Patterns of intracellular and intercellular Ca2+ waves in the longitudinal muscle layer of the murine large intestine in vitro

Ca2+ wave activity was monitored in the longitudinal (LM) layer of isolated murine caecum and proximal colon at 35 degreesC with fluo-4 AM and an iCCD camera. Both intracellular (within LM cells) and intercellular (also spreading from cell to cell) Ca2+ waves were observed. Intracellular Ca2+ waves were associated with a lack of muscle movement whereas intercellular Ca2+ waves, which were five times more intense than intracellular waves, were often associated with localized contractions. Several intracellular Ca2+ waves were present at the same time in individual LM cells. Waves in adjacent LM cells were not coordinated and were unaffected by TTX (I muM) but were blocked by IP3 receptor antagonists xestospongin-C (Xe-C; 2 muM) or 2-aminoethyl diphenylborate (2-APB; 25 mum), and by ryanodine (10 muM). Caffeine (5 mm) restored wave activity following blockade with Xe-C. NiCl2 (1 mm) blocked intracellular Ca2+ waves, and nicardipine (2 muM) reduced their frequency and intensity, but did not affect their velocity, suggesting the sarcoplasmic reticulum may be fuelled by extracellular Ca2+ entry. Intercellular Ca2+ waves often occurred in bursts and propagated rapidly across sizeable regions of the LM layer and were blocked by heptanol (0.5 mm). Intercellular Ca2+ waves were dependent upon neural activity, external Ca2+ entry through L-type Ca2+ channels, and amplification via calcium-induced calcium release (CICR). In conclusion, intracellular Ca2+ waves, which may reduce muscle excitability, are confined to individual LM cells. They depend upon Ca2+ release from internal Ca2+ stores and are likely to be fuelled by extracellular Ca2+ entry. Intercellular Ca2+ waves, which are likely to underlie smooth muscle tone, mixing and propulsion, depend upon neural activity, muscle action potential propagation and amplification by CICR.