Detrusor smooth muscle contractions are elicited by the release of Ca²⁺ from intracellular stores to cause activation of the contractile machinery. The release of the Ca²⁺ normally occurs through a second messenger cascade from activation of the muscarinic M3receptor. It is thought (Olanrewaju et al. 1996) that adenosine may modulate the release of intracellular Ca²⁺ through P1-receptors, of which there are a number of subtypes (A1, A2a, A2b and A3subtypes). The aim of the study was to determine the particular subtypes involved in this modulation. The intracellular [Ca²⁺] was measured in isolated detrusor cells from human bladder samples (obtained with local ethical approval and written patient consent). Tissue was dissociated with a solution of digestive enzymes according to previously determined protocols. (Wu et al. 2002). The isolated cells were loaded with a fluorescent indicator, Fura

2. Cell suspensions were placed onto a microscope stage and were

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superfused at 37C° in HCO/CO₂ Tyrode’s solution (pH7.35).

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Ca²⁺-transients were stimulated with the acetylcholine analogue, carbachol (1-10µM). The effect of P1-receptor agonists and antagonists on these Ca²⁺-transients were investigated. All compounds were from Sigma UK and, with the exception of adenosine, were made as stocks in DMSO and diluted in Tyrode’s solution to working concentrations. Data are expressed as mean±S.D., (n) is number of experiments, Student’s t-test was used to test for significance between data sets (∗p<0.05).1mM adenosine reduced significantly carbachol-mediated Ca²⁺-transients in human detrusor cells [74±11% of control (n=14)]. 10µM NECA (general A1,A2-receptor agonist) reduced the Ca²⁺-transients to a similar extent [60±20% of control (n=6)]. The effect of adenosine was not significantly different in the presence of the A2b-receptor antagonist alloxazine (1 µM) [76±8% of control (n=7)]. 10µM CPA (an A1-specific agonist) and 1µM DPCPX (an A1-specific antagonist) and 10µM IB-MECA (an A3-specific agonist) did not have any significant effects (all n=3). Adenosine, NECA and alloxazine (all n=3) had no significant effects on Ca²⁺-transients elicited by raising extracellular KCl to 80mM. In human detrusor myocytes, adenosine modulation of carbachol-mediated Ca²⁺-transients appears to be via A2-receptors; although the particular subtype remains to be unequivocally determined. The modulation may be through alteration of cAMP levels, as P1-receptors are coupled to adenylate cyclase: alternatively, there may be a direct effect on intracellular stores.