Hemiparkisonism Induced Through Unilateral Expression of Adenovirus-Mediated Clostridial Light Chain Gene in the Substantia Nigra

Jun Yang, Qingshan Teng, Mary E. Garrity-Moses, Thais Federici, John Butler, Shearwood McClelland III and Nicholas M Boulis

Departments of Neuroscience and Neurosurgery, the Cleveland Clinic Foundation/Lerner Research Institute, Cleveland OH 44195

Background: Clostridial light chain (LC) inhibits synaptic transmission by digesting a vesicle-docking protein, synaptobrevin, without altering neuronal health. We have previously reported focal synaptic inhibition through adenovirus-mediated gene transfer of LC (AdLC) in the central nervous system. We report here the feasibility of creating reversible rat hemiparkinsonism (HPD) model through AdLC injection into the substantia nigra (SN).
Methods: 20 adult Sprague Dawley rats were divided into four groups for injection of AdLC, AdGFP (vector control), 6-hydroxydopamine (6-OHDA, positive control), and PBS (negative control). 4 ml of AdLC, AdGFP, PBS and 6-OHDA were injected into right SN. After induction with amphetamine or apomorphine, rotation in 30 minutes duration was assessed before and 8 or 10 days after SN injection. Finally, brains were extracted for histopathological studies.

Results: After challenging with amphetamine, rotation increased from 8.7±7.5 (before) to 89.2±40.7 (after, p=0.04) in the AdLC group. In contrast, amphetamine induced greater rotation in the 6-OHDA group (6.3±7.5, before; 256±118 after, p=0.03). There were no significant differences in PBS group (16.8±9.9, before; 5.6±18.2, after, p=0.36) and AdGFP group (1.2±11.8, before; 4.8±1.3, after, p=0.21). After challenging with apomorphine, a significant increase in ipsilateral rotation was observed in the AdLC group (20.2±18, before; 291.2±119, after, p=0.01). On the contrary, apomorphine induced rotation in 6-OHDA group, but without a statistical difference (16.3±32.3, before; 85.7±138, after, p=0.6). We also detect no significant difference in PBS group (10.2±14.6, before; 11.2±20.9, after, p=0.69) and AdGFP group (2.6±27.2, before; 31±16.8, after, p=0.09). Histological results revealed spatially discrete LC expression with corresponding synaptobrevin depletion in the SN. Immunohistochemistry (IHC) with antibody against tyrosine hydroxylase (TH) showed symmetric staining in bilateral SN and caudate putamen (CPu) in AdGFP rats, but decreased staining in ipsilateral SN and CPu in 6-OHDA rats. Interestingly, IHC with TH antibody also showed decreased staining in ipsilateral CPu, but increased staining in ipsialteral SN in AdLC rats. The histological results are consistent with behavioral results in AdLC and 6-OHDA groups.

Conclusion: Our results demonstrated that it is feasible to block the nigrostratum pathway through nigral LC expression without altering neuronal health. Different from 6-OHDA, LC expression inhibits synaptic activity without killing neurons. This approach therefore represents a potentially reversible approach to dopamine depletion. The typical behavioral characteristic of HPD demonstrated in this model represents a strategy for behavioral study and testing therapeutic strategies following nigrostriatum circuit blocking. We believe that this model would facilitate transient and controlled nigral inhibition for studying striatal recovery, dopaminergic re-innervation, and normalization of striatal receptors following the recovery of striatal dopaminergic input.

Key Words: Hemiparkinsonism; Targeted Gene Expression; Clostridial Toxin; Synaptic

Transmission

Support Contributed By: NINDS K08 NS43305