SUMMARY: Drugs of abuse (like opiates, cocaine, and nicotine) are initially taken because people want to alter how they subjectively feel, but these substances have potentially longer lasting negative consequences on the human brain, because of neurobiological effects that can lead to substance use disorder. The dopamine and serotonin systems have been strongly implicated in these processes, but these systems have not been investigated in humans with the temporal resolution required to connect (1) how these rapid signals alter our behavior as human navigate the world, nor (2) how these neurochemical signals alter how we feel subjectively. Progress has been hindered by lack of technology that permits simultaneous, real-time, measurements of dopamine and serotonin release during conscious decision-making and experience in humans (and for the fact that non-human model organisms cannot report ‘how they feel’). This proposal capitalizes on recent advances in mathematized human decision theory and our group’s first-of- its-kind technological approach, which now allows simultaneous, col-localized, measurements of dopamine and serotonin release in the human brain with sub-second temporal resolution during conscious decision- making and experience. Here, we pursue two specific goals, which uniquely combines our validated, peer- reviewed, and published approach with probabilistic learning tasks that have been specifically designed to probe the interplay between opponent reward and punishment behavioral signals, rapid changes in dopamine and serotonin release, and moment-to-moment changes in how participants feel during sequences of experience. First (aim 1), we will test the hypothesis that sub-second changes in extracellular dopamine and serotonin encode monetary reward/appetitive prediction errors and monetary punishment/aversive prediction errors, respectively, as opponent signals in the human striatum during an instrumental learning task and a passive Pavlovian conditioning task. Second (aim 2), we will test the hypothesis that sub-second changes in extracellular dopamine and serotonin, together, direct moment-to-moment changes in how people feel. The experiments proposed will for the first time directly test the controversial “opponent hypothesis” for serotonin/dopamine function in learning and decision-making in humans; and, we will for the first time begin to expose what role rapid microfluctuations in dopamine and serotonin play in encoding positive and negative subjective feelings in humans. We expect that these first-of-their-kind data will provide new insight into how drugs of abuse may alter how people feel, the impact drugs of abuse may have on critical neurochemical learning systems in the human brain, and may lay a foundation for future work aimed at understanding how these neurochemical systems affect other human brain regions (including nucleus accumbens, prefrontal cortex, amygdala, and the temporal lobes) where access maybe afforded by ...