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Cut-loading: A Useful Tool for Examining the Extent of Gap Junction Tracer Coupling Between Retinal Neurons
Published on: January 12, 2012
Ca(2+) signalling and gap junction coupling within and between pigment epithelium and neural retina in the developing
Rachael A Pearson1, Marina Catsicas, David L Becker
1Department of Physiology, University College London, Gower Street, London, WC1E 6BT.
Insights
Calcium signaling and gap junction coupling between retinal pigment epithelium (RPE) and neural retina progenitor cells influence retinal development. These cell connections may orchestrate proliferation and differentiation.
Area of Science:
- Developmental biology
- Neuroscience
- Cell biology
Background:
- Neural retina development is regulated by the adjacent retinal pigment epithelium (RPE).
- Understanding intercellular communication is crucial for elucidating retinal development mechanisms.
Purpose of the Study:
- Investigate calcium (Ca2+) signaling and gap junctional coupling within and between the RPE and neural retina.
- Determine the role of these interactions in embryonic chick retinal development at embryonic day 5 (E5).
Main Methods:
- Utilized calcium imaging to detect spontaneous Ca2+ transients and waves.
- Employed gap junction blockers and Neurobiotin tracer to assess coupling.
- Performed immunolabeling for connexin 43 (Cx43), beta-tubulin, and vimentin.
Main Results:
- RPE and neural retina ventricular zone (VZ) exhibited spontaneous Ca2+ transients and waves.
- Gap junction blockers reduced Ca2+ wave propagation but not individual cell transients.
- Neurobiotin tracer revealed extensive cell coupling within the neural retina and between RPE and neural retina.
- Cx43 was present at the RPE-VZ junction, suggesting a role in inter-tissue coupling.
- Coupled neural retina cells primarily consisted of progenitor cells.
Conclusions:
- Gap junctions facilitate communication between RPE and neural retina progenitor cells.
- This intercellular coupling may orchestrate retinal proliferation and differentiation through the propagation of Ca2+ or other signaling molecules.
Abstract:
Development of the neural retina is controlled in part by the adjacent retinal pigment epithelium (RPE). To understand better the mechanisms involved, we investigated calcium signalling and gap junctional coupling within and between the RPE and the neural retina in embryonic day (E) 5 chick. We show that the RPE and the ventricular zone (VZ) of the neural retina display spontaneous Ca(2+) transients. In the RPE, these often spread as waves between neighbouring cells. In the VZ, the frequency of both Ca(2+) transients and waves was lower than in RPE, but increased two-fold in its presence. Ca(2+) signals occasionally crossed the boundary between the RPE and VZ in either direction. In both tissues, the frequency of propagating Ca(2+) waves, but not of individual cell transients, was reduced by gap junction blockers. Use of the gap junction permeant tracer Neurobiotin showed that neural retina cells are coupled into clusters that span the thickness of the retina, and that RPE cells are both coupled together and to clusters of cells in the neural retina. Immunolabelling for Cx43 showed this gap junction protein is present at the junction between the RPE and VZ and thus could potentially mediate the coupling of the two tissues. Immunolabelling for beta-tubulin and vimentin showed that clusters of coupled cells in the neural retina comprised mainly progenitor cells. We conclude that gap junctions between progenitor cells, and between these cells and the RPE, may orchestrate retinal proliferation/differentiation, via the propagation of Ca(2+) or other signalling molecules.
