Conservation of structure in ATP-depleted proximal tubules: Role of calcium, polyphosphoinositides, and glycine

Increases of intracellular free Ca²⁺ (Ca_f) may mediate phospholipid hydrolysis and disintegration in energy-compromised cells; on the other hand, glycine and related amino acids preserve structure. We have examined the effects of increased Ca_f on phospholipids and structure in ATP-depleted cells, as well as how these actions may be modified by glycine. Incubation of isolated proximal tubules with antimycin A led to ATP depletion, delayed increases of Ca_f to micromolar levels, polyphosphoinositide (PPI) hydrolysis by phospholipase C, and generalized disintegration of cell structure. Glycine inhibited PPI hydrolysis and preserved cell structure in entirety but did not apparently modify the Ca_f increases. When overwhelming increases of Ca_f were induced by the additional presence of a Ca²⁺ ionophore, glycine did not inhibit either the hydrolysis of PPI or disruption of mitochondria and microvilli. However, the cells remained integrated and unbroken. Incubation in low-Ca²⁺ medium prevented Ca_f increases, inhibited PPI hydrolysis, and preserved the structure of mitochondria and microvilli. Nevertheless, there was lethal damage by disintegration of all other membranes. This damage was prevented specifically and completely by glycine. Thus compartments of cells were shown to be differentially susceptible to injury from increased Ca_f or lack of glycine. Although damage by either factor occurs by distinct mechanisms, glycine also appears to have effects that suppress the deleterious effects of Ca²⁺ so long as Ca_f increases are not overwhelming. Our results also suggest that the PPI have a major structural role, which may be compromised by Ca_f increase during ATP depletion.