Project Abstract Norepinephrine (NE) and serotonin (5-HT) are neurotransmitters released by a small number of neurons in the locus coeruleus (LC) and dorsal raphe (DR), respectively. Each group of neurons provides extensive innervation of most of the brain. Although these groups of cells are often considered to be homogenous, recent data indicates the existence of subclasses of each neuron. This proposal aims to generate multi-scale data on each system, comprising molecular, genetic, anatomical, physiological, and behavioral experiments. Our team is in a unique position to integrate across experiments and develop a new understanding of the subclasses of LC-NE and DR-5-HT neurons and their functions during behavior. Three aims test specific questions about the cell types and functions of LC-NE and DR-5-HT neurons in mice: 1) LC-NE neuron subclasses have distinct biophysical properties and axonal projections; 2) subclasses of DR-5-HT neurons have distinct biophysical properties and axonal projections; and 3) subclasses of LC-NE and DR-5-HT neurons have different functions during dynamic decision making. Our experiments will be used to develop a comprehensive classification of NE and 5-HT neuronal subclasses and will be used to generate reagents to target those cells.