Date of Award

2009

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Biology

First Advisor

Martin-Caraballo, Miguel

Abstract

Ca2+ influx through ionotropic glutamate receptors regulates a variety of developmental processes including neurite outgrowth and naturally occurring cell death. In the CNS, NMDA receptors were originally thought to be the sole source of Ca2+ influx through glutamate receptors; however, AMPA receptors also allow a significant influx of Ca2+ ions. The Ca2+ permeability of AMPA receptors is regulated by the insertion of one or more edited GluR2 subunits into the receptors. Although Ca2+-permeable AMPA receptors are a familiar feature in developing neurons, the developmental function of these receptors during the formation of the nervous system has yet to be established. This study was designed to investigate the expression and functional role of Ca2+-permeable AMPA receptors in developing chicken spinal motoneurons. Our results demonstrate that chicken lumbar motoneurons express functional AMPA receptors as early as embryonic day (E) 5. Electrophysiological recordings of kainate-evoked currents indicate a significant reduction in the Ca2+ permeability of AMPA receptors between E6 and E11. During this developmental period, the Ca2+ permeability of AMPA receptors decreases three-fold. Reduction in the Ca2+ permeability of AMPA receptors is accompanied by increased expression of GluR2 mRNA in the spinal motoneuron pool. Changes in GluR2 mRNA expression occur in parallel to changes in GluR2 protein expression in the chicken ventral spinal cord. Changes in the Ca2+-permeability of AMPA receptors are not mediated by age-dependent changes in the editing pattern of GluR2 subunits. At early stages of development, functional AMPA receptors were composed of a combination of GluR3 and GluR4 subunits. mRNA analysis indicates that GluR4 is the most abundant subunit in the chicken ventral spinal cord between E6 and E11. Immunohistochemistry analysis of spinal cord sections also demonstrated that both GluR3 and GluR4 proteins are expressed at E6 and E11. Expression of Ca2+-permeable AMPA receptors regulates the maturation of dendritic outgrowth in developing spinal motoneurons. Measurements of dendritic length and branching pattern demonstrate significant changes in the dendritic morphology of spinal motoneurons between E6 and E11. Blockade of AMPA receptor activation with CNQX between E5 and E8 causes a significant increase in dendritic outgrowth in lumbar motoneurons, when compared with vehicle-treated embryos. Treatment of chicken embryos with CNQX between E8 and E11, when AMPA receptors become Ca2+-impermeable, has no affect on dendritic morphology. However, blockade of NMDA receptor activation with MK-801 causes a significant reduction in dendritic outgrowth of lumbar motoneurons by E11. These findings indicate that AMPA receptor activation between E5 and E8 limits dendritic outgrowth in developing motoneurons, whereas NMDA receptor activation is involved in dendritic remodeling after the establishment of synaptic contacts with sensory afferents.

Download

Share

COinS