Flexible neuronal participation within a reliable motor sequence

The reliable execution of learned motor sequences poses a challenge: they require reproducible neuron activity patterns for behavioral consistency yet must retain flexibility for modulation that depend on internal states and adaptation to external contingencies. The zebra finch, a songbird, provides an ideal system to examine this problem because their vocalizations are highly stereotyped and are associated with precisely time-locked bursts in HVC, a key nucleus involved in song production. However, their songs are not immutable /totally fixed, as they can be modulated during social interactions and can recover after brain lesions. Here, we used calcium imaging of HVC projection neurons during freely produced songs to track activity across repeated renditions of the same song from seconds to weeks. We first confirmed that song-locked calcium-event timing was stable across days. Surprisingly, not every neuron showed a calcium event on every rendition of the song. We referred to the presence of such an event as participation. This participation was a stable, neuron-specific property that varied with the past and future renditions of the songs within an utterance and depended on social context. Song recovery after brain lesion and refinement during learning further revealed that neural participation could change while sequence timing remained stable. These results suggest that stable motor sequences need not rely on rigid repeated activation of the same cellular ensemble but can preserve temporal order while flexibly selecting which neurons contribute to each expression of the behavior. This provides a cellular solution to the reliability-flexibility tradeoff in learned motor control.