Georgievska Biljana, Kirik Deniz, Björklund Anders
Wallenberg Neuroscience Center, Department of Physiological Sciences, Lund University, BMC A11, 221 84, Lund, Sweden.
Exp Neurol. 2002 Oct;177(2):461-74. doi: 10.1006/exnr.2002.8006.
The effects of sustained (up to 9 months) striatal overexpression of glial cell line derived neurotrophic factor (GDNF) on lesioned nigrostriatal dopamine (DA) neurons was studied using a recombinant lentiviral (rLV) vector to deliver GDNF into the striatum 4 weeks prior to the creation of an intrastriatal 6-hydroxydopamine lesion. The results of the amphetamine-induced rotation suggested an initial partial protection followed by a complete recovery, whereas the spontaneous motor behaviors remained impaired. There was a clear protection of the nigral tyrosine hydroxylase (TH)-positive neurons in the rLV-GDNF group compared to rats injected with the control vector encoding green fluorescent protein (GFP) (70 and 20% of the intact side, respectively). However, the striatal TH+ fiber density was equally reduced (to 20% of the intact side) in both groups. Further morphological analyses indicated that the nigrostriatal projections of the DA neurons were indeed preserved in the GDNF group. The axonal projections were visualized using two independent methods: First, retrograde labeling of the nigral cell bodies by intrastriatal Fluoro-Gold injections showed that the majority of rescued cells in the GDNF group had preserved axonal projections to striatum. Second, injections of a recombinant adeno-associated viral vector expressing GFP into the nigra was used to anterogradely fill the DA neurons and their projections with GFP protein. GFP immunostaining clearly demonstrated that the fibers of the nigral DA cells were preserved along the nigrostriatal pathway and innervated large parts of the striatum, but did not express TH at detectable levels. In addition, fiber sprouting was observed in the globus pallidus, entopeduncular nucleus, and substantia nigra, corresponding to areas where GDNF protein was released. The lack of functional recovery in the spontaneous motor behaviors may, at least in part, be explained by this extensive aberrant fiber sprouting in the downstream striatal target nuclei and/or decreased synthesis of dopamine in the striatum.
利用重组慢病毒（rLV）载体在纹状体内注射6-羟基多巴胺损伤前4周将胶质细胞源性神经营养因子（GDNF）递送至纹状体，研究了纹状体持续（长达9个月）过表达GDNF对损伤的黑质纹状体多巴胺（DA）神经元的影响。苯丙胺诱导旋转试验的结果表明，起初有部分保护作用，随后完全恢复，而自发运动行为仍受损。与注射编码绿色荧光蛋白（GFP）的对照载体的大鼠相比，rLV-GDNF组黑质酪氨酸羟化酶（TH）阳性神经元有明显的保护作用（分别为完整侧的70%和20%）。然而，两组纹状体TH+纤维密度均同样降低（至完整侧的20%）。进一步的形态学分析表明，GDNF组DA神经元的黑质纹状体投射确实得以保留。采用两种独立方法观察轴突投射：第一，通过纹状体内注射荧光金逆行标记黑质细胞体，结果显示GDNF组大多数获救细胞保留了至纹状体的轴突投射。第二，将表达GFP的重组腺相关病毒载体注射到黑质中，用于用GFP蛋白顺行填充DA神经元及其投射。GFP免疫染色清楚地表明，黑质DA细胞的纤维沿黑质纹状体通路得以保留，并支配纹状体的大部分区域，但未检测到TH表达。此外，在苍白球、内囊核和黑质中观察到纤维芽生，对应于GDNF蛋白释放的区域。自发运动行为缺乏功能恢复至少部分可由下游纹状体靶核中这种广泛的异常纤维芽生和/或纹状体中多巴胺合成减少来解释。
