Studies on Ion Accumulation in Muscle Cells

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 25°C, the apparent association constants for K +, Rb +, and Cs + ion are 665, 756, and 488 (mole/liter) -l. 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 t-and "y-carboxyl groups of muscle cell proteins (260 to 288 mmole/kg). I N T R O D U C T I O NIons and other solutes do not, as a rule, distribute themselves within cells at the concentrations anticipated on the basis of simple thermodynamic equilibria (1, 2). Past study of this phenomenon has often been hampered by difficulties in preserving isolated cells in reasonably good condition for a prolonged period of time. Recently, by adapting tissue culture methods, we have overcome this problem and have been able to investigate quantitatively the steady ionic level reached in the muscle cell following changes of the external ionic environment.A priori, a constantly maintained level of an ion or other solute in the cell m a y represent either an equilibrium phenomenon or a steady-state phenomenon. The maintenance of an equilibrium needs no expenditure of free energy. However, in the maintenance of a steady state, which must involve active transport or " p u m p s " of some sort, a continued free energy expenditure is mandatory. Comparison of the total energy available to the muscle cell

NMR Evidence for Complexing of Na+ in Muscle, Kidney, and Brain, and by Actomyosin. The Relation of Cellular Complexing of Na+ to Water Structure and to Transport Kinetics

“…Similar microelectrode results were obtained by Hinke (54) and McLaughlin and Hinke (55). The N M R evidence supports the theoretical and experimental work of Troshin (4, 5), Ling (6)(7)(8)(9)(10)(11)(12), and Simon (13)(14)(15), and the kinetic theory of Cope (1,53). I363…”

“…Prior to the use of N M R , the most direct evidence for Na + complexing in tissues consisted of the equilibrium binding studies of Troshin (4,5) and Ling (12), and of cation-sensitive microelectrode studies (17,18,54,55). The equilibrium binding studies showed that equilibria of Na + between cell and solution conformed to the Langmuir absorption isotherm, with a correction for a minor fraction of Na + dissolved in intracellular water.…”

Coherent Behavior and the Bound State of Water and K+ Imply Another Model of Bioenergetics: Negative Entropy Instead of High-energy Bonds

“…[32] discussed above, indicating that Hill [2, 3] found an exception by only studying urea; (b) the fact that resting muscle cells without an intact cell membrane (EMOC preparations: see further) are able to sustain normal intracellular K + concentrations for at least two days [51]; (c) the demonstration that Artemia cysts survive for four years without energy consumption [52]; (d) reinvestigations of measurements of ionic conductivity in various cell types and of K + mobility in frog muscle cells, giving much lower values than for free K + and providing convincing explanations why some earlier studies gave high values [12, 18, 19, 22]; (e) proof that use of a K + selective electrode readily injures the cell, resulting in much too high activity measurements, so that more confidence can be given to those experimental set-ups giving low values [18, 19, 22, 64]; (f) X-ray absorption-edge fine structure determination of red blood cells giving a result that differs markedly from control measurements on a KCl solution of the same concentration [65]; (g) 39 K + NMR spectra of neurons, muscle cells and E. coli , which produced relaxation times much shorter than those of 0.1 M to 0.4 M KCl solutions and comparable to those of K + adsorbed on a Dowex-50 cation-exchange resin [66, 67, 12, 18, 19, 22]; (h) demonstration that the relationship between internal and external 42 K + concentration follows Langmuir’s adsorption isotherm, whereby real proof of adsorption was given by studying the influence of inhibitors that compete in different ways for the same adsorption sites (unlabeled Cs + and unlabeled K + ) [19, 68]. …”

“…Actin and myosin can be taken as examples of cell-matrix (NUN-) proteins [12, 14, 18, 70, 73, 134]. They possess unfolded regions [14, 75], polarize water [14, 18, 73], adsorb K + [68-73], possess cardinal adsorption sites for ATP and exhibit ATPase activity [12, 14]. It was shown that upon activation of myofibrils weak binding between the S 1 -head of myosin and actin is followed by the release of P i , allowing strong binding, which in turn leads to the power stroke and only then to ADP-release [145].…”

Freeze‐dried and resin‐embedded biological material is well suited for ultrastructure research

“…According to the current view, most of the mobile cellular K + ions are freely dissolved in free cellular water – hence they should follow the cellular water distribution. On the other hand, it has been postulated (Ling & Ochsenfeld, 1966) that the bulk of cellular K + is weakly adsorbed (not completely immobilized) to β‐ and γ‐carboxyl groups of cellular proteins; the K + distribution should follow the distribution of these side chains. The divergent views have been tested with normal K + ‐containing frog muscles and living muscles in which most of the cellular K + was (reversibly) replaced in a mole‐for‐mole fashion with the K + surrogates Rb + , Cs + or Tl + (Ling, 1977).…”