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Research Article

A Large Conductance (BK) Potassium Channel subtype affects both growth and mineralisation of Human Osteoblasts

¹Cardiff University

Submitted 16 July 2009 ; revised 15 September 2009 ; accepted in final form 15 September 2009

The pharmacology of the large conductance BK channel in human osteoblasts is not well defined and its role in bone is speculative. Here we assess BK channel properties in both MG63 cells and primary human osteoblasts and determine whether pharmacological modulation affects cell function. We used RT-PCR and patch-clamp methods to determine the expression of BK channel subunits, and cell number assays in the absence and presence of BK channel modulators. RT-PCR showed the presence of KCNMA1, KCNMB1, KCNMB2, KCNMB3 and KCNMB4 subunits. The BK channel was voltage-dependent with a mean unitary conductance of 228.8 pS (n = 10) in cell-attached patches (140 K⁺/140 K⁺), a conductance of 142.5 pS (n = 16) in excised outside-out and 155 pS (n = 6) in inside-out patches in 3 K⁺/140 K⁺. The selectivity ratio PK/PNa was 15/1. The channel was blocked by tetraethylammonium (TEA) (0.3 mM), iberiotoxin (5-60 nM), tetrandrine (5-30 µM) and paxilline (10 µM) and activated by isopimaric acid (20 µM). BK channel modulators affected MG63 cell numbers: TEA and tetrandrine significantly increased cell numbers at low concentrations (1 mM, 3 µM respectively), and reduced numbers at higher concentrations (>10 mM, >10 µM respectively). Neither iberiotoxin (20-300 nM) nor slotoxin (300 nM) affected cell numbers. The increase in cell number by TEA was blocked by isopimaric acid. TEA (0.1-3.0 mM) significantly increased mineralisation in primary osteoblasts. In conclusion, the BK channel has a distinctive pharmacology and is thus a target for therapeutic strategies aimed at modulating osteoblast proliferation and function.

Bone; MG63 cells; potassium channel; tetrandrine