AIM:To study the origin of calcium necessary for agonist-induced contraction of the distal colon in rats. METHODS:The change in intracel lular calcium concentration ([Ca 2+ ]i) evoked by elevating external Ca 2+ was detected by fura 2/AM fluorescence. Contractile activity was measured with a force displacement transducer. Tension was continuously monitored and recorded using a Powerlab 4/25T data acquisition system with an ML110 bridge bioelectric physiographic amplifier. RESULTS:Store depletion induced Ca 2+ influx had an effect on [Ca 2+ ]i. In nominally Ca 2+ -free medium, the sarco-endoplasmic reticulum Ca 2+ -ATPase inhibitor thapsigargin (1 µmol/L) increased [Ca 2+ ]i from 68 to 241 nmol/L, and to 458 (P < 0.01) and 1006 nmol/L (P < 0.01), respectively, when 1.5 mmol/L and 3.0 mmol/L extracellular Ca 2+ was reintroduced. Furthermore, the change in [Ca 2+ ]i was observed with verapamil (5 µmol/L), La 3+ (1 mmol/L) or KCl (40 mmol/L) in the bathing solution. These channels were sensitive to La 3+ (P < 0.01), insensitive to verapamil, and voltage independent. In isolated distal colons we found that in normal Krebs solution, contraction induced by acetylcholine (ACh) was partially inhibited by verapamil, and the inhibitory rate was 41% (P < 0.05). On the other hand, in Ca 2+ -free Krebs solution, ACh induced transient contraction due to Ca 2+ release from the intracellular stores. The transient contraction lasted until the Ca 2+ store was depleted. Restoration of extracellular Ca 2+ in the presence of atropine produced contraction, mainly due to Ca 2+ influx. Such contraction was not inhibited by verapamil, but was decreased by La 3+ (50 µmol/L) from 0.96 to 0.72 g (P < 0.01). CONCLUSION:The predominant source of activator C a 2 + f o r t h e c o n t ra c t i l e r e s p o n s e t o a g o n i s t i s extracellular Ca 2+ , and intracellular Ca 2+ has little role to play in mediating excitation-contraction coupling by agonists in rat distal colon smooth muscle in vitro . The influx of extracellular Ca 2+ is mainly mediated through voltage-, receptor-and store-operated Ca 2+ channels, which can be used as an alternative to develop new drugs targeted on the dysfunction of digestive tract motility.