Mixed Selectivity of Subthalamic Nucleus Neurons in Encoding Motor and Reward Behaviors

Summary The subthalamic nucleus (STN) plays a critical role in modulating motor and cognitive functions within the basal ganglia, with its dysfunction being implicated in movement disorders such as Parkinson’s disease. However, the behavioral representations of individual STN neurons remain incompletely understood. Using in vivo two-photon calcium imaging in behaving mice, we systematically mapped the activity of single STN neurons across diverse behavioral contexts, including locomotion, licking, and reward-driven actions. Our findings reveal that STN neurons exhibit mixed selectivity, encoding multiple behaviors with distinct temporal dynamics and excitatory or inhibitory response patterns. This mixed selectivity allows the STN to robustly encode motor parameters such as locomotion speed and licking intensity while integrating contextual information from different behavioral states. Comparisons with the adjacent zona incerta (ZI) revealed distinct encoding properties: while both regions represent locomotion, STN neurons more faithfully track motor states, whereas ZI neurons exhibit prolonged calcium events with weaker movement correlations. Population-level analysis showed STN activity in a low-dimensional neural manifold, with components linked to movement velocity and licking intensity. Notably, locomotion encoding in STN was context-dependent, diverging when movements were internally generated versus reward-modulated. Together, these findings highlight the specialized yet flexible role of the STN in integrating motor and reward-related signals, supporting a framework in which STN neurons contribute to motor control through multiplexed and context-dependent encoding. This work provides new insights into the functional organization of basal ganglia circuits and has implications for understanding STN’s role in both physiological and pathological conditions.