Interventions to improve mild cognitive impairment (MCI) are largely limited by the lack of knowledge on the mechanisms underlying the neurological condition. Thus, understanding the factors that initiate and exacerbate the disease would contribute to advancement in treatments. It has been shown that dietary modifications, including a high fiber resistant starch (RS) diet, impact neurocognitive health. In this study, mice were given an RS supplemented diet or an isocaloric diet for 2 months to determine the effect of RS supplementation on neuroinflammation and neurodegeneration. After 2 months, the hippocampus and entorhinal cortex of the brains were analyzed for astrogliosis, microglial activation, and mature neurons. The group on the RS diet had fewer neuroinflammatory markers; however, we did not see a decrease in neurodegeneration. These findings are consistent with previous studies suggesting an interaction between metabolic health and cognitive impairment. Future research investigating this interplay would be beneficial for medical advancements for MCI.

Organophosphate cholinesterase inhibitors (OPs) are a class of neurotoxic chemicals that have been used globally as both pesticides and chemical warfare agents. Most recently, OPs have been used during the Syrian civil war, a political assassination in Asia, and terrorist attacks in Europe. Survivors of high-dose acute OP intoxication can experience convulsions, prolonged seizures, or death if not treated within minutes of exposure. During the months or years that follow, humans who survive acute OP intoxication often experience significant changes in brain structure, general cognition, memory, or electrographic activity. Even those who received standard of care treatment with atropine, oximes, and benzodiazepines may experience chronic neurological deficits or electrographic abnormalities. For this reason, there is an urgent need for more efficacious therapeutics to combat acute OP poisoning. To develop more effective therapeutic strategies, it is critical to better understand how acute OP intoxication impacts the brain and the reasons for the limitations of the current standard of care. A critical question in the field is whether seizure-independent mechanisms contribute to long-term neurological damage following acute OP intoxication, and if so, which mechanisms play a role. Additionally, very few studies have examined the influence of sex and age on the chronic neurological impacts of acute OP intoxication. This dissertation seeks to address these data gaps using a rat model of acute intoxication with OP threat agent diisopropylfluorophosphate (DFP) as a model. Chapter 2 demonstrates that the current standard of care reduced seizure activity but offered only partial neuroprotection at 3 and 6 months post-intoxication, and failed to protect against chronic microglial activation and midbrain mineralization. Studies described in Chapter 3 leveraged a seizure-resistant subset of rats acutely intoxicated with DFP to determine whether DFP induced neurological damage independent of seizure activity. Seizure-resistant animals showed pronounced neuronal degeneration and mineralization, although these outcomes were not as severe or as persistent as in those animals who experienced seizures. Chapter 5 describes the development of a juvenile rat model of acute DFP intoxication that addressed potential sex differences and better reflected the diversity of individuals potentially affected by OP intoxication. Juvenile males had more severe seizures, astrogliosis, and aberrant neurogenesis compared to juvenile females, but both sexes experienced persistent microglial activation and cognitive deficits following DFP intoxication. This model also suggested that juveniles were generally more resistant to OP-induced seizures and experienced a later onset of neuroinflammation compared to their adult counterparts. These findings have important implications for the development of medical countermeasures against OP threat agents. The observation that both benzodiazepine-treated and seizure-resistant animals experienced chronic neuronal degeneration despite having little to no seizure activity suggests that seizure-independent mechanisms contribute to the long-term neurological effects of acute OP intoxication. My findings also suggest that both age and sex are biological variables that influenced OP-induced neurotoxicity, and should, therefore, be important considerations in preclinical assessment of toxicity and therapeutic efficacy. Lastly, the temporal patterns of microglial activation in juvenile animals, as well as the persistence of microglial activation despite antiseizure treatment, point to microglial-mediated neuroinflammation as a likely factor contributing to long-term neurological consequences following acute OP intoxication.

Organophosphorus cholinesterase inhibitors (OPs) are a class of toxic chemicals that includes both pesticides and nerve agent chemical weapons. Every year accidental or intentional exposure to OPs accounts for millions of poisonings globally. Current standard of care (SOC) for treatment of acute OP intoxication improves survival if administered promptly following intoxication but does not prevent development of chronic neuropathology or neurological sequelae in survivors of such intoxications. While acute OP-induced status epilepticus (SE) is known to be a critical driver of subsequent neuropathology and adverse neurological consequences following intoxication, treatment of acute OP intoxication is likely to be delayed beyond the optimal therapeutic window for SOC intervention. This dissertation addresses two limitations in the treatment of acute OP intoxication: 1) identification of biomarkers for identification of populations at-risk of chronic neurological outcomes, and 2) assessment and targeting of the downstream process of neuroinflammation to mitigate adverse neurological consequences following OP poisoning. In Chapter 2, we identify acute and delayed electrophysiologic signatures following acute intoxication with the OP pesticide diisopropylfluorophosphate (DFP) that correlate with the frequency of spontaneous recurrent seizures (SRS). These findings present novel and translatable electrophysiologic biomarkers with potential to assist in triage following a mass casualty OP exposure incident. Chapter 3 characterizes the microglial and astrocytic neuroinflammatory responses following acute DFP intoxication, providing novel information regarding the spatiotemporal progression of glial phenotypes up to 28 days post exposure (DPE). These data point to early targeting of pro-inflammatory microglial activation as a potential strategy to improve outcomes following acute DFP intoxication. Chapter 4 builds upon the findings of Chapter 3 through transient depletion of microglia via administration of the colony stimulating factor 1 receptor (CSF1R) inhibitor PLX5622 to female rats following acute DFP intoxication. We identify a time-dependent anti-epileptogenic effect of CSF1R inhibition post-DFP intoxication; PLX5622 administration 1-3 DPE prevented the development of SRS while PLX5622 from 4-6 DPE did not reduce the incidence of SRS relative to animals intoxicated with DFP and not provided PLX5622. Animals treated with PLX5622 1-3 DPE displayed attenuated chronic microglial and astrocytic activation and a greater density of neurons at 28 DPE. Collectively, Chapters 3 and 4 provide a natural history of glial inflammatory responses and identify the time-dependent epileptogenic effect of microglial activation following acute DFP intoxication. In Chapter 5, we assess the pleiotropic anti-seizure/anti-inflammatory neurosteroid allopregnanolone (ALLO) in a translatable model of acute soman intoxication. ALLO provides dose-dependent control of soman-induced seizures and mitigates acute neurodegeneration and neuroinflammation following soman intoxication. The findings of this dissertation offer tangible, clinically relevant improvements to acute and delayed treatment of OP poisoning, while also highlighting the time-dependency of microglial-targeted anti-inflammatory interventions post-intoxication.

Organophosphate (OP) cholinesterase inhibitors and GABA receptor antagonists represent mechanistically diverse classes of chemicals that can cause seizures and death following acute intoxication at relatively low doses. OPs inhibit acetylcholinesterase (AChE), which results in the over accumulation of acetylcholine (ACh) in the nervous system. Rapid inhibition of > 60-80% of brain AChE triggers seizures that rapidly progress to life-threatening status epilepticus (SE). GABA receptor antagonists, exemplified by tetramethylenedisulfotetramine (TETS), induce seizures and death by reducing the inhibitory tone in the nervous system, which results in neuronal hyperexcitability. The standard first-line therapy for the treatment of TETS- or OP-induced SE, high-dose benzodiazepines, are effective at terminating seizure behavior and reducing mortality if administered within minutes after exposure. However, even with timely treatment, benzodiazepines often fail to adequately protect against subsequent brain damage and neurologic deficits. Preclinical rodent models are essential to elucidating the cellular and molecular mechanisms driving acute seizurogenic events and the spatiotemporal progression of brain damage following SE. The major objective of this dissertation was to characterize acute and chronic neurotoxic outcomes in novel mouse models of TETS- and OP-induced SE to gain a better understanding of the pathogenic mechanisms contributing to SE and the consequential neuropathology. In Chapter 2, in vivo imaging is used to compare the spatiotemporal progression of neuropathology after TETS-SE in two different mouse strains commonly used to study chemical-induced SE. The findings in this study demonstrate that the extent of brain damage observed in the mouse brain after TETS-induced SE varies according to strain and the duration of SE. Chapter 3 focuses on the development and characterization of a mouse model of acute intoxication with diisopropylfluorophosphate (DFP). Specifically, this chapter describes the spatiotemporal progression of AChE inhibition, neuropathology, and behavioral deficits up to 28 days post-exposure. The findings in this chapter demonstrate that this mouse model replicates many of the outcomes observed in rats and humans acutely intoxicated with OPs, suggesting the feasibility of using this model for mechanistic studies and therapeutic screening. The mouse model of acute DFP intoxication was leveraged in Chapter 4 to investigate the role of nicotinic cholinergic receptors (nAChR) in the initiation and propagation of OP-induced SE. The findings described in Chapter 4 identify a role for α4-containing nAChR in the initiation and/or progression of seizures following acute OP intoxication, and support further investigation of nicotinic antagonists as prophylactics for OP-induced SE. The studies presented in this dissertation support the use of mouse models to investigate the mechanisms contributing to the initiation and consequences of SE caused by chemical threat agents. The findings of this work have important mechanistic and therapeutic implications regarding the pathophysiology and treatment of chemical-induced SE. The observation that genetic background and duration of SE influence the neuropathologic consequences of TETS-SE suggests that humans who do not respond to antiseizure medication after acute TETS intoxication are at increased risk for brain injury, and this susceptibility may vary genetically. The observation that nicotinic receptors play a necessary role in the initiation of OP-induced SE highlights a novel therapeutic target, and supports the strategy of using a nicotinic antagonist as a preventative antidote against OP-induced seizures.