The Effects of Diuretics on Intracellular Ca2+ Dynamics of Arteriole Smooth Muscles as Revealed by Laser Confocal Microscopy

摘要

The regulation of cytosolic Ca²⁺ homeostasis is essential for cells, including vascular smooth muscle cells. Arterial tone, which underlies the maintenance of peripheral resistance in the circulation, is a major contributor to the control of blood pressure. Diuretics may regulate intracellular Ca²⁺ concentration ([Ca²⁺]i) and have an effect on vascular tone. In order to investigate the influence of diuretics on peripheral resistance in circulation, we investigated the alteration of [Ca²⁺]i in testicular arterioles with respect to several categories of diuretics using real-time confocal laser scanning microscopy. In this study, hydrochlorothiazide (100 µM) and furosemide (100 µM) had no effect on the [Ca²⁺]i dynamics. However, when spironolactone (300 µM) was applied, the [Ca²⁺]i of smooth muscles increased. The response was considerably inhibited under either extracellular Ca²⁺-free conditions, the presence of Gd³⁺, or with a treatment of diltiazem. After the thapsigargin-induced depletion of internal Ca²⁺ store, the spironolactone-induced [Ca²⁺]i dynamics was slightly inhibited. Therefore, the spironolactone-induced dynamics of [Ca²⁺]i can be caused by either a Ca²⁺ influx from extracellular fluid or Ca²⁺ mobilization from internal Ca²⁺ store, with the former being dominant. As tetraethylammonium, an inhibitor of the K⁺ channel, slightly inhibited the spironolactone-induced [Ca²⁺]i dynamics, the K⁺ channel might play a minor role in those dynamics. Tetrodotoxin, a neurotoxic Na⁺ channel blocker, had no effect, therefore the spironolactone-induced dynamics is a direct effect to smooth muscles, rather than an indirect effect via vessel nerves.