Metabolic Brain Disease

, Volume 2, Issue 3, pp 167–182 | Cite as

Intracellular pH determination by absorption spectrophotometry of neutral red

  • Joseph C. LaManna
Review Article


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Ackerman, J. J. H., Berkowitz, B. A., and Deuel, R. K. (1984). Phosphorus-31 NMR of rat brain in vivo with bloodless perfluorocarbon perfused rat.Biochem. Biophys. Res. Comm. 119: 913–919.Google Scholar
  2. Ahmed, Z., and Connor, J. A. (1980). Intracellular pH changes induced by calcium influx during electrical activity in molluscan neurons.J. Gen. Physiol. 75: 403–426.Google Scholar
  3. Barbosa, P., and Peters, M. (1971). The effects of vital dyes on living organisms with special reference to methylene blue and neutral red.Histochem. J. 3: 71–93.Google Scholar
  4. Battocletti, J. H. (1984). Medical applications of NMR spectroscopy. I. Clinical applications of phosphorus-31 NMR.CRC Crit. Rev. Biomed. Eng. 10: 1–25.Google Scholar
  5. Baylor, S. M., Chandler, W. K., and Marshall, M. W. (1982). Optical measurements of intracellular pH and magnesium in frog skeletal muscle fibres.J. Physiol. 331: 105–137.Google Scholar
  6. Bernard, M., Canioni, P., and Cozzone, P. J. (1983). Etude du metabolisme cellulaire in vivo par resonance magnetique nucleaire du phosphore-31.Biochimie 65: 449–470.Google Scholar
  7. Braun, A. D., and Nemchinskaya, V. L. (1958). The interaction of adenosine triphosphate with dyes.Biokim. 23: 359–365.Google Scholar
  8. Brooks, W. M., and Willis, R. J. (1985). Determination of intracellular pH in the Langendorff-perfused guinea-pig heart by 31P nuclear magnetic resonance spectroscopy.J. Mol. Cell. Cardiol. 17: 747–752.Google Scholar
  9. Bulychev, A., Trouet, A., and Tulkens, D. (1976). Accumulation and localization of neutral red in cultured fibroblasts.Arch. Int. Physiol. Biochem. 84: 1055–1056.Google Scholar
  10. Cady, E. B., Dawson, M. J., Hope, P. L., Tofts, P. S., Costello, A. M. deL., Delpy, D. T., Reynolds, E. O. R., and Wilkie, D. R. (1983). Non-invasive investigation of cerebral metabolism in newborn infants by phosphorus nuclear magnetic resonance spectroscopy.Lancet 1 (8333): 1059–1062.Google Scholar
  11. Caldwell, P. C. (1956). Intracellular pH.Int. Rev. Cytol. 5: 229–277.Google Scholar
  12. Chamberlain, S. C., Battelle, B.-A., and Caiman, B. G. (1984). Subcellular localization of neutral red staining inLimulus photoreceptors.J. Neurobiol. 15: 79–87.Google Scholar
  13. Chester, M., and Nicholson, C. (1985). Regulation of intracellular pH in vertebrate central neurons.Brain Res. 325: 313–316.Google Scholar
  14. Cohen, R. D., and Iles, R. A. (1975). Intracellular pH: Measurement, control, and metabolic interrelationships.CRC. Crit. Rev. Clin. Lab. Sci. 6: 101–143.Google Scholar
  15. Crozier, W. J. (1918). On indicators in animal tissues.J. Biol. Chem. 35: 455–460.Google Scholar
  16. Csiba, L., Paschen, W., and Hossman, K.-A. (1983). A topographical quantitative method for measuring brain tissue pH under physiological and pathophysiological conditions.Brain Res. 289: 334–337.Google Scholar
  17. Dell'Antone, P., Colonna, R., and Azzone, G. F. (1972a). The membrane structure studied with cationic dyes. 1. The binding of cationic dyes to submitochondrial particles and the question of the polarity of the ion-translocation mechanism.Eur. J. Biochem. 24: 553–565.Google Scholar
  18. Dell'Antone, P., Colonna, R., and Azzone, G. F. (1972b). The membrane structure studies with cationic dyes. 2. Aggregation, metachromatic effects and pKa shifts.Eur. J. Biochem. 24: 566–576.Google Scholar
  19. de Wolf, F. A., Groen, B. H., van Houte, L. P. A., Peters, F. A. L. J., Krab, K., and Kraayenhof, R. (1985). Studies on well-coupled photosystem I-enriched subchloroplast vesicles. Neutral red as a probe for external surface charge rather than internal protonation.Biochim. Biophys. Acta 809: 204–214.Google Scholar
  20. Djamgoz, M. B. A., and Laming, P. J. (1981). Intracellular measurement of ionic activity.TINS 4: 280–283.Google Scholar
  21. Federspil, G., Zaccaria, M., Reffo, G., and DePalo, C. (1972). Metabolic effects of neutral red in rats.Acta Diabetol. Latina (Milano) 9: 562–576.Google Scholar
  22. Gadian, D. G., Radda, G. K., Dawson, M. J., and Wilkie, D. R. (1982). pH measurements of cardiac and skeletal muscle using 31P-NMR. In Nuccitelli, R., and Deamer, D. W. (eds.),Intracellular pH: Its Measurement, Regulation, and Utilization in Cellular Functions, Alan R. Liss, New York, pp. 61–77.Google Scholar
  23. Gerson, D. F. (1982). Determination of intracellular pH changes in lymphocytes with 4-methylumbelliferone by flow microfluorometry. In Nuccitelli, R., and Deamer, D. W. (eds.),Intracellular pH: Its Measurement, Regulation, and Utilization in Cellular Functions, Alan R. Liss, New York, pp. 125–133.Google Scholar
  24. Giaume, C., and Kado, R. T. (1983). Application of antimony microelectrodes to intracellular pH monitoring.Biochim. Biophys. Acta 762: 337–343.Google Scholar
  25. Gillies, R. J., and Deamer, D. W. (1979). Intracellular pH: Methods and applications.Curr. Top. Bioenerg. 9: 63–87.Google Scholar
  26. Gillies, R. J., Alger, J. R., den Hollander, J. A., and Shulman, R. G. (1982). Intracellular pH measured by NMR: Methods and results. In Nuccitelli, R., and Deamer, D. W. (eds.),Intracellular pH: Its Measurement, Regulation, and Utilization in Cellular Functions, Alan R. Liss, New York, pp. 79–104.Google Scholar
  27. Gomba, S., Szokoly, V., and Soltesz, B. M. (1967). The effect of basic vital dyes on the acid phosphatase activity of the granulated juxtaglomerular cells.Experientia 23: 422–423.Google Scholar
  28. Gutter, B., Speck, W. T., and Rosenkranz, H. S. (1977). Light-induced mutagenicity of neutral red (3-amino-7-aimethylamino-2-methylphenazine hydrochloride).Cancer Res. 37: 1112–1114.Google Scholar
  29. Harris, J. E., Grubb, L., and Hoskinson, G. (1958). The effect of methylene blue and certain other dyes on cation transport and hydration of the rabbit lens.Am. J. Ophthalmol. 47: 387–395.Google Scholar
  30. Harvey, E. N. (1911). Studies on the permeability of cells.J. Exp. Zool. 10: 507–556.Google Scholar
  31. Haselgrove, J. C., Subramanian, V. H., Leigh, J. S. Jr., Gyulai, L., and Chance, B. (1983). In vivo onedimensional imaging of phosphorus metabolites by phosphorus-31 nuclear magnetic resonance.Science 220: 1170–1173.Google Scholar
  32. Heiple, J. M., and Taylor, D. L. (1982). An optical technique for measurement of intracellular pH in single living cells. In Nuccitelli, R., and Deamer, D. W. (eds.),Intracellular pH: Its Measurement, Regulation, and Utilization in Cellular Functions, Alan R. Liss, New York, pp. 21–54.Google Scholar
  33. Herbst, M., and Piontek, P. (1972). Uber den verlauf des intracellularen pH-wertes des skeletmuskels wahrend der kontraktion.Pfluger's Arch. 335: 213–223.Google Scholar
  34. Hersey, S. J. (1979). Intracellular pH measurements in gastric mucosa.Am. J. Physiol. 237: E82-E89.Google Scholar
  35. Jacobs, M. H. (1922). The influence of ammonium salts on cell reaction.J. Gen. Physiol. 5: 181–188.Google Scholar
  36. Jacobus, W. E., and Saks, V. A. (1982). Creatine kinase of heart mitochondria: Changes in its kinetic properties induced by coupling to oxidative phosphorylation.Arch. Biochem. Biophys. 219: 167–178.Google Scholar
  37. Kobatake, K., Sako, K., Izawa, M., Yamamoto, Y. L., and Hakim, A. M. (1984). Autoradiographic determination of brain pH following middle cerebral artery occlusion in the rat.Stroke 15: 540–547.Google Scholar
  38. Koehring, V. (1930). The neutral red reaction.J. Morphol. Physiol. 49: 45–137.Google Scholar
  39. Kogure, K., Alonso, O., and Martinez, E. (1980). A topographic measurement of brain pH.Brain Res. 195: 95–109.Google Scholar
  40. Kramer, K. J., Childs, C. N., and Speirs, R. D. (1979). Effect of neutral red on carbohydrate levels in the tobacco hornworm, manduca sexta (L.) (Lepidoptera: shingidae).Comp. Biochem. Physiol. 64C: 229–230.Google Scholar
  41. LaManna, J. C., and McCracken, K. A. (1985). The use of neutral red as an intracellular pH indicator in rat brain cortexin vivo. Anal. Biochem.142: 117–125.Google Scholar
  42. LaManna, J. C., Saive, J. J., Macdonald, V. W., and Jobsis, F. F. (1978). Simultaneous monitoring by optical techniques of respiratory chain and intracellular pH in toad ventricle strip.Experientia 34: 203–206.Google Scholar
  43. LaManna, J. C., Saive, J. J., and Snow, T. R. (1980). The relative time course of early changes in mitochondrial function and intracellular pH during hypoxia in the isolated toad ventricle strip.Circ. Res. 46: 755–763.Google Scholar
  44. LaManna, J. C., Lust, W. D., Whittingham, T. S., Selman, W. R., VanDer Veer, C. A., Crumrine, R. C., and Ratcheson, R. A. (1985). Evidence for tissue metabolic stress extending beyond the borders of perfusion deficits in a rat model of focal stroke.J. Cereb. Blood Flow Metab. (Suppl.) 5: S305-S3065.Google Scholar
  45. LaManna, J. C., McCracken, K. A., Whittingham, T. S., and Lust, W. D. (1986). Determination of intracellular pH by color film histophotometry of frozen in situ rat brain. In Longmuir, I. S. (ed.),Oxygen Transport to Tissue VIII, Plenum Press, New York, pp. 253–259.Google Scholar
  46. Lee, H. C., Forte, J. G., and Epel, D. (1982). The use of fluorescent amines for the measurement of pH: Applications in liposomes, gastric microsomes, and sea urchin gametes. In Nuccitelli, R., and Deamer, D. W. (eds.),Intracellular pH: Its Measurement, Regulation, and Utilization in Cellular Functions, Alan R. Liss, New York, pp. 135–160.Google Scholar
  47. Lockwood, A. H., Busto, R., Kogure, K., Finn, R. D., Butler, C. M., Alonso, O. F., and Gutierrez, M. T. (1981). Strategies for measuring brain pH by positron-emission tomography.J. Cereb. Blood Flow Metabol. (Suppl.) 1: S41.Google Scholar
  48. Lutaya, G., Rahim, Z. H. A., Shuttlewood, R. J., Bashford, C. L., and Griffiths, J. R. (1981). Alkalinization of phosphorylase kinase-deficient muscle during tetanic contraction.Biosci. Rep. 1: 177–182.Google Scholar
  49. Macdonald, V. W., and Jobsis, F. F. (1976). Spectrophotometric studies on the pH of frog skeletal muscle.J. Gen. Physiol. 68: 179–195.Google Scholar
  50. Macdonald, V. W., Keizer, J. H., and Jobsis, F. F. (1977). Spectrophotometric measurements of metabolically induced pH changes in frog skeletal muscle.Arch. Biochem. Biophys. 184: 423–430.Google Scholar
  51. Moon, R. B., and Richards, J. H. (1973). Determination of intracellular pH by 31P magnetic resonance.J. Biol. Chem. 248: 7276–7278.Google Scholar
  52. Nair, P. K., Spande, J. I., and Whalen, W. J. (1984). A microelectrode for measuring intracellular pH. In Bruley, D., Bicher, H. I., and Reneau, D. (eds.),Oxygen Transport to Tissue VI, Plenum Press, New York, pp. 881–886.Google Scholar
  53. Okuda, T., and Grollman, A. (1966). Action of neutral red on the secretion of glucagon and glucose metabolism in the rat.Endocrinology 78: 195–203.Google Scholar
  54. Petroff, O. A. C., Prochard, J. W., Behar, K. L., Alger, J. R., den Hollander, J. A., and Shulman, R. G. (1985). Cerebral intracellular pH by 31P nuclear magnetic resonance spectroscopy.Neurology 35: 781–788.Google Scholar
  55. Pick, U., and Avron, M. (1976). Neutral red response as a measure of the pH gradient across chloroplast membranes in the light.FEBS Lett. 65: 348–353.Google Scholar
  56. Prince, R. C., Linkletter, S. J. G., and Dutton, P. L. (1981). The thermodynamic properties of some commonly used oxidation-reduction mediators, inhibitors, and dyes, as determined by polarography.Biochim. Biophys. Acta 635: 132–148.Google Scholar
  57. Roberts, J. K. M., Wade-Jardetzky, N., and Jardetzky, O. (1981). Intracellular pH measurements by 31P nuclear magnetic resonance. Influence of factors other than pH on 31P chemical shifts.Biochemistry 20: 5389–5394.Google Scholar
  58. Roos, A., and Boron, W. F. (1981). Intracellular pH.Physiol. Rev. 61: 296–434.Google Scholar
  59. Roos, A., and Keifer, D. W. (1982). Estimation of intracellular pH from distribution of weak electrolytes. In Nuccitelli, R., and Deamer, D. W. (eds.),Intracellular pH: Its Measurement, Regulation, and Utilization in Cellular Functions, Alan R. Liss, New York, pp. 55–59.Google Scholar
  60. Salhany, J. M., Pieper, G. M., Wu, S., Todd, G. L., Clayton, F. C., and Eliot, R. S. (1979). 31P nuclear magnetics resonance measurement of cardiac pH in perfused guinea-pig hearts.J. Mol. Cell. Cardiol. 11: 601–610.Google Scholar
  61. Schmidtmann, M. (1925). Über die intracellulare wasserstoffionenkonzentration unter physiologischen und einigen pathologischen bedingungen.Z. Ges. Exp. Med. 45: 714–742.Google Scholar
  62. Sick, T. J., and LaManna, J. C. (1986). Changes in intracellular pH in hippocampal slices during anoxia measured with neutral red.Fed. Proc. 45: 903.Google Scholar
  63. Siesjo, B. K., Folbergrova, J., and MacMillan, V. (1972). The effect of hypercapnia upon intracellular pH in the brain, evaluated by the bicarbonate-carbonic acid method and from the creatine phosphokinase equilibrium.J. Neurochem. 19: 2483–2495.Google Scholar
  64. Skoogh, B.-E., Grillo, M. A., and Nadel, J. A. (1983). Neutral red stains ganglia in the vagal motor pathway to ferret trachea without affecting ganglionic transmission.J. Neurosci. Meth. 8: 33–39.Google Scholar
  65. Slavik, J. (1983). Intracellular pH topography: Determination by a fluorescent probe.FEBS 156: 227–230.Google Scholar
  66. Snow, T. R., Saive, J.-J., and Hemstreet, T. M. (1982). A study of the temporal relation between intracellular pH and contractile performance in toad ventricle strips during hypercapnic acidosis.J. Mol. Cell. Cardiol. 14: 1–12.Google Scholar
  67. Stolarsky, F., and Haley, T. J. (1951). Acute and chronic toxicity of toluidine blue and neutral red.Fed. Proc. 10: 337.Google Scholar
  68. Sundt, T. M., and Anderson, R. E. (1980). Umbelliferone as an intra-cellular pH-sensitive fluorescent indicator and blood-brain barrier probe: Instrumentation, calibration, and analysis.J. Neurophysiol. 44: 60–75.Google Scholar
  69. Sundt, T. M., Anderson, R. E., and VanDyke, R. A. (1978). Brain pH measurements using a diffusible, lipid soluble pH sensitive fluorescent indicator.J. Neurochem. 31: 627–635.Google Scholar
  70. Taylor, J. S., and Deutsch, C. (1983). Fluorinated a-methylamino acids as 19F NMR indicators of intracellular pH.Biophys. J. 43: 261–267.Google Scholar
  71. Thomas, R. C. (1974). Intracellular pH of snail neurones measured with a new pH-sensitive glass microelectrode.J. Physiol. 238: 159–180.Google Scholar
  72. Visser, J. W. M., Jongeling, A. A. M., and Tanke, H. J. (1979). Intracellular pH-determination by fluorescence measurements.J. Histo. Cyto. 27: 32–35.Google Scholar
  73. Waddell, W. J., and Bates, R. G. (1969). Intracellular pH.Physiol. Rev. 49: 285–329.Google Scholar
  74. Walz, F. G., Terrena, B., and Rokince, D. (1975). Equilibrium studies on neutral red—DNA binding.Biopolymers 14: 825–837.Google Scholar
  75. Warburg, O. (1910). Uber die Oxydationen in lebenden zellen nach versuchen am seeiglei.Hoppe-Seyler Z. Physiol. Chem. 66: 305–340.Google Scholar

Copyright information

© Plenum Publishing Corporation 1987

Authors and Affiliations

  • Joseph C. LaManna
    • 1
    • 2
  1. 1.Department of NeurologyUniversity HospitalsUSA
  2. 2.Departments of Neurology and Physiology/BiophysicsCase Western University School of MedicineCleveland

Personalised recommendations