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. Author manuscript; available in PMC: 2009 Mar 17.
Published in final edited form as: Mol Ther. 2005 Oct;12(4):618–633. doi: 10.1016/j.ymthe.2005.05.006

FIG. 5. ISH analysis of ppEnk and DARPP-32 transcripts.

FIG. 5

Coronal sections shown on the left of (A), (C), and (E) were hybridized to a ppEnk-specific oligonucleotide probe (Table 1), while sections in (B), (D) and (F) were hybridized to a DARPP-32-specific oligonucleotide probe (Table 1). The arrows above the coronal sections show the approximate vector injection site and the fill patterns of the arrows correspond to the fill patterns of the histograms to show which treatment was received in each hemisphere. To better demonstrate the anatomical pattern of striatal transcripts the coronal sections were false colored. The scale is shown at the bottom. (A) Evaluation of long-term intrastriatal rAAV-siHUNT-1 expression on striatal ppEnk levels in R6/1 mice and wild-type littermate controls. Quantitative analysis of normalized ppENK levels confirm that ppENK levels are reduced in 22-week-old R6/1 mice (F[1,66] = 99.5, *P < 0.0001). rAAV-siHUNT-1 injection into wild-type control striata does not lead to any alteration in striatal ppENK levels compared to untreated control left striata (Fisher’s PLSD post hoc test, P > 0.7). In contrast, in R6/1 mice, striatal rAAV5-siHUNT-1 treatment led to a mild 24% increase in striatal ppEnk levels compared to the ppEnk levels observed in the control striatum (Fisher’s PLSD post hoc test, †P = 0.01). (B) Evaluation of long-term intrastriatal rAAV-siHUNT-1 injection on steady-state striatal DARPP-32 mRNA levels. Similar to ppENK mRNA levels, DARPP-32 mRNA was reduced in R6/1 animals compared to littermate controls (F[1,66] = 114.1, *P > 0.0001). rAAV5-siHUNT-1 injection did not affect striatal DARPP-32 levels in wild-type controls (Fisher’s PLSD post hoc test, *P > 0.1). Striatal rAAV5-siHUNT-1 treatment resulted in a significant 16% increase in DARPP-32 mRNA levels (Fisher’s PLSD post hoc, †P < 0.03) in R6/1 mice compared to DARPP-32 mRNA levels in the untreated striatum. (C) Analysis of the effect of intrastriatal injection of rAAV-siHUNT-2 on striatal ppEnk transcript levels. In stark contrast to the pattern seen with rAAV5-siHUNT-1 injection (A), rAAV5-siHUNT-2 long-term expression led to a 75% reduction of striatal ppENK levels in the injected striatum of wild-type littermate controls (Fisher’s PLSD post hoc test, *P < 0.0001). Likewise, rAAV5-siHUNT-2 treatment also resulted in a further 16% reduction in striatal ppENK levels in R6/1 mice (Fisher’s PLSD post hoc test, †P < 0.05). (D) Analysis of the effect of intrastriatal injection of rAAV-siHUNT-2 on striatal DARPP-32 mRNA levels. Similar to the pattern seen in response to rAAV5-siHUNT-2 expression striatal ppEnk levels (C), rAAV5-siHUNT-2 long-term expression led to a 44% reduction of striatal DARPP-32 mRNA levels in the injected striatum of wild-type littermate controls (Fisher’s PLSD post hoc test, *P = 0.0005). In addition, rAAV5-siHUNT-2 treatment resulted in a further 70% reduction in striatal DARPP-32 mRNA levels in R6/1 mice (Fisher’s PLSD post hoc test, †P < 0.01). This further reduction in the treated striatum of the R6/1 mice is particularly apparent in the right side of the coronal section shown to the left of the histogram. (E) Evaluation of the effect of the long-term expression of the control shRNA, siRho, in the striatum of wild-type littermate controls and R6/1 mice on striatal ppEnk levels. rAAV5-siRho injections had no effect on striatal ppEnk transcript levels in either wild-type controls (Fisher’s PLSD post hoc test, P > 0.9) or R6/1 mice (Fisher’s PLSD post hoc test, P > 0.6). (F) Evaluation of the effect of the long-term expression of the control shRNA, siRho, in the striatum of wild-type littermate controls and R6/1 mice on striatal DARPP-32 mRNA levels. rAAV5-siRho injections had no effect on striatal DARPP-32 mRNA levels in either wild-type controls (Fisher’s PLSD post hoc test, P > 0.7) or R6/1 mice (Fisher’s PLSD post hoc test, P > 0.6). Error bars are +SEM.