Ca2+ Regulation in Detrusor Smooth Muscle From Ovine Fetal Bladder After In Utero Bladder Outflow Obstruction

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Abstract

Purpose

We characterized intracellular Ca2+ regulation in fetal bladders following outflow obstruction by examining the Ca2+ response to agonists in smooth muscle cells.

Materials and Methods

Severe bladder outflow obstruction was induced in male fetal sheep by placing a urethral ring and urachal ligation midway through gestation at 75 days. Fetuses were examined 30 days after surgery. Intracellular Ca2+ in single smooth muscle cells isolated from the bladder wall was measured with epifluorescence microscopy using fura-2(AM) during exposure to agonists, such as carbachol and adenosine triphosphate, and to other activators, such as caffeine and KCl.

Results

Detrusor smooth muscle cells from obstructed bladders had resting intracellular Ca2+ similar to that in sham operated controls. The maximal response to carbachol was decreased following obstruction (p <0.05). Construction of dose-response curves also demonstrated higher EC50 (p <0.05). However, these changes were not mirrored by caffeine evoked Ca2+ release, which was not significantly different between the obstruction group and sham operated controls. Kinetic analysis of carbachol transients further revealed an attenuated maximal rate of increase in obstructed bladders (p <0.01). The magnitude of intracellular Ca2+ to purinergic neurotransmitter adenosine triphosphate was also found to be smaller in cells from obstructed bladders (p <0.05), although transmembrane influx by high K depolarization was not significantly affected.

Conclusions

Muscarinic and purinergic pathways were down-regulated in fetal detrusor muscle following outflow obstruction. These major functional receptors appeared to be more susceptible to obstruction than other Ca2+ regulators. Their impairment may contribute to the compromised contractile function seen in in utero bladder outflow obstruction.

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