The recent attack on civilians in Syria using an organophosphate (OP) nerve agent, sarin, confirms its continued threat to humans. OP nerve agents are potent seizurogenic neurotoxins. OP intoxication, in long-term, will cause irreversible brain damage due to hyperexcitability of neurons, reactive gliosis, and neurodegeneration. If these are not adequately controlled at a very early stage, they will lead to the development of epilepsy, cognitive deficits and other neurological disorders. Currently there is no treatment for long-term effects. The current toxin-specific and symptomatic drugs atropine, oxime, and diazepam (DZP) are inadequate to prevent OP-induced long-term brain injury. DZP controls seizures and could be neuroprotective if it is given within 30 minutes of OP exposure. This is impossible to achieve in real life scenarios of mass OP exposure. Therefore the objective of the project is to test two novel neuroprotectants, diapocynin and 1400W that are not previously tested in OP intoxication models, to prevent long-term brain damage. OP exposure increases the levels of reactive oxygen/nitrogen species (ROS/RNS), which cause long-term brain damage. The diapocynin, an effective NADPH oxidase inhibitor targets ROS. The 1400W, a potent and highly selective inhibitor of inducible nitric oxide synthase targets RNS, and no adverse effects or toxicity are reported when it was tested in humans and rodent models. Both neuroprotectants are blood-brain barrier permeable and ameliorated long-term neuropathology in rodent models of epilepsy, Parkinson's disease, and traumatic brain injury. ROS and RNS are also increased in these models. Therefore, our overarching hypothesis is that diapocynin and/or 1400W, given after the symptomatic drugs, will protect OP-induced long-term brain pathology. To test the hypothesis, we will use our established diisopropylfluorophosphate (DFP, a surrogate for nerve agents) rat model to replicate a real life scenario of nerve agent poisoning. Our preliminary studies from this model revealed an increased production of ROS/RNS and cytokines levels between 24-48h after the DFP exposure. At 1 week, hyperexcitability of neurons (increased epileptiform spiking on the EEG), reactive gliosis (astrocytes/microglia markers), and neurodegeneration (increased number of flurojade-B positive neurons) were observed in the hippocampus. Both diapocynin and 1400W suppressed these DFP-induced changes during the 24-48h and at 1 week, and also reduced the number of seizures during the first 4 weeks. These findings suggest long-term neuroprotective potential of diapocynin and 1400W in OP poisoning. In the proposed study, we will further validate these findings comprehensively by utilizing various histological and biochemical assays from serum and hippocampi samples at 4 week, cognitive function test, and continuous video-EEG monitoring for 12 weeks. The positive outcome from our study will provide proof-of-concept for long-term neuroprotective properties of diapocynin and 1400W in DFP-induced brain injury. This will help us to advance the compounds as potential countermeasures for real nerve agents by optimizing in preclinical testing for potency, toxicity, and pharmacokinetics (ADME studies).
The diisopropylfluorophosphate (DFP) is a potent seizurogenic neurotoxin and a surrogate for organophosphate (OP) nerve agents that are classified as extremely lethal warfare agents likely to be abused in terrorism. The civilian populations are at risk if they are exposed to these chemical agents, which will cause deaths at high concentrations but sublethal concentrations cause acute seizures that will most likely lead to the development of epilepsy for lifelong and/or other neurological disorders if they are not adequately treated immediately. The current treatment with symptomatic drugs alone is inadequate to prevent long-term neurological problems, therefore we propose to test combined effects of symptomatic drugs and neuroprotectants in the rat DFP model to mimic a real life scenario of OP intoxication in humans.
| Sharma, Shaunik; Puttachary, Sreekanth; Thippeswamy, Thimmasettappa (2017) Glial source of nitric oxide in epileptogenesis: A target for disease modification in epilepsy. J Neurosci Res : |
