Julia F. Burne

Programmed cell death and Bcl-2 protection in the absence of a nucleus.

The molecular basis of programmed cell death (PCD) is unknown. An important clue is provided by the Bcl‐2 protein, which can protect many cell types from PCD, although it is not known where or how it acts. Nuclear condensation, DNA fragmentation and a requirement for new RNA and protein synthesis are often considered hallmarks of PCD. We show here, however, that anucleate cytoplasts can undergo PCD and that Bcl‐2 and extracellular survival signals can protect them, indicating that, in some cases at least, the nucleus is not required for PCD or for Bcl‐2 or survival factor protection. We propose that PCD, like the cell cycle, is orchestrated by a cytoplasmic regulator that has multiple intracellular targets.

The role of laminin and the laminin/fibronectin receptor complex in the outgrowth of retinal ganglion cell axons

Chick embryo retinal ganglion cell (RGC) axons grow to the optic tectum along a stereotyped route, as if responding to cues distributed along the pathway. We showed previously that, in culture, RGCs from embryonic Day 6 retina are responsive to the neurite-promoting effects of the extracellular matrix glycoprotein laminin and that this response is lost by RGCs at a later stage of development. Here we report that, before axon outgrowth is initiated in vivo, laminin, is expressed along the optic pathway at nonbasal lamina sites that are accessible to the growth cones of RGC axons. The distribution of laminin within the pathway is consistent with its localization at the end-feet of neuroepithelial cells that line the route, and it continues to be expressed at these marginal sites during the first week of embryonic development. At later stages, concomitant with the loss of response by RGCs in culture, laminin becomes restricted to basal laminae at the retinal inner limiting membrane and pial surface of the optic pathway. Neurofilament-positive RGC axons bind a monoclonal antibody, JG22, which recognizes the laminin/fibronectin receptor complex, and continue to do so throughout embryonic development. We show that, in vitro, the JG22 antigen expressed by RGCs appears to function as a laminin receptor, by demonstrating that JG22 antibody blocks neurite outgrowth on a substrate of laminin. These findings are consistent with the possibility that laminin defines a transient performed pathway specifically recognized by early RGC growth cones as they navigate toward their central target.

Evidence that migratory oligodendrocyte-type-2 astrocyte (O-2A) progenitor cells are kept out of the rat retina by a barrier at the eye-end of the optic nerve.

SummaryThere is evidence that oligodendrocyte-type-2 astrocyte (O-2A) progenitor cells migrate along the developing rat optic nerve from the chiasm toward the eye before differentiating into oligodendrocytes that myelinate the retinal ganglion cell axons in the nerve. Why, then, do these progenitor cells not migrate into the eye, differentiate into oligodendrocytes and myelinate the nerve fibre layer of the retina? Myelination would opacify the neural retina and thereby severely impair vision. Here we provide evidence that there is a barrier at the eye-end of the rat optic nerve that prevents the migration of O-2A progenitor cells into the retina. Our findings in the rat support a previous hypothesis that such a barrier keeps myelin-forming glial cells out of the human retina.

Evidence for large-scale astrocyte death in the developing cerebellum

There is increasing evidence that some glial cells die during normal vertebrate development, but the extent of the death and the types of glial cells that die remain uncertain. We have analyzed pyknotic cells in the developing postnatal rat cerebellum. During the first postnatal week, the majority of pyknotic cells are in the developing white matter where their number peaks at about postnatal day 7 (P7) and then declines sharply. Pyknotic cells in the internal granule cell layer peak at P10, while those in the molecular and external granule cell layers peak later. Both electron microscopy and in situ end labeling of DNA catalyzed by terminal deoxynucleotidyl transferase confirm that the pyknotic cells are undergoing apoptosis. Immunohistochemical staining suggests that 50–70% of the pyknotic cells in the white matter and internal granule cell layer are astrocytes. We estimate that at P7, as many as 50% of the white matter cells die and, of these, more than half appear to be astrocytes.

Quantification of Normal Cell Death in the Rat Retina: Implications for Clone Composition in Cell Lineage Analysis

Naturally occurring cell death complicates the analysis of cell lineage studies by making the surviving members of a clone appear more closely related than they actually are. Here we ask how much normal cell death occurs during rat retinal development, and whether that amount of death is sufficient to confuse the analysis of cell lineage relationships. We measure total cell death in the retina by combining relative counts of dead cells with absolute measurements of total cell loss. For most cell types, but not rods, we find that half of the cells generated die during normal retinal development. We use a computer model to quantify the effects of different amounts of cell death in a simulated lineage study. The simulation indicates that 50% cell death means that clonal variability analysed after the cell death period is not necessarily a good indicator of how much variability actually occurs in the underlying lineage.

Retinal Ganglion Cell Axons Drive the Proliferation of Astrocytes in the Developing Rodent Optic Nerve

We show that the proliferation of astrocytes in the developing rodent optic nerve absolutely depends on axons and that this axonal influence depends on axonal transport but not on axonal electrical activity. We also show that purified retinal ganglion cells stimulate DNA synthesis in optic nerve astrocytes in culture and that the effect can be mimicked by fibroblast growth factor but not by neuregulins or several other growth factors. Taken together with previous findings, our present results indicate that axons promote glial cell proliferation and survival in the developing optic nerve by at least three distinct mechanisms.