ABSTRACT Although behavioral deficits are common in neurological disorders, the genetic pathways and neural circuits underlying behavior are largely unknown. A behavior that is disrupted in numerous disorders including attention deficit disorder, autism spectrum disorders, and schizophrenia is the startle response. Following an intense auditory stimulus, a short latency response occurs wherein rapid muscle activation produces a defensive posture. Disrupted startle responses likely result from a broad defect in genes or circuits controlling behavior. The genetic contribution to behavioral symptoms in humans has been challenging to uncover, in part because they result from multigenic disorders affecting the vastly complex human brain. Genetically tractable animal models have emerged as a valuable system for investigating gene function in the development and function of neural circuits underlying behavior. The Granato lab performed a genetic screen for regulators of the startle response in the larval zebrafish. Molecular cloning of 15 screen candidates identified a number of genes that modulate the startle response in zebrafish and are implicated in human disease. One such gene is solute carrier family 5 member 7a (slc5a7a), which encodes high affinity choline transporter that is required for synthesis of the neurotransmitter acetylcholine. Mutations in slc5a7 have been implicated in psychiatric disorders including attention deficit disorder and major depression. Mice lacking slc5a7 die immediately after birth, precluding behavioral studies. This proposal aims to determine the contribution of slc5a7a to neural circuit development and/or execution of the zebrafish startle response (Aims 1 and 2). In wild type larval zebrafish, the startle response begins with a sharp turn that initiates at the head and progresses towards the tail. In slc5a7a mutants, the turn initiates randomly along the body axis, resulting in an uncoordinated movement. This phenotype suggests neural circuitry between the hindbrain command neuron that initiates the movement and the downstream spinal motoneurons may be disrupted. Circuit defects in slc5a7a mutants will be investigated using in vivo Ca2+ imaging from individual neurons known to be required for the startle response and whole brain activity mapping of additional required neurons. The developmental stage and anatomical region where slc5a7a is required will be investigated using a combination of transgenic rescue constructs and CRISPR/Cas9 generated mutant alleles. Finally, a number of additional mutants that display uncoordinated or weak startle responses will be sequenced to determine the causative genetic lesion and will be characterized using the same techniques (Aim 3). The results of these proposed experiments will determine how slc5a7a promotes a coordinated startle response. More broadly, the results will elucidate acetylcholine’s role in regulating behavior. Finally, characterization of slca5a7a and genes identified in Aim 3 will contribute to models of the genetic program and neural circuitry underlying vertebrate startle behavior.

PROJECT NARRATIVE Behavioral defects are common in a number of neurological and psychiatric disorders, but the genes and neural circuits controlling behavior are largely unknown. This proposal aims to investigate genetic contributions to a simple, conserved behavior, the startle response, which is disrupted in a number of behavioral disorders. By utilizing zebrafish as a model, we aim to uncover how genes direct behavior and how human disease alleles disrupt this behavior.