Intracellular pH transients in rainbow trout tissues measured by dimethadione distribution

Abstract

Intracellular pH transients in response to PCO2 changes were measured in vitro with the weak acid dimethadione (DMO) in gas-equilibrated red blood cells and a perfused trunk preparation (white muscle) of the rainbow trout. Red cell intracellular pH (pHi) was also measured directly on cell lysates. At an extracellular pH (pHe) of 7.81 +/- 0.04, PCO2 = 2 Torr, red cell pHilysate averaged 7.40 +/- 0.02, and [DMO]i/[DMO]e averaged 0.37 +/- 0.02, corresponding to a mean pHiDMO of 7.39 +/- 0.02. Decreasing pHe to 7.53 +/- 0.04 by increasing PCO2 to 8 Torr caused [DMO]i/[DMO]e to increase to 0.50 +/- 0.03 and resulted in a decline in pHi to a mean of 7.19 +/- 0.03 as measured by both techniques. With both methods red cell pHi responded rapidly (less than 5 min) to the PCO2 change, paralleling the response of pHe. In the isolated perfused trunk preparation at a perfusate pHe of 7.79 +/- 0.04 and PCO2 of 2 Torr, [DMO]i/[DMO]e averaged 0.38 +/- 0.03, yielding an average white muscle pHi of 7.35 +/- 0.04. Decreasing pHe to 7.34 +/- 0.02 by elevating PCO2 to 15 Torr caused pHi to drop to a mean of 7.11 +/- 0.03, as indicated by the significant increase in [DMO]i/[DMO]e to 0.58 +/- 0.03. The response of [DMO]i/[DMO]e was complete within 15 min. In both preparations the pHi changes were fully reversible. The DMO distribution method for measuring intracellular pH transients proved to be rapid and reliable in fish tissues.