Abstract
Mechanisms of acid release and intracellular pH (pHi) homeostasis were analysed in goldfish (Carassius auratus) gill cells in primary culture. The rate of acid secretion was measured using a cytosensor microphysiometer, and pHi was determined using the fluorescent probe 2′,7′-bis-(3-carboxypropyl)-5-(and-6)-carboxyfluorescein (BCPCF). Amiloride, a Na+ channel and Na+/H+ exchanger (NHE) inhibitor, had no effect on pHi, but acid secretion of the gill cells was significantly impaired. In the presence of amiloride, the intracellular acidification (achieved using the NH4Cl pulse technique) was more severe than in the absence of amiloride, and recovery from the acidosis was slowed down. Accordingly, acid secretion of gill cells was severely reduced in the absence of extracellular Na+. Under steady-state conditions, 4,4′-diisothiocyanatodihydro-stilbene-2,2′-disulfonic acid (DIDS), a HCO3−-transport inhibitor, caused a slow acidification of pHi, and acid secretion was significantly reduced. No recovery from intracellular acidification was observed in the presence of DIDS. Bafilomycin A1, an inhibitor of V-ATPase, had no effect on steady-state pHi and recovery from an intracellular acidification, whereas the rate of acid secretion under steady-state conditions was slightly reduced. Immunohistochemistry clearly revealed the presence of the V-ATPase B-subunit in goldfish gill lamellae. Taken together, these results suggest that a Na+-dependent HCO3− transport is the dominant mechanism besides an NHE and V-ATPase to control pHi in goldfish gill cells.
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Abbreviations
- DIDS :
-
4,4′-diisothiocyanatodihydro-stilbene-2,2′-disulfonic acid
- NHE :
-
Na+/H+ exchanger
- pH i :
-
intracellular pH
- pH e :
-
extracellular pH
- ANOVA :
-
analysis of variance
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We thank Simone Boesch and Georg Kern for fruitful discussion and technical support.
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Communicated by G. Heldmaier
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Sandbichler, A.M., Pelster, B. Acid–base regulation in isolated gill cells of the goldfish (Carassius auratus). J Comp Physiol B 174, 601–610 (2004). https://doi.org/10.1007/s00360-004-0449-x
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DOI: https://doi.org/10.1007/s00360-004-0449-x