Subcellular localization and axonal transport of the survival motor neuron (SMN) protein in the developing rat spinal cord.

The subcellular localization of the survival motor neuron (SMN) protein, encoded by the spinal muscular atrophy determining gene, was investigated in motor neurons of the developing and adult rat spinal cord by light and electron microscopy immunocytochemistry. The experiments were carried out with a panel of anti-SMN antibodies, all recognizing an SMN-specific protein band at 39 kDa in HeLa cells and rat spinal cord protein extracts. SMN protein expression decreased during postnatal spinal cord development, but it remained unchanged in distribution and intensity in motor neurons at all ages examined. SMN protein was mainly organized in immunoreactive aggregates sparse in the nucleoplasm and cytoplasm of both mature and developing motor neurons, and it was more rarely localized within the endoplasmic reticulum and in apposition to the external mitochondrial membrane. Most strikingly, the SMN protein was found in association with cytoskeletal elements in spinal dendrites and axons, where it was particularly evident during postnatal development. The present findings suggest that SMN protein may be transported via axoplasmic flow in maturing neurons. Given the RNA-binding activity of SMN, the SMN protein could be involved in the transport of specific mRNAs in axons and dendrites of motor neurons. The reduced transport of specific mRNAs within motor neurons during development could play a role in the motoneuronal degeneration and impaired axonal sprouting observed in spinal muscular atrophy.

A short term analysis of the behaviour of conditionally immortalized neuronal progenitors and primary neuroepithelial cells implanted into the fetal rat brain.

Conditionally immortalized (temperature-sensitive) striatal-derived neuronal progenitor cell lines and primary neuroepithelial cells were transplanted into the CNS of gestational day 15-16 rat fetuses using an 'in utero' surgical procedure. Each fetus received 2.5-3 x 10(4) donor cells previously labelled in vitro by incubation with 5-bromo-2'-deoxyuridine (BrdU). At 5 days following transplantation, 69% of the fetuses were still alive. Engrafted cells were detected by BrdU immunohistochemistry, and the appearance of the engrafted cells and the time course of Nestin and PCNA expression were measured at 6, 24, 64 h and 5 days after transplantation. The evolution of Large T-Antigen immunoreactivity in engrafted temperature-sensitive (ts) cells was also evaluated at the above time intervals. The results indicate that the majority of the implanted cells were aggregated into clusters 24 h after transplantation. These clusters were not visible at 6 h, when most of the cells were isolated. The clusters were located in both the ventricles and parenchyma. These findings were common to both ts cells and striatal primary neuroepithelial cells. At 64 h and 5 days, isolated cells associated with the germinal layer and scattered throughout the parenchyma were also found. In the clusters, Nestin expression decreased proportionally with time following transplantation. Furthermore, Large T-Antigen immunoreactivity disappeared from ts cells between 6 and 24 h after transplantation. Finally, measurements of the temporal evolution of PCNA expression within the clusters indicate a progressive reduction in the mitotic activity of the transplanted cells. The results demonstrate that striatal primary neuroepithelial cells and conditionally immortalized neuronal progenitors can survive, migrate and/or compartimentalize into clusters whilst changing their antigenic properties and ability to proliferate.