Deciphering the role of serotonin in brain circuit formation

The neurotransmitter serotonin has been implicated in a range of complex neurological disorders linked to alterations of neuronal circuitry. Serotonin is synthesized in the developing brain before most neuronal circuits become fully functional, suggesting that serotonin might play a distinct regulatory role in shaping circuits prior to its function as a classical neurotransmitter. In this study, we asked if serotonin acts as a guidance cue for axons during development by examining how serotonin alters the motility of neuronal growth cones. Using a growth cone motility assay, we found that extracellular gradients of serotonin acted as both an attractive and repulsive guidance cue through a narrow concentration range. Low concentrations of serotonin (50 ¬¨¬µM) elicited attraction, mediated by the serotonin 5-HT2a receptor whilst high concentrations (100 ¬¨¬µM) elicited repulsion mediated by the 5-HT1b receptor. Importantly, pharmacological interventions and high resolution imaging of growth cones suggested that these receptors signalled through their canonical pathways of endoplasmic reticulum-mediated calcium release and cAMP depletion respectively. This novel characterisation of growth cone motility in response to serotonin gradients provides compelling evidence that secreted serotonin acts as an axon guidance cue to shape neuronal circuit formation during development. In vitro optogenetic manipulation of G˜í¬±q signalling associated with the 5HT2a receptor, reversed growth cone attraction to repulsion from serotonin, further demonstrating the role of serotonin as a bidirectional guidance cue. To examine the function of serotonin in an intact animal, we investigated early circuit development in the zebrafish. While previous work has suggested a non-canonical role for serotonin in early development, it remained unclear whether serotonin modulates the guidance of pioneer axons or rather plays a more instructive role in overall circuit function. We hypothesized that optogenetic manipulation of G˜í¬±q signalling is sufficient to modulate neural connectivity in early zebrafish embryos. Using a range of behavioural assays and time-lapse recordings of axon trajectories, we observed a critical developmental window during which altered G˜í¬±q signalling might lead to structural and functional abnormalities. Specifically, inhibition of G˜í¬±q signalling led to an increased coiling behaviour but decreased sensitivity to the touch-startle response in zebarafish larvae. Moreover, inhibition of G˜í¬±q signalling disrupted axon pathfinding in a subset of serotonergic neurons, leading to several misprojecting and straying axons. Unravelling how serotonin functions to regulate the normal wiring of the brain will allow us to better understand how dysfunction in serotonin signalling contributes to important neurodevelopmental disorders.