Development of a macromolecular diffusion pathway in the lens

Abstract

The mammalian lens consists of an aged core of quiescent cells enveloped by a layer of synthetically active cells. Abundant gap junctions within and between these cell populations ensure that the lens functions as an electrical syncytium and facilitates the exchange of small molecules between surface and core cells. In the present study, we utilized an in vivo mouse model to characterize the properties of an additional pathway, permeable to macromolecules, which co-exists with gap-junction-mediated communication in the lens core. The TgN(GFPU)5Nagy strain of mice carries a green fluorescent protein (GFP) transgene. In the lenses of hemizyous animals, GFP was expressed in a variegated fashion, allowing diffusion of GFP to be visualized directly. Early in development, GFP expression in scattered fiber cells resulted in a checkerboard fluorescence pattern in the lens. However, at E15 and later, the centrally located fiber cells became uniformly fluorescent. In the adult lens, a superficial layer of cells, approximately 100 μm thick, retained the original mosaic fluorescence pattern, but the remainder, and majority, of the tissue was uniformly fluorescent. We reasoned that at the border between the two distinct labeling patterns, a macromolecule-permeable intercellular pathway was established. To test this hypothesis, we microinjected 10 kDa fluorescent dextran into individual fiber cells and followed its diffusion by time-lapse microscopy. Injections at depths of >100 μm resulted in intercellular diffusion of dextran from injected cells. By contrast, when injections were made into superficial fiber cells, the injected cell invariably retained the dextran. Together, these data suggest that, in addition to being coupled by gap junctions, cells in the lens core are interconnected by a macromolecule-permeable pathway. At all ages examined, a significant proportion of the nucleated fiber cell population of the lens was located within this region of the lens.