Kv1.3 modulates neuroinflammation and neurodegeneration in Parkinson’s disease
Souvarish Sarkar, … , Heike Wulff, Anumantha G. Kanthasamy
Souvarish Sarkar, … , Heike Wulff, Anumantha G. Kanthasamy
Published June 29, 2020
Citation Information: J Clin Invest. 2020;130(8):4195-4212. https://doi.org/10.1172/JCI136174.
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Research Article Inflammation Neuroscience

Kv1.3 modulates neuroinflammation and neurodegeneration in Parkinson’s disease

Abstract

Characterization of the key cellular targets contributing to sustained microglial activation in neurodegenerative diseases, including Parkinson’s disease (PD), and optimal modulation of these targets can provide potential treatments to halt disease progression. Here, we demonstrated that microglial Kv1.3, a voltage-gated potassium channel, was transcriptionally upregulated in response to aggregated α-synuclein (αSynAgg) stimulation in primary microglial cultures and animal models of PD, as well as in postmortem human PD brains. Patch-clamp electrophysiological studies confirmed that the observed Kv1.3 upregulation translated to increased Kv1.3 channel activity. The kinase Fyn, a risk factor for PD, modulated transcriptional upregulation and posttranslational modification of microglial Kv1.3. Multiple state-of-the-art analyses, including Duolink proximity ligation assay imaging, revealed that Fyn directly bound to Kv1.3 and posttranslationally modified its channel activity. Furthermore, we demonstrated the functional relevance of Kv1.3 in augmenting the neuroinflammatory response by using Kv1.3-KO primary microglia and the Kv1.3-specific small-molecule inhibitor PAP-1, thus highlighting the importance of Kv1.3 in neuroinflammation. Administration of PAP-1 significantly inhibited neurodegeneration and neuroinflammation in multiple animal models of PD. Collectively, our results imply that Fyn-dependent regulation of Kv1.3 channels plays an obligatory role in accentuating the neuroinflammatory response in PD and identify Kv1.3 as a potential therapeutic target for PD.

Authors

Souvarish Sarkar, Hai M. Nguyen, Emir Malovic, Jie Luo, Monica Langley, Bharathi N. Palanisamy, Neeraj Singh, Sireesha Manne, Matthew Neal, Michelle Gabrielle, Ahmed Abdalla, Poojya Anantharam, Dharmin Rokad, Nikhil Panicker, Vikrant Singh, Muhammet Ay, Adhithiya Charli, Dilshan Harischandra, Lee-Way Jin, Huajun Jin, Srikant Rangaraju, Vellareddy Anantharam, Heike Wulff, Anumantha G. Kanthasamy

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Figure 5

Fyn modulates the posttranslational modification of Kv1.3. (A) Western blot analysis of postmortem human PD and age-matched control brains showing increased phosphorylation of Kv1.3. (B) Immunoprecipitation of Fyn and Kv1.3 showing direct Fyn-Kv1.3 interaction after αSynAgg treatment.

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Fyn modulates the posttranslational modification of Kv1.3. (A) Western b...
(C) Duolink PLA showing αSynAgg-induced interaction between Kv1.3 and Fyn. Scale bar: 25 μm. (D) Western blot of Fyn WT and KO PMCs revealed that Kv1.3 phosphorylation at residue 135 was Fyn dependent. (E) IHC analysis of substantia nigra from Fyn+/+ and Fyn–/– mice showing reduced phosphorylation of Kv1.3 after αSynPFF injection. Scale bars: 100 μm; 60 μm (insets). (F) IHC of substantia nigra from MitoPark mice and their littermate controls showing that pharmacological inhibition of Fyn by saracatinib reduced Kv1.3 phosphorylation. Scale bar: 100 μm. (G–J) Immortalized MMCs were either transfected with WT Kv1.3 or aY135A Kv1.3 plasmid. (G) qRT-PCR analysis and (H) Griess assay showing reduced levels of inducible NOS (iNOS) and nitrite release, respectively, in Y135A Kv1.3-transfected cells compared with WT cells. (I) qRT-PCR analysis showing reduced IL-1β production in Y135A Kv1.3–transfected versus WT Kv1.3–transfected MMCs. (J) Luminex assay showing reduced IL-1β secretion in Y135A Kv1.3–transfected compared with WT Kv1.3–transfected MMCs. A 1-way ANOVA was used to compare multiple groups. In most cases, Tukey’s post hoc analysis was applied. A 2-tailed Student’s t test was used to compare 2 groups in A. Each dot on the bar graphs represents a biological replicate. Data are presented as the mean ± SEM, with 3–4 biological replicates from 2–3 independent experiments. *P ≤ 0.05, **P < 0.01, and ***P < 0.001.