD pathology occurs they will continue to persist. Collectively these data suggest that the CVN-AD neuropathology drastically impacts cognitive function, the brain metabolome and microglial response. Additionally, exercise as an early intervention during the perimenopausal transition period can prevent some, but not all the deleterious effects of the loss of estrogens and AD neuropathology. Overall, these findings will be significant in contributing to the AD field, especially in evaluating AD as a multiomic disease with sex-specific risk factors that can be modulated by early non-invasive exercise intervention.