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

The permeability of the teleost branchial exchanger to oxygen and carbon dioxide is apparently enhanced during exercise by increased blood perfusion of thin walled high surface area pathways in the gills, the secondary lamellae. However this augmented permeability to respiratory gases may well be accompanied by unfavourable elevations of water and electrolyte fluxes between internal and external environments. The object of the present study was to investigate the effect of imposed swimming activity on sodium and water regulation in the fresh water adapted rainbow trout, Salmo gairdneri. Radiotracer methods were used to measure unidirectional components of branchial sodium exchange in fish at rest, during one hour of swimming, and during one hour of recovery from this exercise condition. Sodium fluxes during extended exercise (up to 8 hours) were quantified by similar techniques in a second series of experiments. These long term swimming trials provided flux rate data.at a wide range of external sodium concentrations; analysis of these results helped to elucidate the relative importance of different mechanisms of branchial sodium transfer in the rainbow trout. Finally, determinations of urine flows and body weight changes under controlled exercise conditions in a swimming respirometer permitted an analysis of water regulation and direct measurement of renal electrolyte losses during activity. The sodium uptake system of Salmo gairdneri in the present study had an extremely high affinity for the ion (half saturation concentration = .014 mEq Na+/L). Both unidirectional flux rates at the gills of rainbow trout were greater than those reported for any other fresh water teleost of comparable size, despite external sodium levels much lower than those used by other workers. The presence of an exchange diffusion mechanism for sodium in the trout gill was strongly indicated but not confirmed.. Branchial transport of the electrolyte was tentatively divided into a large exchange diffusion component, and smaller active influx and simple diffusional efflux elements. In resting animals, branchial sodium influx and efflux rates were equal. However short term activity (1 hour) was associated with a 70% increase in efflux of sodium across the gills, creating a net sodium deficit. This effect was quickly reversed (within 5 minutes) upon the termination of swimming. As influx did not vary, these phenomena probably represented changes in the simple diffusional efflux component without disturbance of carrier mediated sodium transport mechanisms. Branchial water entry was also greatly elevated at the start of exercise. These results were interpreted in terms of augmented passive movements of sodium and water caused by increased blood perfusion of the high permeability respiratory pathways of the gills during swimming. The extended exercise experiments revealed that the high sodium efflux rate of the first hour of activity diminished, during the second hour, and had returned to resting levels by the third and subsequent hours of swimming; influx again remained unchanged. The initial high branchial water entry was also apparently curtailed, but over a shorter time interval (15 - 60 minutes after the onset of activity). These reductions in branchial permeability to water and sodium were interpreted as compensations to decrease the osmotic penalty of exercise. As water entry through the gills declined, urinary output was augmented; an elevated renal sodium loss accompanied the diuresis. However sodium efflux through the kidney remained small relative to the efflux of this electrolyte through the gills. A final equilibrium between branchial entry and renal excretion of water was attained, but at a higher turnover rate than during rest. Before this balance, however, urinary elimination had over-compensated for the initial water gain. The resulting net water deficit reduced the blood space below resting volume, causing a slight increase in plasma sodium levels despite enhanced branchial and renal losses of the ion. An ischemia of "white" muscle may also have accompanied the haemoconcentration. In summary, the results indicated that an initial osmoregulatory disturbance was associated with a redistribution of blood flow through the gills during swimming, but that both branchial hydromineral permeability and the functioning of other systems could be modified by compensations necessary to maintain sodium and water balance during extended exercise. / Science, Faculty of / Zoology, Department of / Graduate

A method is described for the preparation of an everted rectal sac of the desert locust. Water and solute absorption by the rectum was determined by measuring changes in hemocoel fluid and rectal tissue. Initial absorption rates of Na, K, Cl, water and trans-rectal potential are comparable to those in vivo under similar conditions. After an initial transient period (1 hour), transport activity of the in vitro rectum remained in a steady state for at least 4- hours. The relationship between osmotic gradient and steady state rate of net water movement across the rectal wall was determined. Absorption of water is partially inhibited by anoxia, malonate (10־² M), dinitrophenol (10־³M), potassium cyanide (10־³ M) plus iodoacetate (10־³ M) and ouabain (10־³ M). Tissue ions and water are secreted into the hemocoel compartment when the rectal sac is incubated in isosmotic pure sucrose solution. Dependence of water movement on solute transport is indicated by the requirement of lumen ions for prolonged maintenance of water absorption. Effects of different ions (Na, K and Cl) in bathing media on absorption rate of water and ions, absorbate concentrations, trans-epithelial electro-potential differences, and tissue compositions were determined. Observed properties of water and solute movement in vitro are discussed and evaluated in relation to possible mechanisms for active absorption of water. Possible locations of transport sites are suggested in a hypothetical scheme based on the ultrastructure of rectal epithelium. / Science, Faculty of / Zoology, Department of / Graduate

by Ho, Chung Shun. / Thesis (Ph.D.)--Chinese University of Hong Kong, 1994. / Includes bibliographical references (leaves [304-328]). / Chapter CHAPTER 1 --- REVIEW ON SODIUM EXCRETION / Chapter L --- Sodium excretion --- p.1-1 / Chapter II. --- Cellular mechanism of sodium reabsorption --- p.1-3 / Chapter III. --- Sensors monitoring ECF volume --- p.1-6 / Chapter IV. --- Factors affecting natriuresis: / Glomerular filtration rate --- p.1-8 / Renal physical forces --- p.1-8 / Sympathetic nervous system --- p.1-10 / Renal dopamine --- p.1-12 / Renin-angiotensin system --- p.1-14 / Aldosterone --- p.1-16 / Renal prostaglandins --- p.1-17 / Renal kallikrein-kinin system --- p.1-18 / Natriuretic peptides --- p.1-19 / Endogenous sodium transport inhibitor --- p.1-21 / Vasopressin --- p.1-22 / Endothelins --- p.1-23 / Endothelin-derived relaxing factor --- p.1-25 / Other hormones --- p.1-25 / Chapter V. --- Conclusion --- p.1-27 / Chapter CHAPTER 2 --- MEASUREMENT OF ENDOGENOUS SODIUM TRANSPORT INHIBITORS / Chapter I. --- Literature review: --- p.2-1 / Pretreatment and purification procedures prior to ESTI measurement --- p.2-1 / Methods of measuring ESTI --- p.2-3 / "Inhibition of purified Na, K-ATPase activity" / Inhibition of sodium pump on intact cells or tissues / Biological effects of sodium pump inhibition / Immunoreactivity with anti-digoxin /anti-ouabain antibodies / Chapter II. --- Method of measurement of ESTI in this study: / Principles of methods --- p.2-11 / Materials and methods --- p.2-14 / Results --- p.2-24 / Discussion --- p.2-53 / Chapter CHAPTER 3 --- MEASUREMENT OF URINARY FREE DOPAMINE / Chapter I. --- Literature review / Properties of dopamine for measurement methods --- p.3-1 / Preservatives used in the urine collection --- p.3-3 / Sample pretreatment procedure before --- p.3-6 / measurement --- p.3-8 / Methods of measurement / Bioassays / Colorimetric method / Fluorometric methods / Radioimmunoassays / Radioenzymatic method / Chromatographic methods --- p.3-16 / Concluding remarks / Chapter II. --- Method of measurement in this study / Principle of the method --- p.3-17 / Materials and methods --- p.3-18 / Results --- p.3-23 / Discussion --- p.3-54 / Chapter CHAPTER 4 --- CROSS SECTIONAL STUDIES IN THE HUMAN / Chapter I. --- Introduction --- p.4-1 / Chapter II. --- Relationship of urinary sodium excretion and plasma ESTI in medical students / Materials and methods --- p.4-2 / Results --- p.4-3 / Discussion --- p.4-6 / Chapter III. --- Excretion of urinary electrolytes and natriuretic factors in young Chinese females / Materials and methods --- p.4-7 / Results --- p.4-8 / Discussion --- p.4-11 / Chapter IV. --- Urinary sodium / DA relationship in Chinese normotensives and hypertensives --- p.4-13 / Materials and methods / Results --- p.4-14 / Discussion --- p.4-17 / Chapter V. --- Urinary DA excretion and plasma ESTI in normotensive and hypertensive NIDDM patients / Materials and methods --- p.4-20 / Results --- p.4-23 / Discussion --- p.4-33 / Chapter CHAPTER 5 --- VOLUME EXPANSION STUDIES IN THE HUMAN / Chapter I --- Introduction --- p.5-1 / Chapter II. --- Volume expansion by headout water immersion / Materials and methods --- p.5-3 / Results --- p.5-5 / Discussion --- p.5-11 / Chapter III. --- Volume expansion by saline infusion / Materials and methods --- p.5-16 / Results --- p.5-19 / Discussion --- p.5-29 / Chapter IV. --- Oral salt loading with free diet / Materials and methods --- p.5-36 / Results --- p.5-38 / Discussion --- p.5-49 / Chapter V. --- Oral salt loading under controlled diet / Materials and methods --- p.5-53 / Results --- p.5-54 / Discussion --- p.5-64 / Chapter CHAPTER 6 --- STUDIES ON THE EFFECTS OF SALT LOADING IN THE RAT / Chapter I. --- Introduction --- p.6-1 / Chapter II. --- Temporal relationship between excretions of DA and ESTI during salt loading in the rat / Materials and methods --- p.6-2 / Results --- p.6-3 / Discussion --- p.6-6 / Chapter III. --- Roles of DA and ESTI in natriuresis in rats treated with carbidopa / Materials and methods --- p.6-8 / Results --- p.6-9 / Discussion --- p.6-14 / Chapter CHAPTER 7 --- CONCLUSION / Measurement of ESTI --- p.7-2 / Measurement of urinary free DA --- p.7-5 / Cross sectional studies in human --- p.7-7 / Volume expansion studies in human --- p.7-11 / Studies on the effects of salt loading in the rat --- p.7-16 / Summary --- p.7-18

Supplementation of a minimal medium with KHCO$ sb3$ and some amino acids reduced the duration of the lag period for the growth of the marine bacterium Deleya aesta 134 at sub-optimal Na$ sp+$ concentrations and decreased the minimal Na$ sp+$ concentration allowing growth, to 8mM. / Na$ sp+$ was required for the transport of most metabolites into D. aesta 134, while some compounds were taken up by both a Na$ sp+$-dependent and a Na$ sp+$-independent system. Phosphate transport was not affected by Na$ sp+.$ Na$ sp+$ was also required for the oxidation of exogenous succinate. Evidence for the presence in D. aesta 134 of a Na$ sp+$-activated NADH: quinone acceptor oxidoreductase was obtained. / A mutant of D. aesta 134 exhibiting a shorter lag period than its parent strain at 10mM Na$ sp+$ was also examined. Enhanced Na$ sp+$/H$ sp+$ antiport activity was obtained in the mutant strain, but the actual mechanism involved in the adaptation to low Na$ sp+$ concentrations is still obscure. / Succinate transport by D. aesta 134 was mediated by a C$ sb4$-dicarboxylate transport system, and exhibited biphasic kinetics, indicating the presence of a high- and a low-affinity transport system. / A partial genomic library of D. aesta was prepared, and DNA from D. aesta complemented mutations from Escherichia coli auxotrophs. Successful introduction of the plasmid pRK404 into D. aesta 134 by electroporation was also obtained.

No description available.

No description available.

<p>Dissolved organic matter (DOM) in natural waters is a heterogeneous mixture of organic molecules with direct and indirect influences on aquatic organisms. Although the influences are usually attributed to DOM quantity (quantified as Dissolved Organic Carbon, DOC), the role of quality (optical and binding characteristics obtained by absorbance and fluorescence spectroscopy and potentiometric titration, respectively) is not well-understood. Through an initial critical review of the literature, followed by experimental geochemical, toxicological, and physiological investigations, a number of conclusions were reached that improve our knowledge in this area. Freshwater DOM sources exhibit source-dependent protection against metal toxicity, in particular copper (Cu). Generally, for this indirect effect, optically-dark terrestrially-derived or allochthonous DOMs offer better protection than microbially-derived or autochthonous sources. Linear regressions revealed that the better ameliorative effect is principally related to a higher aromatic composition (specific absorption coefficient, SAC₃₄₀) and a greater humic-like fluorescent component as quantified by parallel factor analysis (PARAFAC). In addition, the allochthonous DOMs were shown to have relatively higher magnitudes of titration index (TI), a new summary of chemical reactivity of DOM molecules obtained by titration analysis, and closely related to optical properties. TI was strongly correlated with SAC₃₄₀, suggesting greater binding capacities for DOM molecules with higher SAC₃₄₀. Consequently, a method for incorporation of SAC₃₄₀ as a DOM quality measure into the Biotic Ligand Model (BLM) was developed which improved Cu toxicity predictions in experimental tests with natural DOMs. For direct effects, two basic physiological functions (Na⁺ metabolism and nitrogen excretion) of the adult water flea (<em>Daphnia magna</em>, a cladoceran crustacean) and the zebrafish (<em>Danio </em><em>rerio</em>, a teleost fish) were investigated at circumneutral and acidic pH (≥ 7 and ~ 5, respectively). Three previously characterized, chemically-distinct natural DOM sources as well as a commercial humic acid (AHA) were examined. Regardless of the pH conditions, while Na⁺ regulation of <em>D</em>. <em>magna </em>remained unaffected by the presence of all DOMs, the passive diffusive efflux of Na⁺ in zebrafish was attenuated, indicating ameliorative action against unidirectional Na⁺ loss. In addition, only a distinct allochthonous-autochthonous DOM source stimulated the Na⁺ uptake rate of zebrafish at low pH. Ammonia excretion rates of <em>D</em>. <em>magna </em>were reduced at circumneutral pH by the most highly coloured, allochthonous DOM, and at low pH by all three natural DOMs. Both in <em>D. magna </em>and in <em>D. rerio</em>, urea excretion rates at both pH conditions were not influenced by the presence of the various DOMs, and the same was true for ammonia excretion in the zebrafish. A commercially prepared humic acid (Aldrich humic acid, AHA) exerted anomalous actions relative to those of natural DOMs, and does not appear to be representative of their normal effects. In contrast to the actions of DOM in detoxifying metals, these direct effects of DOMs on freshwater organisms appeared highly unpredictable with variable dependencies on the source, pH and species. This thesis has advanced our understanding of the relationships between DOM quality and its indirect and direct effects on aquatic organisms, and points to new directions for future work.</p> / Doctor of Philosophy (PhD)