Untitled

Abstract

1. The monoamines dopamine (DA), norepinephrine (NE), epinephrine (E), and serotonin (5-HT) serve as endogenous neurotransmitters in the nervous system. We recently reported that the neurotransmitter DA can trigger apoptosis (programmed cell death; PCD) in cultured, postmitotic chick embryo sympathetic neurons, suggesting a role for apoptosis in degenerative processes such as Parkinson's disease (PD). However, PD is also associated with involvement of other monoaminergic (MA) neuronal systems (noradrenergic and serotoninergic), though to a lesser extent. 2. We therefore tested the apoptosis-triggering potential of NE, E, and 5-HT in comparison to the DA effect, in cultured postmitotic nerve growth factor (NGF)-dependent chick embryo sympathetic neurons and mouse cerebellar granule cells. 3. In both model systems MA induced neuronal attrition characteristic of apoptosis. MA caused marked morphological alterations: severe neuronal soma shrinkage, membrane blebbing, nuclear condensation and fragmentation, and axonal disintegration. Flow-cytometric analysis of propidium iodide-stained cell nuclei revealed characteristic apoptotic nuclear fragmentation. MA-induced apoptosis could be blocked by SH-group containing antioxidants but not by inhibitors of transcription and translation. 4. Comparison between the two model systems revealed that the cerebellar granule neurons were distinctly more sensitive to the neurotoxic potential of the MA than sympathetic neuronal cells. Significant differences in the dose dependencies and time course of the apoptotic effect were observed among the examined MA, graded as DA > NE approximately E > 5-HT. 5. We conclude that the apoptosis triggering potential, probably mediated by oxidative metabolites, is shared by all MA tested, but with differential time course and dose dependencies. A correlation can be drawn between the effects of DA vs NE vs 5-HT and the relative involvement of dopaminergic/noradrenergic/serotoninergic pathways in PD, which may suggest a common multisystem underlying abnormality in neuronal apoptosis-control mechanisms.

Involvement of T-complex Protein-1δ in Dopamine Triggered Apoptosis in Chick Embryo Sympathetic Neurons

“…In support of this concept, we recently showed that DA can induce apoptosis in cultured neuronal as well as non-neuronal cells, including chick embryo sympathetic neurons, mouse thymocytes, and rat pherochromacytoma cells (PC12) (7)(8)(9)(10). Based on these findings we hypothesized that neuronal degeneration in PD may be caused at least in part by DA-mediated induction of apoptosis in the nigrostriatal neurons.…”

“…Paravertebral sympathetic ganglia at embryonic day 9 (8,24) were dissected out, and the cells were dissociated by trypsinization (0.25% in EDTA for 30 min at 37°C). Cells were then plated on 24 poly-L-lysine-coated wells (4 ϫ 10 5 cells/well) or 35-mm tissue culture plates (10 6 cells/well) (TPP, Trasadingen, Switzerland) and grown in serum-free medium DCCM-1 (Biological Industries, Kibbutz Beit Haemek, Israel) supplemented with 2% chick embryo extract, 0.5% horse serum, 20 g/ml fluorodeoxyuridine, and 20 g/ml uridine (to kill the non-neuronal cells), 2 mM glutamine, penicillin, streptomycin, and amphotericine B, and 10 ng/ml nerve growth factor (NGF).…”

“…CH8502, Sigma) was dissolved directly in the appropriate culture medium. Sympathetic neurons (days 2-3 in culture in the presence of NGF) were treated for a given time with 300 M DA (previously found to be the potent concentration following detailed dose response studies (7,8)). Plates maintained in identical conditions but without exposure to DA served as controls.…”

“…Plates maintained in identical conditions but without exposure to DA served as controls. Neuronal viability was determined by trypan blue exclusion assay, as described by Zilkha-Falb et al (8).…”

“…(b) The requirement of large numbers of neurons precluded studies on the subpopulation of dopaminergic neurons, which comprise only 3% of the total neuronal population of the substantia nigra. (c) Since all the cells we investigated (several types of neurons, PC12 cells, thymocytes including dopaminergic neurons (7)(8)(9)(10)20)) were susceptible to the apoptotic triggering potential of DA, similar cellular mechanisms might be at work. (d) Since dopaminergic neurons synthesize, store, and secrete DA, it is feasible that under certain conditions the secreted and/or accumulated DA becomes toxic and induces apoptotic neuronal death, making it an important mediator of PD.…”