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Differential Kv1.3, KCa3.1, and Kir2.1 expression in "classically" and "alternatively" activated microglia
Hai M Nguyen1, Eva M Grössinger1, Makoto Horiuchi2,3
1Department of Pharmacology, University of California, Davis, California.
Abstract:
Microglia are highly plastic cells that can assume different phenotypes in response to microenvironmental signals. Lipopolysaccharide (LPS) and interferon-γ (IFN-γ) promote differentiation into classically activated M1-like microglia, which produce high levels of pro-inflammatory cytokines and nitric oxide and are thought to contribute to neurological damage in ischemic stroke and Alzheimer's disease. IL-4 in contrast induces a phenotype associated with anti-inflammatory effects and tissue repair. We here investigated whether these microglia subsets vary in their K+ channel expression by differentiating neonatal mouse microglia into M(LPS) and M(IL-4) microglia and studying their K+ channel expression by whole-cell patch-clamp, quantitative PCR and immunohistochemistry. We identified three major types of K+ channels based on their biophysical and pharmacological fingerprints: a use-dependent, outwardly rectifying current sensitive to the KV 1.3 blockers PAP-1 and ShK-186, an inwardly rectifying Ba2+ -sensitive Kir 2.1 current, and a Ca2+ -activated, TRAM-34-sensitive KCa 3.1 current. Both KV 1.3 and KCa 3.1 blockers inhibited pro-inflammatory cytokine production and iNOS and COX2 expression demonstrating that KV 1.3 and KCa 3.1 play important roles in microglia activation. Following differentiation with LPS or a combination of LPS and IFN-γ microglia exhibited high KV 1.3 current densities (∼50 pA/pF at 40 mV) and virtually no KCa 3.1 and Kir currents, while microglia differentiated with IL-4 exhibited large Kir 2.1 currents (∼ 10 pA/pF at -120 mV). KCa 3.1 currents were generally low but moderately increased following stimulation with IFN-γ or ATP (∼10 pS/pF). This differential K+ channel expression pattern suggests that KV 1.3 and KCa 3.1 inhibitors could be used to inhibit detrimental neuroinflammatory microglia functions. GLIA 2016;65:106-121.
Insights
Microglia K+ channel expression differs based on activation state. Inhibiting KV 1.3 and KCa 3.1 channels may reduce harmful neuroinflammation in conditions like stroke and Alzheimer's disease.
Area of Science:
- Neuroimmunology
- Cellular Electrophysiology
- Molecular Biology
Background:
- Microglia, the immune cells of the central nervous system, exhibit plasticity and can adopt different phenotypes.
- Classically activated M1-like microglia (induced by LPS/IFN-γ) promote neuroinflammation, while M2-like phenotypes (induced by IL-4) are associated with repair.
- Understanding the molecular mechanisms, including ion channel expression, underlying these phenotypes is crucial for therapeutic targeting.
Purpose of the Study:
- To investigate the differential expression and function of potassium (K+) channels in distinct microglia phenotypes.
- To determine the role of specific K+ channels (KV 1.3, Kir 2.1, KCa 3.1) in microglia activation and pro-inflammatory responses.
- To assess the therapeutic potential of K+ channel blockers in modulating microglia function.
Main Methods:
- Neonatal mouse microglia were differentiated into M(LPS) and M(IL-4) phenotypes.
- Whole-cell patch-clamp electrophysiology was used to characterize K+ currents.
- Quantitative PCR and immunohistochemistry were employed to analyze K+ channel expression.
- The effects of K+ channel blockers (PAP-1, ShK-186, TRAM-34) on cytokine production and gene expression (iNOS, COX2) were evaluated.
Main Results:
- Three major K+ channel types were identified: KV 1.3, Kir 2.1, and KCa 3.1.
- M(LPS) microglia displayed high KV 1.3 currents and minimal KCa 3.1/Kir currents, whereas M(IL-4) microglia showed large Kir 2.1 currents.
- KV 1.3 and KCa 3.1 channel blockers significantly reduced pro-inflammatory cytokine production and iNOS/COX2 expression in activated microglia.
Conclusions:
- Microglia activation states are characterized by distinct K+ channel expression profiles, with KV 1.3 being prominent in M1-like cells.
- KV 1.3 and KCa 3.1 channels play critical roles in mediating microglia pro-inflammatory responses.
- Targeting KV 1.3 and KCa 3.1 channels represents a promising strategy for mitigating detrimental neuroinflammation.
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