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Scn1bb, a zebrafish ortholog of SCN1B expressed in excitable and nonexcitable cells, affects motor neuron axon morphology and touch sensitivity.

ABSTRACT: Voltage-gated Na(+) channels initiate and propagate action potentials in excitable cells. Mammalian Na(+) channels are composed of one pore-forming alpha-subunit and two beta-subunits. SCN1B encodes the Na(+) channel beta1-subunit that modulates channel gating and voltage dependence, regulates channel cell surface expression, and functions as a cell adhesion molecule (CAM). We recently identified scn1ba, a zebrafish ortholog of SCN1B. Here we report that zebrafish express a second beta1-like paralog, scn1bb. In contrast to the restricted expression of scn1ba mRNA in excitable cells, we detected scn1bb transcripts and protein in several ectodermal derivatives including neurons, glia, the lateral line, peripheral sensory structures, and tissues derived from other germ layers such as the pronephros. As expected for beta1-subunits, elimination of Scn1bb protein in vivo by morpholino knock-down reduced Na(+) current amplitudes in Rohon-Beard neurons of zebrafish embryos, consistent with effects observed in heterologous systems. Further, after Scn1bb knock-down, zebrafish embryos displayed defects in Rohon-Beard mediated touch sensitivity, demonstrating the significance of Scn1bb modulation of Na(+) current to organismal behavior. In addition to effects associated with Na(+) current modulation, Scn1bb knockdown produced phenotypes consistent with CAM functions. In particular, morpholino knock-down led to abnormal development of ventrally projecting spinal neuron axons, defasciculation of the olfactory nerve, and increased hair cell number in the inner ear. We propose that, in addition to modulation of electrical excitability, Scn1bb plays critical developmental roles by functioning as a CAM in the zebrafish embryonic nervous system.

SUBMITTER: Fein AJ

PROVIDER: S-EPMC2741146 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundVoltage-gated Na+ channel beta1 (Scn1b) subunits are multi-functional proteins that play roles in current modulation, channel cell surface expression, cell adhesion, cell migration, and neurite outgrowth. We have shown previously that beta1 modulates electrical excitability in vivo using a mouse model. Scn1b null mice exhibit spontaneous seizures and ataxia, slowed action potential conduction, decreased numbers of nodes of Ranvier in myelinated axons, alterations in nodal architecture, and differences in Na+ channel alpha subunit localization. The early death of these mice at postnatal day 19, however, make them a challenging model system to study. As a first step toward development of an alternative model to investigate the physiological roles of beta1 subunits in vivo we cloned two beta1-like subunit cDNAs from D. rerio.ResultsTwo beta1-like subunit mRNAs from zebrafish, scn1ba_tv1 and scn1ba_tv2, arise from alternative splicing of scn1ba. The deduced amino acid sequences of Scn1ba_tv1 and Scn1ba_tv2 are identical except for their C-terminal domains. The C-terminus of Scn1ba_tv1 contains a tyrosine residue similar to that found to be critical for ankyrin association and Na+ channel modulation in mammalian beta1. In contrast, Scn1ba_tv2 contains a unique, species-specific C-terminal domain that does not contain a tyrosine. Immunohistochemical analysis shows that, while the expression patterns of Scn1ba_tv1 and Scn1ba_tv2 overlap in some areas of the brain, retina, spinal cord, and skeletal muscle, only Scn1ba_tv1 is expressed in optic nerve where its staining pattern suggests nodal expression. Both scn1ba splice forms modulate Na+ currents expressed by zebrafish scn8aa, resulting in shifts in channel gating mode, increased current amplitude, negative shifts in the voltage dependence of current activation and inactivation, and increases in the rate of recovery from inactivation, similar to the function of mammalian beta1 subunits. In contrast to mammalian beta1, however, neither zebrafish subunit produces a complete shift to the fast gating mode and neither subunit produces complete channel inactivation or recovery from inactivation.ConclusionThese data add to our understanding of structure-function relationships in Na+ channel beta1 subunits and establish zebrafish as an ideal system in which to determine the contribution of scn1ba to electrical excitability in vivo.

Project description:BACKGROUND:Tick-borne encephalitis virus (TBEV) is an important human pathogen that can cause the serious illness tick-borne encephalitis (TBE). Patients with clinical symptoms can suffer from severe meningoencephalitis with sequelae that include cognitive disorders and paralysis. While less than 30% of patients with clinical symptoms develop meningoencephalitis, the number of seropositive individuals in some regions indicates a much higher prevalence of TBEV infections, either with no or subclinical symptoms. The functional relevance of these subclinical TBEV infections and their influence on brain functions, such as learning and memory, has not been investigated so far. METHODS:To compare the effect of low and high viral replication in the brain, wildtype and Irf-7-/- mice were infected with Langat virus (LGTV), which belongs to the TBEV-serogroup. The viral burden was analyzed in the olfactory bulb and the hippocampus. Open field, elevated plus maze, and Morris water maze experiments were performed to determine the impact on anxiety-like behavior, learning, and memory formation. Spine density of hippocampal neurons and activation of microglia and astrocytes were analyzed. RESULTS:In contrast to susceptible Irf-7-/- mice, wildtype mice showed no disease signs upon LGTV infection. Detection of viral RNA in the olfactory bulb revealed CNS infections in wildtype and Irf-7-/- mice. Very low levels of viral replication were detectable in the hippocampus of wildtype mice. Although wildtype mice develop no disease signs, they showed reduced anxiety-like behavior and impaired memory formation, whereas Irf-7-/- mice were not affected. This impairment was associated with a significant decrease in spine density of neurons in the hippocampal CA1 region of wildtype mice. Microglia activation and astrogliosis were detected in the hippocampus. CONCLUSION:In this study, we demonstrate that subclinical infections by viruses from the TBEV-serogroup affected anxiety-like behavior. Virus replication in the olfactory bulb induced far-reaching effects on hippocampal neuron morphology and impaired hippocampus-dependent learning and memory formation.

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Scn1bb, a zebrafish ortholog of SCN1B expressed in excitable and nonexcitable cells, affects motor neuron axon morphology and touch sensitivity.

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