Cation sensitive glass microelectrodes were inserted into single striated muscle fibers of the giant barnacle, Balanus nubilus, to measure directly the activities of sodium and potassium in the myoplasm. The total sodium and potassium content of the individual experimental fibers was determined by flame photometry. From these measurements, the percentage of sodium in the fiber which did not affect the microelectrodes and the percentage of water in the fiber which was not available to act as solvent for the potassium ions were calculated. The minimal percentages of "bound" sodium and water were 84% and 42% respectively. It was hypothesized that a significant fraction of this "bound" sodium was involved in ion pair formation with carboxyl moieties on the myosin molecules which comprise the thick filaments, and experiments were designed to test this hypothesis.
In the second series of experiments, the activities of sodium, potassium and hydrogen in the myoplasm were measured as the temperature of the solution bathing the fibers was increased from 7 to 40°C. An irreversible shortening occurred in all fibers between 37 and 40°C. When the fibers shortened in a sodium free Ringer solution, the mean activity of sodium increased by 130%, the mean activity of potassium remained relatively constant, and the pH decreased from 7.17 to 6.77. These experiments provided strong evidence that sodium is bound to myosin in the living fiber, for extracted myosin is known to denature at 37°C and release its associated alkali metal cations.
In the third series of experiments, the optical density, O.D., of the single striated muscle fibers was measured at 50 mµ intervals between 450 and 850 mµ. At all wavelengths, the O.D. decreased markedly when the normal Ringer bathing solution was replaced by sodium free sucrose Ringer. For example, at 850 mµ the O.D. of the fibers, relative to the initial value in normal Ringer, decreased from 1 to 0.21 ± 0.06 in 25 minutes. The corresponding increase in the transmittance, T, (O.D. = -log T) was from 5% to 55%. This change in O.D. could be reversed by returning the normal Ringer bathing solution to the bath. Large, reversible decreases in O.D. were also observed when potassium and tris were used as substitutes for sodium. These changes in O.D. are explained by the theory of light scattering if it is assumed that sodium is bound to the main scattering centers in the myoplasm, the thick filaments. When the fibers were bathed in sodium free, lithium substituted Ringer, a small reversible increase in the O.D. was observed, which may indicate that lithium is complexed more strongly than sodium to the binding sites on the thick filaments.
In the final series of experiments, the number of sodium and potassium ions "bound" to the contractile proteins in a glycerinated fiber was measured. The free concentrations of hydrogen, sodium and potassium were maintained at values similar to those found in an intact fiber. The results indicated that substantial binding of both sodium and potassium occurred, and that proportionally more sodium than potassium ions were "bound". If the results are extrapolated to the intact fiber, they imply that about as much sodium is "bound" to the contractile proteins as is free in the myoplasm. This amount of "bound" sodium is sufficient to explain the results of the denaturation and light scattering experiments, but insufficient to account for the anomalously low activity of sodium in the myoplasm, as measured by a sodium sensitive microelectrode. Thus, it was concluded that either some factor must enhance the binding of sodium to the contractile proteins in a living cell, or that sodium must be sequestered in organelles which are destroyed by the glycerination process. / Medicine, Faculty of / Graduate
| Identifer | oai:union.ndltd.org:UBC/oai:circle.library.ubc.ca:2429/36792 |
| Date | January 1968 |
| Creators | McLaughlin, Stuart Graydon Arthur |
| Publisher | University of British Columbia |
| Source Sets | University of British Columbia |
| Language | English |
| Detected Language | English |
| Type | Text, Thesis/Dissertation |
| Rights | For non-commercial purposes only, such as research, private study and education. Additional conditions apply, see Terms of Use https://open.library.ubc.ca/terms_of_use. |
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