Acute phosphate depletion and in vitro rat proximal tubule injury: Protection by glycine and acidosis

Acute phosphate depletion and in vitro rat proximal tubule injury: Protection by glycine and acidosis.The effects of phosphate (PO4) removal from Krebs Henseleit buffer on freshly isolated rat proximal tubules (rPT) were assessed by measuring Ca2+ uptake (nmol/mg protein), cellular adenosine triphosphate (ATP) (nmol/mg), tissue K+ content (nmol/mg) and lactate dehydrogenase (LDH) as an index of cell integrity. Ca2+ uptake increased by 50% in rPT incubated in zero PO4 medium as compared to control (2.6 ±0.1 vs. 3.9 ±0.19, P < 0.001) and LDH release increased 2.5-fold from 14.2 ±0.6 to 31.6 ±1.6%, P < 0.001. Neither verapamil (200 µM) nor mepacrine (50 µM) reduced Ca2+ uptake or decreased LDH release suggesting that the increased Ca2+ uptake was not occurring through potential operated channels and that phospholipase-induced cell injury was not the cause of increased LDH release. Either glycine (2 mM) or extracellular fluid acidosis (pH 7.06), however, significantly diminished rPT injury and Ca2+ uptake. Specifically, as compared to the increased LDH released in untreated, PO4-depleted rPT, LDH release was diminished significantly by glycine treatment (31.0 ±0.9 vs. 15.5 ±1.6%, P < 0.001) or acidosis (30.3 ±0.04 vs. 19.2 ±0.9%, P < 0.01). Ca2+ uptake did not increase in glycine treated tubules (2.6 ±0.1 vs. 2.8 ±0.2 nmol/mg, NS) or in the presence of acidosis (2.6 ±0.1 vs. 2.97 ±0.17 nmol/mg, NS). ATP concentrations were markedly reduced by PO4 depletion (2.8±0.2 vs. 4.8±0.3 nmol/mg, P < 0.001) and remained at low levels during either acidosis or glycine-induced protection. ATP depletion was accompanied by loss of K+ from rPT and this was only modestly attenuated by either glycine or acidosis. Total cell PO4 was not significantly altered, however, perchloric acid (PCA) extractable free PO4 was reduced significantly (33.3 ±4.5 to 15.9 ±3.5 nmol/mg, P < 0.01). The rPT injury, associated with acute PO4 depletion, may be related to Ca2+ uptake since Ca2+ uptake and LDH release were both attenuated by glycine administration or acidosis

Induced alterations in calcium uptake rate in normoxic rat proximal tubules

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