Novel chloride‐dependent acid loader in the guinea‐pig ventricular myocyte: part of a dual acid‐loading mechanism.

Abstract

1. The fall of intracellular pH (pHi) following the reduction of extracellular pH (pH.) was investigated in guinea-pig isolated ventricular myocytes using intracellular fluorescence measurements of carboxy-SNARF-1 (to monitor pH,). Cell superfusates were buffered either with a 5% CO2-HCO3 system or were nominally C02-HC03-free.2. Reduction of pHo from 7A4 to 6A4 reversibly reduced pHi by about 0 4 pH units, independent of the buffer system used.3. In HC03--free conditions, acid loading in low pHo was not dependent on Na`-H+ exchange or on the presence of Na+. It was unaffected by high-K+ solution, by voltage-clamp depolarization, by various divalent cations (Zn2+, Cd2+, Ni2+ and Ba2P) and by the organic Ca2+ channel blocker diltiazem, thus ruling out proton influx through H+-or Ca2+-conductance channels or influx via a K+-H+ exchanger. The fall also persisted in the presence of glycolytic inhibitors, or the lactate transport inhibitor, a-cyano-4-hydroxy cinnamate.4. In HC03--free conditions, acid loading in low pH0 was reversibly inhibited (by up to 85 %) by Cl-removal and was slowed by the stilbene drug DBDS (dibenzamidostilbene disulphonic acid). In contrast, the Cl--HCO3 exchange inhibitor DIDS (4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid) had no inhibitory effect. Acid loading is therefore mediated by a novel CF-dependent, acid influx pathway. 5. After switching to C02-HC03--buffered conditions, acid loading was doubled. It was still not inhibited by Nae-free or high-K+ solutions but was once again inhibited (by 78 %) in Clfree solution. The HC03--stimulated fraction of acid loading was inhibited by DIDS. 6. We propose a model of acid loading in the cardiomyocyte which consists of two parallel carriers. One is Cl--HCO3-exchange, while we suggest the other to be a novel ClF-OHexchanger (although we do not rule out the alternative configuration of H+-Cl-co-influx). The proposed dual acid-loading mechanism accounts for most of the sensitivity of pHi to a fall of pHo.Intracellular pH is an important physiological modulator of cardiac contraction (see Orchard & Kentish, 1990 for review). This is largely because reducing pH, reduces Ca2+ binding to troponin-C, thereby attenuating the force of contraction. Of the many conditions that can reduce pHi in cardiac cells, a common one is a reduction of extracellular pH. Over the physiological range of pH, intracellular pH is related approximately linearly to pH., with a transfer function (slope of pH1 vs. pHO) of 0 3-0'4 (Ellis & Thomas, 1976;Vaughan-Jones, 1986). Thus, because extracellular pH is an important modulator of pHi in cardiac cells, it becomes an important modulator of contractility in the heart. The mechaWism, however, whereby pHi is linked to pH0 remains unknown (Sun & Vaughan-Jones, 1994). This is the case not only for cardiac cells, but also for many other cell types. In the present work, we have investigated the mechanism linking pHi to pH0 in single enzymically isolated cardiomyocytes, while measuring pH1 using the pH-sensitive fluorophore, ...

Glucagon-Like Peptide-1 Increases cAMP but Fails to Augment Contraction in Adult Rat Cardiac Myocytes

“…We are aware of only one other report 42 3 Ϫ levels and, hence, NBC and AE activities, are negligible. Not surprisingly, acid influx due to CHE is very low at and below normal resting pH i levels and is negligible at pH i 6.95, 43 so it is unlikely to be responsible for acidosis in the present experiments. Even if CHE were somehow activated during ␤-AR or GLP-1 receptor stimulation by a previously unknown cAMP/PKAdependent mechanism, still NHE would be more than capable to substantially compensate and blunt the acidosis.…”

New insights into the physiological role of carbonic anhydrase IX in tumour pH regulation

“…Moreover, under certain circumstances, cells may incur a deficiency of H þ ions and require acid uptake (for example, by Cl Vasseur et al, 1989;Sun et al, 1996;Niederer et al, 2008), rather than acid extrusion.…”

“…Consequently, the acid-releasing cells that are more distant from capillaries will be surrounded by a milieu of lower pH e . As many membrane transporters are sensitive to pH e , this may feed back on acid/base traffic carried by such transporters (Vaughan-Jones and Wu, 1990;Sun et al, 1996;Stewart et al, 2009). Homeostasis of pH in tissue must therefore involve processes that regulate pH i and pH e .…”

Acidosis in models of cardiac ventricular myocytes

“…The regulation of intra-and extracellular pH is achieved by the transport of protons, bicarbonate and hydroxide ions across the cell membrane. The integrated control of this process is achieved by the balance of four separate transport proteins, each of which is specialized to a specific exchange cycle (Sun et al 1996). Two acid extruders, NBC and NHE, use the Na C gradient favouring Na C entry into the cell to extrude H C (in the case of NHE) or co-transport HCO K 3 (for NBC) into the cell.…”