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


METHODS
Human detrusor removed at surgery was dissected into strips and also enzymatically digested and cultured. Strips (5x1x1 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 microM concentrations. Mean data +/- S.E.M. are expressed as a percentage of maximal tension produced by 1 microM 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 microM 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 microM 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+.
Masters
J G
JG
Department of Surgical Sciences, The Medical School, Newcastle University, Newcastle upon Tyne, United Kingdom.
Neal
D E
DE
Gillespie
J I
JI
D002121:Calcium Channel Blockers / Q000494:pharmacology; D002217:Carbachol / Q000494:pharmacology*; D004110:Diltiazem / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D005682:Gadolinium / Q000494:pharmacology; D006801:Humans; D007552:Isotonic Solutions / Q000494:pharmacology*; D009119:Muscle Contraction / Q000187:drug effects*; D009130:Muscle, Smooth / Q000187:drug effects* / Q000502:physiology*; D001743:Urinary Bladder / Q000187:drug effects* / Q000502:physiology*
D002121:Calcium Channel Blockers; D007552:Isotonic Solutions; C037692:Tyrode's solution; D002217:Carbachol; D005682:Gadolinium; D004110:Diltiazem