THE DOMINANT ENVIRONMENTAL STRESSORS THAT MAY AFFECT CARDIAC BIOLOGY DURING SPACE EXPLORATION ARE SPACE RADIATION AND WEIGHTLESSNESS. THE KEY CENTRAL OBJECTIVE OF THIS PROPOSAL IS TO OBTAIN A MULTI OMICS AND PHYSIOLOGICAL UNDERSTANDING OF THE IMPACT OF SPACE RADIATION AND WEIGHTLESSNESS INDIVIDUALLY AND TOGETHER ON THE CARDIOVASCULAR SYSTEM USING TWO RODENT MODELS: RAT NEONATAL CARDIOMYOCYTES (RNNC) AND C57BL/6 MICE. IN STUDIES EVALUATING THE EFFECT OF MICROGRAVITY ON THE PROTEIN CONTENT OF RNNC WE WERE ABLE TO DEMONSTRATE THAT PROTEIN TURNOVER DIMINISHES GLOBALLY IN THESE CELLS. MOREOVER WE WERE ABLE TO PINPOINT MOLECULAR PATHWAYS POTENTIALLY RESPONSIBLE AND POSSIBLY TARGETABLE FOR THIS EFFECT. WE HAVE AN ONGOING STUDY EVALUATING THE CARDIAC PHENOME AND CARDIAC PROTEOME IN C57BL/6 MICE EXPOSED TO GALACTIC COSMIC RAYS (GCR). OUR DATA SUGGESTS THAT IN CONTRAST TO OTHER FORMS OF RADIATION A SINGLE EXPOSURE TO GCR RESULT IN A SIGNIFICANT DETERIORATION OF CARDIAC FUNCTION THAT BECOMES APPARENT LONG AFTER EXPOSURE (9-12 MONTHS POST GCR). BIOINFORMATICS ANALYSES OF THE CARDIAC PROTEOME OF THESE MICE HAVE IDENTIFIED THAT KERATINIZATION AND CORNIFICATION PATHWAYS ARE UP REGULATED IN HEARTS FROM GCR IRRADIATED MICE. THESE PATHWAYS ARE PRESUMABLY UPREGULATED AS A RESPONSE TO SUSTAINED INFLAMMATION AND PERSISTENT OXIDATIVE DAMAGE POST RADIATION. THE IMPACT OF BOTH STRESSORS SIMULTANEOUSLY (GCR AND WEIGHTLESSNESS) ON CARDIAC PHYSIOLOGY AND CARDIAC PROTEOMICS HAS NOT BEEN EXPLORED AT ALL. WE WILL LEVERAGE OUR EXPERTISE USING THESE MODELS FROM TWO SPECIES (MOUSE AND RAT) AS WELL AS ACCUMULATED DATA SETS AND BIOSPECIMENS ACCRUED FROM OTHER STUDIES TO ADDRESS THE FOLLOWING TWO AIMS: AIM 1. TEST THE HYPOTHESIS THAT THE COMBINATION OF GCR EXPOSURE AND WEIGHTLESSNESS WILL RESULT IN ADDITIVE OR EVEN SYNERGISTIC ENHANCEMENTS IN CELL DEATH INCREASES IN REACTIVE OXYGEN SPECIES PROTEIN TURNOVER AND MITOCHONDRIAL DYSFUNCTION OVER THAT OBSERVED WITH EITHER STRESSOR ALONE. THIS AIM WILL UTILIZE THE RNNC MODEL AND THE SYNTHECON ROTARY CELL CULTURE SYSTEM FOR WEIGHTLESSNESS. GCR WITH AND WITHOUT WEIGHTLESS WILL BE ADMINISTERED TO RNNC AT NSRL. FUNCTIONAL READOUTS WILL INCLUDE ASSESSMENTS OF CELL VIABILITY CELL DEATH (APOPTOSIS) MITOCHONDRIAL INTEGRITY AND FUNCTIONALITY AS WELL AS MULTI OMICS COMPARATIVE ASSESSMENTS (TRANSCRIPTOMICS AND MASS SPECTROMETRY-BASED PROTEOMICS).