Effect of calcium and calcium chelator on the response of the bladder to in vitro ischaemia

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Abstract
Objective To examine the effect of different concentrations of calcium on the contractile responses of isolated strips of rabbit bladder detrusor to various forms of stimulation after 2 h incubation in the presence of substrate and oxygen depletion (in vitro ischaemia), followed by 1 h of recovery. The resultant contractile responses were correlated with the level of lipid peroxidation as determined by malonedialdehyde (MDA) concentration. Materials and methods Isolated strips of rabbit bladder detrusor smooth muscle were incubated in Tyrode’s solution containing different concentrations of calcium (0–5.4 mmol/L). The effect of 2 h of incubation in oxygen‐ and substrate‐free medium (in vitro ischaemia), followed by a 1‐h incubation in the presence of oxygen and substrate, on the contractile responses to field stimulation, carbachol and KCl were determined. The effects of repetitive stimulation (15 s of stimulation at 32 Hz applied every 5 min during the 2‐h experimental period) were also assessed. Results The contractile responses to all stimuli increased as the extracellular calcium concentration was increased from 0.6 to 5.4 mmol/L. A 2 h exposure to in vitro ischaemia, followed by a return to normal solution, resulted in a diminished response to all stimuli. This contractile dysfunction was least in the presence of calcium chelator (EGTA) and greatest in the presence of 5.4 mmol/L calcium. Repetitive stimulation during in vitro ischaemia also exacerbated the contractile dysfunction. Lipid peroxidation increased during in vitro ischaemia in proportion to the calcium concentration and was enhanced by repetitive stimulation during this period. Regardless of the incubation conditions, the reduction in the contractile response was significantly greater for field‐stimulated tissues than for those stimulated with carbachol or KCl. Conclusions These results show that the magnitude of contractile dysfunction induced by incubation in the presence of substrate and oxygen depletion is reduced in the presence of low calcium concentrations, increased in the presence of high calcium levels and increased in the presence of repetitive stimulation. In addition, the level of lipid peroxidation after the recovery period was proportional to the magnitude of contractile dysfunction present.

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