Abstract
Cytosolic-free [Ca 2+] was evaluated in freshly dissociated smooth muscle cells from mouse thoracic aorta by the ratio of Fura Red and Fluo 4 emitted fluorescence using confocal microscopy. The role of intercellular communication in forming and shaping ATP-elicited responses was demonstrated. Extracellular ATP (250 μM) elicited [Ca 2+] i transient responses, sustained [Ca 2+] i rise, periodic [Ca 2+] i oscillations and aperiodic repetitive [Ca 2+] i transients. Quantity of smooth muscle cells in the preparation responding to ATP with periodical [Ca 2+] i oscillations depended on the density of isolated cells on the cover slip. ATP-elicited bursts of [Ca 2+] i spikes in 66 ± 7% of cells in dense and in 33 ± 8.5% of cells in non-dense preparations. The number of cells responding to ATP with bursts of [Ca 2+] i spikes decreased from 55 ± 5% ( n = 84) to 14 ± 3% ( n = 141) in dense preparations pretreated with carbenoxolone. Simultaneous measurement of [Ca 2+] i and ion currents revealed a correlation between [Ca 2+] i and current oscillations. ATP-elicited bursts of current spikes in 76% of cells regrouped in small clusters and in 9% of isolated cells. Clustered cells responding to ATP with current oscillations had higher membrane capacity than clustered cells with transient and sustained ATP-elicited responses. Lucifer Yellow (1% in 130 mM KCl) injected into one of clustered cells was transferred to the neighboring cell only when ATP-elicited oscillations. Fast application of carbenoxolone (100 μM) inhibited ATP (250 μM) elicited Ca 2+-dependent current oscillations. Taken together these results suggest that the probability of ATP (250 μM) triggered cytosolic [Ca 2+] i oscillations accompanied with K + and Cl − current oscillations increased with the coupling of smooth muscle cells.
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