kv1 3 (Alomone Labs)


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Kv1 3, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Average 93 stars, based on 1 article reviews
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1) Product Images from "Kv1.3 activity perturbs the homeostatic properties of astrocytes in glioma"
Article Title: Kv1.3 activity perturbs the homeostatic properties of astrocytes in glioma
Journal: Scientific Reports
doi: 10.1038/s41598-018-25940-5

Figure Legend Snippet: Kv1.3 channel activity controls tight junction protein expression on bEnd.3. ( a ) Immunofluorescence analysis of Kv1.3 (in red, Hoechst in blue) on endothelial bEnd.3 cells. On the right, cells stained only with secondary Ab as control. ( b ) Astrocytes were co-cultured with bEnd.3 in the absence (−GL261) or presence (+GL261) of GL261 cells (as depicted in the inset), and treated or not with PAP-1 (50 nM). Trans-endothelial electric resistance (TEER, in Ωcm2) was measured at the indicated time points. ( c ) bEnd.3 in the absence (−GL261) or presence (+GL261) of GL261 cells (as depicted in the inset), and treated or not with PAP-1 (50 nM) were assayed for TEER (in Ωcm 2 ) at the indicated time points. ( d ) RT-PCR gene expression of claudin-5, occludin and zo-1 in untreated (C) or PAP-1 (50 nM) treated bEnd.3 co-cultured or not with astrocytes. Data are expressed as fold increase in co-cultures vs bEnd.3 alone (no astrocytes) and are the mean ± s.e.m., n = 4, *p = 0.001, Dunn’s method One Way ANOVA.
Techniques Used: Activity Assay, Expressing, Immunofluorescence, Staining, Cell Culture, Reverse Transcription Polymerase Chain Reaction

Figure Legend Snippet: Kv1.3 activity modulates microglia functions. ( a – b ) Non-conditioned medium (NCM)- and glioma conditioned medium (GCM)-treated microglia, in the absence (C) or presence of PAP-1 (50 nM) assayed for phagocytosis ( a ) and migration ( b ). Data are expressed as the % of phagocytosing ( a ) and migrated ( b ) cells ± s.e.m. *p = 0.001vs NCM; n = 4, Kruskal-Wallis One Way ANOVA on Ranks. ( c ) Coronal brain sections of GL261-bearing mice treated with PAP-1 (40 mg/kg/die) or vehicle were stained for Iba1 (red) and Hoechst (blue), scale bar 20 µm. On the right, % of Iba1 + cell area normalized for tumor area, *p = 0.002, unpaired t -test, n = 6. ( d , e ) RT-PCR for pro- ( cd86, tnfα, il1α, il15 ) and anti- ( arg1, ym1, cd163, cd206 ) inflammatory genes expressed by CD11b + cells extracted from ipsilateral hemisphere of brains of GL261-bearing mice treated with vehicle (C) or PAP-1 (40 mg/kg/die). Data are expressed as fold change of PAP-1-treated vs vehicle-treated samples (C, dashed lines) and are the mean ± s.e.m., *p
Techniques Used: Activity Assay, Migration, Mouse Assay, Staining, Reverse Transcription Polymerase Chain Reaction

Figure Legend Snippet: The inhibition of Kv1.3 channels induces neuroprotection against the toxic effects of glioma. ( a ) Cortical neurons (CN) co-cultured with GL261 cells (grey bars) or alone (black bars) were treated with PAP-1 (50 nM, 18 h) or vehicle (C) and analyzed for neuronal viability. Results are expressed as number of viable cells/field. *p = 0.001 vs C; n = 4, unpaired t -test. ( b ) Hippocampal neurons (HN) pre-treated or not with empty or clodronate-filled liposomes for 24 h, co-cultured as in ( a ) for a further 18 h in presence of PAP-1 (50 nM) or vehicle (C), were analyzed for neuronal viability. Results are expressed as number of viable cells/field. **p = 0.001 and *p
Techniques Used: Inhibition, Cell Culture

Figure Legend Snippet: Kv1.3 is expressed by glioma cells and modulates their migration. ( a ) Typical current traces in response to repeated voltage ramps from −120 to +50 mV (holding potential −70 mV) in Ctrl and PAP-1 (100 nM) treated GL261 cells. ( b ) Bar graph representing PAP-1 sensitive current amplitude in GL261 cells, n = 14, *p = 0.01, t -test. ( c ) Migration assay on untreated (Ctrl) and PAP-1 (50 nM, 4 h) treated GL261, GL-15 and GBM18 cells; data are the mean ± s.e.m., n = 4, *p = 0.001, # p = 0.05, @ p = 0.001, unpaired t -test.
Techniques Used: Migration
![Kv1.3 activity modulates glutamate buffering on astrocytes. ( a ... Kv1.3 activity modulates glutamate buffering on astrocytes. ( a ) Time course of fluorescence ratio (ΔF/F0) changes induced by a puff of glutamate (1 mM for 0.5 sec, Glut puff ) onto astrocytic cultures loaded with BCECF-AM (10 μM, 45 min) and pre-treated with vehicle (n = 71) or PAP-1 (100 nM, n = 69). At peak ΔF/F0 in PAP1 = −0.08 ± 0.008 vs −0.046 ± 0.014 in Ctrl p = 0.0009, unpaired Student’s t -test). (b) Astrocytes were treated with PAP-1 (50 nM, grey circles) or not (black circles) with or without DHK (500 μM, triangles) for different times (from 2 to 45 min) and analyzed for intracellular D-[ 3 H]Asp, as described in the Methods section. Results are expressed as pCi/μg proteins and are the mean ± s.e.m. of at least 5 triplicate experiments. *p = 0.001 vs C of the correspondent time point, Holm-Sidak method One Way ANOVA. ( c ) Confocal images of astrocytes, untreated (C) or treated with PAP-1 (50 nM, 25 min), stained for plasma membrane GLT-1 (red, Hoechst in blue), scale bar 10 μm. On the right, data represent the mean fluorescence intensity of red signals per field ± s.e.m. n = 4, *p = 0.042 vs C, unpaired t -test. ( d ) Astrocytes untreated (−) or treated (+) with PAP-1 (50 nM, 25 min) were immunoprecipitated for Sumo-1 or control IgG and immmuno-blotted for GLT-1; total lysate (input) is shown. On the right, data represent the mean ± s.e.m. of optical density of sumoylated GLT-1 expressed as % of the input, *p = 0.028 vs C, unpaired t -test. ( e ) Representative time course of spontaneous Ca 2+ oscillation (F/F0) in cultured astrocytes loaded with Fluo4-AM in CTRL (left panel) and after PAP-1 application (right panel). Each trace in the panel represent a single ROI in the field. ( f ) Quantification of Ca 2+ transients before and after PAP-1 treatment. ( g ) Average ΔF/F0 of Ca 2+ transient in CTRL condition and after PAP-1 application. ( h ) Coronal brain sections of GL261-bearing mice treated with vehicle or PAP-1 (40 mg/kg/die) were stained for GFAP (green; Hoechst, in blue) and visualized at the border of the tumor (white dashed line), scale bar 20 μm. Right, data represent the mean area (in pixels) covered by GFAP + cells present at a distance up to 100 μm from the tumor border (mean ± s.e.m. n = 6, *p = 0.015, unpaired t -test).](https://storage.googleapis.com/bioz_article_images/PMC5955950/41598_2018_25940_Fig2_HTML.jpg)
Figure Legend Snippet: Kv1.3 activity modulates glutamate buffering on astrocytes. ( a ) Time course of fluorescence ratio (ΔF/F0) changes induced by a puff of glutamate (1 mM for 0.5 sec, Glut puff ) onto astrocytic cultures loaded with BCECF-AM (10 μM, 45 min) and pre-treated with vehicle (n = 71) or PAP-1 (100 nM, n = 69). At peak ΔF/F0 in PAP1 = −0.08 ± 0.008 vs −0.046 ± 0.014 in Ctrl p = 0.0009, unpaired Student’s t -test). (b) Astrocytes were treated with PAP-1 (50 nM, grey circles) or not (black circles) with or without DHK (500 μM, triangles) for different times (from 2 to 45 min) and analyzed for intracellular D-[ 3 H]Asp, as described in the Methods section. Results are expressed as pCi/μg proteins and are the mean ± s.e.m. of at least 5 triplicate experiments. *p = 0.001 vs C of the correspondent time point, Holm-Sidak method One Way ANOVA. ( c ) Confocal images of astrocytes, untreated (C) or treated with PAP-1 (50 nM, 25 min), stained for plasma membrane GLT-1 (red, Hoechst in blue), scale bar 10 μm. On the right, data represent the mean fluorescence intensity of red signals per field ± s.e.m. n = 4, *p = 0.042 vs C, unpaired t -test. ( d ) Astrocytes untreated (−) or treated (+) with PAP-1 (50 nM, 25 min) were immunoprecipitated for Sumo-1 or control IgG and immmuno-blotted for GLT-1; total lysate (input) is shown. On the right, data represent the mean ± s.e.m. of optical density of sumoylated GLT-1 expressed as % of the input, *p = 0.028 vs C, unpaired t -test. ( e ) Representative time course of spontaneous Ca 2+ oscillation (F/F0) in cultured astrocytes loaded with Fluo4-AM in CTRL (left panel) and after PAP-1 application (right panel). Each trace in the panel represent a single ROI in the field. ( f ) Quantification of Ca 2+ transients before and after PAP-1 treatment. ( g ) Average ΔF/F0 of Ca 2+ transient in CTRL condition and after PAP-1 application. ( h ) Coronal brain sections of GL261-bearing mice treated with vehicle or PAP-1 (40 mg/kg/die) were stained for GFAP (green; Hoechst, in blue) and visualized at the border of the tumor (white dashed line), scale bar 20 μm. Right, data represent the mean area (in pixels) covered by GFAP + cells present at a distance up to 100 μm from the tumor border (mean ± s.e.m. n = 6, *p = 0.015, unpaired t -test).
Techniques Used: Activity Assay, Fluorescence, Size-exclusion Chromatography, Staining, Immunoprecipitation, Cell Culture, Mouse Assay
2) Product Images from "Targeting Kv1.3 channels to reduce white matter pathology after traumatic brain injury"
Article Title: Targeting Kv1.3 channels to reduce white matter pathology after traumatic brain injury
Journal: Experimental neurology
doi: 10.1016/j.expneurol.2016.06.011

Figure Legend Snippet: Callosal Kv1.3 channel protein in axons and glia is altered after injury and with CFZ treatment A. Confocal overlay showing Kv1.3 ( red ) and Kv1.2 ( green ) in rat corpus callosum 24h following midline fluid percussion TBI. Low magnification shows that each channel protein is found in reactive glia around callosal vessels (arrowheads) and along axon bundles (arrows). Inset shows paired paranodal distribution of Kv1.3 and Kv1.2 channels, some nodes with co-localization (yellow arrow), others with single channel expression (green, red arrows). B . Confocal overlays showing Kv1.3 in callosal astrocytes of sham injured (GFAP+, left-arrows; inset shows cell body and perivascular co-localization) and microglia of 24h postinjury cases (IBA1+, right-arrows). C. Western blot (WB) of protein extracts from 24h postinjury corpus callosum revealed that TBI reduced 67kD Kv1.3 levels and that CFZ treatment reversed loss of Kv1.3 expression. Data expressed as percent of paired untreated sham controls run on same blot. Lanes representative of group effects are shown in each panel. (ANOVA, *p
Techniques Used: Expressing, Western Blot

Figure Legend Snippet: Corpus callosum mixed glial cultures grown in MatTek dishes and subjected to confocal dual labeling with antibodies to Kv1.3 (green) and microglial marker protein IBA1 (red) or astrocyte marker protein GFAP (red) A. Microglia are predominantly ramified in untreated cultures (arrowhead), shifting to reactive rounded cells with lobular processes after LPS (arrows). Kv1.3 protein is localized within the majority of microglia in each field and shifts from a normal concentration around nuclei (inset, UNT), to a more uniform distribution after inflammatory stimulation (inset, LPS). B. Astrocytes show mixed flat and fibrous morphologies. Kv1.3 signal is much reduced in astrocytes relative to surrounding microglia (not stained in these images) and predominantly found in small aggregates around cell nuclei (arrow). Astrocytes do not show LPS group differences in Kv1.3 expression. Bars= 20 µm.
Techniques Used: Labeling, Marker, Concentration Assay, Staining, Expressing
3) Product Images from "Kv1.3 modulates neuroinflammation and neurodegeneration in Parkinson’s disease"
Article Title: Kv1.3 modulates neuroinflammation and neurodegeneration in Parkinson’s disease
Journal: The Journal of Clinical Investigation
doi: 10.1172/JCI136174

Figure Legend Snippet: 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 αSyn Agg treatment. ( C ) Duolink PLA showing αSyn Agg -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 αSyn PFF 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
Techniques Used: Modification, Western Blot, Immunoprecipitation, Proximity Ligation Assay, Immunohistochemistry, Mouse Assay, Injection, Inhibition, Transfection, Plasmid Preparation, Quantitative RT-PCR, Griess Assay, Luminex

Figure Legend Snippet: Kv1.3 inhibition protects against αSyn PFF -induced behavior deficit and dopaminergic neuronal loss. ( A ) Treatment paradigm corresponding to the αSyn PFF mouse model of PD. ( B ) Representative movement tracks showing that PAP-1 rescued movement deficits induced by αSyn PFF . ( C – E ) A VersaMax open-field test showed decreased ( C ) rest time and increased ( D ) horizontal activity and ( E ) total distance traveled for αSyn PFF mice treated with PAP-1. ( F and G ) HPLC showing that PAP-1 treatment protected against loss of ( F ) dopamine and ( G ) DOPAC induced by αSyn PFF . ( H ) Western blot analysis of TH showing loss of TH induced by αSyn PFF in the SNpc region. A 1-way ANOVA was used to compare multiple groups. In most cases, Tukey’s post hoc analysis was applied. Each dot on the bar graphs represents a biological replicate. Data are presented as the mean ± SEM, with 3–7 animals per group. * P ≤ 0.05, ** P
Techniques Used: Inhibition, Activity Assay, Mouse Assay, High Performance Liquid Chromatography, Western Blot

Figure Legend Snippet: Upregulated expression of the potassium channel Kv1.3 upon aggregated αSyn stimulation in ex vivo slices and B cells derived from patients with PD. ( A ) Midbrain slice cultures were treated with 1 μM αSyn Agg for 24 hours. qRT-PCR shows upregulated Kv1.3 mRNA expression. ( B ) Western blot shows upregulated Kv1.3 protein level in midbrain slice cultures treated with 1 μM αSyn Agg for 24 hours. ( C ) qRT-PCR of midbrain slice cultures treated with 1 μM αSyn Agg for 24 hours, revealing upregulation of the proinflammatory factors Nos2 , Csf2 , IL-6 , IL-1β , and Tnfa . ( D ) qRT-PCR shows increased Kv1.3 mRNA expression in B cell lymphocytes isolated from patients with PD compared with expression in B cell lymphocytes from age-matched controls. ( E ) Whole-cell patch clamping of B cell lymphocytes isolated from patients with PD showed higher Kv1.3 channel activity compared with that observed in age-matched controls ( n = 3 control and n = 3 PD). A 1-way ANOVA was used to compare multiple groups in C and D . Tukey’s post hoc analysis was applied. A 2-tailed Student’s t test was used to compare 2 groups. Each dot on the bar graphs represents a biological replicate. Data are presented as the mean ± SEM, with 3–7 biological replicates from 2–3 independent experiments unless otherwise indicated. * P ≤ 0.05 and ** P
Techniques Used: Expressing, Ex Vivo, Derivative Assay, Quantitative RT-PCR, Western Blot, Isolation, Activity Assay

Figure Legend Snippet: Kv1.3 expression is highly induced in microglial cells in experimental models of PD and postmortem PD brains. ( A ) Western blot showing increased Kv1.3 protein levels in the substantia nigra of the Syn-AAV mouse model of PD. ( B ) qRT-PCR analysis of 8- to 24-week-old nigral tissues from the MitoPark mouse model of PD showing Kv1.3 induction compared with age-matched littermate controls. ( C ) Western blot of 24-week-old nigral tissues from the MitoPark mouse model of PD (MP) showing induction of Kv1.3 protein expression compared with age-matched littermate control mice (LM). ( D ) IHC in 24-week-old nigral tissues from the MitoPark mouse model of PD showing higher Kv1.3 protein levels (red) in IBA1-positive microglial cells (green) compared with age-matched controls as revealed by their colocalization (yellow). Scale bar: 20 μm. ( E ) qRT-PCR analysis of nigral tissues from the MPTP mouse model revealing induction of Kv1.3 mRNA expression. ( F ) Western blot showing increased Kv1.3 protein levels in substantia nigra of the MPTP mouse model of PD. ( G ) qRT-PCR analysis of postmortem human PD brains showing elevated Kv1.3 mRNA expression. ( H ) Western blot of the SN region of postmortem human PD brain showing induction of Kv1.3 protein expression compared with age-matched controls. n = 6–8. ( I ) Immunostaining revealing higher Kv1.3 levels in the prefrontal cortex of postmortem human PD brains compared with age-matched controls. Lower panel shows the deconvoluted binary image used for analysis. Three regions per brain were analyzed. Scale bar: 200 μm. ( J ) Dual DAB staining showing induction of Kv1.3 expression in HLA-DR–positive microglial cells in patients with DLBs compared with age-matched controls. Scale bars: 100 μm; 20 μm (enlarged insets). A 1-way ANOVA was used to compare multiple groups. Tukey’s post hoc analysis was applied B . A 2-tailed Student’s t test was used to compare 2 groups. Each dot on the bar graphs represents a biological replicate. Data are presented as the mean ± SEM, with 3–9 biological replicates from 2–3 independent experiments unless otherwise indicated. * P ≤ 0.05, ** P
Techniques Used: Expressing, Western Blot, Quantitative RT-PCR, Mouse Assay, Immunohistochemistry, Immunostaining, Staining

Figure Legend Snippet: Fyn modulates the transcriptional regulation of Kv1.3 in microglial cells through the Fyn/PKCδ kinase signaling cascade. ( A ) In silico analysis of the promoter sequence of Kv1.3 revealed probable Nf-κB– and SP1-binding sites. ( B ) qRT-PCR analysis of immortalized MMCs cotreated with αSyn Agg and either SN50 (100 μg/mL) or SB203580 (1 μM), showing that both compounds attenuated αSyn Agg -induced Kv1.3 expression. ( C ) Western blot of Fyn WT and KO PMCs treated with αSyn Agg , showing that Fyn KO reduced the induction of the p38 MAPK pathway. ( D ) qRT-PCR analysis revealed that Fyn KO reduced αSyn Agg -induced Kv1.3 mRNA levels. ( E ) Whole-cell patch-clamp recording showing that Fyn KO attenuated αSyn Agg - and LPS-induced Kv1.3 activity compared with Fyn WT PMCs (WT control n = 24, WT αSyn Agg n = 12, WT LPS n = 29, Fyn KO αSyn Agg n = 20, Fyn KO LPS n = 15). ( F ) ICC showing that Fyn KO reduced αSyn Agg -induced Kv1.3 protein levels in PMCs. Scale bar: 15 μm. ( G ) ICC of PMCs revealed that αSyn Agg -induced Kv1.3 protein expression was reduced by PKCδ KO. Scale bar: 15 μm. ( H ) qRT-PCR analysis of PMCs showing that PKC KO reduced the expression of αSyn Agg -induced Kv1.3 mRNA. ( I ) Whole-cell patch clam recording of PMCs showing that PKC KO attenuated αSyn Agg - and LPS-induced Kv1.3 activity compared with PKC WT PMCs (WT control n = 24, WT αSyn Agg n = 12, WT LPS n = 20, PKC-KO αSyn Agg n = 29, PKC-KO LPS n = 35). Data are presented as the mean ± SD. A 1-way ANOVA was used to compare multiple groups. In most cases, Tukey’s post hoc analysis was applied. 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 unless otherwise indicated. * P ≤ 0.05, ** P
Techniques Used: In Silico, Sequencing, Binding Assay, Quantitative RT-PCR, Expressing, Western Blot, Patch Clamp, Activity Assay, Immunocytochemistry

Figure Legend Snippet: Kv1.3 modulates neuroinflammation in a cell culture model of PD. ( A – C ) Kv1.3 WT and KO PMCs were treated with 1 μM αSyn Agg for 24 hours. Luminex analysis shows that Kv1.3 KO reduced the release of the αSyn Agg -induced proinflammatory factors ( A ) TNF-α, ( B ) IL-12, and ( C ) IL-1β. ( D – H ) Immortalized MMCs were transfected with WT a Kv1.3 plasmid, and then 48 hours after transfection, cells were treated with 1 μM αSyn Agg for 24 hours. ( D – F ) qRT-PCR analysis showing that Kv1.3 overexpression aggravated αSyn Agg -induced production of the proinflammatory factors ( D ) Nos2 , ( E ) pro– IL-1β , and ( F ) TNF-α . ( G and H ) Luminex analysis showing that Kv1.3 overexpression potentiated the release of the proinflammatory factors ( G ) IL-6 and ( H ) IL-12. ( I ) Voltage ramp from –120 mV to 40 mV elicited a characteristic outward rectifying current in αSyn Agg -treated microglia that was sensitive to the Kv1.3-selective inhibitor PAP-1. ( J ) LDH assay showing that PAP-1 reduced αSyn Agg -induced LDH release from microglial cells. ( K – M ) Luminex assay revealing that PAP-1 attenuated the αSyn Agg -induced proinflammatory factors ( K ) IL-12, ( L ) TNF-α, and ( M ) IL-6. ( N ) Western blot analysis demonstrating that PAP-1 reduced αSyn Agg -induced NLRP3 expression. ( O ) ICC analysis revealed that PAP-1 reduced NLRP3 expression induced by αSyn Agg . Scale bar: 25 μm. A 1-way ANOVA was performed to compare multiple groups. In most cases, Tukey’s post hoc analysis was applied. 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
Techniques Used: Cell Culture, Luminex, Transfection, Plasmid Preparation, Quantitative RT-PCR, Over Expression, Lactate Dehydrogenase Assay, Western Blot, Expressing, Immunocytochemistry

Figure Legend Snippet: Upregulated expression of the potassium channel Kv1.3 upon aggregated αSyn stimulation in microglial cells in vitro. ( A ) Whole-cell patch-clamp recordings of PMCs treated with 1 μM αSyn Agg for 24–48 hours, showing αSyn Agg -induced increased Kv1.3 activity (control n = 24 and αSyn Agg n = 12). Kv1.3 was identified by its characteristic use dependence, which was revealed when applying a train of ten 200-ms pulses from –80 to 40 mV at 1-second intervals (1 Hz). ( B ) qRT-PCR showing that αSyn Agg induced Kv1.3 mRNA expression without significantly altering other potassium channels. ( C ) Western blot of αSyn Agg -induced Kv1.3 protein expression in PMCs. ( D ) ICC of αSyn Agg -induced Kv1.3 protein expression in PMCs. Scale bar: 100 μm. ( E ) Flow cytometric analysis of immortalized MMCs treated with 1 μM αSyn Agg for 24 hours, showing αSyn Agg -induced Kv1.3 surface expression. ( F ) qRT-PCR of human microglia treated with LPS (1 μg/mL) and IL-4 (20 ng/mL) for 6 hours, showing LPS-induced Kv1.3 expression. A 1-way ANOVA was used to compare multiple groups. In most cases, Tukey’s post hoc analysis was applied in B and F . A 2-tailed Student’s t test was used for all other figures when comparing 2 groups. Each dot on the bar graphs represents a biological replicate. Data are presented as the mean ± SEM, with 3–5 biological replicates from 2–3 independent experiments unless otherwise noted. * P ≤ 0.05, ** P
Techniques Used: Expressing, In Vitro, Patch Clamp, Activity Assay, Quantitative RT-PCR, Western Blot, Immunocytochemistry