Target-dependent cell death is an important embryogenic mechanism for regulating and sculpting the developing motor system. Efficient characterization of apoptosis has been more difficult in the nervous system than in other systems due to the use of several different primary culture systems as well as with heterogeneity of neuronal cell populations. We have developed a simple in vitro model of apoptosis with the motor neuron hybrid NSC34, a cell line which expresses much of the motor neuron phenotype (Cashman et al. 1992). Serum-deprived NSC 34 cells in bulk culture undergo cell death, likely from the withdrawal of the growth factors and/or hormones present in fetal calf serum medium supplements. This cell death is accompanied by fragmentation of chromatin into nucleosome multimers, heterochromatization of the nucleus, and other ultrastructural changes reminiscent of apoptotic death. Cell death is inhibitable by addition of agents which block new gene expression ( e.g. cycloheximide) or inhibit endonuclease activity (e.g. aurintricarboxylic acid). / We report similar findings with primary embryonic rat motor neurons identified by surface immunoreactivity for p75 LA NGFR, the low-affinity neurotrophin receptor (Bloch-Gallego et al. 1991; Camu and Henderson 1992; Chao and Hempstead 1995). The p75+ motor neuron population could be maintained for more than 48 hours in mixed suspension cultures supplemented with 10% fetal calf serum. However, the p75+ cell population was rapidly depleted in serum-deprived cultures, a phenomenon accompanied by the appearance of oligonucleosomal ladders. Serum-deprived p75+ cells were supported by the motor neuron-relevant factors BDNF, CNTF, GDNF and IGF-1, but not the non-relevant factor NGF. Serum-deprived p75 + cells were also protected by cycloheximide, suggesting a role for apoptosis in the cell death. / We have investigated the role of reactive oxygen species in acquired and genetic motor neuron diseases. Interestingly, a rapid burst of reactive oxygen species is observable within one hour of serum deprivation in both NSC34 and rat motor neuron systems. This burst precedes measurable cell death by at least one day, indicating that oxygen species generation may be an initial hallmark of target-dependent death. The amplitude and temporal nature of this burst may be altered by manipulating various cellular ROS defence mechanisms. Such manipulations also alter cell death progression, suggesting that the apoptotic cascade may be dependent upon this early ROS burst. The identity, source and activity of the relevant ROS may provide insight into the etiology and treatment of human motor neuron diseases.
| Identifer | oai:union.ndltd.org:LACETR/oai:collectionscanada.gc.ca:QMM.35486 |
| Date | January 1998 |
| Creators | Shaw, Ivan Ting-kun, 1966- |
| Publisher | McGill University |
| Source Sets | Library and Archives Canada ETDs Repository / Centre d'archives des thèses électroniques de Bibliothèque et Archives Canada |
| Language | English |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Format | application/pdf |
| Coverage | Doctor of Philosophy (Department of Neurology and Neurosurgery.) |
| Rights | All items in eScholarship@McGill are protected by copyright with all rights reserved unless otherwise indicated. |
| Relation | alephsysno: 001652048, proquestno: NQ50259, Theses scanned by UMI/ProQuest. |
Page generated in 0.0021 seconds