Mutant huntingtin enhances activation of dendritic Kv4 K+ channels in striatal spiny projection neurons
- Northwestern University, United States
- National Autonomous University of Mexico, Mexico
- Stony Brook University School of Medicine, United States
- Washington University School of Medicine, United States
- Weill Cornell Medical College, United States
Figures
Figure 1 with 2 supplements
Distal dendritic excitability was reduced in iSPNs from HD models.
(a) Characterization of Ca2+ transients induced by bAPs in dendritic spines of SPNs. Left: Maximum intensity Z projection images of an iSPN filled with Alexa Fluor 568 (wild-type mouse >2 months; …
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Figure 1—source data 1
Source data for Figure 1.
- https://doi.org/10.7554/eLife.40818.007
Figure 1—figure supplement 1
bAP-evoked distal but not proximal dendritic fluorescence measurements were diminished in BACHD iSPNs.
Left, boxplot of peak fluorescence measurements (G/R) in proximal dendrites of BACHD and WT iSPNs (p=1.0, Mann-Whitney U, Two-Tailed; n = 9); right, boxplot of peak fluorescent measurements in …
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Figure 1—figure supplement 1—source data 1
Source data for Figure 1—figure supplement 1.
- https://doi.org/10.7554/eLife.40818.004
Figure 1—figure supplement 2
Expression of Cav3 mRNAs was not significantly altered in HD iSPNs.
(a FACS-qPCR was performed to determine transcript expression in Q175 ±iSPNs relative to their WT controls. Data are presented as ΔCT. A modest elevation in Cav3.1 expression (lower ΔCT) was …
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Figure 1—figure supplement 2—source data 1
Source data for Figure 1—figure supplement 2.
- https://doi.org/10.7554/eLife.40818.006
Figure 2 with 3 supplements
Upregulation of Kv4 channel activity suppressed dendritic excitability in HD iSPNs.
(a) Viral delivery of Kv4.2 shRNA into the striatum of BACHD mice. Z projection images of an iSPN filled with Alexa Fluor 568 and transfected with Kv4.2 shRNA for 4 weeks. Infected neurons are …
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Figure 2—source data 1
Source data for Figure 2.
- https://doi.org/10.7554/eLife.40818.014
Figure 2—figure supplement 1
AAV Kv4.2-shRNA mediated knockdown in striatum increased iSPN excitability.
(a) Confocal images of a coronal brain section 4 weeks after stereotaxic delivery of AAV Kv4.2 shRNA-eGFP expression construct to the striatum. DAPI was used to stain nuclei. (b) The knockdown …
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Figure 2—figure supplement 1—source data 1
Source data for Figure 2—figure supplement 1.
- https://doi.org/10.7554/eLife.40818.010
Figure 2—figure supplement 2
4-AP (2 mM) increased EPSP amplitude only when 5–10 spines were stimulated.
(a) MNI-Glu was uncaged directly adjacent to individual spine heads (small spot, red dots) or by optogenetically stimulating release of Glu onto a group of 5–10 spines (large spot, blue dots) before …
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Figure 2—figure supplement 2—source data 1
Source data for Figure 2—figure supplement 2.
- https://doi.org/10.7554/eLife.40818.012
Figure 2—figure supplement 3
Computation of the Kv4-index.
Schematic showing how the Kv4 index was computed. The amplitude of the EPSP before (A0) and after (A1) increasing 4-AP concentration was measured. The ratio (A0/A1) was subtracted from one to get …
Figure 3 with 4 supplements
Functional upregulation of Kv4 channels depended upon KChIPs.
(a) Ca2+ transients in response to bAPs (arrows) at proximal and distal dendritic spines in iSPNs from BACHD mice with the internal perfusion of KChIP2 antibody (1:50); perfusion rescued dendritic …
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Figure 3—source data 1
Source data for Figure 3.
- https://doi.org/10.7554/eLife.40818.023
Figure 3—figure supplement 1
Expression of Kv4 and KChIP mRNAs was unchanged in HD iSPNs.
FACS-qPCR was performed to isolate and profile iSPNs. Data are presented as boxplots of ΔCT where smaller numbers reflect higher transcript abundance (see Materials and methods). (a) Boxplots of ΔCT …
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Figure 3—figure supplement 1—source data 1
Source data for Figure 3—figure supplement 1.
- https://doi.org/10.7554/eLife.40818.017
Figure 3—figure supplement 2
Colocalization of Kv 4.2 immunoreactivity and sparse-labeled ISPN dendrites.
(a) Red labeled ISPNs after striatal injection of AAV-EF1-Flex-tdTomato in the wild type and (b) Q175 mouse. Middle panels show Kv 4.2 immunoreactivity in the striatum and right panels are overlays …
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Figure 3—figure supplement 2—source data 1
Source data for Figure 3—figure supplement 2.
- https://doi.org/10.7554/eLife.40818.019
Figure 3—figure supplement 3
Pan-KChIP antibody normalized HD iSPN dendritic excitability.
Boxplot of the dendritic index in wild-type iSPN dendrites and BACHD iSPN dendrites after dialysis with a pan-KChIP antibody (p=0.49, Mann-Whitney U, Two tailed; n = 14 control, n = 4 BACHD). …
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Figure 3—figure supplement 3—source data 1
Source data for Figure 3—figure supplement 3.
- https://doi.org/10.7554/eLife.40818.021
Figure 3—figure supplement 4
Cav1.3 channel was not associated with Kv4.2.
Kv4.2 antibody was unable to pulldown Cav1.3 in mouse striatum lysate.
Figure 4 with 1 supplement
TrkBR signaling regulated Kv4 channel association with KChIPs.
(a) KChIPs co-immunoprecipitated with Kv4.2 in mice striata, KChIP1 and KChIP2 more robustly associated with Kv4.2 than KChIP3/4. To the right is a schematic of the Kv4.2/KChIP/Cav3.2 membrane …
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Figure 4—source data 1
Source data for Figure 4.
- https://doi.org/10.7554/eLife.40818.027
Figure 4—figure supplement 1
The co-immunoprecipitation experiments detected specific proteins.
(a) Kv4.2 was pulled-down by KChIP antibody, further verification of Kv4.2-KChIP interaction in mouse striatum. (b) Other similar Kv channels such as Kv1.2 did not interact with KChIPs, as Kv1.2 …
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Figure 4—figure supplement 1—source data 1
Source data for Figure 4—figure supplement 1.
- https://doi.org/10.7554/eLife.40818.026
Figure 5
Enabling TrkBR signaling restored dendritic excitability.
(a) Ca2+ transients evoked by bAPs (arrows) at proximal and (b) distal dendritic spines in iSPNs from WT mouse in the presence of BDNF (40 ng/ml). (c) Dendritic index was significantly increased …
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Figure 5—source data 1
Source data for Figure 5.
- https://doi.org/10.7554/eLife.40818.029
Figure 6 with 1 supplement
Suppression of mHtt normalized iSPN excitability.
(a) ZFP vector map 1646 bp (EcoR1/HindIII) inserts from pVAX-30645 (Sigma) sub-cloned into pAAV-CMV-SV40-WPRE vector and packaged by Virovek. The expression of N-term NLS and C-term FLAG tagged …
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Figure 6—source data 1
Source data for Figure 6.
- https://doi.org/10.7554/eLife.40818.033
Figure 6—figure supplement 1
ZFP-30645 construct selectively suppressed mHtt transcription in fibroblasts.
A striatal derived cell line ST HDH Q7/111 (Coriell, CH00096 1) from a knock-in transgenic mouse containing heterozygous Huntingtin (Htt) loci with a humanized Exon 1 with 111 polyglutamine repeats …
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Figure 6—figure supplement 1—source data 1
Source data for Figure 6—figure supplement 1.
- https://doi.org/10.7554/eLife.40818.032
Figure 7
Suppression of mHtt restored TrkB receptor-mediated spine LTP in distal iSPN dendrites.
(a) Maximum projection image of ZFP +Q175±iSPN with a high magnification image of a distal dendrite where 2PLSM spot uncaging of MNI-Glu was conducted (uEPSCs, yellow dots). (b) Schematic of the LTP …
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Figure 7—source data 1
Source data for Figure 7.
- https://doi.org/10.7554/eLife.40818.035
Additional files
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Transparent reporting form
- https://doi.org/10.7554/eLife.40818.036
