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J T Tupper

Publications and source records attributed to J T Tupper.

32 records · Page 2Linked to original sources

Potassium transport in normal and transformed mouse 3T3 cells.

The components of unidirectional K influx and efflux have been investigated in the 3T3 cell and the SV40 transformed 3T3 cell in expontntial and stationary growth phase. Over the cell densities used for transport experiments the 3T3 cell goes from exponential growth to density dependent inhibition of growth (4 X 10(4) to 4 X 10(5) cell cm-2) whereas the SV40 3T3 maintains exponential or near exponential growth (4 X 10(4) to 1 X 10(6) cell cm-2). In agreement with previous observations, volume per cell and mg protein per cell decrease with increasing cell density. Thus, transport measurements have been expressed on a per volume basis. Total unidirectional K influx and efflux in the 3T3 cell is approximately double that of the SV40 3T3 cell at all cell densities investigated. Both cell types have similar volumes initially and show similar decreases with increasing cell density. Thus, in this clone of the 3T3 cell SV40 transformation specifically decreases unidirectional K flux. The magnitude of the total K flux does not change substantially for either cell line during transition from sparse to dense cultures. However, the components of the K transport undergo distinct changes. Both cell lines possess a ouabain sensitive component of K influx, presumably representing the active inward K pump. Both also possess components of K influx and efflux sensitive to furosemide. The data suggest this component represents a one-for-one K exchange mechanism. The fraction of K influx mediated by the ouabain sensitive component is reduced to one half its value when exponential versus density inhibited 3T3 cells are compared (63% versus 31% of total influx). No comparable drop occurs in the SV40 3T3 cell at equivalent cell densities (64% versus 56% of total influx). Thus, the pump mediated component of K influx would appear to be correlated with growth. In contrast, the furosemide sensitive component represents approximately 20% of the total unidirectional K influx and efflux in both cell lines in sparse culture. At high cell densities, where growth inhibition occurs in the 3T3 cell but not the SV40 3T3, the furosemide sensitive component doubles in both cell lines. Thus, the apparent K-K exchange mechanism is density dependent rather than growth dependent.

Biological Transport, Active↗

Cation flux in the ehrlich ascites tumor cell. Evidence for Na+-for-Na+ and K+-for-K+ exchange diffusion.

In a previous study, evidence was presented for an external Na+-dependent, ouabain-insensitive component of Na+ efflux and an external K+-dependent component of K+ efflux in the Ehrlich ascites tumor cell. Evidence is now presented that these components are inhibited by the diuretic furosemide and that under conditions of normal extracellular Na+ and K+ they represent Na+-for-Na+ and K-+for-K+ exchange mechanisms. Using 86Rb to monitor K+ movements, furosemide is shown to inhibit an ouabain-insensitive component of Rb+ influx and a component of Rb+ efflux, both representing approx. 30 percent of the total flux. Inhibition of Rb+ efflux is greatly reduced by removal of extracellular K+. Furosemide does not alter steady-state levels of intracellular K+ and it does not prevent cells depleted of K+ by incubation in the cold from regaining K+ upon warming. Using 22Na to monitor Na+ movements, furosemide is shown to inhibit an ouabain-insensitive component of unidirectional Na+ efflux which represents approx. 22 percent of total Na+ efflux. Furosemide does not alter steady-state levels of intracellular Na+ and does not prevent removal of intracellular Na+ upon warming from cells loaded with Na+ by preincubation in the cold. The ability of furosemide to affect unidirectional Na+ and K+ fluxes but not net fluxes is consistent with the conclusion that these components of cation movement across the cell membrane represent one-for-one exchange mechanisms. Data are also presented which demonstrate that the uptake of alpha-aminoisobutyrate is not affected by furosemide. This indicates that these components of cation flux are not directly involved in the Na+-dependent amino acid transport system A.

Amino Acids↗

Cation permeability and ouabain-insensitive cation flux in the Ehrlich ascites tumor cell.

The components of Na and K flux across the plasma membrane have been investigated in the Ehrlich ascites tumor cell. At intracellular K levels of approximately 100 mM, unidirectional K influx is composed of a ouabain-sensitive component, a ouabain-insensitive, nondiffusional component and a diffusional component. Unidirectional K efflux is composed of an external K-dependent component and a diffusional component. Upon reduction of intracellular K to approximately 50 mM, the external K-dependent component becomes maximal and diminishes upon further reduction of intracellular K. Unidirectional Na efflux is composed of a ouabain-sensitive component, a diffusional component and a saturable, external Na-dependent, ouabain-insensitive component. Unidirectional Na influx may be accounted for by a diffusional component, based on estimates of membrane permeability to Na, membrane potential and Na distribution. This would suggest that the ouabain-insensitive, external Na-dependent Na efflux is not Na--Na exchange. The origin of the cell membrane potential has not been previously established in the Ehrlich ascites cell. From the diffusional components of Na and K flux determined in these experiments, the membrane permeabilities to Na and K have been estimated. These permeabilities, in conjunction with the Na and K distributions across the plasma membrane, predict a cell membrane potential of - 18mV (inside negative). Passive Cl distributions in these cells predict a cell membrane potential of - 21 mV, which is in agreement with previous microelectrode measurements and dibenzyldimethylammonium distributions. The results are therefore consistent with the conclusion that the magnitude and polarity of the cell membrane potential in the Ehrlich ascites cell is dictated primarily by Na and K.

Animals↗

Microelectrode studies on the membrane properties of isolated mitochondria.

The membrane potentials and resistances of isolated Drosophila mitochondria were measured after impalement with piezoelectric-driven microelectrodes. The potential is approximately +10 mv (positive inside), and the membrane resistance is approximately 2Omega.cm(2) under state 4 conditions. Experiments involving changes in external osmolality at constant ionic strength have suggested that the measured potential is across the mitochondrial semipermeable membrane. The mitochondria generally collapse seconds after impalement. In cases of successive impalements, potential and resistance values show slight decreases from the initial impalements.

Animals↗

Microelectrode studies on the membrane properties of isolated mitochondria. II. Absence of a metabolic dependence.

Mitochondrial membrane potentials and resistances have been determined under various metabolic conditions by the use of piezoelectric-driven microelectrodes. The mean state 3 potentials (about +19 mv) are higher than the potentials in state 4 (about +9 mv). DNP in uncoupling concentrations or KCN have no significant effect on membrane resistance or potential. Previous experiments have shown the potential to be sensitive to mitochondrial swelling and have suggested that it depends predominantly on anionic distribution.

Adenine Nucleotides↗