Adaptation to metabolic acidosis by turtle urinary bladder.

Abstract

We utilized the turtle urinary bladder to study the mechanisms responsible for adaptation to metabolic acidosis. Bladders removed from acidotic turtles had a higher rate of H+ secretion in vitro than bladders from control turtles, despite identical extracellular pH. HCO3 secretion, however, was not different between the two groups. The increase in H+ secretion could be mediated by a decrease in intracellular pH and/or by an increase in the number of cells thought to be responsible for H+ secretion. To study this issue, we measured intracellular pH with the fluorescent dye 6-carboxyfluorescein diacetate and quantified the number of cells by fluorescence microscopy utilizing acridine orange, rhodamine 123, and 6-carboxyfluorescein diacetate in turtles receiving different acid loads. Urinary acidification measured in vivo was increased in turtles fed a low-acid load for 48 h and in turtles fed a high-acid load for 24-48 h. Intracellular pH was lower in bladders from turtles fed a high-acid load for 48 h but it was not different from controls in the other groups, indicating that intracellular pH cannot account for the adaptive increase in H+ secretion. Bladders from all groups fed an acid load had a higher number of cells with positive staining for acridine orange compared with controls. Double labeling with acridine orange and the mitochondrial stain rhodamine 123 or 6-carboxyfluorescein showed a significant increase in the number of mitochondria-rich cells between control and bladders from turtles fed an acid load. The increase in the number of rhodamine 123- or 6-carboxyfluorescein-positive cells was lower than the increase in acridine orange-positive cells, suggesting that the apparent increase in the number of acridine orange-positive cells is due to an increase in the number of acidic vesicles in the mitochondria-rich cells and in the granular cells rather than solely to an increase in the number of mitochondria-rich cells. Plasma membrane fraction prepared from control and acidotic bladders failed to disclose an increase in the putative H+-ATPase as assessed by enzymatic activity and transport studies. In conclusion, the present study suggests that the adaptive increase in H+ secretion in metabolic acidosis is associated both with an increase in the number of mitochondria-rich cells as well as with an increase in the number of acidic vesicles in these cells.