TY - JOUR
T1 - Dopaminergic neurotoxins require excitotoxic stimulation in organotypic cultures
AU - Kress, Geraldine J.
AU - Reynolds, Ian J.
N1 - Funding Information:
This work was supported by USAMRMC grant DAMD 17-98-1-8627. We thank Julianne Jaumotte for valuable advice on the preparation of the cultures and Dr. Teresa Hastings for helpful discussion.
PY - 2005/12
Y1 - 2005/12
N2 - We have investigated the properties of the dopaminergic neurotoxins 6-hydroxydopamine, 1-methyl-4-phenylpyridinium and rotenone using an organotypic culture that included slices of substantia nigra, striatum and cortex maintained for about 20 days in vitro. At this age, the organotypic culture contains dopaminergic neurons, visualized using tyrosine hydroxylase (TH) immunohistochemistry, that project into the striatal slice and extend up to 1 mm into the cortical slice. Using TH immunohistochemistry to assess survival of dopaminergic neurons, we found that the three dopaminergic toxins alone were not selectively neurotoxic. However, the addition of a low concentration of N-methyl-d-aspartate together with each individual toxin resulted in profound injury to the dopaminergic neurons, reflected by the loss of cell bodies and the fragmentation of processes. The combined toxicity was completely blocked by MK801. To assess the specificity of the injury, we measured the diameter of cell nuclei in the organotypic culture stained with Hoechst 33342 because the nucleus shrinks when neurons are injured. These measurements showed that the combined toxin treatment selectively injured only the TH immunoreactive cells. Thus, in a model culture system where dopaminergic neurons innervate appropriate targets, excitotoxicity appears to be essential for the manifestation of the toxic actions of 6-hydroxydopamine, 1-methyl-4-phenylpyridinium and rotenone.
AB - We have investigated the properties of the dopaminergic neurotoxins 6-hydroxydopamine, 1-methyl-4-phenylpyridinium and rotenone using an organotypic culture that included slices of substantia nigra, striatum and cortex maintained for about 20 days in vitro. At this age, the organotypic culture contains dopaminergic neurons, visualized using tyrosine hydroxylase (TH) immunohistochemistry, that project into the striatal slice and extend up to 1 mm into the cortical slice. Using TH immunohistochemistry to assess survival of dopaminergic neurons, we found that the three dopaminergic toxins alone were not selectively neurotoxic. However, the addition of a low concentration of N-methyl-d-aspartate together with each individual toxin resulted in profound injury to the dopaminergic neurons, reflected by the loss of cell bodies and the fragmentation of processes. The combined toxicity was completely blocked by MK801. To assess the specificity of the injury, we measured the diameter of cell nuclei in the organotypic culture stained with Hoechst 33342 because the nucleus shrinks when neurons are injured. These measurements showed that the combined toxin treatment selectively injured only the TH immunoreactive cells. Thus, in a model culture system where dopaminergic neurons innervate appropriate targets, excitotoxicity appears to be essential for the manifestation of the toxic actions of 6-hydroxydopamine, 1-methyl-4-phenylpyridinium and rotenone.
KW - Excitotoxicity
KW - NMDA receptors
KW - Organotypic culture
KW - Parkinson's disease
KW - Substantia nigra
KW - Tyrosine hydroxylase
UR - http://www.scopus.com/inward/record.url?scp=27744434081&partnerID=8YFLogxK
U2 - 10.1016/j.nbd.2005.04.019
DO - 10.1016/j.nbd.2005.04.019
M3 - Article
C2 - 15996475
AN - SCOPUS:27744434081
SN - 0969-9961
VL - 20
SP - 639
EP - 645
JO - Neurobiology of Disease
JF - Neurobiology of Disease
IS - 3
ER -