Tyrosine hydroxylase mRNA expression by dopaminergic neurons in culture: effect of 1-methyl-4-phenylpyridinium treatment.
To enable us to study expression of tyrosine hydroxylase [TH; tyrosine 3-monooxygenase; L-tyrosine tetrahydropteridine:oxygen oxidoreductase (3-hydroxylating); EC 1.14.16.2] as a measure of dopaminergic neuron function in future experiments, methods were developed to quantify TH mRNA levels in cultures of dopaminergic mesencephalic cells. The model of selective dopaminergic toxicity of 1-methyl-4-phenylpyridinium (MPP+) was used to verify the specificity of our methods. Fetal (embryonic day 15) rat ventral mesencephalic cell cultures were treated with 15 microM MPP+ for 48 h, conditions previously shown to reduce the number of TH-immunoreactive neurons, TH activity, and dopamine uptake to 5-10% of control values. This treatment decreased the number of neurons labeled by TH in situ hybridization to 9% of untreated controls and caused a strong reduction of the abundance of TH mRNA in Northern blots. Our findings establish TH mRNA expression as a parameter for future studies of toxic and trophic effects on cultured dopaminergic neurons, and they support the view that MPP+ destroys dopaminergic neurons.
Beck
K D
KD
Andrus Gerontology Center, University of Southern California, Los Angeles 90089-0191.
Knusel
B
B
Pasinetti
G
G
Michel
P P
PP
Zawadzka
H
H
Goldstein
M
M
Hefti
F
F
D015655:1-Methyl-4-phenylpyridinium / Q000494:pharmacology*; D000818:Animals; D015152:Blotting, Northern; D001921:Brain / Q000201:enzymology*; D002478:Cells, Cultured; D004298:Dopamine / Q000378:metabolism*; D004622:Embryo, Mammalian; D015870:Gene Expression / Q000187:drug effects; D008636:Mesencephalon / Q000166:cytology / Q000201:enzymology; D009474:Neurons / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D012333:RNA, Messenger / Q000187:drug effects / Q000235:genetics*; D051381:Rats; D011919:Rats, Inbred Strains; D014446:Tyrosine 3-Monooxygenase / Q000235:genetics* / Q000378:metabolism
D012333:RNA, Messenger; D014446:Tyrosine 3-Monooxygenase; D015655:1-Methyl-4-phenylpyridinium; D004298:Dopamine