AIM: To study the origin of calcium necessary for agonist-induced contraction of the distal colon in rats.METHODS: The change in intracellular calcium concentration ([Ca2+]i) evoked by elevating external Ca2+ 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 Ca2+ influx had an effect on [Ca2+]i. In nominally Ca2+-free medium, the sarco-endoplasmic reticulum Ca2+-ATPase inhibitor thapsigargin (1 μmol/L) increased [Ca2+]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 Ca2+ was reintroduced. Furthermore, the change in [Ca2+]i was observed with verapamil (5 μmol/L), La3+ (1 mmol/L) or KCl (40 mmol/L) in the bathing solution. These channels were sensitive to La3+ (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 Ca2+-free Krebs solution, ACh induced transient contraction due to Ca2+ release from the intracellular stores. The transient contraction lasted until the Ca2+ store was depleted. Restoration of extracellular Ca2+ in the presence of atropine produced contraction, mainly due to Ca2+ influx. Such contraction was not inhibited by verapamil, but was decreased by La3+ (50 μmol/L) from 0.96 to 0.72 g (P < 0.01). CONCLUSION: The predominant source of activator Ca2+ for the contractile response to agonist is extracellular Ca2+, and intracellular Ca2+ has little role to play in mediating excitation-contraction coupling by agonists in rat distal colon smooth muscle in vitro. The influx of extracellular Ca2+ is mainly mediated through voltage-, receptor- and store-operated Ca2+ channels, which can be used as an alternative to develop new drugs targeted on the dysfunction of digestive tract motility.
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