Iron is a biologically essential metal, but excess iron can cause damage to the cardiovascular and nervous systems. We examined the effects of extracellular Fe 2ϩ on permeation and gating of Ca V 3.1 channels stably transfected in HEK293 cells, by using whole-cell recording. Precautions were taken to maintain iron in the Fe 2ϩ state (e.g., use of extracellular ascorbate). With the use of instantaneous I-V currents (measured after strong depolarization) to isolate the effects on permeation, extracellular Fe 2ϩ rapidly blocked currents with 2 mM extracellular Ca 2ϩ in a voltage-dependent manner, as described by a Woodhull model with K D ϭ 2.5 mM at 0 mV and apparent electrical distance ␦ ϭ 0.17. Extracellular Fe 2ϩ also shifted activation to more-depolarized voltages (by ϳ10 mV with 1.