Research Article: 4.1Ba is necessary for glutamatergic synapse formation in the sensorimotor circuit of developing zebrafish

Date Published: October 4, 2018

Publisher: Public Library of Science

Author(s): Javier Fierro, Dylan R. Haynes, Philip Washbourne, Brian Key.


During the process of synapse formation, thousands of proteins assemble at prospective sites of cell-cell communication. Although many of these proteins have been identified, the roles they play in generating functional connections during development remain unknown. 4.1 scaffolding proteins have been implicated in synapse formation and maturation in vitro, but in vivo studies for some family members have suggested these proteins are not important for this role. We examined the role of family member 4.1B because it has been implicated in glutamatergic synaptogenesis, but has not been described in vivo. We identified two 4.1B genes in zebrafish, 4.1Ba and 4.1Bb, by sequence comparisons and synteny analysis. In situ hybridization shows these genes are differentially expressed, with 4.1Ba expressed primarily in the nervous system and 4.1Bb expressed in the nervous system and muscle, but not the spinal cord. We focused our studies on 4.1Ba in the spinal cord. 4.1Ba knockdown reduced the number of glutamatergic synapses at caudal primary motor neurons and caused an increase in the duration of touch-evoked coiling. These results suggest 4.1Ba is important for the formation of functional glutamatergic synapses in the developing zebrafish spinal cord.

Partial Text

The central nervous system (CNS) is composed of trillions of synapses, important cellular structures that together coordinate the multitude of functions of the brain. Each of these cell-cell junctions is assembled from thousands of different synaptic proteins important for establishing, operating, and maintaining these synapses over time [1]. Genetic mutations that disrupt these proteins can change how the macromolecular complexes form and function at the synapse, ultimately leading to neurological disorders such as autism and schizophrenia [2, 3]. To further our understanding of the mechanisms that underlie these disorders, a complete characterization of the synaptic proteins involved in normal brain activity is necessary. In the present study, we investigated the role of protein 4.1B during neuronal development in zebrafish (Danio rerio) embryos.

4.1B is a membrane-bound scaffolding molecule suggested to be involved in synapse formation in vitro [16]. To understand its role during synaptogenesis in vivo, we studied 4.1B in developing zebrafish embryos. We identified two 4.1B gene orthologs in zebrafish, 4.1Ba, expressed in the CNS, and 4.1Bb, expressed in the brain and myotome. We identified 4.1Ba as important for synaptic integrity at PMNs, with trans-synaptic capabilities necessary for organizing the presynaptic specialization. We also found a loss of 4.1Ba altered touch-evoked coiling at 26 hpf after MO knockdown, which correlated with loss of synapses at PMNs. Together, these data confirm 4.1B’s role in glutamatergic synapse formation and establishes its importance in mediating fine control of touch-evoked coiling. These data also highlight the use of zebrafish for studying 4.1 proteins during synaptogenesis.




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