Meeting Abstract
Zebrafish (Danio rerio) have increasingly become a popular organism in biological research fields, especially in neuroscience, as a paradigmatic vertebrate model. The mature adult zebrafish model can be used to study neural circuits and the physiological processes within the spinal cord. Building upon previous zebrafish research regarding the spinal cord, we developed a novel in vitro primary cell culture for spinal neurons from adult zebrafish. After dissecting and isolating neurons from an intact spinal cord, we cultured robust cells displaying distinct neuronal morphology and behavior with large cell somas, long axons and dendrites, and highly active growth cones. To characterize cells in culture as neurons, immunofluorescence labeling, calcium imaging, and patch clamping were performed. The staining of the cells with NeuN antibody, neuronal nuclear marker, and axonal marker acetyltubulin confirmed neuronal characteristics. Morphologic analysis was supported by calcium imaging which confirmed an excitable cell type. Lastly, analysis from the recordings of whole cell patch clamping revealed strong voltage-dependent currents, both inward and outward, as well as tetrodotoxin-blocked sodium conductance, consistent with a neuronal phenotype. Altogether, these tests successfully and effectively differentiated the spinal neurons from adult zebrafish from other surrounding cells in culture confirming successful isolation. The cultured neurons represent a powerful new system to investigate modulators of endogenous ion channel targets in the zebrafish model system.
