Whole-cell patch-clamp analysis revealed a resting membrane potential of −60 mV in primary osteoblasts and in the MG-63 osteoblast-like cells. Depolarization-induced action potentials were characterized by duration of 60 ms, a minimal peak-to-peak distance of 180 ms, a threshold value of −20 mV and a repolarization between the spikes to −45 mV. Expressed channels were characterized by application of voltage pulses between −150 mV and 90 mV in 10 mV steps, from a holding potential of −40 mV. Voltages below −60 mV induced an inward current. Depolarizing voltages above −30 mV evoked two currents: (a) a fast activated and inactivated inward current at voltages between −30 and 30 mV, and (b) a delayed-activated outward current that was induced by voltages above −30 mV. Electrophysiological and pharmacological parameters indicated that hyperpolarization activated strongly rectifying K+ (Kir) channels, whereas depolarization activated tetrodotoxin sensitive voltage gated Na+ (Nav) channels as well as delayed, slowly activated, non-inactivating, and tetraethylammonium sensitive voltage gated K+ (Kv) channels. In addition, RT-PCR showed expression of Nav1.3, Nav1.4, Nav1.5, Nav1.6, Nav1.7, and Kir2.1, Kir2.3, and Kir2.4 as well as Kv2.1. We conclude that osteoblasts express channels that allow firing of action potentials.