Generating a comprehensive toolkit for functional studies of associative conditioning circuitry in larval Drosophila

The circuit for associative learning reconstructed in Drosophila larva during the course of this visitor project offers an excellent opportunity to determine the functional roles of distinct microcircuit motifs within a learning center and to determine how a complement of a few hundred first and second-order mushroom body output neurons orchestrate distinct aspects of a conditioned response in distinct contexts. Of particular interest are functional roles of different types of feedback neurons - MBONs that synapse onto DANs and MBONs. We will test whether they might be involved in encoding "prediction error" signals or in encoding information about timing of conditioned stimulus (CS) and unconditioned stimulus (US). We will optogenetically pair the CS and US pathways in varying temporal sequences and with varying timing intervals, and monitor the activation of DAN and MBON in response to US or CS during and after training. We will also manipulate the activity of the feedback neurons during the different pairing protocols and determine the effect of these manipulations on DAN and MBON activation in fictive preparation and on memory formation and retrieval in freely behaving animals.
We will also test whether CS and US pairing can induce plasticity at additional CS and US convergence sites upstream and downstream of the MB. For example, we will monitor the activity of the second-order MBON "convergence" neurons that receive MBON, CS and US inputs, following repeated CS and US presentation while inactivating the input MBON.