Left/right asymmetry disruptions and mirror-image reversals to behavior and brain anatomy in Ciona

Background Left-right asymmetries are a common feature of metazoan nervous systems. This is particularly pronounced in the comparatively simple larval central nervous system (CNS) of the tunicate Ciona , whose swimming tadpole larva shows a clear chordate ground plan. While common pathway elements for specifying the left-right axis are found in the chordates, particularly a requirement for Nodal signaling, Ciona differs from its vertebrate cousins by specifying its axis at the neurula stage, rather than at gastrula. Additionally, Ciona , and other ascidians, have a requirement for an intact chorionic membrane for proper left/right specification. Results We present here results showing that left-right asymmetry disruptions caused by removal of the chorion (dechorionation) are highly variable and present throughout the Ciona larval nervous system. While previous studies have documented disruptions to the conspicuously asymmetric sensory systems in the anterior brain vesicle, we document asymmetries in seemingly symmetric structures such as the posterior brain vesicle and motor ganglion. Moreover, defects caused by dechorionation include misplaced or absent neuron classes, loss of asymmetric gene expression, aberrant synaptic connectivity, and abnormal behaviors. In the motor ganglion, a brain structure that has been equated with the vertebrate hindbrain, we find that despite the apparent left/right symmetric distribution of interneurons and motor neurons, AMPA receptors are expressed exclusively on the left side, which equates with asymmetric swimming behaviors. We also find that within a population of dechorionated larvae, there is a small percentage with apparently normal left-right specification, and approximately equal population with inverted (mirror-image) asymmetry. We present a method based on a behavioral assay for isolating these larvae. When these two classes of larvae (normal and inverted) are assessed in a light dimming assay they display mirror-image behaviors, with normal larvae responding with counterclockwise swims, while inverted larvae respond with clockwise swims. Conclusions Our findings highlight the importance of left-right specification pathways not only for proper CNS anatomy, but also for correct synaptic connectivity and behavior. ### Competing Interest Statement The authors have declared no competing interest.