Biomedical subjects

J A Hinke

Publications and source records attributed to J A Hinke.

At least 19 recordsLinked to original sources

PubMed1988

Potassium, sodium, water, and Donnan potential changes in the myofibrils of intact muscle fibres after exposure to ouabain and K-free Ringer.

Frog sartorius muscles were superfused for 40 min with solutions of K-free Ringer, normal Ringer containing ouabain, or K-free Ringer containing ouabain. Changes in myoplasmic K and Na were measured with ion-selective microelectrodes; changes in total fibre K and Na were measured by means of atomic absorption spectroscopy; and changes in total fibre water content were obtained from wet and dry weights. Application of a two-compartment model permitted one to calculate (i) the K, Na, and water changes in the myofibrils and in the surrounding myoplasm (extramyofibrillar space); (ii) the changes in the transmyofibrillar Donnan potential (ED); and (iii) the changes in the ratio of the apparent association constants (kNa/kK) of the myofilament charge sites to Na and K. In the resting fibres, the K, Na, and water content of the myofibrils were calculated to be 82, 87, and 80% of total fibre content, respectively; ED was calculated as -4.5 mV; kNa/kK was calculated as 1.4. After a 40-min ouabain treatment, 12 mmol (per kg fibre water) of intrafibre K exchanged with 7.5 mmol of extrafibre Na, 6.4 mmol of myofibrillar K exchanged with an equal amount of extramyofibrillar Na, ED increased to -8.3 mV, and kNa/kK remained relatively constant. After a 40-min K-free treatment, the fibres gained 5.5 mmol of Na without any change in fibre K or water, the myofibrils shifted 9.3% of their water into the extramyofibrillar space instead of exchanging K for Na, ED increased to -10.7 mV, and kNa/kK decreased to 0.47.(ABSTRACT TRUNCATED AT 250 WORDS)

Algorithms↗

PubMed1987

Thirty years of ion-selective microelectrodes: disappointments and successes.

The need to know the intracellular activity of an ion and how it changes under controlled conditions is as important today as it was 30 years ago. In 1956, one could fabricate only a H+-selective microelectrode and with a tip size not much smaller than 100 micron. Today, one can fabricate microelectrodes selective to H+, Na+, K+, Cl-, HCO3-, Ca2+, or Mg2+ (plus others) and with active tips less than 1 micron. The reduction of active tip size can be attributed mainly to the introduction of liquid ion exchanger (LIX) and neutral carrier ligands. Unfortunately, the LIX microelectrodes, as currently fabricated, do not yet function optimally as reliable and stable electrochemical measuring devices. A durable bond between the active membrane and its insulated container continues to remain the major design problem even after 30 years of development.

Animals↗

PubMed1987

Transmembrane K+, Na+, and Cl- permeability and conductance changes in the frog sartorius fibres following ouabain and zero [K+]o treatment.

Changes in the K+, Na+, and Cl- permeabilities (P) and conductances (g) of the intact frog sartorius fibre membrane following ouabain or zero [K+]o treatment were calculated from intrafibre activity and whole muscle electrolyte changes. Conventional equations relating ionic fluxes to resting potential (E), ionic gradient potential, and internal and external ionic activities were used. Both treatments produced a three- to five-fold increase in PNa and gNa. In addition, ouabain produced a fivefold increase in PK (and gK) and a small decrease in PCl (and gCl), whereas zero [K+]o produced a 60% reduction in PK, a 90% reduction in gK, and a threefold increase in PCl (and gCl). When the two treatments were combined, the P and g changes were paradoxical, suggesting that the ouabain-induced increase in gK and the zero [K+]o-induced decrease in gK were occurring but in different channels (or carriers). During ouabain treatment, E reflects mainly the transmembrane K+ gradient potential; during zero [K+]o treatment, E reflects mainly the Cl- gradient potential. Despite channel (or carrier) specificity, it appears that all three ionic permeabilities are altered during the perturbations.

Animals↗

PubMed1982

Water in barnacle muscle. IV. Factors contributing to reduced self-diffusion.

The relative self-diffusion coefficients D/Do, of water in various solutions, in fresh barnacle muscle fibers, and in membrane-damaged fibers equilibrated with several media have been estimated from NMR relaxation rates in the presence of applied field gradients. A model has been developed to account for the contributions to the observed reduction in D/Do from small organic solutes, and from the hydration and obstruction effect of both soluble macromolecules and myofilament proteins. Intracellular ions do not affect D/Do, but all tested organic solutes do. Solute effects are additive. When artificially combined in the proportions found in barnacle muscle ultracentrifugate (measured D/Do = 0.77), organic acids, small nitrogenous solutes, and proteins give D/Do = 0.77. After correcting the D/Do measured in fibers for this value, we calculate the myofilament hydration, Hm, in fresh muscle to be 0.65 g H2O/g macromolecule. Only in membrane-damaged fibers, highly swollen by salt-rich media, was this significantly increased. Because our earlier NMR relaxation measurements indicate only 0.07 g H2O bound/g myofilament protein, we conclude that the "hydration" water measured by reduction of D/Do cannot be described by stationary layers of water molecules; instead, we propose that nonpolar groups on the proteins cause extensive, hydrophobically-induced interactions among a large fraction of solvent molecules, slowing their translational motion.

Animals↗

PubMed1981

Water in barnacle muscle. III. NMR studies of fresh fibers and membrane-damaged fibers equilibrated with selected solutes.

Water in barnacle muscle has been studied using NMR techniques. Fresh fibers are compared with membrane-damaged fibers treated with solutes that greatly alter fixed charge and total water content. Both water (97%) and solute (3%) protons are visible in continuous wave spectra of oriented fresh fibers. No local field inhomogeneities were detected, nor are cell solutes significantly bound. In pulse experiments, all cell water is visible and exhibits a single exponential decay. In fresh fibers, T2 approximately or equal to 40 ms; faster decaying signals are assigned to immobile and mobile protons on macromolecules. T1 and T1p are frequency dependent. Using equations derived for a two-compartment model with fast exchange, we calculate the following: tau b, the correlation time for anisotropic rotational motion of bound water; Sb, its order parameter; tau ex, the correlation time for exchange between bound and free fractions; f, the fraction of water bound; and Hr, the grams of water bound per gram of macromolecule. Whereas f varies inversely with total water content, the other parameters are virtually constant, with values: tau b approximately or equal to 1.3 X 10(-8) S; tau ex approximately or equal to 8 X 10(-6) s; Sb approximately or equal to 0.06; and Hr approximately or equal to 0.1g H2O/g macromolecule. Thus, the NMR relaxation detectable properties of water bound to macromolecules are unaffected by solutes that greatly alter the macromolecular surface charge.

Animals↗

PubMed1981

Intracellular Cl activity, Cl binding, and 36Cl efflux in rabbit papillary muscle.

Chloride liquid ion-exchange microelectrodes were used to measure intracellular chloride activity (aCl)i or rabbit papillary muscle in vitro. In HCO3- -- CO2 buffered Ringer (aCl)i was 15.2 +/- 0.6 mM (n = 40) at pH 7.3 and 15.4 +/- 0.3 mM (n = 29) at pH 6.8. In HEPES-buffered Ringer (aCl)i was 24.1 +/- 0.6 mM (n = 21) at pH 7.3 and 20.0 +/- 0.3 mM (n = 17) at pH 6.9. If Cl was distributed passively across the membrane (aCl)i should be about 3 mM. From total muscle Cl analysis and [14C]sorbitol extracellular space measurements we calculate an apparent intracellular Cl concentration of 71 mM. As this estimate is three times larger than the measured free Cl in the cytoplasm we conclude that a significant fraction of muscle Cl must be bound or compartmentalized. Analysis of 36Cl efflux from the papillary muscle indicates the presence of two compartments. The compartment with the slower efflux rate (2.0 X 10(-2) min-1) contains about 25% of the intracellular Cl, which compares with the estimation (21%) of the free intracellular CL fraction obtained from the Cl microelectrode measurements. Our data permit us to conclude that about 37% of the muscle Cl is bound and that it is probably located extracellularly.

Animals↗

PubMed1981

The influence of "nonmyoplasmic" sodium on the measured value of the sodium efflux from barnacle muscle.

In single striated muscle cells of the giant barnacle Balanus nubilus, the sodium content of the myoplasm was measured with an intracellular microelectrode, and the total sodium content of the cell was measured by flame photometry, during immersion of the cells in sodium-free solution. The sodium content of the myoplasm declined slowly but steadily from ca. 10 mmol/kg water to ca. 4 mmol/kg water during immersions lasting up to 16 h. The "nonmyoplasmic" sodium content of the cells, defined as the sodium content after subtraction of the sodium in the extracellular (sorbitol) space and in the myoplasm, declined rapidly from ca. 15 mmol/kg water to ca. 3 mmol/kg water during the first 30 min of immersion in sodium-free solution, but remained constant thereafter. The rapidly lost portion of the nonmyoplasmic sodium (ca. 123 mmol/kg water) was ascribed to the extracellular space but the location of the inexchangeable portion was not discovered. The behavior of the efflux of 22Na which was loaded into the myoplasm by microinjection was consistent with this interpretation. It was concluded that the nonmyoplasmic sodium does not have an appreciable influence on the measured value of the sodium efflux from the myoplasm of barnacle muscle cells.

Animals↗

PubMed1980

Water and electrolyte content of the myofilament phase in the chemically skinned barnacle fiber.

Muscle fibers from the giant barnacle, Balanus nubilus, were placed inside the lumen of a porous glass capillary and equilibrated for 48 h in an electrolyte solution containing 2% Tween. The glass capillary prevented the chemically "skinned" fiber from swelling with a water content beyond 80%. Isotope exchange studies using 22Na, 42K, and 36Cl indicated the existence of an intermediate rate constant and compartment which varied with pH. This intermediate rate was attributed to counter-ions and co-ions in the myofilament phase. Analysis of the electrolyte composition of the fiber at pH 8 predicts that the myofilaments contain about 0.3 of the fiber water, and that a -15 mV Donnan potential exists at the myofilament surface. An open-tipped (1-micrometer) microelectrode in the skinned fiber measured a potential (similar in magnitude to the Donnan potential), which decreased and reversed sign as the pH was lowered. The measured cation contents of the fiber between pH 5 and 8 were found to be similar to the cation contents predicted from the measured Donnan potentials. The net negative charge of the myofilaments at pH 7.5 and at ionic strength 0.56 is estimated to be 41 eq per 10(5) g of dry weight.

Animals↗

PubMed1978

Evaluation of the DMO method for measuring intracellular pH.

Intracellular pH in single muscle fibres from Balanus nubilus was determined, both by the weak acid distribution method (using 5,5-dimethyl-2,4-oxazolidine dione) and by means of a glass pH microelectrode, following prolonged equilibrations in normal, acid or alkaline Ringer solutions. In the DMO method [3H]inulin was combined with [14C]DMO so that an extracellular space could be calculated for each 'DMO space' on each fibre. Less than ideal uptake curves were obtained both for [14C]DMO and [3H]inulin, yet the calculated pHi compared favourably with the microelectrode pHi, especially when the DMO-pHi was calculated from [14C]DMO and [3H]inulin values obtained by extrapolating the 'plateaus' of the uptake curves to time zero. The Waddell and Butler equation was examined to determine why acceptable pHi values should be forthcoming from less than ideal uptake data.

Acid-Base Equilibrium↗

PubMed1976

Intracellular pH of single crustacean muscle fibres by the DMO and electrode methods during acid and alkaline conditions.

1. The intracellular pH of intact single muscle fibres of the giant barnacle was measured directly with a glass micro-electrode following prolonged (2-5 hr) equilibration in one of three solutions: normal Ringer, CO2 Ringer and NH4+ Ringer. 2. The intracellular pH of identically-prepared fibres from the same specimen was measured indirectly from the distribution of DMO following prolonged equilibration in the same solutions. 3. The DMO-pH compared favourably with the electrode-pHi provided DMO-pHi was calculated from the values of the indicator compounds, [14C]DMO and [3H]inulin, obtained by extrapolating the slow uptake phase to time zero. 4. Following prolonged equilibration, the transmembrane H+ ion distribution was found to vary with the membrane potential but not in accordance with a simple Gibbs-Donnan equilibrium. 5. A model which recognizes the existence of two independent net fluxes for H+ across the membrane in developed to explain the results. One of the fluxes represents passive diffusion and the other represents the so called H+-pump. The model predicts the H+-pump rate increases by two orders of magnitude when pHi is reduced from 7-2 to 6-7.

Animals↗

PubMed1974