The sequestration of Na+, K+ and Cl− in the cellular nucleus and its energetic consequences for the gradient hypothesis of amino acid transport in Ehrlich cells

Pietrzyk, C.; Heinz, Erich

doi:10.1016/0005-2736(74)90231-4

Cited by 53 publications

(29 citation statements)

References 9 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…It is evident that in all of these studies, there are only small variations in N͞C values for Cl Ϫ and Na ϩ . Exceptions to these types of results, where N͞C ratios were assessed by analyzing nuclei in nonaqueous media, were seen in rat liver, where N͞C ϭ 7.9 for Cl Ϫ and 11.0 for Na ϩ (48) and in Ehrlich acites cells, where N͞C Ͼ Ͼ 1 for both Cl Ϫ and Na ϩ (49). These studies may be flawed due to redistribution of cellular ions during the preparative procedure.…”

Section: CLmentioning

confidence: 99%

Fluorescent imaging of Cl ^- in Amphiuma red blood cells: How the nuclear exclusion of Cl ^- affects the plasma membrane potential

Hoffman

Geibel²

2005

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

In this work, we test the idea that most, if not all, cellular Cl ؊ of Amphiuma red blood cells is contained in the cytoplasm. If true, this could resolve the difference between the measured plasma membrane potential (E m) and that expected from the Donnan equilibrium distribution of Cl ؊ . We studied the changes in the fluorescence intensity of the Cl ؊ -sensitive dye, MQAE, entrapped in red cells that occurred when intracellular Cl ؊ was exchanged with NO 3 ؊ . We could thus monitor the distribution of Cl ؊ between the nuclear and cytoplasmic compartments. We found that essentially all of the cell's Cl ؊ resides in the cytoplasm. Knowing the volume of the cell occupied by the nucleus, we could accordingly correct the measured values of cell Cl ؊ . This resulted in establishing a concordance between the measured values of E m and those calculated from the corrected values of the Cl ؊ ratio, thus explaining the discrepancy. The exclusion of Cl ؊ from the nucleus may result from its unusually high content of ''excess'' DNA that imposes an imbalance of net negative charge.MQAE ͉ nuclear͞cytoplasmic ratios T he equilibrium distribution of permeable ions such as Cl Ϫ across the plasma membrane of red blood cells should be determined by the membrane potential (E m ) that is set by the relative permeability to Na ϩ and K ϩ and their concentration gradients. This paper addresses the E m of Amphiuma red blood cells and the reason why the measured E m appears to differ significantly from that expected from the equilibrium distribution of Cl Ϫ . The idea that the E m of red blood cells in general and of human red cells in particular could be described by a Donnan potential is based on analyses (1-4) of the passive equilibrium distribution of permeable ions between the cytoplasm (Cl i Ϫ ) and the external medium (Cl o ). The E m of human red cells, measured indirectly with the use of fluorescent dyes (5), was found to vary quantitatively with the Donnan ratio of Cl i ͞Cl o (r Cl ). This was so when r Cl was varied either by changes in external pH (pH o ) (5-7) or by altering the concentration of Cl o by substitution with an impermeable anion (5). Validation that the fluorescence technique provided an accurate assessment of E m in human red cells stemmed from the concordance of E m determined directly in Amphiuma red blood cells by impalement with microelectrodes (8, 9, †) with those determined by the use of fluorescent dyes (5). These results established the basis for expecting measurements of E m in Amphiuma red blood cells to conform to a Donnan potential. However, analyses showed that, although the E m of Amphiuma responded to changes in Cl i produced by changes in pH o , the calculated values of E m based on measured values of Cl i and Cl o were significantly higher than the observed E m values. E m was calculated by use of the Nernst equation, E m ϭ 58 log Cl i ͞Cl o , where 58 is the value of RT͞F at 20°C., with R being the gas constant, T the temperature, and F, the Faraday constant. It turns out that the problem li...

show abstract

Section: CLmentioning

confidence: 99%

Fluorescent imaging of Cl ^- in Amphiuma red blood cells: How the nuclear exclusion of Cl ^- affects the plasma membrane potential

Hoffman

Geibel²

2005

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

show abstract

“…(12)), because the measured Vm which may be regarded as the initial value after a sudden change in external K+ concentration, was found to be reasonably constant with time and because the Nernst potentials for C1-(Ec1) and Na+ (ENa) are essentially constant in this region (see Christoffersen, 1973;Lassen, 1977;Lassen, Pape & Vestergaard-Bogind, 1978 Lassen, 1977;Lassen & Rasmussen, 1978). Although there is increasing evidence for the existence of an electrogenic Na+-K+ pump in Ehrlich ascites cells (Heinz et al 1975(Heinz et al , 1977 concentrations of K+, Na+, and Cl-, assuming equal activity coefficients in cell water and in the extracellular medium, and neglecting the available evidence ofintracellular compartmentation of Na+ and Cl- (Smith & Adams, 1977), or nuclear sequestration of these ions (Pietrzyk & Heinz, 1974 (Mills & Tupper, 1975;Tupper, 1975). This component would appear to be small, however, in cells with high intracellular K+ concentration, and suspended in ordinary Ringer solutions (see Fig.…”

Section: Saturation Of the Chloride Transportmentioning

confidence: 99%

Membrane potential, chloride exchange, and chloride conductance in Ehrlich mouse ascites tumour cells.

Hoffmann¹,

Simonsen²,

Sjøholm³

1979

The Journal of Physiology

109

View full text Add to dashboard Cite

SUMMARY1. The steady-state tracer exchange flux of chloride was measured at 10-150 mM external chloride concentration, substituting either lactate or sucrose for chloride. The chloride flux saturates in both cases with a K1 about 50 and 15 mm, respectively.2. The inhibitory effect of other monovalent anions on the chloride transport was investigated by measuring the 3601-efflux into media where either bromide, nitrate, or thiocyanate had been substituted for part of the chloride. The sequence of increasing affinity for the chloride transport system was found to be: Br-< Cl1 < SCN-= NO3-.3. The chloride steady-state exchange flux in the presence of nitrate can be described by Michaelis-Menten kinetics with nitrate as a competitive inhibitor of the chloride flux.4. The apparent activation energy (EA) was determined to be 67 + 6-2 kJ/mole, and was constant between 7 and 38 0C.5. The membrane potential (Vm) was measured as a function of the concentration of external K+, substituting K+ for Na+. The transference number for K+ (tK) was estimated from the slope of Vm vs. log10 (K+)e, and tcl and tNa were calculated, neglecting current carried by ions other than Cl-, K+, and Na+. The diffusional net flux of K+ was calculated from the steady-state exchange flux of 42K+, assuming the flux ratio equation to be valid. From this value the K+ conductance and the Na+ and Cl-conductances were calculated. The experiments showed that %t, GN., and GK are all about 14 ,US/cm2. 6. The net (conductive) chloride permeability derived from the chloride conductance was 4 x 10-8 cm/sec compared with the apparent permeability of 6 x 10-7 cm/ sec as calculated from the chloride tracer exchange flux. These data suggest that about 95 % of the chloride transport is mediated by an electrically silent exchange diffusion.7. Comparable effects of phloretin (0.25 mM) on the net (conductive) permeability and the apparent permeability to chloride (about 80 % inhibition) may indicate that the chloride exchange and conductance pathways are not completely separate and Experiments were performed at both addresses.

show abstract

“…Since the changes in Na1 and K1 produced by prolonged stimulation were much greater than those produced by 3 hr exposure to ouabain it seems unlikely that the ouabain-induced reduction in amino acid influx is a result of changes in Na1 and K1 and in the gradients of these two ions. A sodium binding effect like that found by Pietrzyk & Heinz (1974) is most unlikely in these experiments since Hinke (1961) found with Na-sensitive micro-electrodes that much of the axoplasmic Na (76 %) is unbound and remains so on stimulation.…”

Section: Effects Of Ouabain On Na and K Concentrations In The Axoplasmmentioning

confidence: 55%

The effect of ouabain on amino acid and orthophosphate influxes in squid giant axons.

Caldwell¹,

Lea²

1979

The Journal of Physiology

View full text Add to dashboard Cite

4. A detailed investigation of the time course of the inhibition of amino acid uptake by ouabain in which intracellular scintillator techniques were used, showed that there was no inhibition for 30-100 min, and then it occurred fairly quickly. This course of events was not affected by the protein synthesis inhibitor cycloheximide. It is unlikely that the inhibition of orthophosphate uptake is subject to a similar delay since this had reached a maximum after 40 min.5. It is concluded that the inhibitory effects of ouabain on amino acid uptake in squid axons are not due to effects on the Na and K gradients brought about by inhibition of the Na-K pump. The delay in the inhibition may reflect the operation of an intracellular regulatory mechanism, which is initiated when ouabain inhibits the pump but which takes time to operate. If the formation of an inhibitory second messenger molecule is involved this is unlikely to be a protein.

show abstract

The sequestration of Na+, K+ and Cl− in the cellular nucleus and its energetic consequences for the gradient hypothesis of amino acid transport in Ehrlich cells

Cited by 53 publications

References 9 publications

Fluorescent imaging of Cl ^- in Amphiuma red blood cells: How the nuclear exclusion of Cl ^- affects the plasma membrane potential

Fluorescent imaging of Cl ^- in Amphiuma red blood cells: How the nuclear exclusion of Cl ^- affects the plasma membrane potential

Membrane potential, chloride exchange, and chloride conductance in Ehrlich mouse ascites tumour cells.

The effect of ouabain on amino acid and orthophosphate influxes in squid giant axons.

Contact Info

Product

Resources

About

The sequestration of Na+, K+ and Cl− in the cellular nucleus and its energetic consequences for the gradient hypothesis of amino acid transport in Ehrlich cells

Cited by 53 publications

References 9 publications

Fluorescent imaging of Cl - in Amphiuma red blood cells: How the nuclear exclusion of Cl - affects the plasma membrane potential

Fluorescent imaging of Cl - in Amphiuma red blood cells: How the nuclear exclusion of Cl - affects the plasma membrane potential

Membrane potential, chloride exchange, and chloride conductance in Ehrlich mouse ascites tumour cells.

The effect of ouabain on amino acid and orthophosphate influxes in squid giant axons.

Contact Info

Product

Resources

About

Fluorescent imaging of Cl ^- in Amphiuma red blood cells: How the nuclear exclusion of Cl ^- affects the plasma membrane potential

Fluorescent imaging of Cl ^- in Amphiuma red blood cells: How the nuclear exclusion of Cl ^- affects the plasma membrane potential