PGE2 stimulates Mg2+ uptake in mouse distal convoluted tubule cells.
Prostaglandins have diverse effects on renal electrolyte reabsorption, inhibiting NaCl absorption in the thick ascending limb and modulating sodium and calcium transport in cortical collecting cells. It is unclear what effect, if any, prostaglandins have on tubular magnesium handling. The effects of prostaglandin E2 (PGE2) were studied on immortalized mouse distal convoluted tubule (MDCT) cells by measuring cellular cAMP formation with radioimmunoassays and Mg2+ uptake with fluorescence techniques. Intracellular free Mg2+ concentration ([Mg2+]i) was measured on single MDCT cells using microfluorescence with mag-fura 2. To assess Mg2+ uptake, MDCT cells were first Mg2+ depleted to 0.22 +/- 0.01 mM by culturing in Mg2+-free media for 16 h and then placed in 1.5 mM MgCl2, and the changes in [Mg2+]i were determined. [Mg2+]i returned to basal levels, 0.53 +/- 0.02 mM, with a mean refill rate, d([Mg2+]i)/dt, of 173 +/- 8 nM/s. Indomethacin, 5 microM, diminished basal Mg2+ uptake, suggesting that endogenous prostaglandins may stimulate Mg2+ entry in control cells. PGE2 stimulated Mg2+ entry in a concentration-dependent manner with maximal response of 311 +/- 12 nM/s, at a concentration of 10(-7) M, which represented an 80 +/- 3% increase in uptake rate above control values. This was associated with a sixfold increase in intracellular cAMP generation. PGE2-stimulated Mg2+ uptake was completely inhibited with the Rp diastereoisomer of adenosine 3',5'-cyclic monophosphothionate (Rp-cAMPS), a protein kinase A inhibitor, and U-73122, a phospholipase C inhibitor, and partially by chelerythrine, a protein kinase C inhibitor. Accordingly, PGE2-mediated Mg2+ entry rates involve multiple intracellular signaling pathways. These studies demonstrate that PGE2 stimulates Mg2+ uptake in a cell line of MDCT.
Dai
L J
LJ
Department of Medicine, University of British Columbia, Vancouver Hospital and Health Sciences Centre, Koerner Pavilion, Vancouver, British Columbia, Canada V6T 1Z3.
Bapty
B
B
Ritchie
G
G
Quamme
G A
GA
D000818:Animals; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000378:metabolism; D015232:Dinoprostone / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D005453:Fluorescence; D007686:Kidney Tubules, Distal / Q000378:metabolism*; D008274:Magnesium / Q000378:metabolism*; D051379:Mice; D011863:Radioimmunoassay
D000242:Cyclic AMP; D008274:Magnesium; D015232:Dinoprostone