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pHi in piglet cerebral microvascular endothelial cells: recovery from an acid load

, : pHi in piglet cerebral microvascular endothelial cells: recovery from an acid load. Proceedings of the Society for Experimental Biology and Medicine. Society for Experimental Biology and Medicine 212(3): 256-262

Cerebral microvascular endothelial cells form a barrier between the blood and brain, which is critical for normal neuronal functions. These endothelial cells can be challenged by metabolic and respiratory acidosis, especially in newborn babies. We investigated mechanism(s) by which cerebral endothelial cells recover intracellular pH (pH-i) when challenged with an intracellular acid load. pH-i in piglet cerebral microvascular endothelial cells in primary culture was monitored using the pH-sensitive fluorescent dye BCECF (2',7'-bis-2-carboxyethyl-5(6)-carboxy-fluorescein acetoxymethyl ester), with dual wavelength fluorescence spectroscopy. Endothelial cells attached to coverslips and continuously superfused with HCO-3-/CO-2 containing medium (25 MM HCO-3-, 5% CO-2; pH 7.40) have a steady state of pH-i of 7.18 +- 0.02. Under basal conditions, amiloride (100 mu-M) and H-2DIDS (0.5 mM) decreased pH-i 0.12 +- 0.01 and 0.05 +- 0.01 pH units, respectively. Removal of external Na+ lowered pH-i 0.18 +- 0.02 pH units, while Cl--free medium decreased pH-i 0.16 +- 0.03 pH units. These data suggest the presence of an amiloride-sensitive Na+-H+ exchanger and a Na+dependent HCO-3--Cl- anion exchanger In endothelial cells. Propionate and high PCO-2 cause rapid intracellular acidification at constant pH-e. The cells recover to control pH-i over 10 min. Recovery from propionate was largely inhibited by amiloride, slightly inhibited by H-2DIDS, and completely prevented by the combination. pH-i recovery during elevated PCO-2 was blocked by amiloride, H-2DIDS, or Na+-free media. These results indicate that recovery from intracellular acidosis can involve amiloride-sensitive Na+H+ exchange and a Na+-dependent HCO-3-/Cl- anion exchange. Relative contributions of pumps and their Independence appears to depend on the nature of the acid load.


PMID: 8677271

DOI: 10.3181/00379727-212-44014

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