Project Summary/Abstract Reducing vector populations will help decrease the global impact of mosquito-borne diseases. Over reliance on insecticide classes repurposed from agriculture has culminated in widespread resistance, resulting in a critical need to investigate physiological processes that can provide selective targets for insecticide development. Insect muscarinic acetylcholine receptors (mAChRs) have been validated as an insecticide target, but no molecules with this mode of action are currently used for pest control. The long-term goal of the proposed research is to exploit pharmacological differences between insects and mammals to develop mAChR-selective insecticides to be integrated into mosquito control programs. The overall objective of the proposed research is to investigate the pharmacology and physiology of Anopheles gambiae mAChRs using chemical probes and novel nutrient conjugates to enhance oral delivery of these toxicants. The central hypothesis of the proposed research is that insect mAChR subtypes display unique pharmacology and play critical roles in the mosquito central nervous system (CNS). The rationale for the proposed research is based on the global health burden that mosquitoes impose, with a focus on the malarial vector Anopheles gambiae. Two specific aims will be performed to test the central hypothesis. Specific Aim #1 will interrogate the pharmacology of An. gambiae mAChR subtypes using a cholinergic chemical library with defined biological and pharmaceutical activity. Specific Aim #2 will focus on the synthesis of a series of analogs of a lead molecule to probe the function of mosquito muscarinic receptor activity, as well as absorption pathways for its dietary delivery. The proposed research is conceptually innovative as it is a departure from the status quo by i) investigating the muscarinic system as an unexploited mode of action to break insecticide resistance, and ii) leveraging midgut nutrient transporters to facilitate dietary uptake of pro- insecticides. This project will provide significant advances to the fields of mosquito biology, as well as insecticide toxicology and development.