[Show abstract][Hide abstract] ABSTRACT: Fully-hydrated Ehrlich carcinoma ascites cells under the protective action of DMSO fully survived exposure to near-absolute zero temperature provided by liquid helium.
Physiological chemistry and physics and medical NMR 01/2008; 40:115-8.
[Show abstract][Hide abstract] ABSTRACT: In 1889 Abderhalden reported his discovery that there is no (or as shown later, little) sodium ion (Na+) in human red blood cells even though these cells live in a medium rich in Na+. History shows that all major theories of the living cell are built around this basic phenomenon seen in all the living cells that have been carefully examined. One of these theories has been steadily evolving but is yet-to-be widely known. Named the association-induction hypothesis (AIH), it has been presented thus far in four books dated 1962, 1984, 1992 and 2001 respectively. In this theory, the low Na+ in living cells originates from (i) an above-normal molecule-to-molecule interaction among the bulk-phase cell water molecules, in consequence of (ii) their (self-propagating) polarization-orientation by the backbone NHCO groups of (fully-extended) cell protein(s), when (iii) the protein(s) involved is under the control of the electron-withdrawing cardinal adsorbent (EWC), ATP. A mature human red blood cell (rbc) has no nucleus, nor other organelle. 64% of the rbc is water; 35% belongs to a single protein, hemoglobin (Hb). This twofold simplicity allows the concoction of an ultra-simple model (USM) of the red blood cell's cytoplasmic protoplasm, which comprises almost entirely of hemoglobin, water, K+ and ATP. Only in the USM, the ATP has been replaced by an artificial but theoretically authentic EWC, H+ (given as HCl). To test the theory with the aid of the USM, we filled dialysis sacs with a 40% solution of pure (ferri-) hemoglobin followed by incubating the sacs till equilibrium in solutions containing different amounts of HCI (including zero) but a constant (low) concentration of NaCl. We then determined the equilibrium ratio of the Na+ concentration inside the sac over that in the solution outside and refer to this ratio as qNaCl. When no H+ was added, the qNaCl stayed at unity as predicted by the theory. More important (and also predicted by the theory,) when the right amount of H+ had been added, qNaCl fell to the 0.1- 0.3 range found in living red blood (and other) cells. These and other findings presented confirm the AIH's theory of life at the most basic level: in the resting living state, microscopic, or nano-protoplasm, is the ultimate physical basis of life. (See Post Script on page 111.)
Physiological chemistry and physics and medical NMR 01/2008; 40:89-113.
[Show abstract][Hide abstract] ABSTRACT: We determined the equilibrium distribution of twenty-one nonmetabolized nonelectrolytes in frog muscle cells. In all cases, plots of the equilibrium intracellular concentrations of a solute in the cell water against the external concentrations of the solute yielded straight lines in agreement with the prediction of such a rectilinear plot by the polarized multilayer (PM) theory. The slopes of these straight lines yield the equilibrium distribution coefficients or q-value of that solute. It was shown that, again in agreement with the PM theory, the q-values of fourteen nonelectrolytes vary with the molecular volumes of the nonelectrolytes, obeying the "size rule", i.e., the larger the solute, the lower its q-value. The q-values of the remaining seven nonelectrolytes also decrease with their molecular volumes but on a separate curve. These q-value vs. molecular volume plots (q-v plots) show strong resemblance to similar q-v plots of solutes in dialysis sacs containing proteins and polymers assuming the fully-extended conformation (extrovert models) but no, or only weak, resemblance to q-v plots of solutions containing native globular proteins (introvert models). These findings also support the PM theory, according to which some protein(s) pervasively present in cells are in the fully-extended conformation; and that these fully extended cell protein(s) polarize(s) in multilayers all or virtually all cell water. The relationship between the q-values of the nonelectrolytes and the solutes' respective molecular volume are described by two sets of theoretical curves, calculated from an equation introduced in the preceding paper. Both curves were computed on the basis of the same exclusion intensity (Uvp = 126 cal/mole). This factor measures the extra water-to-water interaction of the polarized water which acts to keep solute out of the cell water in degree according to the size of the solute. The two curves are computed on the basis of two different values of U(s), which represents the surface or solute component of the polarization energy, describing the affinity of the solute for the dynamic water structure: one curve which roughly predicts the q-value of fourteen of the nonelectrolytes studied was computed with a U(s) equal to 119 cal/mole. The second curve which roughly predicts the q-value of the remaining seven solutes was computed on the basis of a higher U(s) (156 cal/mole).(ABSTRACT TRUNCATED AT 400 WORDS)
Physiological chemistry and physics and medical NMR 02/1993; 25(3):177-208.
[Show abstract][Hide abstract] ABSTRACT: High-molecular weight poly(ethylene glycol) (PEG-8000) in the bathing medium prolongs the survival of 2-mm-wide frog muscle segments with open ends. In a PEG-8000-containing medium Rb+, K+, and Na+ in the muscle segments reached new diffusion equilibrium in 2-4 hours. At this new equilibrium, the cell's preference of K+ over Na+ was preserved but very much weakened. Studies of the influence of pH on the equilibrium distribution of labelled Na+ in 2-mm-wide muscle segments confirmed the prediction that beta- and gamma-carboxyl groups, carried respectively on aspartic and glutamic acid residues of intracellular proteins, adsorb K+, Na+ and other monovalent cations. These carboxyl groups have a characteristic pKa between 3.65 and 4.25. A pKa of 3.85 was observed. These findings, when seen in the light of other relevant information available, led to the conclusion that beta- and gamma-carboxyl groups on myosin molecules adsorb--in a close contact one-ion-one-site fashion--the majority (67% to 80%) of K+ in resting muscle cells. Other evidence suggests that in muscle contraction, the K(+)-adsorbing beta- and gamma-carboxyl groups on myosin heads form salt linkages with cationic sites on actin, displacing and releasing the adsorbed K+. Present and earlier findings together offer support for an earlier suggestion that the formation and dissociation of these salt-linkages may underlie the force-generating, cyclic formation and dissociation of cross-bridges during muscle contraction.
Physiological chemistry and physics and medical NMR 02/1991; 23(3):133-60.
[Show abstract][Hide abstract] ABSTRACT: In this report, we describe the result of an extensive investigation of the effects of the conformations of proteins on the solvency of the bulk-phase water in which the proteins are dissolved. The concentrations of the proteins used were usually between 20 to 40%; the temperature was 25 degrees +/- 1 degree C. To probe the solvency of the water, the apparent equilibrium distribution coefficients (or p-values) of 4 solutes were studied: Na+ (sulfate), glycine, sucrose, and urea. From 8 to 14 isolated proteins in three types of conformations were investigated: native; denatured by agents that unravel the secondary structure (e.g., alpha-helix, beta-pleated sheet) of the protein (i.e., 9 M urea, 3 M guanidine HCl); denatured by agents that only disrupt the tertiary structure but leave the secondary structure intact or even strengthened (i.e., 0.1 M sodium dodecylsulfate or SDS, 2 M n-propanol). The results are as follows: (1) as a rule, native proteins have no or weak effect on the solvency of the water for all 4 probes; (2) exposure to 0.1 M SDS and to 2 M n-propanol, as a rule, does not significantly decrease the p-value of all 4 probes; (3) exposure to 9 M urea and to 3 M guanidine HCl consistently lowers the p-values of sucrose, glycine and Na+ (sulfate) and equally consistently produces no effect on the p-value of urea. Sucrose, glycine, and Na+ are found in low concentrations in cell water while urea is not. These experiments were designed and carried out primarily to test two subsidiary theories of the AI hypotheses: the polarized multilayer (PM) theory of cell water; and the theory of size-dependent solute exclusion.(ABSTRACT TRUNCATED AT 400 WORDS)
Physiological chemistry and physics and medical NMR 02/1989; 21(1):19-44.
[Show abstract][Hide abstract] ABSTRACT: Studies of the metabolism of four pentoses (D-, and L-arabinose; D-, and L-xylose) in frog muscle at 0 degree C revealed that all are metabolized at extremely slow rates. As a result, the metabolic degradation of these pentoses does not significantly affect their equilibrium distribution in muscle cells at this temperature at least. Of the four stereoisomers, three (L-xylose, D-arabinose, L-arabinose) were found exclusively or almost exclusively in the cell water, demonstrating a rectilinear distribution pattern; the equilibrium distribution coefficients (q-values) obtained from the slopes of these rectilinear distribution curves are 0.256, 0.274, and 0.271 respectively. The fourth pentose, D-xylose, is apparently partially adsorbed. With the aid of the equation for solute distribution according to the association-induction (AI) hypothesis, the data for this sugar can also be fitted with a theoretical curve calculated on the basis of a q-value close to those for the other three pentoses. The close similarity of the q-values of pentoses which are sterically different, but have identical molecular weights, provides further confirmation of the "size rule", a prediction of the polarized multilayer (PM) theory of cell water, according to which, the q-values are as a rule primarily determined by the molecular weights of the solute and to a lesser extent dependent on its stereospecificity.
Physiological chemistry and physics and medical NMR 02/1988; 20(4):309-17.
[Show abstract][Hide abstract] ABSTRACT: In this paper we studied the volume changes of dialysis sacs containing concentrated solutions of native and denatured proteins and of oxygen-containing polymers after immersion in aqueous solution of Na-citrate, D-glucose, and sorbitol of varying concentration. The results confirm the theory of cell volume regulation: volume changes of living cells in different solutions represent a balance between the tendency of intracellular proteins -- which exist in the fully extended conformation -- to polarize, sorb, and draw into the sac or cell more water and the opposite tendency to lose water from the sac or cell created by the lower level of the solutes in the cell or sac water than in the external medium. The lower level of the solutes is the consequence of the reduced solvency of the polarized water in the sac or cell water for large and complex solutes like sugar and free amino acids. This study adds another important physico-chemical attribute of the living cell that can be duplicated by aqueous solutions of gelatin, oxygen-containing polymers like PEO and PEG as well as urea-denatured proteins but not, or only weakly so, by aqueous solutions of native proteins or SDS-denatured proteins.
Physiological chemistry and physics and medical NMR 02/1987; 19(3):177-92.
[Show abstract][Hide abstract] ABSTRACT: The effects of 10(-7) M valinomycin, nonactin, and monactin on human erythrocytes, frog sartorius muscle, and frog ovarian oocytes in the presence of varying external K+ concentration were studied. The results showed essentially a consistent but relatively modest increase of the K+ permeability constant in cm/sec with all three antibiotics on human erythrocytes. No change in response to any one of the antibiotics was observed in frog muscles or in frog ovarian eggs. These results and reports of similar failure to demonstrate ionophore-mediated increase of K+ permeability in squid axon and inner membrane of the liver mitochondria led to the conclusion that lipid membrane barrier to ionic traffic may be significant in the human erythrocytes but even here one must regard the evidence as tentative. In contrast, for the majority of other cell types studied, the data indicate the primary, if not exclusive route of ion traffic, is via the nonlipid component of the cell membrane. The evidence that these nonlipid paths are the fixed charge-polarized water layer complex and that they cover much of the cell surface of many types of living cells was discussed.
Physiological chemistry and physics and medical NMR 02/1986; 18(2):109-24.
[Show abstract][Hide abstract] ABSTRACT: Incubation of frog sartorius muscles for 18 hours at constant external concentrations of K+ and Na+, but with the external concentration of Mg++ varying from 1.2 to 73.2 mM, brought about no change in the resting potential which remained constant at 88 mV. Results were the same in MgCl2 as in MgSO4. However, these cells are nearly as permeable to Mg++ as they are to K+. These results contradict the membrane theory of the cellular potential but are compatible with the surface adsorption model of the cellular potential.
Physiological chemistry and physics and medical NMR 02/1983; 15(5):379-90.
[Show abstract][Hide abstract] ABSTRACT: The quantitative relationships between the concentrations of solutions of gelatin, polyvinylpyrrolidone, poly(ethylene oxide), polyvinylmethylether, and poly(ethylene glycol), and their ability to reduce the solubility of water for Na citrate are presented. The data in general are in harmony with the polarized multilayer theory of protein (and polymer) dominated water in vitro and in living cells.
Physiological chemistry and physics and medical NMR 02/1983; 15(2):127-36.
[Show abstract][Hide abstract] ABSTRACT: 1. The effluxes of labeled Na+, D-arabinose, and sucrose from normal muscle and muscle poisoned with low concentrations of iodoacetate were studied. The procedure involved repeated loading with isotope, followed by washing of the same muscle while still normal and at different states of dying. 2. The rates of Na+ efflux in both the fast and slow fraction remained either quite constant or showed some unpredictable, minor fluctuations. This was true for both Na+ and the two sugars studied, confirming earlier conclusions that the steady levels of these solutes were not maintained by pumps. 3. In all cases studied, the efflux curves showed at least two fractions. It is the fast-exchanging fraction that steadily and consistently increased in magnitude as the muscles were dying, until finally the concentration of solute in this fraction reached and sometimes surpassed the labeled solute concentrations in the original labeled solutions in which the muscles were equilibrated. The slow fractions showed only a transient increase or none at all. These observations show that it is the fast fraction that represents solute dissolved in cell water and rate-limited by passage through the cell surface and that the partial exclusion of Na+ and the sugars have a unitary cause—a reduced solubility in the cell water which in the presence of ATP exists in the state of polarized multilayers.
[Show abstract][Hide abstract] ABSTRACT: The effluxes of labeled Na+ and labeled ~arabinose from single frog ovarian eggs were simultaneously recorded from normal eggs and eggs in various stages of deterioration following exposure to the rnetabolic poison Na iodoacetate. Findings were: I. At least two fractiorls of Na+ occrrr in these egg cells: one fast-exchanging fraction and one slowly exchanging fraction. Also, at least two fractions of ~arabinose occrrr: one fast exchanging, the other slowly exchanging. 2. Apparently the fast fractions of both Na+ and D-arabinose represent free Na+ or ~arabinose in the cell water with rates of efflux limited by passage through the cell surface. The slow fractions appear to represent Nu+ or D-arabinose adsorbed on rnacromolec~rlar sites; their rates of loss are limited by the rates of desorption. 3. While the poisoned cells steadily gain Na+, neither the rates of eflrrx of the fast-exchanging Na+ nor the rates of loss of the slowly exchanging Na-4- show evidence of a slow-down. This contradicts the Na-pump theory. 4. The most outstanding and consistent observation is a steady gain in con- centration of both the fast-exchanging Na+ and the fast-exchanging ~arabinose as the poison takes effect. Maximum concentrations are reached in dead eggs. 5. A correlation coefficient of +0.92 is demonstrated between the concen- trations of fast-exchanging Na+ and fast-exchanging ~arabinose in the eggs studied. This finding offers srrpport for the "universality rule" according to which the changing levels of No+ and D-arabinose, as well as other solutes nor- rnally excluded from the satlie cells, all /lave a unitary origin-the changing physical state of the cell water. Therefore changes of all these solutes should proceed pari passu and show positive correlation.
Physiological chemistry and physics 02/1977; 9(4-5):405-26.
[Show abstract][Hide abstract] ABSTRACT: The Na+ efflux curves of single ovarian eggs are separable into two fractions. The magnitude of the slow fraction increases slowly with time of exposure of the eggs to labeled Na+, long after the fast fraction has reached equilibrium. The data agree with the theory that the fast fraction is rate-limited by surface permeation and that the slow fraction is rate-limited by desorption from intracellular adsorption sites.
Physiological chemistry and physics 02/1977; 9(4-5):427-31.
[Show abstract][Hide abstract] ABSTRACT: At 25 degrees C, frog sartorius muslces rapidly gained Na+ and lost K+ in iodoacetamide and pure nitrogen. Beginning at normal levels, the concentrations of these ions in the cells reached those in the surrounding Ringer solution in 140 min. Yet during that time the Na+ efflux rate showed no sign of the slowing down demanded by Na-pump theory. The data support the view that maintenance and alterations of N1+ levels in frog muslce cells reflect adsorption on protein sites and the solubility property of bulk phase water and are independent of the rate at which Na+ leaves the cell surface.
Physiological chemistry and physics 02/1976; 8(5):389-95.
[Show abstract][Hide abstract] ABSTRACT: The diffusion coefficient of potassium ion (DK) in frog muscles was studied by a new method. In normal cytoplasm DK averaged one-eighth of the value in the free solution. Arrest of metabolism or injury caused an increase in DK. In muscles killed with iodoacetate, DK rose to three-quarters of the value in free solution. The data support the association-induction hypothesis.
[Show abstract][Hide abstract] ABSTRACT: With the use of a newly introduced technique, the "influx profile analysis," we studied the diffusion of tritiated water in and out of frog ovarian eggs at 25 degrees C. The results show that the rate-limiting step in the exchange of labeled water is not permeation through the cell membrane but diffusion in the bulk of the intracellular water.
The Journal of General Physiology 08/1967; 50(6):1807-20. · 4.57 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: A comparison is made between the quantitative predictions of equilibrium ionic distribution in living cells according to the membrane theory (Donnan equilibrium) and according to the association-induction hypothesis. This comparison shows that both theories predict competitive effects of one permeant ion on the equilibrium concentration of another permeant ion; but within the limit of experimental accuracy only the association-induction model predicts quantitatively significant specific competition of one specified ion with the accumulation of another specified ion. The equilibrium distributions of K(+), Rb(+), and Cs(+) ions in frog sartorius muscle were studied and quantitatively significant specific competition was demonstrated; these results favor the association-induction hypothesis (adsorption on cell proteins and protein complexes and partial exclusion from cell water). Based on this model we estimated that at 257deg;C, the apparent association constants for K(+), Rb(+), and Cs(+) ion are 665, 756, and 488 (mole/liter)(-1). We found that the total concentration of adsorption sites (no less than 240 mmole/kg of fresh cells) agrees with the analytically determined concentrations of beta- and gamma-carboxyl groups of muscle cell proteins (260 to 288 mmole/kg).
The Journal of General Physiology 04/1966; 49(4):819-43. · 4.57 Impact Factor