Evidence for Mitoxantrone-induced Block of Inwardly Rectifying K⁺Channels Expressed in the Osteoclast Precursor RAW 264.7 Cells Differentiated with Lipopolysaccharide

Abstract

Background/Aims: Mitoxanthrone (MX) is an anthracenedione antineoplastic agent. Whether this drug and other related compounds have any effects on ion currents in osteoclasts remains largely unclear. Methods: In this study, the effects of MX and other related compounds on inwardly rectifying K⁺ current (I_K(IR)) were investigated in RAW 264.7 osteoclast precursor cells treated with lipopolysaccharide. Results: The I_K(IR)in these cells are blocked by BaCl₂(1 mM). MX (1-100 µM) decreased the amplitude of I_K(IR)in a concentration-dependent manner with an IC₅₀value of 6.4 µM. MX also slowed the time course of I_K(IR)inactivation elicited by large hyperpolarization. Doxorubicin (10 µM), 17β-estradiol (10 µM) and tertiapin (1 µM) decreased the I_K(IR)amplitude in these cells. In bafilomycin A₁-treated cells, MX-mediated block of I_K(IR)still existed. In cell-attached configuration, when the electrode was filled with MX (10 µM), the activity of inwardly rectifying K⁺ (Kir) channels was decreased with no change in single-channel conductance. MX-mediated reduction of channel activity is accompanied by a shortening of mean open time. Under current-clamp conditions, addition of MX resulted in membrane depolarization. Therefore, MX can interact with the Kir channels to decrease amplitude and to depolarize the membrane in these cells. Conclusion: The block by ^{the I}K(IR)this drug of Kir2.1 channels appears to be one of the important mechanisms underlying its actions on the resorptive activity of osteoclasts, if similar results occur in vivo. Targeting at Kir channels may be clinically useful as an adjunctive regimen to anti-cancer drugs (e.g., MX or doxorubicin) in influencing the resorptive activity of osteoclasts.

The Inhibition of Inwardly Rectifying K+Channels by Memantine in Macrophages and Microglial Cells

“…In this study, unlike astrocytes or T lymphocytes, BV2 microglial cells were clearly found to exhibit the activity of Kir channels as described recently in RAW 264.7 macrophages [23]. Addition of MEM was found to inhibit the amplitude of inwardly rectifying K + current ( I K(IR) ) though the Kir channels in RAW 264.7 macrophages and BV2 microglial cells.…”

“…Their resistances commonly ranged between 3 and 5 MΩ when the pipettes were filled with different internal solutions described above. Patch-clamp recordings under whole-cell, cell-attached or inside-out configuration were performed in standard patch-clamp technique using either an RK-400 (Bio-Logic, Claix, France) or an Axopatch 200B patch-clamp amplifier (Molecular Devices, Sunnyvale, CA) [23,25]. The offset potential between the pipette and bath solution was generally compensated with amplifier after the pipette entered the bath but immediately before a seal was made.…”

“…For further analyses, the digital data were exported to Origin 8.0 (OriginLab, Northampton, MA) or Excel 2007 (Microsoft, Redmond, WA). The inactivation time courses of I K(IR) in response to large hyperpolarizations with or without addition of MEM were adequately measured by fitting an one-exponential function [23]. …”

“…7 macrophages [20]. On the other hand, the activity of Kir channels has been previously described in macrophages and microglial cells [5,21,22,23]. These Kir channels in microglial cells can play a role in the maintenance of extracellular K + by absorbing excess K + ions released from excited neurons.…”

Ion channels in cancer: future perspectives and clinical potential

“…On the other hand, the inhibition of Kir channels has been shown to slow down cell proliferation [105,106].…”