Normal formation and function of spinal cord circuits requires differentiation of sensory, motor and interneuron subtypes. Our work addresses the role that voltage-gated potassium (Kv) channels play in neuronal subtype differentiation in the embryonic vertebrate spinal cord. We propose that a select subset of Kv channels regulate neuronal differentiation by determining electrical membrane properties that control spontaneous elevations of intracellular calcium, known as calcium (Ca) spikes. Long-duration action potentials occur spontaneously and trigger Ca spikes during a restricted ~6 hr period prior to synapse formation (Stages [St] 20-28). The frequency of Ca spikes determines downstream effects on differentiation programs. We have found that a subset of Kv channels show patterned expression in the embryonic spinal cord and regulate Ca spike properties differently in the dorsal versus ventral embryonic spinal cord. We test the roles of dorsally-expressed Kv channel (Kv1.1) in differentiation of spinal interneurons (Aim 1) and ventrally-expressed Kv channel (Kv2.2) in differentiation of motor neurons (Aim 2).
For Aim 3, we focus on subtypes within the motor neuron population and test whether differences in Kv currents account for subtype-specific Ca spike properties and encoded developmental signals Our studies take advantage of the experimental strengths of the Xenopus and zebrafish embryo models and our experience using (1) antisense (AS), morpholino (MO) and dominant negative (DN) overexpression methods in Xenopus and zebrafish embryos, (2) zebrafish genetic mutants and transgenic lines, (3) electrophysiological recording and Ca imaging from neurons in vivo, and (4) morphological methods to analyze spinal cord development. The results of the proposed experiments will provide new insights into molecular mechanisms that allow early spinal cord neuron activity to direct generation of the diverse neuronal identities required for formation of functional circuits.

Public Health Relevance

The developing nervous system not only generates a staggering number of neurons but also endows them with diverse identities. Neuronal circuit formation requires neurons of diverse identities. Our studies seek to identify novel mechanisms generating neuronal diversity in the embryonic spinal cord.

National Institute of Health (NIH)
National Institute of Neurological Disorders and Stroke (NINDS)
Research Project (R01)
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Special Emphasis Panel (ZRG1-MDCN-J (02))
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Silberberg, Shai D
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University of Colorado Denver
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Moreno, Rosa L; Ribera, Angeles B (2014) Spinal neurons require Islet1 for subtype-specific differentiation of electrical excitability. Neural Dev 9:19
Carlisle, Tara C; Ribera, Angeles B (2014) Connexin 35b expression in the spinal cord of Danio rerio embryos and larvae. J Comp Neurol 522:861-75
McKeown, Kelly Anne; Moreno, Rosa; Hall, Victoria L et al. (2012) Disruption of Eaat2b, a glutamate transporter, results in abnormal motor behaviors in developing zebrafish. Dev Biol 362:162-71
Moreno, Rosa L; Ribera, Angeles B (2009) Zebrafish motor neuron subtypes differ electrically prior to axonal outgrowth. J Neurophysiol 102:2477-84
Pineda, Ricardo H; Ribera, Angeles B (2008) Dorsal-ventral gradient for neuronal plasticity in the embryonic spinal cord. J Neurosci 28:3824-34
Gravagna, Nicole G; Knoeckel, Christopher S; Taylor, Alison D et al. (2008) Localization of Kv2.2 protein in Xenopus laevis embryos and tadpoles. J Comp Neurol 510:508-24
Pineda, Ricardo H; Knoeckel, Christopher S; Taylor, Alison D et al. (2008) Kv1 potassium channel complexes in vivo require Kvbeta2 subunits in dorsal spinal neurons. J Neurophysiol 100:2125-36
Lazaroff, M A; Hofmann, A D; Ribera, A B (1999) Xenopus embryonic spinal neurons express potassium channel Kvbeta subunits. J Neurosci 19:10706-15
Blaine, J T; Ribera, A B (1998) Heteromultimeric potassium channels formed by members of the Kv2 subfamily. J Neurosci 18:9585-93
Ribera, A B (1996) Homogeneous development of electrical excitability via heterogeneous ion channel expression. J Neurosci 16:1123-30

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