Inhibiting Inducible Nitric Oxide Synthase with 1400W Reduces Soman (GD)-Induced Ferroptosis in Long-Term Epilepsy-Associated Neuropathology: Structural and Functional Magnetic Resonance Imaging Correlations with Neurobehavior and Brain Pathology.

Organophosphate (OP) nerve agent (OPNA) intoxication leads to long-term brain dysfunctions. The ineffectiveness of current treatments for OPNA intoxication prompts a quest for the investigation of the mechanism and an alternative effective therapeutic approach. Our previous studies on 1400W, a highly selective inducible nitric oxide synthase (iNOS) inhibitor, showed improvement in epilepsy and seizure-induced brain pathology in rat models of kainate and OP intoxication. In this study, magnetic resonance imaging (MRI) modalities, behavioral outcomes, and biomarkers were comprehensively investigated for brain abnormalities following soman (GD) intoxication in a rat model. T1 and T2 MRI robustly identified pathologic microchanges in brain structures associated with GD toxicity, and 1400W suppressed those aberrant alterations. Moreover, functional network reduction was evident in the cortex, hippocampus, and thalamus after GD exposure, and 1400W rescued the losses except in the thalamus. Behavioral tests showed protection by 1400W against GD-induced memory dysfunction, which also correlated with the extent of brain pathology observed in structural and functional MRIs. GD exposure upregulated iron-laden glial cells and ferritin levels in the brain and serum, 1400W decreased ferritin levels in the epileptic foci in the brain but not in the serum. The levels of brain ferritin also correlated with MRI parameters. Further, 1400W mitigated the overproduction of nitroxidative markers after GD exposure. Overall, this study provides direct evidence for the relationships of structural and functional MRI modalities with behavioral and molecular abnormalities following GD exposure and the neuroprotective effect of an iNOS inhibitor, 1400W. SIGNIFICANT STATEMENT: Our studies demonstrate the MRI microchanges in the brain following GD toxicity, which strongly correlate with neurobehavioral performances and iron homeostasis. The inhibition of iNOS with 1400W mitigates GD-induced cognitive decline, iron dysregulation, and aberrant brain MRI findings.

The NADPH Oxidase Inhibitor, Mitoapocynin, Mitigates DFP-Induced Reactive Astrogliosis in a Rat Model of Organophosphate Neurotoxicity.

NADPH oxidase (NOX) is a primary mediator of superoxides, which promote oxidative stress, neurodegeneration, and neuroinflammation after diisopropylfluorophosphate (DFP) intoxication. Although orally administered mitoapocynin (MPO, 10 mg/kg), a mitochondrial-targeted NOX inhibitor, reduced oxidative stress and proinflammatory cytokines in the periphery, its efficacy in the brain regions of DFP-exposed rats was limited. In this study, we encapsulated MPO in polyanhydride nanoparticles (NPs) based on 1,6-bis(p-carboxyphenoxy) hexane (CPH) and sebacic anhydride (SA) for enhanced drug delivery to the brain and compared with a high oral dose of MPO (30 mg/kg). NOX2 (GP91phox) regulation and microglial (IBA1) morphology were analyzed to determine the efficacy of MPO-NP vs. MPO-oral in an 8-day study in the rat DFP model. Compared to the control, DFP-exposed animals exhibited significant upregulation of NOX2 and a reduced length and number of microglial processes, indicative of reactive microglia. Neither MPO treatment attenuated the DFP effect. Neurodegeneration (FJB+NeuN) was significantly greater in DFP-exposed groups regardless of treatment. Interestingly, neuronal loss in DFP+MPO-treated animals was not significantly different from the control. MPO-oral rescued inhibitory neuronal loss in the CA1 region of the hippocampus. Notably, MPO-NP and MPO-oral significantly reduced astrogliosis (absolute GFAP counts) and reactive gliosis (C3+GFAP). An analysis of inwardly rectifying potassium channels (Kir4.1) in astroglia revealed a significant reduction in the brain regions of the DFP+VEH group, but MPO had no effect. Overall, both NP-encapsulated and orally administered MPO had similar effects. Our findings demonstrate that MPO effectively mitigates DFP-induced reactive astrogliosis in several key brain regions and protects neurons in CA1, which may have long-term beneficial effects on spontaneous seizures and behavioral comorbidities. Long-term telemetry and behavioral studies and a different dosing regimen of MPO are required to understand its therapeutic potential.