Abstract
The cytosolic pH (pHi) regulatory mechanisms of peripheral blood human lymphocytes and the effect of okadaic acid on the activity of these mechanisms were studied by means of fluorescence imaging microscopy of 2',7'-bis-(carboxyethyl)-5(6')-carboxyfluorescein (BCECF)-loaded individual cells. Lymphocytes recover from a CO(2)-induced acid load in an extracellular Na+-dependent, intracellular Cl- -independent fashion. This pHi recovery is highly sensitive to the anion exchange inhibitor 4,4'-diisothiocyanato-stilbene-2,2'-disulfonic acid (DIDS) and minimally sensitive to the Na+/H+ exchange inhibitor amiloride, suggesting that it is mostly due to the action of a Na+-dependent HCO3- transporter. Extracellular Cl- and Na+ removal experiments point to the existence of a DIDS-sensitive Na+-independent Cl-/HCO3- exchanger. Preincubation with okadaic acid stimulates the pHi recovery rate from a CO2-induced acid load in the presence of DIDS (0.002 pHu/min vs. 0.065 pHu/min), but not in the presence of amiloride. Okadaic acid also accelerates the pHi elevation induced by Cl- removal (0.039 pHu/min vs. 0.067 pHu/min). In summary, these results indicate that okadaic acid stimulates the activity of Na+/H+ and Na+-independent Cl-/HCO3- exchangers, but has no effect on the activity of the Na+-dependent HCO3- transporter of human lymphocytes.
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