2--7--bis(carboxyethyl)-5(6)-carboxyfluorescein and Acidosis--Renal-Tubular

The effect of acid-base disturbances on sodium/proton (Na+/H+) exchange has been examined in animal models; however, few data are available from human studies. To test the effect of metabolic acidosis on Na+/H+ exchange in man, as well as to examine the relationship between Na+/H+ exchange and cytosolic calcium ([Ca2+]i), we measured both variables in patients with decreased renal function with mild metabolic acidosis (pH 7.34 +/- 0.06), in normal control subjects (pH 7.41 +/- 0.02), and in subjects before (pH 7.40 +/- 0.01), and after (pH 7.26 +/- 0.04) ammonium chloride (NH4Cl) 15 g for 5 d. Lymphocytes and platelets were loaded with the cytosolic pH (pHi) indicator 2'-7'-bis(carboxyethyl)-5,6-carboxyfluorescein and acidified to pH approximately 6.6 with propionic acid. To quantitate Na+/H+ exchange, dpHi/dt was determined at 1 min. [Ca2+]i was measured with fura-2. Na+/H+ exchange was significantly increased only in lymphocytes of patients with renal insufficiency. Neither intracellular pH (pHi) nor [Ca2+]i was different from controls. NH4Cl resulted in a significant increase in Na+/H+ exchange in lymphocytes, but not in platelets of normal subjects. Values of pHi and [Ca2+]i in either cell type remained unaffected. Since metabolic acidosis influenced Na+/H+ only in lymphocytes, but not in platelets, it is possible that protein synthesis may be involved in increasing Na+/H+ exchange.

Topics: Acidosis; Acidosis, Renal Tubular; Acute Disease; Blood Platelets; Carbon Dioxide; Carrier Proteins; Chronic Disease; Electrolytes; Fluoresceins; Fluorescent Dyes; Humans; Hydrogen-Ion Concentration; In Vitro Techniques; Kinetics; Lymphocytes; Male; Sodium; Sodium-Hydrogen Exchangers

The focus of this investigation was to compare data from isolated tubules and mitochondria of rat and dog kidneys subjected to altered medium pH (pHe) in vitro and to determine the effects of pH on ammonium production. Cytosolic pH (pHi) was determined using the fluoroprobe BCECF-AM; mitochondrial matrix pH (pHm) was estimated from the distribution of a labeled weak acid. The data indicate differences between rat and dog. pHi was consistently lower in rat than dog proximal tubules. In rat, pHm decreased in parallel with pHe and phi, accounting, at least in part, for the accelerated alpha-ketoglutarate dehydrogenase (alpha KGDH) flux, which is important in triggering increased ammoniagenesis. In dog, pHm varied to only a small extent with pHe and pHi changes. The results suggest that changes in pHm are unlikely to explain an increase in alpha KGDH flux in acute acidosis in the dog. However, pHe and pHi could conceivably lead to alterations in other factors than pHm, i.e. matrix Ca2+, which may facilitate accelerated ammonium production.

Topics: Acidosis, Renal Tubular; Ammonia; Animals; Calcium; Dogs; Fluoresceins; Fluorescent Dyes; Hydrogen-Ion Concentration; In Vitro Techniques; Ketoglutarate Dehydrogenase Complex; Kidney Cortex; Mitochondria; Rats; Substrate Specificity