Mg2+-sensitive non-capacitative basolateral Ca2+ entry secondary to cell swelling in the polarized renal A6 epithelium

• Published in: The Journal of physiology 2002-05, Vol.541 (1), p.91-101
• Main Authors: Jans, Danny, Weer, Paul, Srinivas, S. P., Larivière, Els, Simaels, Jeannine, Driessche, Willy
• Format: Article
• Language: English
• Subjects: Original
• ISSN: 0022-3751; 1469-7793
• DOI: 10.1113/jphysiol.2001.013199

Polarized renal A6 epithelia respond to hyposmotic shock with an increase in transepithelial capacitance ( C T ) that is inhibited by extracellular Mg 2+ . Elevation of free cytosolic [Ca 2+ ] ([Ca 2+ ] i ) is known to increase C T . Therefore, we examined [Ca 2+ ] i dynamics and their sensitivity to extracellular Mg 2+ during hyposmotic conditions. Fura-2-loaded A6 monolayers, cultured on permeable supports were subjected to a sudden reduction in osmolality at both the basolateral and apical membranes from 260 to 140 mosmol (kg H 2 O) −1 . Reduction of apical osmolality alone did not affect [Ca 2+ ] i . In the absence of extracellular Mg 2+ , the hyposmotic shock induced a biphasic rise in [Ca 2+ ] i . The first phase peaked within 40 s and [Ca 2+ ] i increased from 245 ± 12 to 606 ± 24 n m . This phase was unaffected by removal of extracellular Ca 2+ , but was abolished by activating P2Y receptors with basolateral ATP or by exposing the cells to the phospholipase C (PLC) inhibitor U73122 prior to the osmotic shock. Suramin also severely attenuated this first phase, suggesting that the first phase of the [Ca 2+ ] i rise followed swelling-induced ATP release. The PLC inhibitor, the ATP treatment or suramin did not affect a second rise of [Ca 2+ ] i to a maximum of 628 ± 31 n m . The second phase depended on Ca 2+ in the basolateral perfusate and was largely suppressed by 2 m m basolateral Mg 2+ . Acute exposure of the basolateral membrane to Mg 2+ during the upstroke of the second phase caused a rapid decline in [Ca 2+ ] i . Basolateral Mg 2+ inhibited Ca 2+ entry in a dose-dependent manner with an inhibition constant ( K i ) of 0.60 m m . These results show that polarized A6 epithelia respond to hyposmotic shock by Ca 2+ release from inositol trisphosphate-sensitive stores, followed by basolateral Ca 2+ influx through a Mg 2+ -sensitive pathway. The second phase of the [Ca 2+ ] i response is independent of the initial intracellular Ca 2+ release and therefore constitutes non-capacitative Ca 2+ entry.