An in vitro model of 1-methyl-4-phenyl-pyridinium (MPP+) toxicity: incubation of rabbit caudate nucleus slices with MPP+ followed by biochemical and functional analysis.
1. Slices of rabbit caudate nucleus were preincubated for up to 24 h in vitro in the presence of the neurotoxic compound 1-methyl-4-phenyl-pyridinium (MPP+). Subsequently the levels of endogenous monoamines in the slices were determined by h.p.l.c. with electrochemical detection. MPP+, in concentrations higher than 32 nM significantly diminished the dopamine levels within the slices in a concentration- and time-dependent manner; at 32 microM the depletion was more than 95%. The concentration of the major metabolite of dopamine, dihydroxyphenyl acetic acid (DOPAC) was decreased at concentrations of MPP+ that did not alter dopamine levels. Thus, MPP+ increased the dopamine/DOPAC ratio. 2. In contrast, both 5-hydroxytryptamine (5-HT) levels and 5-HT/5-hydroxyindolacetic acid (5-HIAA) ratios were increased at nanomolar concentrations of MPP+. 5-HT was significantly reduced only at 32 microM. 3. The dopamine uptake inhibitor nomifensine reduced the depletory effect of MPP+ on dopamine and DOPAC content. 4. Following 24 h pretreatment with MPP+, the uptake of [3H]-dopamine into rabbit caudate nucleus slices was either enhanced (at 0.32 microM, 1 microM and 3.2 microM MPP+) or reduced (at 32 microM MPP+). 5. Preincubation of slices with 10 microM MPP+ for only 1 h increased their 3H-labelling (in contrast to 24 h pretreatment) whereas after 9 h no net increase was detectable. After 1 and 9 h MPP+ pretreatment, much less deaminated metabolites of [3H]-dopamine were found in the incubation medium of MPP+ treated slices than in the medium of control slices. These findings suggest that MPP+ strongly inhibits the enzyme monoamine oxidase (MAO) within dopaminergic (and 5-hydroxytryptaminergic) terminals before destroying them. 6. To validate the proposed in vitro model functionally, the electrically evoked release of [3H]-acetylcholine ([3H]-ACh) was investigated in MPP+ treated slices and controls. MPP+ reduced both the facilitatory effect of the D2-receptor antagonist domperidone and the inhibitory effect of the catecholamine uptake inhibitor nomifensine on [3H]-ACh release; effects compatible with a diminished inhibitory dopaminergic input on cholinergic neurones. 7. These findings also show that the terminal region of dopaminergic neurones, the caudate nucleus, is a site for MPP+ toxicity. The present in vitro model may be useful for investigating the effects of MPP+ and its interaction with other drugs under defined conditions.
Feuerstein
T J
TJ
Neurologische Universitätsklinik, Freiburg, Federal Republic of Germany.
Hedler
L
L
Jackisch
R
R
Hertting
G
G
D015655:1-Methyl-4-phenylpyridinium; D000818:Animals; D015306:Biogenic Monoamines / Q000378:metabolism; D002421:Caudate Nucleus / Q000187:drug effects / Q000378:metabolism / Q000502:physiology*; D002794:Choline / Q000378:metabolism; D003641:Deamination; D004298:Dopamine / Q000378:metabolism; D004558:Electric Stimulation; D005260:Female; D066298:In Vitro Techniques; D008297:Male; D009498:Neurotoxins / Q000633:toxicity*; D009627:Nomifensine / Q000494:pharmacology; D011726:Pyridinium Compounds / Q000633:toxicity*; D011817:Rabbits
D015306:Biogenic Monoamines; D009498:Neurotoxins; D011726:Pyridinium Compounds; D009627:Nomifensine; D002794:Choline; D015655:1-Methyl-4-phenylpyridinium; D004298:Dopamine