Cultured bladder cell responses to hypo-osmolar solution; evidence for stretch activated channels?
Masters JG., Neal DE., Gillespie JI.
Introduction: The myogenic contribution to changes in bladder compliance, both normally and in disease, is controversial. As the bladder fills, one mechanism by which muscles may be activated and compliance altered is via stretch-activated channels (SACs). SACs open in response to mechanical deformation allowing an influx of ions across the cell membrane which may lead to cell contraction. SACs have been detected in guinea-pig detrusor but not in man. Here we present the effects of hypo-osmolar solutions (HOS) (which produce cell deformation and can be used to study SACs) on cultured human detrusor smooth muscle cells. Materials and methods: With ethical approval, human detrusor was enzymatically disaggregated, cultured in flasks, passaged and placed on glass coverslips. Once confluent, the cells were incubated with the Ca++ sensitive fluorochrome Fura-2AM. Coverslips were placed in a bath on the stage of an EPI-fluoresence microscope and solutions were perfused through the bath (5 mL/min, 35° C, pH 7.4). Changes in fluorescence emission ratio (proportional to changes in cytosolic Ca ++ ) were measured. HOS was produced by reducing the NaCl content of the basic salt solution (BS). The Wilcoxon signed-rank test was used for statistical analysis. Results: HOS application produced a rise in the cytosolic Ca++ (336/350 cells, 13 bladders). In paired experiments. 10 nmol/L Gdi+, a specific blocker of SACs, very significantly reduced this response (median 0 vs. 0.38, 62 cells, seven bladders), as did Ca ++ -free BS (median 0 vs. 0.44, 46 cells, seven bladders). Diltiazem (10 μmol/L, an L-type Ca ++ channel antagonist) did not influence the response to HOS (P = 0.14, median 0.5 vs. 0.54, 31 cells, four bladders). Conclusion: It appears that in cultured human detrusor cells SACs are involved in elevating the cytosolic Ca ++ in response to membrane deformation by HOS. This response depends in part on extracellular Ca++. Therefore, during filling, as the bladder stretches, Ca ++ influx through SACs may lead to cell contraction, increased bladder tone and altered compliance, and may contribute to instability in abnormal bladders. © 1998 British Journal of Urology.