Project Grant R01NS133584

MECHANISMS OF NEURODEGENERATION IN A NERVE AGENT MODEL - PROJECT SUMMARY/ABSTRACT ORGANOPHOSPHATE (OP) NERVE AGENTS (OPNA) ARE INCREASINGLY USED TO ATTACK CIVILIANS WORLDWIDE. OPNA POISONING IS A GLOBAL HEALTH PROBLEM. THERE IS NO EFFECTIVE TREATMENT FOR OPNA SURVIVORS. THE LIFE-LONG HEALTH CONSEQUENCES OF OPNA SURVIVORS ARE BEGINNING TO EMERGE. HOWEVER, THE MECHANISMS OF OPNA-INDUCED LONG-TERM BRAIN INJURY ARE LARGELY UNKNOWN. ACUTE EXPOSURE TO OPNA INDUCES SEIZURES (NEURAL EXCITABILITY) AND STATUS EPILEPTICUS (SE). IN THE LONG TERM, SE-INDUCED BRAIN CHANGES ALTER THE SIGNALING MOLECULES IN NEURONS AND GLIA. WE HYPOTHESIZE THAT OPNA-INDUCED SE PROMOTES KEY MOLECULAR INTERACTIONS AND EXACERBATES NEURODEGENERATION, REACTIVE GLIOSIS, AND THE DEVELOPMENT OF EPILEPSY. IN RECENT YEARS, NOVEL PATHWAYS OF NEUROINFLAMMATION AND NEURODEGENERATION ARE EMERGING AS MECHANISTIC TARGETS FOR THERAPEUTIC DEVELOPMENT. OUR CURRENT FINDINGS AND OTHERS SUGGEST THAT A NON-RECEPTOR SRC FAMILY TYROSINE KINASE FYN AND A SERINE/THREONINE CYCLIN-DEPENDENT KINASE 5 (CDK5) ARE THE CRITICAL KINASES ACTIVATED IN BOTH NEURONS AND GLIA IN RESPONSE TO STATUS EPILEPTICS (SE) THAT PROMOTE NEUROINFLAMMATION, HYPEREXCITABILITY, AND NEURODEGENERATION. THE ACTIVATED FYN AND CDK5 CAN TRIGGER A SELF-PERPETUATING PATHWAY IN THE GLIA AND INTERACT WITH PHOSPHORYLATED TAU, NR2B, AND PSD95 IN NEURONS TO PROMOTE AND MAINTAIN THE DISEASE STATE. THEREFORE, OUR OVERARCHING HYPOTHESIS IS THAT THE SEIZURES INDUCED BY ACUTE EXPOSURE TO OPNA FACILITATE FYN-TAU INTERACTIONS IN NEURONS AND CDK5 AND FYN ACTIVATION IN BOTH GLIA AND NEURONS. THESE, IN TURN, ACTIVATE NR2B-PSD95 INTERACTIONS TO CAUSE NEURONAL HYPEREXCITABILITY (EPILEPTIFORM SPIKING AND SPONTANEOUS SEIZURES), REACTIVE GLIOSIS AND THE PRODUCTION OF PROINFLAMMATORY CYTOKINES (NEUROINFLAMMATION), NITROOXIDATIVE STRESS AND NEURODEGENERATION ("DISEASE PROMOTERS"), AND PROMOTE BRAIN PATHOGENESIS IN THE LONG TERM. WE WILL TEST THE HYPOTHESIS IN A WELL-CHARACTERIZED OPNA (DIISOPROPYLFLUOROPHOSPHATE (DFP)) RAT MODEL. IN SPECIFIC AIM 1 (SA1), AFTER ACUTE EXPOSURE TO DFP, WE WILL CHARACTERIZE THE CHANGES IN FYN, TAU, AND CDK5 AND THEIR INTERACTIONS AT VARIOUS TIME POINTS IN KEY BRAIN REGIONS. WE WILL FRACTIONATE BRAIN LYSATES TO ISOLATE SYNAPTOSOMAL MEMBRANES, CYTOSOL, AND NUCLEAR FRACTIONS AND CONDUCT CO-IMMUNOPRECIPITATION (CO-IP)/WB. WE WILL USE BRAIN SECTIONS FOR PROXIMITY LIGATION ASSAY (PLA) TO DETERMINE INTERACTING COMPLEXES AND IMMUNOHISTOCHEMISTRY FOR CELL-SPECIFIC LOCALIZATION OF SIGNALING MOLECULES, GLIOSIS, AND NEURODEGENERATION. IN SA2, WE WILL VALIDATE THESE INTERACTIONS USING FYN-TAU INTERACTIONS BLOCKING PEPTIDE AND A CDK5 INHIBITOR. WE WILL INVESTIGATE THE INTERACTIONS OF FYN AND CDK5 WITH NR2B AND PSD95 AND THEIR IMPACT ON HYPEREXCITABILITY, NEURODEGENERATION, AND PROTECTION BY PATHWAY INHIBITORS AND BLOCKING PEPTIDES. WE WILL INVESTIGATE PERIPHERAL BIOMARKERS OF NEURODEGENERATION IN THE SERUM AND CSF. THIS PROPOSAL IS IN RESPONSE TO THE CCRP INITIATIVE FOA (PAR-23-027), "WHICH IS EXPECTED TO GENERATE DATA THAT ELUCIDATE THE MECHANISMS OF OPNA-INDUCED BRAIN TOXICITY AND POTENTIAL NEW TARGETS FOR THERAPEUTIC DEVELOPMENT."