Carbonic acid buffer species measured in real time with an intracellular microelectrode array

• Main Authors: Wietasch, Kristina, Kraig, Richard P.
• Format: Text
• Language: English
• Published: 1991
• Subjects: Article; Carbonic acid
• Online Access: http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2826694; http://www.ncbi.nlm.nih.gov/pubmed/1653544

Carbonic acid buffer anions, HCO3−andCO32−, play an instrumental role in a host of vital processes in animal cells and tissues. Yet study of carbonic acid buffer species is hampered because no means are available to simultaneously monitor them at a cellular level in a rapid and dynamic fashion. An ion-selective cocktail, previously reported to measure changes in bicarbonate activity (αHCO3−), was instead shown to be principally selective for αCO32−. Ion-selective micropipettes (ISMs) based on this exchanger and consisting of a 3:1:6 (volume) mixture of tri-n-octylpropylammonium chloride, 1-octanol, and trifluoroacetyl-p-butylbenzene showed no significant interference from bicarbonate, chloride, phosphate, ascorbate, lactate, glutamate, acetate, or hydroxyl ions at concentrations expected in vivo. Intracellular and triple-barrel ISMs, consisting of a CO32−-sensitive, pH-sensitive, and reference barrel, were fabricated. Skeletal muscle cells (n = 17) were penetrated in vivo and showed values of 74 ± 7 mV for membrane potential, 6.94 ± 0.09 pHi, and 11 ± 5 µM intracellular αCO32−, from which intracellular αHCO3− of 25 ± 10 mM and CO2 tension of 120 ± 55 Torr were calculated. All ion measurements reached a new steady state in 9 ± 2 s after cell penetration. Thus measurements of intracellular αCO32− and pH and associated levels of αHCO3 and CO2 tension can be determined in biological tissues and cells with a spatial and temporal resolution previously unattainable.