Caption: FIGURE 3: (a) CA IX and CA II and (b) CA XII activities were assessed using MIMS, under normoxic or hypoxic conditions for 16 h in the presence or absence of the impermeant
sulfonamide inhibitor N-3500.
Instantly after adding red membrane impermeant
dyes, either EtBr (25 [micro]g/ml, Sigma) or EthD2 (25 [micro]g/ml, Life Technologies) was added at 0.5 ul per well.
We measured intercellular coupling by exposing freshly cut hearts to a solution containing the GJ-permeant dye, Lucifer Yellow, and the GJ impermeant
dye, Alexa 647 dextran.
Cells were subsequently labeled with 2.5 [micro]M of the cytoplasmic gap junction permeant calcein red-orange AM (Cell Trace; Molecular Probes/Invitrogen, C34851) for 1 hour, 5 [micro]M of the cytoplasmic gap junction impermeant
chloromethyl fluorescein diacetate (Cell Tracker Green CMFDA; Molecular Probes/Invitrogen, C7025) for 1 hour (with a media replacement after 30 minutes), or 5 [micro]M of the membrane labeling carbocyanine dyes DiO or DiD (Vibrant Cell Labeling Solution; Molecular Probes/Invitrogen, V22886, V-22887) for 1 hour.
Ethidium Bromide and Propidium Iodide are classic cell impermeant
DNA stains that are structurally similar to phenanthridium intercalators.
 presented a theoretical model that determined membrane pore size as a function of calcein (a cell impermeant
dye) uptake where calcein uptake is directly related to pore size (i.e.
Since propidium iodide, a dye highly impermeant
to intact plasma membranes, is unable to stain living cells, K562 cells exerting propidium fluorescence are dead or have compromised membranes.
Lithium uptake was measured in artificial seawater (ASW) solutions with Na replaced with the impermeant
cation choline and lithium (100% ASW assay solution: 9.4 mM KCl, 9.0 mM Ca[Cl.sub.2], 22.1 mM Mg[Cl.sub.2], 25.6 mM MgS[O.sub.4], 5.4 mM KHC[O.sub.3], 4.2 mM LiCl, and 420.8 mM choline chloride).
The gating currents were isolated by replacing permeant ions with impermeant
ones, thus reducing ionic currents, subtracting away symmetrical currents and, in most cases, blocking the ionic sodium current with tetrodotoxin [1, 2].
The medium was then replaced by fresh culture medium without serum nor antibiotics and the cells - with exception of the SH-SY5Y which were directly submitted to the next step - were further incubated for 30 min to allow full hydrolysis of Fura-2/acetoxymethyl by cellular esterases, resulting in cytosolic capture of the membrane impermeant
The inhibition of hexose transport by permeant and impermeant
sulfhydryl agents in rat adipocytes.
Although an in vivo calibration following application of quinidine could not be performed, an estimate of the change in intracellular sodium as a result of quinidine application, based on the calibration of the instrumentation with the cell impermeant
form of SBFI, reveals an increase of approximately 102 [+ or -] 6 mM (SE) intracellular sodium over baseline levels.