You might want to review this article by Guy E. Abraham, MD & Jorge Flechas, MD http://www.mgwater.com/gafibro.shtml

If you're interested, I can put you in touch w/ Dr. Abraham.

Mg may bind to albumin as well as calcium and then will be taken by receptors on the cells.endocytosis mediated by receptors

I haven't had to think about it recently, but it is a good question. You have to distinguish between in vitro and in vivo.

[1] There is ionic balance maintained between cations and anions on both sides of the cell membrane. 90% of calcium is in the bone matrix, and the balance is in plasma/serum depending on whether you keep the clotting factors.

[2] Mg++ is intracellular.

[3] the total body Mg++ is not measured in serum, and the level measured in serum may well not indicate Mg++ depletion.

[4] Now that you are in the cell, you want to know the balance between the cytoplasm and the inner or outer membrane of the mitochondrion.

[5] If you are talking about muscle, which is an energy generating cell with the contractile apparatus, it is another thing than liver cells, which are anabolic, that is, engaged in the synthesis of proteins and lipids, which is tied to NADP_NADPH, whereas muscle is more dependent on NAD_NADH.

[6] it is not generally discussed, but this is where the NAD_NADP transhydrogenase would really become important. The role of the transhydrogenases was a classic paper by Kaplan, Colowick and .... in JBC in the late 1940s, and it was the subject of the Harvey Lecture at Harvard in 1971.

You may be able to incubate your cells with MgCl2 and glucose together and after 1-2h

major both Atp and Mg content of the cell.This is what we have already done it with Al

Addition of glucose and Mg to incubation medium and measurement both levels following 1-2h..you may also add minor amount of BSA

It is not such straitforward. About Mg, knowing that it is a competitor of Calcium, increasing external Mg wil disturbe calcium available at the surface of the cells and the transmembrane calcium exchange. In short, changing Mg will influence calcium flux and signaling. About ATPsince ATP is involve in the regulation of glu transporters, there is little chance that increasing glu will significantly increase internal ATP, if ATP is normal ,in control situation; moreover glacose excess is cytotoxic. It must be aslo considered whether culture medium provides lipids...

If you had the right equipment, could acutely and rapidly increase intracellular Mg2+ using a "caged" Mg2+, using DM-nitrophen AM. See, for example:

Ellis-Davies, G.C. (2006) Cell Calcium. Jun;39(6):471-473.

Thanks a lot. Gentlemen. This is good for us. We originally want to increase the cellular content of ATP and Mg to favor lipid kinase reaction. However, direct addition of ATP seems not accessible to cell inside. Ali, would you pls offer a more detailed reference? It will grateful to me.

5-10 mmolar og glucose and icreasing level of MgCl2 from physiological level

The total nucleotide levels (ATP+ADP+AMP) within cells are well preserved. Under basic physiological states the concentrations of ADP and AMP compared to ATP are quite low. Therefore, increase of glucose levels would not help, albeit energetics of your cells is significantly compromised. Elevated glucose may only support intracellular ATP, since glycolysis-driven ATP production, as well as mitochondria aerobic ATP synthesis, is fairly ADP dependent, i.e. and as soon as you phosphorylate all ADP to ATP the benefit of additional substrates will be neglected.

Asking this question, do you mean to increase total intracellular nucleotide levels? Such an increase, in contrast to support intracellular ATP, would require promoting pathways for de novo ATP synthesis. To this end I am curious, why in general you need to elevate intracellular ATP? Its concentration within cells is usually well above Km values for any ATP-dependent processes? Having saturating ATP levels for cellular metabolism what you may expect increasing intracellular ATP?

I agree with Alexey, who explains in details what I saied previously. In short, ATP cellular content is esquisitely regulated and normal cells consume exactly the amount of substate they need, even if you provide substates in excess, glucose or FFA.

Agree with Alexey. Assuming ATP synthase is normal, you add more ADP to stimulate more ATP production than glucose.

Hi, Alexey. Thanks for your kind explanation. In fact, I do not mean to increase the total intracellular nucleotide levels. I am just considering increasing the ATP level, to increase the substrate level for lipid kinase reaction, which will resulted in increase of membrane PIP2, when this lipid is depleted. I found people were doing so in patch clamp by dialyse cell with Mg2+ and ATP through the patch pipette. However, we cannot do this, because we are dealing with intact cells. We do not want to have any damage to cell integrity. In considering your explanation, we probably need to change our target from ATP. Probably, the inhibition of phospholipase (that hydrolyze PIP2) is a selection.

Dear Dr. Alexey, Dr. Pierre, I tried 8 Bromo-ATP to increase intracellular ATP levels but it did not work. Does ATP regulates Katp channel expression? Is there any relationship between channel activity and its expression?

Dear khaja shameem Mohammed Abdul, if I correctly understood, you have applied 8-bromo-ATP as a membrane-permeable ATP analog. If so, I would doubt that the modification of adenine ring by addition of Br to the 8th position can significantly improve the extremely restricted membrane permeability of ATP molecule that anyway bears strong electrical charge (in contrast to 8-bromo-cAMP). Therefore, I would not expect detectable changes in intracellular ATP level in the presence of extracellular 8-bromo-ATP.

To the best of my knowledge, intracellular ATP does not affect the expression of K-ATP channels, yet it can be altered by certain signaling pathways involved in adaptive response to stress (e.g. Zingman et al., JMCC 51:72,2011; Zhu et al., BBRS 415:637,2011; Sierra et al., JBC 288:1568,2013).