Anoctamin1 (Ano1, TMEM16A) is a calcium-activated chloride channel specifically expressed in interstitial cells of Cajal (ICC) of the gastrointestinal (GI) tract muscularis propria. Ano1 is necessary for normal electrical slow waves and ICC proliferation. The full length human Ano1 sequence includes an additional exon, exon "0," at the N-terminus. Ano1 with exon "0" (Ano1₍₀₎) had a lower EC₅₀ for intracellular calcium ([Ca²⁺]_i) and faster chloride current (I_Cl) kinetics. The Ano1 alternative splice variant with segment "c" encoding exon 13 expresses on the first intracellular loop four additional amino acid residues, EAVK, which alter I_Cl at low [Ca²⁺]_i. Exon 13 is expressed in 75-100% of Ano1 transcripts in most human tissues but only 25% in human stomach. Our aim was to determine the effect of EAVK deletion on Ano1₍₀₎ I_Cl parameters. By voltage-clamp electrophysiology, we examined I_Cl in HEK293 cells transiently expressing Ano1(0) with or without the EAVK sequence (Ano1(0)EAVK). The EC50 values of activating and deactivating ICl for [Ca²⁺]_i was 438±7 and 493±9 nM for Ano1₍₀₎ but higher for Ano1₍₀₎EAVK at 746±47 and 761±26 nM, respectively. Meanwhile, the EC50 values for the ratio of instantaneous to steady-state I_Cl were not different between variants. Congruently, the time constant of activation was slower for Ano1₍₀₎EAVK than Ano1₍₀₎ currents at intermediate [Ca²⁺]_i. These results suggest that EAVK decreases the calcium sensitivity of Ano1₍₀₎ current activation and deactivation by slowing activation kinetics. Differential expression of EAVK in human stomach may function as a switch to increase sensitivity to [Ca²⁺]_i via faster gating of Ano1.