Gap junctions modulate interkinetic nuclear movement in retinal progenitor cells.

During early retinal development, progenitor cells must divide repeatedly to expand the progenitor pool. During G(1) and G(2) of the cell cycle, progenitor cell nuclei migrate back-and-forth across the proliferative zone in a process termed interkinetic nuclear movement. Because division can only occur at the ventricular surface, factors that affect the speed of nuclear movement could modulate the duration of the cell cycle. Gap-junctional coupling and gap junction-dependent Ca(2+) activity are common features of proliferating cells in the immature nervous system. Furthermore, both gap-junctional coupling and changes in [Ca(2+)](i) have been shown to be positively correlated with the migration of a number of immature cell types. Using time-lapse confocal microscopy, we describe the nature and rate of progenitor cell interkinetic nuclear movement. We show that nuclear movement is usually, but not always, associated with Ca(2+) transients and that buffering of these transients with BAPTA slows movement. Furthermore, we show for the first time that gap-junctional communication is an important requirement for the maintenance of normal nuclear movement in retinal progenitor cells. Conventional blockers of gap junctions and transfection of cells with dominant-negative constructs of connexin 43 (Cx43) and Cx43-specific antisense oligodeoxynucleotides (asODNs) all act to slow interkinetic nuclear movement. The gap junction mimetic peptide Gap26 also acts to slow movement, an effect that we show may be attributable to the blockade of gap junction hemichannels.

Ca(2+) signalling and gap junction coupling within and between pigment epithelium and neural retina in the developing chick.

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.