title: beta-Adrenergic agonists stimulate Mg(2+) uptake in mouse distal convoluted tubule cells.
abstract: beta-Adrenergic agonists influence electrolyte reabsorption in the proximal tubule, loop of Henle, and distal tubule. Although isoproterenol enhances magnesium absorption in the thick ascending limb, it is unclear what effect, if any, beta-adrenergic agonists have on tubular magnesium handling. The effects of isoproterenol were studied in immortalized mouse distal convoluted tubule (MDCT) cells by measuring cellular cAMP formation with radioimmunoassays and Mg(2+) uptake with fluorescence techniques. Intracellular free Mg(2+) concentration ([Mg(2+)](i)) was measured in single MDCT cells by using microfluorescence with mag-fura-2. To assess Mg(2+) uptake, MDCT cells were first Mg(2+) depleted to 0.22 +/- 0.01 mM by culturing in Mg(2+)-free media for 16 h and then placed in 1.5 mM MgCl(2), and the changes in [Mg(2+)](i) were determined. [Mg(2+)](i) returned to basal levels, 0.53 +/- 0.02 mM, with a mean refill rate, d([Mg(2+)](i))/dt, of 168 +/- 11 nM/s. Isoproterenol stimulated Mg(2+) entry in a concentration-dependent manner, with a maximal response of 252 +/- 11 nM/s, at a concentration of 10(-7) M, that represented a 50 +/- 7% increase in uptake rate above control values. This was associated with a sixfold increase in intracellular cAMP generation. Isoproterenol-stimulated Mg(2+) uptake was completely inhibited with RpcAMPS, a protein kinase A inhibitor, and U-73122, a phospholipase C inhibitor, and partially blocked by RO 31-822, a protein kinase C inhibitor. Accordingly, isoproterenol-mediated Mg(2+) entry rates involve multiple intracellular signaling pathways. Aldosterone potentiated isoproterenol-stimulated Mg(2+) uptake (326 +/- 31 nM/s), whereas elevation of extracellular Ca(2+) inhibited isoproterenol-mediated cAMP accumulation and Mg(2+) uptake, 117 +/- 37 nM/s. These studies demonstrate that isoproterenol stimulates Mg(2+) uptake in a cell line of mouse distal convoluted tubules that is modulated by hormonal and extracellular influences.
authors.affiliation: Department of Medicine, University of British Columbia, Vancouver Hospital and Health Sciences Centre, Vancouver, British Columbia, Canada V6T 1Z3.
authors.forename: H S
authors.initials: HS
authors.lastname: Kang
authors.forename: D
authors.initials: D
authors.lastname: Kerstan
authors.forename: L J
authors.initials: LJ
authors.lastname: Dai
authors.forename: G
authors.initials: G
authors.lastname: Ritchie
authors.forename: G A
authors.initials: GA
authors.lastname: Quamme
mesh_terms: D000318:Adrenergic beta-Agonists / Q000494:pharmacology*; D000450:Aldosterone / Q000494:pharmacology; D000818:Animals; D002460:Cell Line; D000242:Cyclic AMP / Q000096:biosynthesis / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D003593:Cytoplasm / Q000187:drug effects / Q000378:metabolism; D007545:Isoproterenol / Q000494:pharmacology; D007686:Kidney Tubules, Distal / Q000187:drug effects / Q000378:metabolism*; D008274:Magnesium / Q000378:metabolism*; D051379:Mice; D010738:Type C Phospholipases / Q000378:metabolism
chemical_list: D000318:Adrenergic beta-Agonists; D000450:Aldosterone; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D010738:Type C Phospholipases; D008274:Magnesium; D007545:Isoproterenol