title: Mg(2+)-sensitive non-capacitative basolateral Ca(2+) entry secondary to cell swelling in the polarized renal A6 epithelium.
abstract: 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 nM. 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 nM. The second phase depended on Ca(2+) in the basolateral perfusate and was largely suppressed by 2 mM 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 mM. 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.
authors.affiliation: Laboratory of Physiology, K. U. Leuven, Campus Gasthuisberg O/N, B-3000 Leuven, Belgium.
authors.forename: Danny
authors.initials: D
authors.lastname: Jans
authors.forename: Paul
authors.initials: P
authors.lastname: De Weer
authors.forename: S P
authors.initials: SP
authors.lastname: Srinivas
authors.forename: Els
authors.initials: E
authors.lastname: Larivière
authors.forename: Jeannine
authors.initials: J
authors.lastname: Simaels
authors.forename: Willy
authors.initials: W
authors.lastname: Van Driessche
mesh_terms: D000255:Adenosine Triphosphate / Q000494:pharmacology; D000465:Algorithms; D002118:Calcium / Q000378:metabolism; D015220:Calcium Channels / Q000187:drug effects*; D002460:Cell Line; D048429:Cell Size / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D004594:Electrophysiology; D004847:Epithelial Cells / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure; D006801:Humans; D007668:Kidney / Q000166:cytology*; D008274:Magnesium / Q000494:pharmacology*; D008564:Membrane Potentials / Q000502:physiology; D009997:Osmotic Pressure; D018408:Patch-Clamp Techniques; D013498:Suramin / Q000494:pharmacology; D010738:Type C Phospholipases / Q000037:antagonists & inhibitors
chemical_list: D015220:Calcium Channels; D013498:Suramin; D000255:Adenosine Triphosphate; D010738:Type C Phospholipases; D008274:Magnesium; D002118:Calcium