Identification of initially appearing glycine‐immunoreactive neurons in the embryonic zebrafish brain

Abstract

Glycine is a major inhibitory neurotransmitter in the central nervous system of vertebrates. Here, we report the initial development of glycine-immunoreactive (Gly-ir) neurons and fibers in zebrafish. The earliest Gly-ir cells were found in the hindbrain and rostral spinal cord by 20 h post-fertilization (hpf). Gly-ir cells in rhombomeres 5 and 6 that also expressed glycine transporter 2 (glyt2) mRNA were highly stereotyped; they were bilaterally located and their axons ran across the midline and gradually turned caudally, joining the medial longitudinal fascicles in the spinal cord by 24 hpf. Gly-ir neurons in rhombomere 5 were uniquely identified, since there was one per hemisegment, whereas the number of Gly-ir neurons in rhombomere 6 were variable from one to three per hemisegment. Labeling of these neurons by single-cell electroporation and tracing them until the larval stage revealed that they became MiD2cm and MiD3cm, respectively. The retrograde labeling of reticulo-spinal neurons in Tg(glyt2:gfp) larva, which express GFP in Gly-ir cells, and a genetic mosaic analysis with glyt2:gfp DNA construct also supported this notion. Gly-ir cells were also distributed widely in the anterior brain by 27 hpf, whereas glyt2 was hardly expressed. Double staining with anti-glycine and anti-GABA antibodies demonstrated distinct distributions of Gly-ir and GABA-ir cells, as well as the presence of doubly immunoreactive cells in the brain and placodes. These results provide evidence of identifiable glycinergic (Gly-ir/glyt2-positive) neurons in vertebrate embryos, and they can be used in further studies of the neurons' development and function at the single-cell level.

Sensory gating of an embryonic zebrafish interneuron during spontaneous motor behaviors

“…By this same logic, it is unlikely that the glycinergic commissural local (CoLo) interneurons responsible for providing reflex inhibition to Mauthner cells during larval escape behaviors are the source of glycinergic inhibition to CoPAs since their axons span only one somite in length (Satou et al, 2009). A recent study by Moly et al (2014) showed that a small number of glycine immunoreactive neurons are present in the hindbrain and rostral somites of the spinal cord at 20 hpf, coinciding with the arrival of glycinergic inputs to spinal neurons (Saint-Amant and Drapeau, 2000). One class of spinal neurons, the pacemaker IC cells, is located exclusively in the rostral spinal cord (Mendelson, 1986), suggesting that this region may contain specialized neurons not present in more caudal regions.…”

Xylazine regulates the release of glycine and aspartic acid in rat brain

“…In brain tissue, Gly is an important inhibitory neurotransmitter. Gly release by binding of the presynaptic membrane with postsynaptic membrane receptors increases the permeability of the postsynaptic membrane to K + and Cl -, which hyperpolarises the postsynaptic membrane and produces an inhibitory postsynaptic potential, leading to the production of inhibitory action potentials by postsynaptic neurons (17,18). Excess Asp levels can activate its receptors, which can lead to excitatory neurotoxicity in the central nervous system.…”

Large scale analysis of the diversity and complexity of the adult spinal cord neurotransmitter typology

“…The antisera in this study have been widely and successfully used previously in zebrafish to identify and describe the transmitter phenotype of neurons (anti-ChAT: Berg et al, 2018;Clemente et al, 2004;Mueller et al, 2004;Moly et al, 2014;Ohnmacht et al, 2016;Reimer et al, 2008;anti-GABA: Berg et al, 2018;Djenoune et al, 2017;Higashijima et al, 2004;Moly et al, 2014;Montgomery et al, 2016;Mueller et al, 2006;anti-Glycine: Berg et al, 2018;Moly et al, 2014;anti-Serotonin: Berg et al, 2018;Kuscha et al, 2012;McPherson et al, 2016;Montgomery et al, 2016;anti-TH: Ampatzis et al, 2008;Ampatzis and Dermon 2010;Rink and Wullimann, 2004;anti-DβΗ: Ampatzis et al, 2008), and the green fluorescent protein (anti-GFP: Barreiro-Iglesias et al, 2013;Böhm et al, 2016;Kuscha et al, 2012).…”