Defects of the Glycinergic Synapse in Zebrafish

Abstract

Glycine mediates fast inhibitory synaptic transmission. Physiological importance of the glycinergic synapse is well established in the brainstem and the spinal cord. In humans, the loss of glycinergic function in the spinal cord and brainstem leads to hyperekplexia, which is characterized by an excess startle reflex to sudden acoustic or tactile stimulation. In addition, glycinergic synapses in this region are also involved in the regulation of respiration and locomotion, and in the nociceptive processing. The importance of the glycinergic synapse is conserved across vertebrate species. A teleost fish, the zebrafish, offers several advantages as a vertebrate model for research of glycinergic synapse. Mutagenesis screens in zebrafish have isolated two motor defective mutants that have pathogenic mutations in glycinergic synaptic transmission: bandoneon (beo) and shocked (sho). Beo mutants have a loss-of-function mutation of glycine receptor (GlyR) β-subunit b, alternatively, sho mutant is a glycinergic transporter 1 (GlyT1) defective mutant. These mutants are useful animal models for understanding of glycinergic synaptic transmission and for identification of novel therapeutic agents for human diseases arising from defect in glycinergic transmission, such as hyperekplexia or glycine encephalopathy. Recent advances in techniques for genome editing and for imaging and manipulating of a molecule or a physiological process make zebrafish more attractive model. In this review, we describe the glycinergic defective zebrafish mutants and the technical advances in both forward and reverse genetic approaches as well as in vivo visualization and manipulation approaches for the study of the glycinergic synapse in zebrafish.

Expansion microscopy of zebrafish for neuroscience and developmental biology studies

“…As another example, the scaffolding protein gephyrin localizes glycine (and GABA A ) receptors to inhibitory synapses ( 96 ). Using ExM in zebrafish, it would be possible to examine the shapes of glycine receptor clusters in mutants where the functions of genes encoding synapse scaffolding components ( 97 , 98 ) or of other proteins involved in the regulation of such clustering are compromised ( 99 – 101 ).…”

The presynaptic glycine transporter GlyT2 is regulated by the Hedgehog pathway in vitro and in vivo

“…Zebrafish express homologs for more than 70% of human genes and their molecular underpinnings controlling neurotransmission are conserved with higher vertebrates 98 . For example, properties of glycinergic biology in zebrafish mimic well the functions observed in mammals, as for example 1) GlyT1 zebrafish mutants present excessive glycinergic transmission due to the lack of glycine recapture 74,99 , 2) loss of glycine receptor function results in an hyperekplexia-like phenotypes 100,101 and 3) GlyT2 can be used as a reliable marker for glycinergic neurons 40,41 . To explore whether Hedgehog control of GlyT2 is conserved in vivo we treated dechorionated zebrafish embryos at 24 hpf with cyclopamine for 24 h, determining at 48 hpf a robust increase of GlyT2 expression measured by both western blot and immunofluorescence of zebrafish spinal cord (Fig.…”

A Great Catch for Investigating Inborn Errors of Metabolism—Insights Obtained from Zebrafish

“…The zebrafish “ bandoneon ” mutant has absent glycinergic neurotransmission and concurrent abnormal tactile response in form of bilateral contractions [ 83 ]. Other mutants such as one termed “ shocked ”, carrying mutations in the glycinergic transporter 1 gene ( slc6a9 ), appear with similar phenotypes [ 84 ]. Detailed analysis of the “ bandoneon ” mutant has shown that all known 7 alleles act with varying sensitivity to glycine [ 85 ].…”