Regulation of intracellular pH in LLC-PK1 cells studied using 31P-NMR spectroscopy

Biochim Biophys Acta. 1987 Feb 18;927(2):203-12. doi: 10.1016/0167-4889(87)90136-4.


31P-NMR spectroscopy was used to monitor intracellular pH (pHi) in a suspension of LLC-PK1 cells, a renal epithelial cell line. The regulation of intracellular pH (pHi) was studied during intracellular acidification with 20% CO2 or intracellular alkalinization with 30 mM NH4Cl. The steady-state pHi in bicarbonate-containing Ringer's solution (pHo 7.40) was 7.14 +/- 0.04 and in bicarbonate-free Ringer's solution (pHo 7.40) 7.24 +/- 0.04. When pHo was altered in nominally HCO3(-)-free Ringer's, the intracellular pHi changed to only a small extent between pHo 6.6 and pHo 7.6; beyond this range pHi was linearly related to pHo. Below pHo 6.6 the cell was capable of maintaining a delta pH of 0.2 pH unit (inside more alkaline), above pH 7.6 a delta pH of 0.4 unit could be generated (inside more acid). During exposure to 20% CO2 in HCO3(-)-free Ringer's solution, pHi dropped initially to 6.9 +/- 0.05, the rate of realkalinisation was found to be 0.071 pH unit X min-1. After removal of CO2 the pHi increased by 0.65 and the rate of reacidification was 0.056 pH unit X min-1. Exposure to 30 mM NH4Cl caused a raise of pHi by 0.48 pH unit and an initial rate of re-acidification of 0.063 pH unit X min-1, after removal of NH4Cl the pHi fell by 0.58 pH unit below the steady-state pHi, followed by a subsequent re-alkalinization of 0.083 pH unit X min-1. Under both experimental conditions, the pHi recovery after an intracellular acidification, introduced by exposure to 20% CO2 and by removal of NH4+, was found to be inhibited by 53% and 63%, respectively, in the absence of sodium and 60% and 72%, respectively, by 1 mM amiloride. These studies indicate that 31P-NMR can be used to monitor steady-state intracellular pH as well a pHi transients in suspensions of epithelial cells. The results support the view that LLC-PK1 cells use an Na+-H+ exchange system to readjust their internal pH after acid loading of the cell.

MeSH terms

  • Ammonium Chloride / pharmacology
  • Animals
  • Body Fluids / metabolism*
  • Carbon Dioxide / pharmacology
  • Carrier Proteins / metabolism
  • Cell Line
  • Epithelium / drug effects
  • Epithelium / metabolism
  • Hydrogen-Ion Concentration
  • Intracellular Fluid / metabolism*
  • Isotonic Solutions / pharmacology
  • Kidney / drug effects
  • Kidney / metabolism*
  • Magnetic Resonance Spectroscopy
  • Ringer's Solution
  • Sodium-Hydrogen Exchangers
  • Swine


  • Carrier Proteins
  • Isotonic Solutions
  • Sodium-Hydrogen Exchangers
  • Ammonium Chloride
  • Carbon Dioxide
  • Ringer's Solution