Evidence for an electrogenic Na+-HCO3- symport in rat cardiac myocytes.
1. The perforated whole-cell configuration of patch clamp and the pH fluorescent indicator SNARF were used to determine the electrogenicity of the Na+-HCO3- cotransport in isolated rat ventricular myocytes. 2. Switching from Hepes buffer to HCO3- buffer at constant extracellular pH (pHo) hyperpolarized the resting membrane potential (RMP) by 2.9 +/- 0.4 mV (n = 9, P < 0.05). In the presence of HCO3-, the anion blocker SITS depolarized RMP by 2.6 +/- 0.5 mV (n = 5, P < 0.05). No HCO3--induced hyperpolarization was observed in the absence of extracellular Na+. The duration of the action potential measured at 50 % of repolarization time (APD50) was 29.2 +/- 6.1 % shorter in the presence of HCO3- than in its absence (n = 6, P < 0.05). 3. Quasi-steady-state currents were evoked by voltage-clamped ramps ranging from -130 to +30 mV, during 8 s. The development of a novel component of Na+-dependent and Cl--independent steady-state outward current was observed in the presence of HCO3-. The reversal potential (Erev) of the Na+-HCO3- cotransport current (INa,Bic) was measured at four different levels of extracellular Na+. A HCO3-:Na+ ratio compatible with a stoichiometry of 2:1 was detected. INa,Bic was also studied in isolation in standard whole-cell experiments. Under these conditions, INa,Bic reversed at -96.4 +/- 1.9 mV (n = 5), being consistent with the influx of 2 HCO3- ions per Na+ ion through the Na+-HCO3- cotransporter. 4. In the presence of external HCO3-, after 10 min of depolarizing the membrane potential (Em) with 45 mM extracellular K+, a significant intracellular alkalinization was detected (0.09 +/- 0. 03 pH units; n = 5, P < 0.05). No changes in pHi were observed when the myocytes were pre-treated with the anion blocker DIDS (0.001 +/- 0.024 pH units; n = 5, n.s.), or when exposed to Na+-free solutions (0.003 +/- 0.037 pH units; n = 6, n.s.). 5. The above results allow us to conclude that the cardiac Na+-HCO3- cotransport is electrogenic and has an influence on RMP and APD of rat ventricular cells.
Aiello
E A
EA
Centro de Investigaciones Cardiovasculares, Facultad de Ciencias Medicas, Universidad Nacional de La Plata, La Plata 1900, Argentina.
Petroff
M G
MG
Mattiazzi
A R
AR
Cingolani
H E
HE
D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / Q000494:pharmacology; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid / Q000494:pharmacology; D000818:Animals; D001578:Benzopyrans; D001639:Bicarbonates / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism*; D002478:Cells, Cultured; D005456:Fluorescent Dyes; D006531:HEPES; D006321:Heart / Q000502:physiology*; D006352:Heart Ventricles; D006863:Hydrogen-Ion Concentration; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D009206:Myocardium / Q000166:cytology / Q000378:metabolism; D018408:Patch-Clamp Techniques; D051381:Rats; D012964:Sodium / Q000378:metabolism / Q000494:pharmacology; D027982:Sodium-Bicarbonate Symporters; D013050:Spectrometry, Fluorescence
D001578:Benzopyrans; D001639:Bicarbonates; D002352:Carrier Proteins; D005456:Fluorescent Dyes; C063917:SNARF dye; D027982:Sodium-Bicarbonate Symporters; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; D012964:Sodium; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D006531:HEPES