Contractions in human detrusor smooth muscle induced by hypo-osmolar solutions

Purpose: The aim of this study was to investigate stretch activated channels in human detrusor using hypo-osmolar solutions to produce cell deformation. Stretch activated channels could provide another mechanism by which detrusor myocytes may be coupled. Materials And Methods: Human detrusor removed at surgery was dissected into strips and also enzymatically digested and cultured. Strips (5 x 1 x 1 mm.) were mounted in an organ bath and perfused with gassed Tyrode's. Hypo-osmolar solutions were made by removal of NaCl. Gadolinium (Gd³⁺), a blocker of stretch activated channels (SACs), and diltiazem, an L-type Ca²⁺ channel antagonist were used at 10 μM concentrations. Mean data ± S.E.M. are expressed as a percentage of maximal tension produced by 1 μM carbachol for each patient. Enzymatically disaggregated, human detrusor was cultured in flasks, passaged and placed on glass coverslips. Once confluent the cells were incubated with the Ca²⁺ sensitive fluorochrome Fura-2AM. Coverslips were placed in a bath on the stage of EPI-fluorescence microscope and solutions were perfused through the bath (5 ml. per minute, 35C, pH 7.4). Changes in fluorescence emission ratio (proportional to changes in cytosolic Ca²⁺) were measured. Results: Hypo- osmolar solutions produced a tension increase in the strips and a Ca²⁺ influx in the cells. In the strips in paired experiments Gd³⁺ and diltiazem significantly reduced the response to hypo-osmolar solution (87% ± 16% v 51% ± 12.5%, p = 0.003, n = 10 for Gd³⁺), and (69% ± 11% v 37% ± 9%, p = 0.001, n = 9 for diltiazem). In Ca²⁺ free solution responses were significantly reduced (65% ± 10% v 21% ± 8%, p = 0.001, n = 9). In the cells in paired experiments, 10 μM Gd³⁺ significantly reduced the elevation of cytosolic Ca²⁺ in response to hypo-osmolar solutions (median 0 v 0.38 (62 cells, n = 7 bladders)), as did Ca²⁺ free hypo-osmolar solution (median 0 v 0.44 (46 cells, n = 7)). 10 μM diltiazem (L-type Ca²⁺ channel antagonist) did not influence the response to hypo-osmolar solution (p = 0.14, median 0.5 v 0.54 (31 cells, n = 4)). Conclusions: Hypo-osmolar solutions produced a tension increase in human detrusor that appears to be dependent on upon influx of Ca²⁺ through stretch activated channels (SACs), influx of Ca²⁺ through L-type Ca²⁺ channels and also on release of intracellular Ca²⁺.