ABSTRACT
Although hyperactivity is associated with a wide variety of neurodevelopmental disorders, the early embryonic origins of locomotion have hindered investigation of pathogenesis of these debilitating behaviors. The earliest motor output in vertebrate animals is generated by clusters of early-born motor neurons that occupy distinct regions of the spinal cord, innervating stereotyped muscle groups. Gap junction electrical synapses drive early spontaneous behavior in zebrafish, prior to the emergence of chemical neurotransmitter networks. We use a genetic model of hyperactivity to gain critical insight into the consequences of errors in motor circuit formation and function, finding that Fragile X syndrome (FXS) model mutant zebrafish are hyperexcitable from the earliest phases of spontaneous behavior, show altered sensitivity to blockade of electrical gap junctions, and have increased expression of the gap junction protein Connexin 34/35. We further show that this hyperexcitable behavior can be rescued by pharmacological inhibition of electrical synapses. We also use functional imaging to examine motor neuron and interneuron activity in early embryogenesis, finding genetic disruption of electrical gap junctions uncouples activity between mnx1 + motor neurons and interneurons. Taken together, our work highlights the importance of electrical synapses in motor development and suggests that the origins of hyperactivity in neurodevelopmental disorders may be established during the initial formation of locomotive circuits.Significance Statement The origins of hyperactivity in neurodevelopmental disorders are difficult to pinpoint in vertebrate systems. Zebrafish locomotive circuits initiate in early embryogenesis, with defined motor neurons and interneurons driving the earliest locomotive movements. Using a genetic model of hyperactivity, we show that Fragile X syndrome model fmr1 mutant embryos display hyperexcitable behavior and express excess gap junction connexin proteins on motor circuit neurons. We further show that this hyperexcitable behavior can be rescued by pharmacological inhibition of electrical synapses. Taken together, this data suggests hyperactive behavior initiates in the earliest phases of neurodevelopment.
ABSTRACT
Motor behavior requires the balanced production and integration of a variety of neural cell types. Motor neurons are positioned in discrete locations in the spinal cord, targeting specific muscles to drive locomotive contractions. Specialized spinal interneurons modulate and synchronize motor neuron activity to achieve coordinated motor output. Changes in the ratios and connectivity of spinal interneurons could drastically alter motor output by tipping the balance of inhibition and excitation onto target motor neurons. Importantly, individuals with Fragile X syndrome (FXS) and associated autism spectrum disorders often have significant motor challenges, including repetitive behaviors and epilepsy. FXS stems from the transcriptional silencing of the gene Fragile X Messenger Ribonucleoprotein 1 (FMR1), which encodes an RNA binding protein that is implicated in a multitude of crucial neurodevelopmental processes, including cell specification. Our work shows that Fmrp regulates the formation of specific interneurons and motor neurons that comprise early embryonic motor circuits. We find that zebrafish fmr1 mutants generate surplus ventral lateral descending (VeLD) interneurons, an early-born cell derived from the motor neuron progenitor domain (pMN). As VeLD interneurons are hypothesized to act as central pattern generators driving the earliest spontaneous movements, this imbalance could influence the formation and long-term function of motor circuits driving locomotion. fmr1 embryos also show reduced expression of proteins associated with inhibitory synapses, including the presynaptic transporter vGAT and the postsynaptic scaffold Gephyrin. Taken together, we show changes in embryonic motor circuit formation in fmr1 mutants that could underlie persistent hyperexcitability.
