Microglial K+ and Ca2+ channels in neuroinflammation

In neurodegenerative disorders such as Alzheimers Disease and ischemic stroke, microglia contribute to neuronal damage and cognitive impairment through their inflammatory activity. However, beneficial microglia functions such as phagocytosis of amyloid plaques in Alzheimers Disease and cellular debris in stroke, as well as release of growth factors and anti-inflammatory mediators constitute a critical element in neuronal recovery and neurogenesis. The diversity of microglia functions is widely associated with specific phenotypes roughly denoted as M1 and M2 microglia. Classically activated M1 cells mainly perform deleterious neurotoxic actions, while alternatively activated M2 microglia are supposed to mediate the beneficial neuronal support. These cellular phenotypes can be distinguished by specific function-related markers, such as scavenger receptor expression. For lymphocytes it has been demonstrated that different subtypes change their potassium channel expression profile upon activation in a subtype-specific manner. Potassium channels are regulating calcium signaling in immune cells via controlling the membrane potential, which constitutes the prerequisite for gene expression, proliferation and various effector functions. The main potassium channels that regulate calcium signaling in immune cells are KCa3.1 and Kv1.3. Cellular subtype-specific differences in their expression levels enable a targeted drug-mediated inhibition, thereby interfering only with certain cellular functions. This strategy has successfully been demonstrated by Heike Wulff and others in various models of autoimmune disorders and malignancies. If such differences in potassium channel expression also exist in microglia subtypes, a targeted inhibition of certain effector functions might provide a new therapeutic strategy for treatment of neuroinflammation-associated diseases. Our preliminary data on activated microglia form MCAO stroke mice suggest that there is indeed a variety in potassium channel expression in microglia from the infarcted area and that microglia activity and effector functions can be modulated by potassium channel blockade. Moreover, mouse post-natal in vitro induced M1 versus M2 microglia also show significant differences in their potassium channel expression. We thus aim to (1) identify the potassium channel profile of microglia subtypes in murine Alzheimer and stroke models, (2) screen for a potential variability in calcium channel expression between M1 and M2 (3) verify our findings in human brain tissue by immunohistochemistry, (4) modulate calcium signaling of microglia subtypes by specific ion channel inhibition with small organic molecules, si-RNA transfection and genetic knock-out models, and (5) validate the effect of targeted inhibition of several microglial effector functions, neuronal viability, -differentiation and neurogenesis. Potassium channel inhibitors are considered as mild immunomodulators with a low side effect profile. The outcomes of this project might have therapeutic implications in the field of microglia- induced neurodegeneration, such as in Alzheimers Disease and stroke. Erwin-Schrödinger-Fellowship

In inflammatory bone disease such as Rheumatoid Arthritis (RA), the increased activity of bone resorbing cells, so called osteoclasts, causes bone lesions and thereby indirectly contributes to inflammation. In our project we were specifically interested, whether, which and how potassium channels could contribute to the calcium signaling-dependent maturation of osteoclasts from macrophage-precursor cells (a type of immune cells) in the presence of the soluble pro-inflammatory mediator TNF-?. Since the inhibition of TNF-? currently still a major target in RA therapy is connected to general weakening of the immune system, our goal was to evaluate a scientific rational for the potential use of clinically tested and less immunosuppressive potassium channel inhibitors as innovative new treatment strategies for inflammatory bone diseases. Ion channels are channel-shaped proteins located in cell membranes that allow the passive exchange of ions (charged particles) between two compartments of a cell (e.g. inside and outside) following an electrochemical gradient. Thereby, potassium exchange can influence other ions such as calcium fluxes which are crucial regulators for many cellular functions, including cell survival, proliferation, migration and differentiation (maturation of a cell). Through genetic analysis of mouse cell cultures, we identified one main potassium channel, named KCa3.1, to be highly abundant in osteoclasts compared to precursor cells. Of interest, precursors that genetically lacked KCa3.1 were less likely to differentiate into osteoclasts than normal, fully functional cells when TNF-? was present in the solution. Similar results were found when KCa3.1 was inhibited with the drug TRAM-34. Furthermore, we discovered that when KCa3.1 activity was inhibited, the diminished formation of osteoclasts was accompanied by decreased cellular calcium signals, as well as a reduction of gene transcripts and proteins that are required for bone resorption. Overall, we identified KCa3.1 as a major regulator for calcium-dependent signaling in physiological, as well as in inflammatory osteoclast maturation pathways. Therefore, KCa3.1 may serve as a future therapeutic target for inflammatory bone diseases.