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Gap junctional communication in ectodermally derived tissues of the developing rat

Potter, Karina Nicole; (1996) Gap junctional communication in ectodermally derived tissues of the developing rat. Doctoral thesis (Ph.D), UCL (University College London). Green open access

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Abstract

The aim of my PhD project was to examine the developmental expression and functional significance of gap junctional communication in ectodermally derived embryonic tissues (nervous system and epidermis) of the rat, with particular emphasis on elucidating the role of gap junctions and putative mechanisms of gap junctional regulation in the development of dorsal root ganglion (DRG) neurons. In this study, immunohistochemistry in combination with the technique of the laser scanning confocal microscopy (LSCM) with its PC-IMAGE image analysis programme was used to determine the developmental expression of gap junction proteins (connexins). Antisera specific to certain peptide sequences of the three major connexin (Cx) types (Cx26, Cx32 and Cx43) were used. Gap junction-mediated intercellular communication was assessed by observing the transfer of the low molecular weight fluorescent dye, Lucifer Yellow CH (LY), from an intracellularly microinjected cell to neighbouring cells under epifiuorescence optics. In thick tissue, the extent of dye transfer was visualized subsequently on the LSCM. These immunohistochemical and dye-injection techniques were used to study connexin expression and functional gap junctional communication in developing non-neural ectoderm (predominantly flank epidermis) and in neural derivatives of ectoderm (predominantly DRG neurons). A developmental study of flank epidermis revealed a temporal increase in Cx26 and Cx43 from embryonic days 10 to 14 (E10-E14). LY dye-coupling assays carried out in parallel were consistent with the immunohistochemical analysis. The observed increases in connexin expression and functional gap junctional communication coincide with the time period in which the epidermis is transformed from a simple unilayer of ectoderm into an epidermal and peridermal bilayer. DRG neurons had previously been shown to be dye-coupled in small groups from E13-E15, but during subsequent embryonic development, both the number of cells that transfer dye and the mean number of cells in a coupled group decreased steadily. Immunohistochemical studies of these developing neurons established that this decrease in dye-coupling was accompanied by a decrease in expression of Cx26 and Cx43. I also carried out in vitro experiments using explant cultures of DRGs and transverse slices of embryos containing intact DRGs, to address questions about the mechanisms controlling developmental regulation of gap junction expression. These results suggest that nerve growth factor and possibly other target derived factors may play a role in gap junctional protein regulation in developing DRG neurons.

Type: Thesis (Doctoral)
Qualification: Ph.D
Title: Gap junctional communication in ectodermally derived tissues of the developing rat
Open access status: An open access version is available from UCL Discovery
Language: English
Additional information: Thesis digitised by ProQuest.
URI: https://discovery.ucl.ac.uk/id/eprint/10101114
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