Excitation of human esophageal smooth muscle (SM) involves release of Ca²⁺ from intracellular stores and influx. The lower esophageal sphincter (LES) shows the distinctive property of tonic contraction; however the mechanisms by which this is maintained are incompletely understood. We examined Ca²⁺ channels in human esophageal muscle and investigated their contribution to LES tone. Functional effects were examined with tension recordings, currents recorded with patch-clamp electrophysiology, channel expression explored by RT-PCR and intracellular Ca²⁺ concentration ([Ca²⁺]_i) monitored by fura-2 fluorescence. LES muscle strips developed tone that was abolished by removal of extracellular Ca²⁺, reduced by application of the L-type Ca²⁺ channel blocker nifedipine (to 13±6% of control), but was unaffected by inhibition of the sarcoplasmic reticulum Ca²⁺-ATPase by cyclopiazonic acid (CPA). Carbachol increased tension above basal tone, and this effect was attenuated by treatment with CPA and nifedipine. Voltage-dependent inward currents were studied using patch clamp techniques and dissociated cells. Similar inward currents were observed in esophageal body (EB) and LES smooth muscle cells. The inward currents in both tissues were blocked by nifedipine, enhanced by Bay K8644, and transiently suppressed by acetylcholine. The molecular form of Ca²⁺ channel was explored using RT-PCR, and similar splice variant combinations of the pore forming 1_C subunit were identified in EB and LES. This is the first characterization of Ca²⁺ channels in human esophageal smooth muscle, and we establish that L-type Ca²⁺ channels play a critical role in maintaining lower esophageal sphincter tone.