Identification and optimization of verapamil as a novel neuroprotective and anti-inflammatory agent for reducing long-term neurological morbidities following organophosphate-induced status epilepticus

点击翻译按钮获取中文摘要

ABSTRACT Organophosphate (OP) compounds include pesticides and chemical warfare nerve agents. They are highly toxic and can produce a cholinergic crisis that rapidly progresses into status epilepticus (SE) and even death without emergency care. The standard-of-care (SOC) treatment with atropine, pralidoxime, and midazolam has dramatically improved survival after OP intoxication. Yet, many survivors of OP-SE exhibit brain injury, cognitive impairments, and spontaneous recurrent seizures (SRS). In addition, both acute and protracted neuro-inflammation and increased expression of pro-inflammatory cytokines have been reported following OP SE. These persistent neuroinflammatory changes are thought to underlie neurodegeneration, network hyper- excitability, and maladaptive plasticity, leading to cognitive dysfunction and SRS. Thus, mitigating neuro- inflammation is a primary target in alleviating neuronal injury and behavioral morbidities following OP SE. Verapamil (VPM) is a water-soluble calcium-channel blocker for high blood pressure and angina treatment. Recent findings have also demonstrated the potent neuroprotective and anti-inflammatory action of VPM in various CNS injuries. Our preliminary results showed that intramuscular VPM (10 mg/kg, i.m.) was safe and produced significant neuroprotection and decreased neuroinflammation in multiple brain regions when administered after the termination of DFP SE. It was also associated with decreased pro-inflammatory and upregulation of anti-inflammatory cytokines. Finally, this effect had a functional outcome since VPM improved anxiety and cognitive impairment at eight weeks post-DFP SE. Thus, this UG3-UH3 will investigate and optimize a VPM therapy as a potential countermeasure for treating OP SE that could be rapidly administered in the field. Studies will employ DFP-SE rat model to conduct the Specific Aims: In Aim 1, the safety of repeated i.m. injections will be assessed along with an assessment of pharmacokinetic parameters and the stability of VPM formulation. In Aim 2, the effects of VPM treatment on reducing neuronal death and neuroinflammation after DFP SE will be evaluated utilizing Fluoro-Jade C along with Glial Fibrillary Acidic Protein and Ionized calcium-binding adaptor molecule-1 immunohistochemical staining, respectively. We will also assess the effect of VPM therapy on pro- and anti-inflammatory cytokine expression. In Aim 3, VPM therapy will be optimized by studying the effects of various VPM doses and treatment durations on neurodegradation and neuroinflammation. In Aim 4, the functional outcomes of optimized VPM therapy on long-term anxiety, cognitive impairment, and SRS will be tested using a battery of rodent behavioral assays and EEG techniques. We will also draft a preliminary target product profile (TPP) by the end of our studies. These studies will provide further insight into the role of neuroinflammatory mechanisms in mediating OP morbidities and optimizing a VPM-based anti-inflammatory therapy as a novel countermeasure drug to alleviate OP toxicities.