Abstract: In order to verify the inotropic effect of berberine (Ber) on myocardial tissues, the effect of Ber on cytosolic free calcium Ca²⁺i in cultured rat single myocardial cells were examined by using Fura-2/AM and AR-CM-MIC cation measurement system. Ber (10～400 μmol·L^-1) was found to increase the resting Ca²⁺i and saturate at about 200 μmol·L^-1. The Ca²⁺i increase was not depressed by atropine, phentolamine, propranolol and tetrodotoxin, but was inhibited by verapamil (Ver) 3 μmol·L^-1 or CoCl₂ 10 mmol·L^-1. The Ca²⁺i increase induced by Ber (30 μmol·L^-1) added to stimulating agents as KCl, CaCl₂, NE or Oua was stronger than that induced by stimulating agents only and Ver 3 μmol·L^-1 still showed inhibitory effects on the Ca²⁺i increase. Under Ca²⁺free or Ca²⁺, K⁺, Na⁺-free conditions, Ber (30～200 μmol·L^-1) still showed ascending effects on Ca²⁺i, and Ver 3 μmol·L^-1 only inhibited the Ber effects in Ca²⁺-free condition. The results suggest that Ber might induce moderate Ca²⁺i increase via Ca²⁺ influx and intracellular Ca²⁺ release, and show positive inotropic effect.