Integration of cooperative and opposing molecular programs drives learning-associated behavioral plasticity

Author summaryHabituation is an evolutionarily ancient form of learning in which responses to repeated stimuli decline over time. While seemingly simple, habituation is nevertheless regulated by multiple molecular mechanisms. This surprising complexity led us to ask: do these diverse mechanisms act independently of one another or do they work in the same pathways to drive habituation? To answer this question, we mapped how individual regulators of habituation alter activity throughout the brain and perturbed habituation-regulating molecular mechanisms in combination to see if they act in the same or different pathways. Our experiments grouped eight molecular regulators of habituation into five different pathways with their own brain activity signatures, indicating that multiple independent pathways regulate even simple learning mechanisms. Habituation is a foundational learning process critical for animals to adapt their behavior to changes in their sensory environment. Although habituation is considered a simple form of learning, the identification of a multitude of molecular pathways including several neurotransmitter systems that regulate this process suggests an unexpected level of complexity. How the vertebrate brain integrates these various pathways to accomplish habituation learning, whether they act independently or intersect with one another, and whether they act via divergent or overlapping neural circuits has remained unclear. To address these questions, we combined pharmacogenetic pathway analysis with unbiased whole-brain activity mapping using the larval zebrafish. Based on our findings, we propose five distinct molecular modules for the regulation of habituation learning and identify a set of molecularly defined brain regions associated with four of the five modules. Moreover, we find that in module 1 the palmitoyltransferase Hip14 cooperates with dopamine and NMDA signaling to drive habituation, while in module 3 the adaptor protein complex subunit Ap2s1 drives habituation by antagonizing dopamine signaling, revealing two distinct and opposing roles for dopaminergic neuromodulation in the regulation of behavioral plasticity. Combined, our results define a core set of distinct modules that we propose act in concert to regulate habituation-associated plasticity, and provide compelling evidence that even seemingly simple learning behaviors in a compact vertebrate brain are regulated by a complex and overlapping set of molecular mechanisms.