Intracellular pH regulation in isolated trout gill mitochondrion-rich (MR) cell subtypes: Evidence for Na+/H+ activity

We have studied intracellular pH (pHi) recovery in isolated trout gill mitochondrion-rich (MR) cells following acidification by the NH4Cl pre-pulse technique. Within a mixed MR cell population, one cell type displayed Na+-independent pHi recovery while the other cell type lacked a Na+-independent pHi recovery. Cells displaying Na+ independent recovery exhibited a significantly higher buffering capacity compared to cells lacking Na+-independent pHi recovery. Cells displaying Na+ independent recovery were identified as PNA+ (peanut lectin agluttinin binding) MR cells while those unable to recover were identified as PNA− (non-peanut lectin agluttinin binding) MR cells. Therefore, recovery from acidification in the absence of Na+ provides a direct functional marker for PNA+ and PNA− MR cells. Re-addition of Na+ to acidified cells resulted in a transient pHi recovery in both cell types. This event was abolished by amiloride (500 µM) but it was insensitive to phenamil (50 µM). The phorbol ester PMA (1 µM) potentiated the Na+ induced pHi recovery suggesting that activation by PKC is required for continuous Na+/H+ exchanger activity in trout gill MR cells. This study is the first functional description of pHi recovery in lectin-identified trout gill MR cells and provides insight into a putative cellular signaling mechanism that may control pHi regulation in the gill epithelium.