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Department of Medicine, University of British Columbia, Vancouver Hospital and Health Sciences Centre, Vancouver, British Columbia, Canada V6T 1Z3
Glucagon and
arginine vasopressin (AVP) enhance renal magnesium
conservation through actions within the loop of Henle and the distal
tubule. Studies were performed on an immortalized mouse distal
convoluted tubule (MDCT) cell line to characterize the cellular actions
of these hormones on Mg2+
transport in this segment of the distal tubule. Glucagon and AVP
increased cellular cAMP concentrations by about fivefold above basal
levels in normal and Mg2+-depleted
cells. Intracellular free Mg2+
concentration
([Mg2+]i)
was determined on single MDCT cells using microfluorescence with
mag-fura 2. To assess Mg2+ uptake,
MDCT cells were first Mg2+
depleted (0.22 ± 0.01 mM) by culturing in
Mg2+-free media for 16 h and then
placed in 1.5 mM MgCl2, and the [Mg2+]i
was determined.
[Mg2+]i
returned to basal levels, 0.53 ± 0.02 mM, with a mean
refill rate,
d([Mg2+]i)/dt,
of 164 ± 5 nM/s. Both glucagon and AVP stimulated
Mg2+ uptake into MDCT cells, 196 ± 11 and 189 ± 6 nM/s, respectively, at concentrations of 3 × 10
7 M and
107 M, respectively.
Enhanced Mg2+ uptake for each of
the hormones was concentration dependent and inhibited by the channel
blocker, nifedipine. Hormone stimulation of
Mg2+ entry was not dependent on
protein synthesis. 8-Bromo-cAMP,
104 M, enhanced
Mg2+ uptake (225 ± 13 nM/s),
whereas phorbol esters were without effect. Finally, protein kinase A
inhibition prevented glucagon and AVP stimulation of
Mg2+ uptake, supporting the notion
that the cAMP pathway is important as expected in the hormone action.
These studies demonstrate that glucagon and AVP stimulate
Mg2+ uptake in MDCT cells and
suggest that these hormones act to control magnesium conservation in
the convoluted segment of the distal tubule.
intracellular magnesium; fluorescence; channel blockers; intracellular adenosine 3',5'-cyclic monophosphate
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