Clinically available antidepressants share the same core mechanisms of blocking serotonin and noradrenaline reuptake in the brain. However, ~50% of patients do not fully respond to available treatments, suggesting other neurotransmitters are also involved in depression. The mesolimbic dopamine system governs reward and motivation, and dysfunction of dopaminergic transmission could account for anhedonia, lack of motivation and other symptoms of depression. The overall goal of this application is to understand how dopaminergic transmission is altered in depressive-like states and how this alteration contributes to depressive behavior. Mesolimbic dopamine is synthesized by dopamine neurons in the ventral tegmental area (VTA), and dopamine release is triggered by action potential (AP) firing. Using chronic unpredictable stress (CUS) as a mouse model of depression, our preliminary studies indicate that CUS decreased AP firing and an increase in the activity of phosphodiesterase 4 (PDE4), the enzyme that catalyzes the hydrolysis of cAMP, in the VTA. Spontaneous AP firing in midbrain dopamine neurons is driven primarily by the pacemaker channel--the hyperpolarization-activated cyclic nucleotide-gated channel (HCN). As its name indicates, HCN senses chemical (cAMP) and electrical (voltage) signals. We hypothesize that the CUS-induced decrease in cAMP in the VTA impairs the activation of HCN, leading to decreases in AP firing and dopamine release; the impaired dopaminergic transmission may contribute to anhedonia and other depressive-like behavior. Two Specific Aims are proposed to test this hypothesis. The objective of Aim I is to investigate the mechanisms for CUS-induced decrease in action potential firing in VTA dopamine neurons. We will examine whether changes in activation of HCN and other voltage-gated channels and excitatory and inhibitory synaptic inputs contribute to CUS-induced decrease in AP firing in VTA dopamine neurons. The objective of Aim II is to test the hypothesis that HCN channelopathy in the VTA contributes to CUS-induced depressive-like behaviors. Specifically, we will examine the cellular and behavioral effects of loss- and gain-of-function of HCN2 in the VTA. Completion of this project is expected to provide key mechanisms linking chronic stress to deficiency in dopaminergic transmission and depressive-like behaviors. Furthermore, this study has the potential to uncover novel therapeutic targets for pharmacotherapy of depression.