Endocardial endothelium modulates subendocardial pHi of rabbit papillary muscles: role of transendothelial HCO3? transport

OBJECTIVE: This study investigated whether endocardial endothelial cells contribute to intracellular pH (pH(i)) regulation of subjacent cardiomyocytes. METHODS: Right ventricular rabbit papillary muscles were loaded with the pH-sensitive dye 2',7'-bis-(2-carboxyethyl)-5-(and-6)-carboxyfluorescein, acetoxymethyl ester (BCECF-AM) to measure pH(i) and HCO(3)(-) equivalent influx or efflux in muscles with intact endocardial endothelium (+EE) and in muscles with the endocardial endothelium removed (-EE). RESULTS: In steady-state conditions, pH(i) was consistently higher in +EE than in -EE muscles (7.38+/-0.03, n=39 vs. 7.27+/-0.04, n=20, p<0.05). In +EE muscles, removal of HCO(3)(-) from the buffer solution or adding 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS), an inhibitor of HCO(3)(-) transport, reduced pH(i) from 7.38 to 7.24+/-0.06 (n=14) and to 7.16+/-0.10 (n=10), respectively, whereas in -EE muscles pH(i) decreased slightly from 7.27 to 7.15+/-0.05 (n=14) and to 7.13+/-0.04 (n=7). In addition, HCO(3)(-) equivalent efflux during alkali loads by NH(4)Cl pulses was smaller in +EE muscles than in -EE muscles (0.89+/-0.50 vs. 1.99+/-0.12 mmol/l/min, n=5, p<0.05) and was inhibited by DIDS. HCO(3)(-) equivalent influx during recovery from acid load imposed upon wash out of NH(4)Cl, was larger in +EE muscles than in -EE muscles (2.15+/-0.54 mmol/l/min, n=6, vs. 1.06+/-0.20 mmol/l/min, n=5). 5-(N-Ethyl-N-isopropyl)amiloride (EIPA), an inhibitor of Na(+)/H(+) exchange, decreased HCO(3)(-) equivalent influx by 50% in both muscle groups but influx was still significantly higher in +EE than in -EE muscles (1.18+/-0.23 vs. 0.57+/-0.07 mmol/l/min, p<0.05). Finally, endocardial endothelial cells cultured on collagen-coated inserts established a DIDS-sensitive transendothelial HCO(3)(-)gradient. CONCLUSION: These data suggest that the endocardial endothelium maintains transendothelial fluxes of HCO(3)(-) from luminal (blood) to basal (muscle) side of the cells, which modulate pH(i) regulation in subjacent cardiomyocytes.