Journal of Neurophysiology, Vol 74, Issue 3 942-949, Copyright © 1995 by APS

ARTICLES

Calcium influx but not pH or ATP level mediates glutamate-induced changes in intracellular magnesium in cortical neurons

S. Rajdev and I. J. Reynolds
Department of Pharmacology, University of Pittsburgh, School of Medicine, Pennsylvania 15261, USA.

1. We have recently shown that glutamate increases [Mg2+]i in cultured rat cortical neurons. However, the mechanism of this increase in [Mg2+]i is not well understood. We used fluorescence microscopic methods to measure [Mg2+]i, [Ca2+]i, and pHi in single neurons. Intracellular ATP analysis was performed by high-performance liquid chromatography (HPLC). 2. A 25-mM NH4Cl pulse followed by Na(+)-free wash rapidly acidified the cytosol. In 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF)-loaded neurons, the pHi was reduced by 2.46 units, and in magfura-2-loaded neurons the [Mg2+]i was increased by 0.62 mM. Five-minute treatment with 100 microM glutamate, on the other hand, reduced the cytosolic pH by 0.73 units and increased the [Mg2+]i by 7.24 mM in rat cortical neurons. These results indicate that change in pHi does not play a significant role in the glutamate-induced [Mg2+]i elevation. 3. The metabolic inhibition (5 mM KCN and 1 mM iodoacetate) for 30 min significantly reduced the intracellular ATP levels. However, 5-min treatment with 100 microM glutamate did not significantly deplete intracellular ATP in cultured cortical neurons. When tested under similar conditions in magfura-2-loaded neurons, glutamate increased [Mg2+]i to a significantly larger extent than metabolic inhibition. This suggests that ATP depletion and subsequent release of Mg2+ from Mg(2+)-ATP complex is not the primary source of [Mg2+]i elevation observed during glutamate stimulation. 4. To further study the role of glutamate-induced Ca2+ influx in subsequent [Mg2+]i elevation, extracellular Ca2+ was elevated from 1.4 to 3.0 mM during glutamate application in magfura-2-loaded neurons.(ABSTRACT TRUNCATED AT 250 WORDS)

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