Voltage-dependent L-type calcium channels (VDCC) play important roles in many cellular processes. The interaction of the actin cytoskeleton with the channel in nonexcitable cells is less well understood. We performed whole-cell patch-clamp surface biotinylation and calcium imaging on different osteoblast cells to determine channel kinetics, amplitude, surface abundance, and intracellular calcium, respectively. Patch-clamp studies showed that actin polymerization by phalloidin increased the peak current density of I Ca, whereas actin depolymerization by cytochalasin D (CD) significantly decreased the current amplitude. This result is consistent with calcium imaging, which showed that CD significantly decreased Bay K8644-induced intracellular calcium increase. Surface biotinylation studies showed that CD is not able to affect the surface expression of the pore-forming subunit α 1C. Interestingly, application of CD caused a significantly negative shift in the steady-state inactivation kinetics of I Ca. There were decreases in the voltage at half-maximal inactivation that changed in a dosedependent manner. CD also reduced the effect of activated vitamin D 3 (1α,25-D3) on VDCC and intracellular calcium. Weconclude that in osteoblasts the actin cytoskeleton affects α 1C by altering the channel kinetic properties, instead of changing the surface expression, and it is able to regulate 1α,25-D3 signaling through VDCC. Our study provides a new insight into calcium regulation in osteoblasts, which are essential in many physiological functions of this cell.
- Actin cytoskeleton
- Voltage-dependent L-type calcium channel
ASJC Scopus subject areas
- Endocrinology, Diabetes and Metabolism
- Orthopedics and Sports Medicine