PESTICIDES IN AGRICULTURAL ENVIRONMENTS CAN HAVE NON-TARGET EFFECTS, INCLUDING SUB-LETHAL IMPACTS ON POLLINATORS, POTENTIALLY CAUSING DAMAGE TO INSECTS AND ECOSYSTEMS. INSECTS THAT FEED ON TOXIC HOST PLANTS MAY RETAIN TOXIC COMPOUNDS (SEQUESTRATION), AND THIS MECHANISM MAY TRANSLATE DIFFERENTIAL SENSITIVITY WHEN CONFRONTED WITH PESTICIDES. AN OVERARCHING GOAL OF THIS PROPOSAL IS TO LINK INSECT PHYSIOLOGY WITH ENVIRONMENTAL PRESSURES (I.E., NATURAL OR APPLIED TOXINS). INSECT WING PHYSIOLOGY REPRESENTS A NEXUS BETWEEN ENVIRONMENTAL TOXINS AND THE INSECT CIRCULATORY SYSTEM. LITTLE IS KNOWN HOW CIRCULATION (AND THUS SENSORY DEMANDS) CAN BE DISRUPTED BY AGRICULTURAL PESTICIDES SUCH AS NEONICOTINOIDS. SEQUESTRATION OF CARDENOLIDES, OCCURRING THROUGH HEMOLYMPH CIRCULATION IN WINGS, COULD ALLOW INSECTICIDES TO "HITCHHIKE" INTO THE WINGS, CAUSING DAMAGE. USING A WELL-RECOGNIZED MODEL SYSTEM, THE MONARCH BUTTERFLY (DANAUS PLEXIPPUS) AND ITS MILKWEED PLANTS (GENUS ASCLEPIAS), WE FOCUS ON ACCUMULATION OF CARDENOLIDES, WHICH ARE DYNAMICALLY PRODUCED IN THE PLANT AND SEQUESTERED BY MONARCHS. FOR STRONG COMPARISON, THIS PROPOSAL MEASURES BOTH CARDENOLIDE AND PESTICIDE ACCUMULATION (SPECIFICALLY NEONICOTINOID CLOTHIANIDIN) IN PAIRS OF CLOSELY RELATED (SEQUESTERING AND NON-SEQUESTERING) LEPIDOPTERA THAT FEED ON MILKWEED: D. PLEXIPPUS AND EUPLOEA CORE, AND CYCNIA TENERA AND EUCHAETES EGLE. SPECIFICALLY, AS CARDENOLIDES AND PESTICIDES ACCUMULATE: 1) DETERMINE LOCALIZATION OF TOXIC COMPOUNDS; 2) CHARACTERIZE CHANGES IN HEMOLYMPH FLOW IN WINGS; 3) MEASURE TOXICITY IN MONARCH POLLINATORS/HOST PLANTS NEAR NATURAL AND AGRICULTURAL AREAS. THIS PROPOSAL ADVANCES OUR KNOWLEDGE OF CARDENOLIDE AND PESTICIDE ACCUMULATION, SEQUESTRATION, AND CRITICAL EFFECTS ON INSECT PHYSIOLOGY IN BOTH NATIVE AND PEST SPECIES.