Mechanism of TRPM7 Channel Inhibition by 2-Aminoethyl Diphenyl Borinate (2-APB)

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Conference Proceeding

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2-APB is a widely used chemical compound in ion channel research. It inhibits numerous channels that include inositol 1,4,5-trisphosphate receptors, store-operated calcium channels, connexins and TRP family cation channels TRPC3, TRPC5, TRPC6, TRPM2, TRPM3 and TRPM7. Native and overexpressed TRPM7 channels are inhibited by 2-APB with IC50-s in the 70-170 μM range. A characteristic of TRPM7 channels is their inhibition by intracellular Mg2+ and acidic pH. using patch-clamp electrophysiology, we recorded native TRPM7 channel activity in Jurkat T lymphocytes and tested 2-APB at 10-300 μM for its ability to inhibit TRPM7 currents. When internal HEPES buffer concentration was low (1 mM), 100-300 μM 2-APB inhibited 60-70% of TRPM7 current with a slow time course. This inhibition was voltage-independent and reversible. Increasing the pH buffering capacity of internal solution to 140 mM HEPES abolished the inhibitory 2-APB effect. Simply making the internal recording solution alkaline, greatly diminished 2-APB inhibition. using single-cell pHi imaging with the fluorescent pH indicator BCECF, we found that at concentrations of 100 μM and higher, 2-APB potently acidifies the cytoplasm. A similar 2-APB-induced acidification was also observed in HEK293 cells, often used for TRPM7 channel overexpression. In contrast to 2-APB, its analog 2,2-diphenyltetrahydrofuran (DPTHF) did not produce cytosolic acidification, when applied at concentrations of 150-300 μM. We also found that Kv1.3 channels expressed in Jurkat T cells are modulated by 2-APB: the apparent voltage dependence of these channels was reversibly altered by the compound. Interestingly, this 2-APB effect on Kv1.3 channels did not depend on cytosolic pH. In summary, these experiments suggest that inhibition of TRPM7 channels by 2-APB is not direct but rather, can be explained by cytoplasmic acidification and resulting channel inhibition.


Poster presented at the 57th Annual Meeting of the Biophysical Society, Philadelphia, PA.

Poster Presentation Number 2330-Pos. Board Number B349.