Intracellular pH in isolated rat renal papillary thin limbs of Henle's loop

Abstract

Intracellular pH (pHi) was measured in isolated, nonperfused and perfused rat papillary thin limbs of Henle's loops in N-2-hydroxyethylpiperazine-N'-2-ethansulfonic acid (HEPES)- or HEPES/bicarbonate-buffered medium at pH 7.4 using the pH-sensitive fluorescent dye 2',7'-bis(2-carboxyethyl)-5,6-carboxyfluorescein (BCECF). Resting pHi was about 6.7 in descending thin limbs (DTL) and about 6.9 in ascending thin limbs (ATL), even with a medium pH of 7.4. These values appeared to reflect the acid pH of the blood in the neighboring vasa recta found in vivo. The resting pHi did not differ whether or not the medium contained bicarbonate although the total buffering capacity of the tubule cells was increased in the presence of bicarbonate. In nonperfused DTL and ATL, pHi was further acidified following an NH4Cl pulse. The rate of recovery of pHi from this level to the resting pHi was reduced by Na+ removal from the bath in both DTL and ATL and by the addition of ethylisopropylamiloride (EIPA) to the bath in the presence of Na+ in DTL. The rate of recovery was not affected by Cl- removal from the bath or K+ (75 mM) or 4,4'-diisothiocyanostilbene-2,2'-disulfonate (DIDS) addition to the bath in either DTL or ATL. These results suggest that the common, amiloride-sensitive, basolateral Na+/H+ exchanger plays a role in the regulation of pHi in rat papillary DTL but that a different basolateral Na+/H+ exchanger or a luminal Na+/H+ exchanger is important in rat papillary ATL.

Regulation of intracellular pH in rat renal inner medullary thin limbs of Henle’s loop

“…This acid pH in the vasa recta reflects an increase in pCO 2 and a decrease in [HCO 3 -] compared to the systemic blood [31]. We also found that the resting pH i of the thin limbs from this inner medullary region is significantly acid, even when these thin limbs are maintained in a standard buffer at pH 7.4 in vitro [18]. In addition, the resting pH i of the descending thin limbs (DTL) is significantly more acid than that of the ascending thin limbs (ATL) [18].…”

“…We also found that the resting pH i of the thin limbs from this inner medullary region is significantly acid, even when these thin limbs are maintained in a standard buffer at pH 7.4 in vitro [18]. In addition, the resting pH i of the descending thin limbs (DTL) is significantly more acid than that of the ascending thin limbs (ATL) [18]. We speculated that this difference between the limbs might reflect a cellular memory of the pH of the surrounding vasa recta blood in vivo [18].…”

“…The kidneys were removed and placed in one of two ice-cold buffers. For the initial series of experiments (see Results), the buffer was the same as that used previously [18]. It consisted of 280 mM sucrose and 10 mM HEPES.…”

“…In previous studies of the rat renal inner medulla, we measured the pH of the blood in the vasa recta in vivo and the intracellular pH (pH i ) of the thin limbs of Henle's loop in vitro [18,27,31]. We found that the pH of blood in the inner medullary vasa recta in vivo is significantly acid compared with that of the systemic blood entering the kidney through the renal artery [27].…”

“…In addition, the resting pH i of the descending thin limbs (DTL) is significantly more acid than that of the ascending thin limbs (ATL) [18]. We speculated that this difference between the limbs might reflect a cellular memory of the pH of the surrounding vasa recta blood in vivo [18]. Study of the recovery of pH i from the increased acidification produced by an NH 4 Cl pulse indicated that the pH i is probably regulated in part by one of the Na + /H + exchangers, but the data were far from clear [18].…”