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Sodium and potassium channels are ion-selective pores that span the cell membrane and serve to regulate and establish membrane potential. They are typically classified into two types:
Sodium ions (Na+) and potassium ions (K+) are particularly important in excitable cells, such as neurons and myocytes, as they function to create the action potential and to rest the cell's resting membrane potential.
There is a major difference in the sodium ion concentration inside and outside the cell (5-40 mM intracellular; 120-450 mM extracellular, depending on the organism). This concentration gradient is essential for nutrient uptake, to regulate concentrations of other intracellular ions and solutes, and to generate and transmit electrical impulses in excitable cells. The low intracellular sodium concentration requires the use of sensitive dyes able to register any small variation. Since intracellular potassium concentration is typically higher than sodium (in excess of 100 mM), Na+ indicators should respond selectively to Na+, not K+.
|Excitation (nm)||Emission (nm)||Kd (mM)|
|Ion NaTRIUM Green - TMA Salt / AM||488-517||540||20|
Ion NaTRIUM Green is a visible wavelength fluorescent indicator with a useful dynamic range for measuring cytosolic Na+ concentrations. The dye is excited by visible light. Although Ion NaTRIUM Green excites maximally at 517 nm, its exceptional brightness enables excitation at the standard 488 nm settings. Moreover, it works well for 2-photon excitation with near-infrared light. Ion NaTRIUM Green is also remarkable in its resistance to photobleaching and leakage.
The TMA salt is the water-soluble salt form of Ion NaTRIUM Green available for calibration purposes or for invasive loading, such as microinjection into cells or loading through a whole-cell patch electrode, as well as for in vitro calibration of the dye.
The acetoxymethyl (AM) ester form of Ion NaTRIUM Green is non-invasive and is the most popular method for loading fluorescent ion indicators into cells. The phenolic and carboxylic acid functions of the molecule are derivatized as AM esters. These esters make the molecule hydrophobic enough to be membrane permeant. Once inside the cell, non-specific esterases found in almost all cell types, hydrolyze the esters back to the polyanionic form necessary for water-solubility, retention in the cell, and sensing ions. Loading is usually performed in a serum-free culture medium with the AM ester at a final concentration ranging from 1-10 μM.
|Indicator||Excitation (nm)||Emission (nm)||Kd (mM)|
|Ion Potassium Green||488||540||18|
Ion Potassium Green is a fluorescent indicator with a useful Kd for measuring cytosolic K+ concentration. It loads readily and is excited by visible light. Although non-ratiometric, its large fluorescence dynamic range allows sensing of even small changes in K+ concentration. Optimal excitation occurs at 517 nm, but the indicator can also be excited at the conventional wavelength of 488 nm. Ion Potassium Green works well with 2-photon excitation at near-infrared wavelengths, and although not fully resistant to photobleaching, it is more robust than other dyes. Like its Na+ counterpart, it is useful for confocal microscopy, flow cytometry, and screening.
The Kd of PBFI for K+ is strongly dependent on whether Na+ is present, with a value of 5.1 mM in the absence of Na+ and 44 mM in solutions with a combined Na+ and K+ concentration of 135 mM (which approximates physiological ionic strength). Although the selectivity of PBFI for K+ is less than that of Ca2+ indicators such as Fura-2, it is sufficient for the detection of physiological concentrations of K+ in the presence of other monovalent cations. The spectral response of PBFI upon ion binding permit excitation ratio measurements, and this indicator can be used with the same optical filters and equipment used for Fura-2.
Ionophores facilitate the transport of Na+ ions through lipid membranes, with the net effect of allowing Na+ to flow from a compartment with higher Na+ concentration, through the membrane, into a compartment with lower Na+ concentrations. These ionophores can be used to equilibrate intracellular and extracellular Na+ concentrations.
Monensin sodium salt - Na+-selective carboxylic ionophore
Valinomycin is a natural, lipid-soluble molecule that binds potassium ions (K+) and facilitates their transfer across lipid bilayers. It exhibits a 10,000-fold selectivity for K+ over Na+. Valinomycin is a potent antibiotic. It is commonly used as an agent to induce apoptosis and can also be used to calibrate potentiometric responses in membrane experiments involving potential-sensitive dyes.
Nigericin is selective for both K+ and hydrogen ions (H+). Therefore it is commonly used in the fluorescence calibration of cytosolic pH indicators.
Monovalent cation ionophore that displays selectivity for K+ and NH4+ (K+ = NH4+ > Na+ > Mg2+ > Li+>> Ca2+). Induces cation transport across artificial membranes.