Professor
Department of Physiology and Biophysics
UCD at Fitzsimons
RC-1 North Tower, P18-7117. PO Box 6511, Mail Stop F8307. Tel (303) 724-4517. Fax (303) 724-4501.
Email angie.ribera@UCHSC.edu

Angie's Curriculum vitae

SUMMARY

Our laboratory is interested in determining the mechanisms that direct differentiation of electrical excitability in neurons, and, in turn, how activity regulates neuronal development. Our studies span the period from when neurons exit the cell cycle and begin terminal differentiation and to when synaptic interactions emerge. In order to have access to the relevant early stages of development, we use two classic vertebrate embryological systems - the frog, Xenopus laevis [In the movie above, the embryo develops from a 1-cell animal to a 2-day larva.], and zebrafish, Danio rerio. Studies using the Xenopus system concern stages of development prior to synapse formation. In contrast, studies using the zebrafish model focus on stages when synaptic interactions occur and behaviors are evident at the level of the organism.

Developmental regulation and function of potassium current in developing spinal neurons (Xenopus laevis).

Potassium currents play a major role in determining the emerging properties of excitability in amphibian embryonic spinal neurons. The overall goal of research in this area is to identify molecular mechanisms that regulate potassium channel function and its contribution to electrical excitability during initial stages of neuronal differentiation. Embryonic amphibian spinal neurons acquire electrical excitability a few hours after they exit the cell cycle, prior to neurite extension. At this time, impulses are of long duration and provide transient elevations of intracellular calcium that trigger developmental signaling cascades. During the next 24 hours, voltage-dependent potassium current (IKv) density gradually triples, leading to decreases in the duration of the action potential and the associated calcium influx. By the time synapses begin to form, impulse durations have undergone profound developmental regulation and are now brief, as is characteristic of adult neurons. During the next day, no further increase in IKv density occurs, indicating establishment of a set point for this current.

We are identifying potassium channel genes that are essential for proper development of excitability. Using molecular, physiological and embryological methods, we manipulate functional expression of these potassium channel genes and uncover underlying mechanisms. Our data so far suggest that transcriptional mechanisms contribute to the rapid initial increase in potassium current density. However, post-translational and post-transcriptional mechanisms determine the set points for potassium current density in mature neurons. A combination of physiological, molecular, histochemical and embryological methods are used for these studies.

[The picture depicts in situ hybridization for the Xenopus Kv1.1a gene in the developing spinal cord. A transverse section through the spinal cord of a 2-day embryo is shown and the blue-purple signal represent the hybridization signal hat is found specifically in dorsal spinal sensory neurons known as Rohon-Beard cells.]

• Ribera AB (1990) A potassium channel gene is expressed at neural induction. Neuron 5:691-701.
  
• Ribera AB and Spitzer NC (1991) The differentiation of potassium current in embryonic amphibian myocytes. Dev. Biol. 144:119-128.
  
• Ribera AB and Nguyen DA (1993) Primary sensory neurons express a Shaker like potassium channel gene. J. Neurosci. 13:4988-4996.
  
• Jones SM and Ribera AB (1994) Overexpression of a potassium channel gene perturbs neural differentiation. J. Neurosci. 14:2789-2799.
  
• Jones SM, Hofmann AD, Lieber JL and Ribera AB (1994) Overexpression of potassium channel RNA: In vivo development rescues neurons from suppression of morphological differentiation in vitro. J. Neurosci. 15: 2867-2874.
  
• Ribera AB (1996) Homogeneous development of electrical excitability via heterogeneous ion channel expression. J. Neurosci. 16:1123-1130.
  
• Burger C and Ribera AB (1996) Xenopus spinal neurons express Kv2 potassium channel transcripts during embryonic development. J. Neurosci. 16:1412-1421.
  
• Gurantz D, Ribera AB and Spitzer NC (1996) Temporal expression of Shaker- and Shab-like potassium channel gene expression in single embryonic spinal neurons during K+ current development. J. Neurosci. 16:3287-3295.
  
• Ribera AB, Pacioretty LM and Taylor RS (1996) Probing identity of native channels with dominant negative mutants. Neuropharm. 35:1007-1016.
  
• Blaine, J. T. and Ribera, A.B. (1998) Heteromultimeric potassium channels formed by members of the Kv2 subfamily. J. Neurosci. 18:9585-9593.
  
• Lazaroff, M.A., Hofmann, A.D. and Ribera, A.B. (1999) Xenopus embryonic spinal neurons express potassium channel Kvb-subunits. J. Neurosci. 19:10706-10715.
  
• Nick, T.A. and Ribera, A.B. (2000) Synaptic activity modulates presynaptic excitablity. Nature Neurosci. 3:142-149.
  
• Blaine, J.T. and Ribera, A. B. (2001) Kv2 channels form delayed-rectifier potassium channels in situ. J. Neurosci. 21:1473-1480.
  
  
  Electrical excitability in wild type and mutant zebrafish (Danio rerio).
  
  Forward genetic strategies uncover genes with irreplaceable functions. Because ion channel activity is developmentally-regulated and essential for generation of organismal behavior, zebrafish motility mutants in which specific behaviors fail to appear may exhibit abnormal developmental expression/function of ion channels. For example, one group of mutants does not respond to touch, although these embryos are motile and can swim (Granato et al., 1996). The specific behavioral phenotype suggests that the defect may originate in mechanosensory Rohon-Beard (RB) neurons. In wildtype embryos, the action potential of RB cells undergoes developmental regulation while the embryo acquires touch sensitivity. The changes in action potential waveform are due to underlying changes in voltage-gated sodium (INa) and potassium currents (Ribera and Nüsslein-Volhard, 1998). The INa of RB cells of the mao touch-insensitive mutant have reduced amplitudes and action potentials are not generated. Further, although the normal developmental changes in potassium current occur, upregulation of INa is absent. These data suggest that developmental regulation of RB INa may underlie stage-specific acquisition of touch sensitivity. We will identify functional, pharmacological and molecular properties of RB INa that are developmentally regulated. In addition, our preliminary data indicate that RB cells of mao mutants exhibit abnormal morphology. Accordingly, we will quantify differences in RB morphology of touch-insensitive and touch-sensitive fish to assess possible contributions of changes in peripheral innervation and central projections to normal developmental acquisition of touch sensitivity. We have recently found that activity of RB cells regulates their programmed cell death. A combination of genetic, molecular, anatomical and physiological methods will be used in these studies.
  
  In related work, we have collaborated with Dr. Winfried Denk (MPI Heidelberg) to examine the appearance of spontaneous calcium transients in the developing zebrafish embryo (/physiology/abr2/movie.htm). Future work will test the possibility that these spontaneous evens influence subsequent development of spinal neurons.
  
  Over the long-term, the studies will provide a framework for analysis of other behavioral mutants and identification of ion channels with essential functions during embryonic development of the nervous system and emergence of stage-specific behaviors.
  
  Representative Publications:
  
  
• Ribera, A.B. and Nüsslein-Volhard, C. (1998) Zebrafish touch-insensitive mutants reveal an essential role for developmental regulation of sodium current. J. Neurosci. 18:9181-9191.
  
  
• Svoboda, K.R., Linares, A.E., and Ribera, A.B. (2001) Activity regulates programmed cell death of zebrafish Rohon-Beard neurons. Development 128(18):3511-3520.
  
  REVIEWS:
  
  
• Ribera, A.B. and Blaine, J.T. (1998) Construction, Characterization and applications of potassium channel dominant negative subunits. Methods in Mol. Biol. in press.
  
  
• Ribera, A.B. (1998) Potassium currents in developing neurons. NYAS 868:399-405.
  
  
• Spitzer, N.C. and Ribera, A.B. (1998) Development of electrical excitability in embryonic neurons: mechanisms and roles. J. Neurobiol 37:190-197.
  
  
• Ribera, A.B. (1998) Perspective: Ion channel activity drives ion channel expression. J. Physiol. 511.3:645.
  

                                            

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