ZFIN ID: ZDB-PUB-050201-2
Developmental, molecular and genetic dissection of INa in vivo in embryonic zebrafish sensory neurons
Pineda, R.H., Heiser, R.A., and Ribera, A.B.
Date: 2005
Source: Journal of neurophysiology   93(6): 3582-3593 (Journal)
Registered Authors: Ribera, Angie
Keywords: voltage-dependent sodium current, zebrafish embryo, antisense, Rohon-Beard neuron, nucleated-patch, Nav1
MeSH Terms:
  • Age Factors
  • Analysis of Variance
  • Animals
  • Cell Nucleus/drug effects
  • Cell Nucleus/physiology
  • Embryo, Nonmammalian
  • Gene Expression/physiology
  • Gene Expression Regulation, Developmental/drug effects
  • Gene Expression Regulation, Developmental/physiology*
  • In Vitro Techniques
  • Ion Channel Gating/drug effects
  • Ion Channel Gating/physiology
  • Ion Channel Gating/radiation effects
  • Membrane Potentials/drug effects
  • Membrane Potentials/physiology
  • Membrane Potentials/radiation effects
  • Microinjections
  • Molecular Biology/methods
  • Mutation/physiology
  • Neurons, Afferent/cytology
  • Neurons, Afferent/drug effects
  • Neurons, Afferent/physiology*
  • Oligodeoxyribonucleotides, Antisense/pharmacology
  • Patch-Clamp Techniques/methods
  • Reaction Time/physiology
  • Sodium Channel Blockers/pharmacology
  • Sodium Channels/genetics*
  • Sodium Channels/metabolism
  • Spinal Cord/cytology*
  • Tetrodotoxin/pharmacology
  • Zebrafish/genetics*
  • Zebrafish/growth & development
PubMed: 15673553 Full text @ J. Neurophysiol.
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ABSTRACT
The presence of multiple Nav1 isotypes within a neuron and the lack of specific blockers hamper identification of the in vivo roles of sodium current (INa) components, especially during embryonic stages. To identify the functional properties of INa components in vivo in developing neurons, we took a molecular genetic approach. Embryonic zebrafish Rohon-Beard (RB) mechanosensory neurons express two different sodium channel isotypes: Nav1.1 and Nav1.6. To examine the properties of Nav1.1 and Nav1.6 encoded currents in RB cells at different developmental stages, we eliminated the contribution of Nav1.6 and Nav1.1 channels, respectively, using an antisense morpholino (MO) approach. MOs were injected into 1-cell stage embryos, and RB sodium currents were recorded using patch-clamp techniques in both conventional whole-cell mode as well from nucleated-patches. Only a subset of RB cells appeared to be affected by the Nav1.1MO. Overall, the effect of the Nav1.1MO was a small 25% average reduction in current amplitude. Further, Nav1.1MO effects were most pronounced in RB cells of younger embryos. In contrast, the effects of the Nav1.6 MO were observed in all cells and increased as development proceeded. These results indicated that developmental upregulation of RB INa entailed an increase in the number of functional Nav1.6 channels. In addition, analysis of voltage-dependent steady-state activation and inactivation parameters revealed that specific functional properties of channels were also developmentally regulated. Finally, analysis of macho mutants indicated that developmental upregulation of INa was absent in RB cells. These results indicate that MOs are a useful tool for the molecular dissection and analysis of ion channel function in vivo.
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