Analysis of NG2-glia in the striatum in 6-OHDA-lesioned
Parkinsonian rats treated with L-DOPA by immunoconfocal microscopy
Firstly, we examined the spatial distribution of NG2-glia in the rat
striatum of the experimental groups No lesion (control), 6OHDA
lesion (6OHDA), 6OHDA+DOXY, 6OHDA+Vehicle+L-DOPA and 6OHDA+DOXY+L-DOPA(Fig. 2B-E; see methods). The four subdivisions of the striatum
(dorsomedial, dorsolateral, ventromedial, ventrolateral) were separately
quantified, due to their functional properties, which reflect the
diversity of the striatal dopaminergic system connections (Fig. 2F-I).
Parkinsonian rat brains were analyzed 32 days after the neurotoxin
microinjection, corresponding to 6OHDA-lesion establishment and
L-DOPA/vehicle time therapy. The analysis of the dorsolateral and
ventrolateral striatal regions showed a slight increase (15%) in
NG2-glia density when compared to the No lesion group. This minor
increase was maintained after doxycycline treatment (experimental group6OHDA+DOXY , p<0.05; Fig. 2G and I).
Parkinsonian rats receiving L-DOPA and reporting LID presented a robust
NG2-glia density reduction in the dorsolateral, ventrolateral and
dorsomedial striatum when compared to the No lesion (control),
6OHDA lesion (6OHDA), 6OHDA+DOXY and 6OHDA+DOXY+L-DOPA groups
(6OHDA+Vehicle+L-DOPA , p<0.001; Fig. 2F, G and I).
Doxycycline administration before L-DOPA prevented dyskinesia and lead
reduced NG2-glia density in the striatum regions (6OHDA+DOXY
+L-DOPA ; p<0.001; Fig. 2F, G and I). There was a negative
correlation of NG2-glia density with the dyskinesia score (dorsomedial
r=-0.83, dorsolateral r=-0.88, and ventrolateral r=-0.64;
p<0.0001; Fig. 2J, K and M). There was no NG2-glia
immunolabelling alteration in the ventromedial striatum of the animals.
The profile of NG2-glia, analyzed by protein expression immunoblotting
in dorsolateral striatum extract, revealed one band of about 26kDa
corresponding to NG2-glia in both the control and the lesioned striatum
(Fig. 2N). Lesioned rats treated or not with doxycycline showed a slight
increase (17%) in NG2-glia expression when compared to No lesionones (6OHDA and 6OHDA+DOXY , p<0.05, Fig. 2N).
The NG2-glia expression decreased in the lesioned striatum of
L-DOPA-treated parkinsonian rats compared to all other groups (50-60%,6OHDA+Vehicle+L-DOPA , p<0.05, Fig. 2M). Doxycycline
treatment induced a significant increase (76%) of NG2-glia
immunoreactivity compared to the intensity detected in animals
presenting LID (6OHDA+DOXY+L-DOPA ; p<0.05, Fig. 2N).