Subthalamic Nucleus Deep Brain Stimulation Does Not Modify the Functional Deficits or Axonopathy Induced by Nigrostriatal α-Synuclein Overexpression - PubMed (original) (raw)
. 2017 Nov 27;7(1):16356.
doi: 10.1038/s41598-017-16690-x.
D Luke Fischer 1 2 3, Christopher J Kemp 1, Allyson Cole-Strauss 1, Jack W Lipton 1 4, Megan F Duffy 1 3, Nicole K Polinski 1 3, Kathy Steece-Collier 1 4, Timothy J Collier 1 4, Sara E Gombash 1, Daniel J Buhlinger 1, Caryl E Sortwell 5 6
Affiliations
- PMID: 29180681
- PMCID: PMC5703955
- DOI: 10.1038/s41598-017-16690-x
Subthalamic Nucleus Deep Brain Stimulation Does Not Modify the Functional Deficits or Axonopathy Induced by Nigrostriatal α-Synuclein Overexpression
D Luke Fischer et al. Sci Rep. 2017.
Abstract
Subthalamic nucleus deep brain stimulation (STN DBS) protects dopaminergic neurons of the substantia nigra pars compacta (SNpc) against 6-OHDA and MPTP. We evaluated STN DBS in a parkinsonian model that displays α-synuclein pathology using unilateral, intranigral injections of recombinant adeno-associated virus pseudotype 2/5 to overexpress wildtype human α-synuclein (rAAV2/5 α-syn). A low titer of rAAV2/5 α-syn results in progressive forelimb asymmetry, loss of striatal dopaminergic terminal density and modest loss of SNpc dopamine neurons after eight weeks, corresponding to robust human-Snca expression and no effect on rat-Snca, Th, Bdnf or Trk2. α-syn overexpression increased phosphorylation of ribosomal protein S6 (p-rpS6) in SNpc neurons, a readout of trkB activation. Rats received intranigral injections of rAAV2/5 α-syn and three weeks later received four weeks of STN DBS or electrode implantation that remained inactive. STN DBS did not protect against α-syn-mediated deficits in forelimb akinesia, striatal denervation or loss of SNpc neuron, nor did STN DBS elevate p-rpS6 levels further. ON stimulation, forelimb asymmetry was exacerbated, indicating α-syn overexpression-mediated neurotransmission deficits. These results demonstrate that STN DBS does not protect the nigrostriatal system against α-syn overexpression-mediated toxicity. Whether STN DBS can be protective in other models of synucleinopathy is unknown.
Conflict of interest statement
The authors declare that they have no competing interests.
Figures
Figure 1
Experimental Design. Rats in Experiment 1 (n = 5) underwent intranigral administration of viral vector and were assessed using the cylinder task at 16 and 55 days after surgery. Rats in Experiment 2 (n = 12) underwent the same viral vector surgery and cylinder testing at Day 16 as in Experiment 1, but on Day 18 were implanted with an electrode in the STN. Rats were randomly assigned to receive continuous stimulation through Day 54 (‘Active’, n = 6) or to not have the electrode activated (‘Inactive’, n = 6). On Day 54, the cylinder task was conducted, and afterward, stimulation was ceased for 24 hours for forelimb asymmetry examination in the stimulation ‘off’ state at Day 55. After the cylinder task was completed on Day 55, stimulation was restarted for Active rats and continued until sacrifice. All rats in Experiments 1 and 2 were sacrificed on Day 56.
Figure 2
Overexpression of α-Syn Vector Model Recapitulates Early-Stage PD. Intranigral administration of rAAV2/5-α-syn (n = 5) results in unilateral transgene expression in the nigrostriatal system when examined for hu-α-syn immunoreactivity (A,B,D,G) and TH immunoreactivity (C,G) with extensive co-localization (E,G). Overexpression of α-syn results in modest but significant forepaw use asymmetry at sixteen days and eight weeks (F, p = 0.006 and p = 0.005, respectively), as well as robust, unilateral striatal α-syn immunoreactivity (p < 0.001) and a ~25% reduction in striatal TH immunoreactivity (G and H, p = 0.042). Overexpression of α-syn over eight weeks resulted in a significant 10% decrease of THir SNpc neurons in this specific cohort (I–K, p = 0.0258). Scale bar in J is 50 μm.
Figure 3
Overexpression of α-syn Does Not Alter SN Gene Expression. qPCR of the SN (ipsilateral vs. contralateral hemispheres) in the No Electrode group (n = 5) was conducted for the following transcripts: human Snca (hu-Snca, to confirm transduction), rat Snca (rt-Snca), Th, Bdnf and Trk2. No significant differences between hemispheres were revealed for rt-Snca, Th, Bdnf or Trk2 (p > 0.05). b.d.l. = below detectable limits.
Figure 4
Effects of α-syn Overexpression on Total and Phosphorylated Levels of rpS6. Representative, low- and high-magnification images (A–D and E–H, respectively) from the SNpc ipsilateral to rAAV2/5 α-syn injection displaying triple label immunofluorescence for tyrosine hydroxylase (TH, green, A,E), ribosomal protein S6 (rpS6, red, B,F) and human alpha-synuclein (α-syn, blue, C,G). Merged images showing numerous SNpc neurons co-expressing all three proteins are shown at low (D) and high (H) magnification. From the same rat in an adjacent section, the ipsilateral SN is shown at low and high magnification for TH (L,P), p-rpS6 (M,Q), human α-syn (N,R) and merged images (O,S). Quantification of the No Electrode group (n = 5) was assessed between hemispheres to determine the effect of α-syn overexpression on TH (I), rpS6 (J) and p-rpS6 (K). A difference between hemispheres was observed for TH (I, p = 0.003) and p-rpS6 (K, p = 0.004). Scale bars are 200 μm and 50 μm for low- and high-magnification images, respectively.
Figure 5
STN Stimulation Does Not Alter α-syn Transgene Expression. Active (A,B) and Inactive (C,D) groups showed robust α-syn transgene expression as revealed by immunohistochemical brightfield (A,C with high-magnification inset) or immunofluorescent (B,D with high-magnification inset) labeling methods in the SNpc. Striatal α-syn immunoreactivity in the ipsilateral hemisphere was not significantly different between the No Electrode, Inactive and Active groups (n = 5, 6 and 6, respectively; E, p > 0.05). Electrode-implanted rats included in this study were examined post mortem for electrode placement targeting the STN (F). The tract previously occupied by the electrode is appreciable dorsal and slightly medial to the STN.
Figure 6
STN Stimulation Does Not Alter Symptom Progression Nor Neuropathology. Both Inactive and Active groups (n = 6 per group) exhibited forepaw use asymmetry after eight weeks compared to baseline or just prior to electrode placement surgery (A, p = 0.004 and p = 0.011, respectively); though, there was no interaction between group and time, so stimulation did not significantly alter the development of symptoms at eight weeks. THir SNpc neurons were not different between Active (shown in B at low magnification and in C and D at high magnification for the ipsilateral and contralateral SNpc, respectively) and Inactive groups (E, p = 0.314). (F) Striatal THir was not significantly different between groups (p > 0.05) but was significantly different between hemispheres for Inactive (p = 0.0002) and Active groups (p = 0.0087). Inspection of the α-syn-ir or THir striatum (G,H and I,J, respectively, for low and high magnification photomicrographs of an Active rat) did not reveal any apparent differences to a blinded investigator in α-syn-ir aggregates (G,H) or THir axonal swellings (I,J) between Active and Inactive rats. Scale bars are 50 μm.
Figure 7
Effects of STN DBS on Phosphorylated and Total Levels of rpS6. Quantification of the Inactive and Active groups was assessed between hemispheres and treatments to determine the effect of stimulation on p-rpS6 (A) and rpS6 (B). (A) THir SN neurons expressing hu-α-syn exhibited significantly more p-rpS6 immunoreactivity (Inactive, n = 6: t(5) = 2.805, p = 0.0378; Active, n = 6: t(5) = 3.040, p = 0.0287). However, p-rpS6 appeared to increase slightly within α-syn expressing THir SN neurons in association with stimulation, whereas STN DBS overall had no significant impact on p-rpS6 immunoreactivity in either the ipsilateral or contralateral SNpc (F(3,22) = 5.715, p > 0.05). (B) Total rpS6 was significantly increased in THir SN neurons expressing hu-α-syn in Inactive rats (n = 4; p = 0.0491) but not in Active Rats (n = 6; p > 0.05). Active STN DBS did not affect rpS6 immunoreactivity in hu-α-syn expressing neurons (p > 0.05) but did increase rpS6 immunoreactivity in the contralateral SNpc THir neurons (p = 0.004).
Figure 8
Acute STN Stimulation Exacerbates Forelimb Asymmetry. Unilateral overexpression of α-syn results in DA neurotransmission and handling dysfunction in the ipsilateral striatum and a functional impairment that was exaggerated by unilateral STN stimulation and an asymmetric release of striatal DA (A). (B) Forelimb use asymmetry was augmented by stimulation when compared to asymmetry in an off-stimulation cylinder task (n = 6; p = 0.047) but asymmetry was not exacerbated in the Inactive group (n = 6; p > 0.05).
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