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Abstract Magnesium Transport in the Renal Distal Convoluted Tubule. Physiol Rev 81: 51–84, 2001.—The distal tubule reabsorbs ;10% of the filtered Mg21, but this is 70–80% of that delivered from the loop of Henle. Because there is little Mg21 reabsorption beyond the distal tubule, this segment plays an important role in determining the final urinary excretion. The distal convoluted segment (DCT) is characterized by a negative luminal voltage and high intercellular resistance so that Mg21 reabsorption is transcellular and active. This review discusses recent evidence for selective and sensitive control of Mg21 transport in the DCT and emphasizes the importance of this control in normal and abnormal renal Mg21 conservation. Normally, Mg21 absorption is load dependent in the distal tubule, whether delivery is altered by increasing luminal Mg21 concentration or increasing the flow rate into the DCT. With the use of microfluorescent studies with an established mouse distal convoluted tubule (MDCT) cell line, it was shown that Mg21 uptake was concentration and voltage dependent. Peptide hormones such asparathyroid hormone, calcitonin, glucagon, and arginine vasopressin enhance Mg21 absorption in the distal tubule and stimulate Mg21 uptake into MDCT cells. Prostaglandin E2 and isoproterenol increase Mg21 entry into MDCT cells. The current evidence indicates that cAMP-dependent protein kinase A, phospholipase C, and protein kinase C signaling pathways are involved in these responses. Steroid hormones have significant effects on distal Mg21 transport. Aldosterone does not alter basal Mg21 uptake but potentiates hormone-stimulated Mg21 entry in MDCT cells by increasing hormone-mediated cAMP formation. 1,25-Dihydroxyvitamin D3, on the other hand, stimulates basal Mg21 uptake. Elevation of plasma Mg21 or Ca21 inhibits hormone-stimulated cAMP accumulation and Mg21 uptake in MDCT cells through activation of extracellular Ca21/Mg21-sensing mechanisms. Mg21 restriction selectively increases Mg21 uptake with no effect on Ca21 absorption. This intrinsic cellular adaptation provides the sensitive and selective control of distal Mg21 transport. The distally acting diuretics amiloride and chlorothiazide stimulate Mg21 uptake in MDCT cells acting through changes in membrane voltage. A number of familial and acquired disorders have been described that emphasize the diversity of cellular controls affecting renal Mg21 balance. Although it is clear that many influences affect Mg21 transport within the DCT, the transport processes have not been identified.

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Abstract An immortalized cell line (designated MDCT) has been extensively used to investigate the cellular mechanisms of electrolyte transport within the mouse distal convoluted tubule. Mouse distal convoluted tubule cells possess many of the functional characteristics of the in vivo distal convoluted tubule. In the present study, we show that MDCT cells also possess a polyvalent cation-sensing mechanism that is responsive to extracellular magnesium and calcium. Southern hybridization of reverse transcribed-polymerase chain reaction (RT-PCR) products, sequence determination and Western analysis indicated that the calcium-sensing receptor (Casr) is expressed in MDCT cells. Using microfluorescence of single MDCT cells to determine cytosolic Ca21 signaling, it was shown that the polyvalent cation-sensing mechanism is sensitive to extracellular magnesium concentration ([Mg21]o) and extracellular calcium concentration ([Ca21]o) in concentration ranges normally observed in the plasma. Moreover, both [Mg21]o and [Ca21]o were effective in generating intracellular Ca21 transients in the presence of large concentrations of [Ca21]o and [Mg21]o, respectively. These responses are unlike those observed for the Casr in the parathyroid gland. Finally, activation of the polycationsensitive mechanism with either [Mg21]o or [Ca21]o inhibited parathyroid hormone-, calcitonin-, glucagon- and arginine vasopressin-stimulated cAMP release in MDCT cells. These studies indicate that immortalized MDCT cells possess a polyvalent cation-sensing mechanism and emphasize the important role this mechanism plays in modulating intracellular signals in response to changes in [Mg21]o as well as in [Ca21]o.

]]> http://jamesdai.com/longjundai/2009/09/extracellular-mg2-and-ca2-sensing-in-mouse-distal-convoluted-tubule-cells/feed/ 0 http://jamesdai.com/longjundai/2009/09/intracellular-mg2-and-magnesium-depletion-in-isolated-renal-thick-ascending-limb-cells/ http://jamesdai.com/longjundai/2009/09/intracellular-mg2-and-magnesium-depletion-in-isolated-renal-thick-ascending-limb-cells/#comments Thu, 17 Sep 2009 22:36:15 +0000 greedy http://jamesdai.com/longjundai/?p=118 Dai LJ, and Quamme GA: Intracellular Mg²⁺ and magnesium depletion in isolated renal  thick ascending limb cells. Journal of Clinical Investigation 88:1255-1264, 1991.

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Abstract Magnesium reabsorption and regulation within the kidney occur principally within the cortical thick ascending limb (cTAL) cells of the loop of Henle. Fluorometry with the dye, mag-fura- 2, was used to characterize intracellular Mg2″ concentration (IMg2j11) in single cTAL cells. Primary cell cultures were prepared from porcine kidneys using a double antibody technique (goat anti-human Tamm-Horsfall and rabbit anti-goat IgG antibodies). Basal [Ig2+"] was 0.52±0.02 mM, which was – 2% of the total cellular Mg. Cells cultured (16 h) in high magnesium media (5 mM) maintained basal [Mg2j1i, 0.48±0.02, in the normal range. However, cells cultured in nominally magnesium- free media possessed [Mg2J1j, 0.27±0.01 mM, which was associated with a significant increase in net Mg transport, (control, 0.19±0.03 and low Mg, 0.35±0.01 nmol. mg-' protein min-') as assessed by 'Mg uptake. Mg2'-depleted cells were subsequently placed in high Mg solution (5 mM) and the Mg2" refill rate was assessed by fluorescence. [Mg2"1 returned to normal basal levels, 0.53±0.03 mM, with a refill rate of 257±37 nM/s. Mg2" entry was not changed by 5.0 mM Ca2" or 2 mM Sr2+, Cd2+, Co2+, nor Ba2+ but was inhibited by Mn2+ = La3+ .,Gd3+ Ni2+ , Zn2+ Be2+ at 2 mM. Intracellular Ca2` and "Ca uptake was not altered by Mg depletion or Mg2+ refill, indicating that the entry is relatively specific to Mg2e. Mg2+ uptake was inhibited by nifedipine (117±20 nM/s), verapamil (165±34 nM/s), and diltiazem (194±19 nM/s) but enhanced by the dihydropyridine analogue, Bay K 8644 (366±71 nM/s). These antagonists and agonists were reversible with removal and IMg2+Jj subsequently returned to normal basal levels. Mg2+ entry rate was concentration and voltage dependent and maximally stimulated after 4 h in magnesium-free media. Cellular magnesium depletion results in increases in a Mg2+ refill rate which is dependent, in part, on de novo protein synthesis. These data provide evidence for novel Mg2+ entry pathways in cTAL cells which are specific for Mg2` and highly regulated. These entry pathways are likely involved with renal Mg2` homeostasis. (J. Clin. Invest. 1991. 88:1255-1264.)