The effects of water hardness upon the uptake, accumulation and excretion of zinc in the brown trout, Salmo trutta L.

The effects of water hardness (9 and 220 mgl⁻¹ as CaCO₃) upon zinc exchange in brown trout exposed to 0.77 μmol Zn 1⁻¹ have been investigated using artificial soft water (<49.9 μmol Ca ^l-1, <40.1 μmol Mg 1⁻¹) and mains hard water (1671.7 μmol Ca 1⁻¹, 493.6 μmol Mg 1⁻¹) of known composition. Both hard and soft water-adapted fish exhibited a bimodal pattern of net zinc influx. Net zinc influxes during both fast and slow uptake phases were significantly greater ( P <0.001) in soft (82.9 and 6.2 μmol Zn 100 g⁻¹ h⁻¹) than in hard water (46.3 and 2.4 μmol Zn 100 g h⁻¹). Zinc efflux (- 0.2 μmol Zn 100 g⁻¹ h⁻¹) was enhanced only in hard water during the slow net influx phase.
Brown trout exposed to zinc in hard water and placed in metal-free media exhibited a greater net efflux (- 25.6 μmol Zn 100 g⁻¹ h⁻¹) of the metal than did fish in soft water (-4.2 μmol Zn 100 g⁻¹ h⁻¹) treated in the same manner. Tissue ⁶⁵Zn activities reflected both the differences in uptake and excretion rates of the metal between hard and soft water fish. During zinc exposure (0.77 μmol Zn 1⁻¹) high water hardness reduced tissue burdens of the metal by reducing net branchial influx, and enhancing efflux of the metal in hard water fish.     

Scaling of oxygen consumption of Lake Magadi tilapia, a fish living at 37°C

Rates of oxygen consumption were measured in the geothermal, hot spring fish, Oreochromis alcalicus grahami by stopped flow respirometry. At 37° C, routine oxygen consumption followed the allometric relationship: V o₂=0.738 M ^0.75, where V o₂ is ml O₂ h ⁻¹ and M is body mass (g). This represents a routine metabolic rate for a 10 g fish at 37° C of 0.415 ml O₂ g⁻¹ h ⁻¹ (16.4 μmol O₂ g ⁻¹ h ⁻¹). Acutely increasing the temperature from 37 to 42° C significantly elevated the rate of O₂ consumption from 0.739 to 0.970 ml O₂ g ⁻¹ h ⁻¹ ( Q ₁₀=l.72). In the field, O. a. grahami was observed to be 'gulping' air from the surface of the water especially in hot springs that exceeded 40° C. O. a. grahami may utilize aerial respiration when O₂ requirements are high.     

Effects of Arachidonic Acid on Dopamine Synthesis, Spontaneous Release, and Uptake in Striatal Synaptosomes from the Rat

Abstract: Arachidonic acid (AA) markedly stimulated, in a dose-dependent manner, the spontaneous release of [³H]dopamine ([³H]DA) continuously synthesized from [³H]tyrosine in purified synaptosomes from the rat striatum. As estimated by simultaneous measurement of the rate of [³H]H₂O formation (an index of [³H]tyrosine conversion into [³H]DOPA), the AA response was associated with a progressive and dose-dependent reduction of [³H]DA synthesis. In contrast to AA, arachidic acid, oleic acid, and the methyl ester of AA (all at 10⁻⁴ M ) did not modify [³H]DA release. The AA (3 × 10⁻⁵ M )-evoked release of [³H]DA was not affected by inhibiting AA metabolism, with either 5,8,11,14-eicosatetraynoic acid or metyrapone, suggesting that AA acts directly and not through one of its metabolites. AA also inhibited in a dose-dependent manner [³H]DA uptake into synaptosomes, with a complete blockade observed at 10⁻⁴ M . However, AA (10⁻⁴ M ) still stimulated [³H]DA spontaneous release in the presence of either nomifensine or other DA uptake inhibitors, indicating that AA both inhibits DA reuptake and facilitates its release process. Finally, the AA (10⁻⁴ M )-evoked release of [³H]DA was not affected by protein kinase A inhibitors (H-89 or Rp -8-Br-cAMPS) but was markedly reduced in the presence of protein kinase C inhibitors (Ro 31-7549 or chelerythrine).     

Transformation of Methanococcus maripaludis and identification of a Pst I-like restriction system

Abstract An optimized polyethylene glycol (PEG) method of transformation was developed for Methanococcus maripaludis using the pKAS102 integration vector. The frequency of transformation with 0.8 μg of plasmid and 3×10⁹ cells was 4.8×10⁻⁵ transformants cfu⁻¹, or 1.8×10⁵ transformants μg⁻¹, which was four orders of magnitude greater than with the natural transformation method. A Pst I restriction activity in M. maripaludis was also identified. Methylation of the plasmid with Pst I methylase increased the methanococcal transformation frequency at least four-fold. Also, chromosomal DNA from M. maripaludis was resistant to digestion by the Pst I endonuclease.     

Transport of G AB A, β-Alanine and Glutamate into Perikarya of Postnatal Rat Cerebellum

Abstract: Cells dissociated from the postnatally developing rat cerebellum retain their high-affinity carrier-mediated transport systems for [³H]GABA ( K _t=1.9 μM, V = 1.8 pmol/10⁶ cells/min) and [³H]glutamate ( K _t= 10 μM, V = 7.9 pmol/10⁶ cells/min). Using a unit gravity sedimentation technique it was demonstrated that [3H]GABA was taken principally into fractions that were enriched in inhibitory neurons (Purkinje, stellate and basket cells). [³H]β-alanine (which is taken up specifically by the glial GABA transport system) and [³H]glutamate were concentrated by glial-enriched fractions. However [³H]glutamate uptake was minimal in fractions enriched in precursors of granule cells, which may utilise this amino acid as their neurotransmitter. These results are discussed in relation to reports of high-affinity [³H]glutamate uptake by glia. The role of glutamate transport in glutamatergic cells is also considered. The data suggest that high-affinity glutamate transport is a property of glial cells but not granule neurons.     

CADMIUM AND ZINC BIOAVAILABILITY TO SELENASTRUM CAPRICORNUTUM (CHLOROPHYCEAE): ACCIDENTAL METAL UPTAKE AND TOXICITY IN THE PRESENCE OF CITRATE

Toxicities of cadmium (Cd) and zinc (Zn) to the green alga Selenastrum capricornutum Printz were determined over 72 h in defined synthetic media buffered by citrate (FRAQ_CIT ; [citrate] = 100 μM or 5 μM) or nitrilotriacetate (FRAQ_NTA ; [NTA] = 5 μM). Algal sensitivity to free Cd²+ or free Zn²+ in FRAQ_CIT was much higher than in FRAQ_NTA. In parallel experiments, short-term intracellular uptake of radiolabeled ¹⁰⁹Cd was measured as a function of time (0–30 min) in FRAQ_CIT and FRAQ_NTA; for a given free Cd²+ concentration (8, 250, or 610 nM), intracellular accumulation of Cd was consistently higher in FRAQ_CIT than in FRAQ_NTA. Under the same conditions, the alga accumulated ¹⁴C-labeled citrate almost linearly over a 2-h period. Loss of ¹⁰⁹Cd from algal cells that had been prelabeled with the radionuclide occurred slowly, and the loss rate was insensitive to the presence or absence of citrate, indicating that the overall permeability of the algal membrane to Cd was unaffected by citrate. The enhanced bioavailability of Cd in the presence of citrate could be explained by membrane transport of a charged Cd–citrate complex, presumably by accidental transport.     

Selenium uptake by Butyrivibrio fibrisolvens and Bacteroides ruminicola

Abstract The uptake and incorporation of ⁷⁵[Se]selenite by Butyrivibrio fibrisolvens and Bacteroides ruminicola were by constitutive systems. Rates of uptake were higher in chemostat culture than in batch culture and there may be some inducible component. Uptake of [⁷⁵Se]selenite was distinct from sulphate or selenate transport, since sulphate and selenate did not inhibit selenite uptake, nor could sulphate or selenate uptake be demonstrated in these organisms. Selenite uptake in B. fibrisolvens had and apparent K _m of 1.74 mM and a V _max of 109 ng Se · min⁻¹· (mg protein)⁻¹. An apparent K _m of 1.76 mM and V _max of 1.5 μg Se · min⁻¹· (mg protein)⁻¹ was obtained for B. ruminicola . [⁷⁵Se]Selenite uptake by both organisms was partially sensitive to inhibition by 2,4-DNP. Uptake by B. fibrisolvens was also partially inhibited by azide and arsenate and in B. ruminicola it was partially inhibited by fluoride. CCCP, CPZ, DCCD or quinine did not inhibit uptake in either B. fibrisolvens or B. ruminicola . Selenite transport by both organisms was sensitive to IAA and NEM and was strongly inhibited by sulphite and nitrite. [⁷⁵Se]Selenite was converted to selenocystine, selenohomocystine and selenomethionine by B. fibrisolvens. B. ruminicola did not incorporate [⁷⁵Se]selenite into organic compounds, but did reduce it to red elemental selenium.     

[14C]GLUTAMATE UPTAKE AND COMPARTMENTATION IN GLIA OF RAT DORSAL SENSORY GANGLION

Abstract— The characteristics of the uptake of l -[U-¹⁴C] glutamate into rat dorsal sensory ganglia were investigated. The uptake was mediated by two distinct kinetic systems, with apparent K_m values of the order of 10⁻³ M (low affinity) and 10⁻⁵ m (high affinity). The high affinity uptake system was strongly dependent upon temperature and sodium ion concn, and was depressed by a number of metabolic inhibitors. Following uptake, [¹⁴C] glutamate was extensively metabolized, primarily to glutamine, although this was not so with cultured ganglia, where in addition to an increased uptake of [¹⁴C] glutamate, the specific radioactivity of glutamate was increased and that of glutamine decreased. The labelled substrates [U-¹⁴C]pyruvate and [U-¹⁴C] acetate were used to investigate this phenomenon and the results are discussed in relation to current knowledge of metabolic compartmentation in nervous tissue.     

Blockade by Neurotransmitter Antagonists of Veratridine-Activated Ion Channels in Neuronal Cell Lines

Abstract: The voltage-dependent Na⁺ ionophore of various neuronal cells is permeable not only to Na⁺ ions but also to guanidinium ions. Therefore, the veratridine-(or aconitine-) stimulated influx of [¹⁴C]guanidinium in neuroblastoma × glioma hybrid cells was measured to characterize the Na⁺ ionophore of these cells. Half-maximal stimulation of guanidinium uptake was seen at 30 μ M veratridine. At 1 m M guanidinium, the veratridine-stimulated uptake of guanidinium was lowered to 50% by approximately 60 m M Li⁺, Na⁺, or K⁺ and by a few millimolar Mn²⁺, Co²⁺, or Ni²⁺. The basal, as well as the veratridine-stimulated, uptake of guanidinium was inhibited by the cholinergic antagonists (+)-tubocurarine ( K_i = 50 to 500 n M ) and atropine ( K_i = 5 to 30 μ M ) and the adrenergic antagonists phentolamine ( K_i = 5 μ M ) and propranolol ( K_i = 60 μ M ). The specificity of the inhibitory effects of these agents is stressed by the ineffectiveness of various other neurotransmitter antagonists. However, the corresponding ionophore in neuroblastoma cells (clone N1E-115) seems to be regulated differently. While phentolamine and propranolol inhibit the veratridine-activated uptake as in the hybrid cells, (+)-tubocurarine and atropine exert only a slight effect.     

Availability, uptake and turnover of glycerol in hypersaline environments

Abstract A sensitive assay for glycerol and other polyols was developed, based on periodate oxidation to formaldehyde, followed by a colorimetric assay with 3-methyl-2-benzothiazolone hydrazone. Apparent glycerol concentrations thus measured in saltern crystallizer ponds were around 20–36 μM, while in the Dead Sea, during a Dunaliella bloom, values were up to 27 μM. However, these values probably overestimate the glycerol concentrations present, as shown by labeled glycerol uptake experiments. Values of [K + S_n] (natural concentration + affinity constant) in saltern ponds were as low as 0.76–1.4 μM, with V_max values of 193–303 nmol 1⁻¹h⁻¹, and turnover times between 2.6–7.2 h at 35°C. Similar measurements in the Dead Sea were: [K + S_n] 0.07–1.41 μM, V_max values 160–426 nmol 1⁻¹h⁻¹, and turnover times in the range of 0.45–3.3 h.     

Sodium Transport from Blood to Brain: Inhibition by Furosemide and Amiloride

Abstract: Brain sodium uptake in vivo was studied using a modified intracarotid bolus injection technique in which the uptake of ²²Na ⁺ was compared with that of the relatively impermeable molecule, [³H]l-glucose. At a Na ⁺ concentration of 1.4 m M , Na ⁺ uptake was 1.74 ± 0.07 times greater than l -glucose uptake. This decreased to 1.34 ± 0.04 at 140 m M Na ⁺, indicating saturable Na ⁺ uptake. Relative Na ⁺ extraction was not affected by pH but was inhibited by amiloride ( K _i= 3 ± 10⁻⁷ M ) and by 1 m M furosemide. The effects of these two inhibitors were additive. Brain uptake of ⁸⁶Rb ⁺, a K ⁺ analogue, was measured to study interaction of K ⁺ with Na ⁺ transport systems. Relative ⁸⁶Rb ⁺ extraction was also inhibited by amiloride; however, it was not inhibited by furosemide. The results suggest the presence of two distinct transport systems that allow Na ⁺ to cross the luminal membrane of the brain capillary endothelial cell. These transport systems could play an important role in the movement of Na ⁺ from blood to brain.     

Cysteine Sulfinic Acid in the Central Nervous System: Uptake and Release of Cysteine Sulfinic Acid by a Rat Brain Preparation

Abstract: Uptake and release of cysteine sulfinic acid by synaptosomal fractions (P₂) and slices of rat cerebral cortex were investigated. The P₂ fraction had a Na⁺-dependent high-affinity uptake system for cysteine sulfinic acid (K_m, 12μM), which was restricted to the synaptosomes. High-affinity uptake of cysteine sulfinic acid was competitively inhibited by glutamate, aspartate, and cysteic acid. None of the various centrally acting drugs tested specifically inhibited this transport system. Release of [¹⁴C]cysteine sulfinic acid from preloaded cortical slices or P₂ fractions was examined by a superfusion method, which avoided reuptake of released [¹⁴C]cysteine sulfinic acid. High K⁺ (56 m M ) and veratridine (10μM) stimulated the release of cysteine sulfinic acid from slices and the P₂ fraction in a partly Ca²⁺-dependent manner. Diazepam at concentrations of 10 and 100 μM markedly inhibited the stimulated release, but not the spontaneous release, by cortical slices. On the contrary, it had no effect on the stimulated release of cysteine sulfinic acid from the P₂ fraction.     

The inhibition of photosynthesis and photorespiration in isolated mesophyll cells of Phaseolus and Lycopersicon by reduced osmotic potentials

Mesophyll cells isolated from Phaseolus vulgaris and Lycopersicon esculentum show decreasing photosynthetic rates when suspended in media containing increasing concentrations of osmoticum. The photosynthetic activity was sensitive to small changes in osmotic potential over a range of sorbitol concentrations from 0.44 M (−1.08 MPa) to 0.77 M (−1.88 MPa). Photorespiration assayed by ¹⁴CO₂ release in CO₂-free air and by ¹⁴CO₂ release from the oxidation of [1–¹⁴C] glycolate also decreased as the osmotic potential of the incubation medium was reduced. The CO₂ compensation points of the cells increased with increasing concentration of osmoticum from approximately 60 μ I⁻¹1 at −1.08 MPa to 130 μl 1⁻¹ for cells stressed at −1.88 MPa. Changes in photosynthetic and photorespiratory activities occurred at moderate osmotic potentials in these cells suggesting that in whole leaves during a reduction in water potential, non- stomatal inhibition of CO₂ assimilation and glycolate pathway metabolism occurs simultaneously with stomatal closure.     

Kinetics of Entry of P0 Protein into Peripheral Nerve Myelin

Abstract: Sciatic nerves from 9-day-old rat pups were removed, sliced into 0.4-mm sections, and incubated with [³H]fucose or [¹⁴C]glycine precursors. The nerve slice system gave nearly linear incorporation of [³H]fucose as a function of time for 3 h, after an initial lag of ˜30 min for homogenate and ˜60 min for myelin. Incorporation of [³H]fucose at constant specific radioactivity was directly proportional to exogenous fucose levels over the range 3.0 × 10⁻⁸ m to 1.5 × 10⁻⁶ m . Analysis of labeled proteins by sodium dodecyl sulfate polyacrylamide gel electrophoresis showed that greater than 50% of labeled glycoprotein was P₀, with no other major constituents. This system was used in fucose-chase experiments to determine that a period of ˜20 min elapses between fucosylation and assembly of P₀ into myelin. Cycloheximide inhibition of protein synthesis was used to determine that a period of ˜33 min elapses between protein synthesis and appearance of P₀ myelin.     

Potassium-Stimulated Release of Radiolabelled Taurine and Glycine from the Isolated Rat Retina

Abstract: The release of preloaded [³H]glycine and [³H]taurine in response to a depolarising stimulus (12.5-50 m M KCl) has been studied in the superfused rat retina. High external potassium concentration immediately increased the spontaneous efflux of [3H]glycine, the effect of 50 m M K⁺ apparently being abolished by omitting calcium from the superfusing medium. In contrast, although high potassium concentrations increased the spontaneous emux of [³H]taurine from the superfused rat retina, this release was not evident until the depolarising stimulus was removed from the superfusing medium. The magnitude of this "late" release of [³H]taurine was dependent on external K⁺ concentrations, and appeared immediately after cessation of the stimulus irrespective of whether it was applied for 4, 8, or 12 min. Potassium (50 m M )-induced release of taurine appeared partially calcium-dependent, being significantly reduced (p < 0.01) but not abolished by replacing calcium with 1 mM EDTA in the superfusate. High-affinity uptake systems for both [³H]glycine and [³H]taurine were demonstrated in the rat retina in vitro ( K _m values, 1.67 μ M and 2.97 μ M ; V_max values, 19.3 and 23.1 nmol/g wet weight tissue/h, respectively). The results are discussed with respect to the possible neuro-transmitter roles of both amino acids in the rat retina.     

Opioid Effects on [3H]Norepinephrine Release from Dissociated Embryonic Locus Coeruleus Cell Cultures

Abstract: The acute and chronic effects of opioid exposure on [³H]norepinephrine ([³H]NE) release were examined in cell cultures of embryonic rat locus coeruleus (LC). Initial morphological and biochemical characterization of the cultures indicated that the cells exhibited properties similar to those observed in situ. Specific [³H]NE uptake was saturable with a K _m value of 222 ± 52 n M . [³H]NE accumulated by LC cells was released in response to 20 m M K⁺ stimulation, in a calcium-dependent manner. Both components of neurotransmitter release, spontaneous and K⁺ evoked, were significantly inhibited by β-endorphin, with the latter being maintained in the presence of tetrodotoxin. The pharmacology of the opioid effect was consistent with that of μ-receptor activation. The effect of chronic exposure to the μ-selective agonist fentanyl (1 μ M ) was examined following 4 days of drug treatment. Although there was no significant effect of fentanyl on K⁺-evoked [³H]NE release, these cells were tolerant to the acute inhibitory effect of β-endorphin. These results indicate that this is an appropriate system for examining the effects of acute and chronic opioid treatment on noradrenergic cells in vitro. In addition, this system may be useful as a CNS model for examining mechanisms that underlie tolerance and dependence following chronic opioid exposure.     

Identification of Strychnine Binding Sites in the Rat Retina

Abstract: [³H]Strychnine specifically binds to membrane fractions isolated from rat retinae. The binding is saturable, with an apparent dissociation constant, K _D, of 14.3 × 10⁻⁹ M and 205 fmol bound/mg protein. Specific binding is time-dependent and proportional to protein concentration. Glycine and taurine are equally potent inhibitors of [³H]strychnine binding ( K _i= 4 × 10⁻⁵ M); no other amino acids endogenously present in the retina inhibited [³H]strychnine binding.     

Oxygen uptake in developing eggs and larvae of the cod, Gadus morhua L.

Unfertilised cod eggs showed a mean oxygen uptake rate at 5°C of 0.089 μl O₂, dry wt.⁻¹ h⁻¹; this gradually rose to 0.768 μl O₂ mg dry wt.⁻¹ h⁻¹ in eggs about to hatch. From hatching to complete yolk absorption larvae respired at 1.6 μl O₂, mg dry wt.⁻¹ h⁻¹. During starvation following yolk absorption, uptake fell significantly to 1.1 μl O₂, mg dry ⁻¹ h⁻¹. Much of this decrease in oxygen consumption was shown to be caused by reduction in activity. Loss of weight during the embryo and larval phases could not easily be reconciled with total oxygen consumption; it is suggested that cod embryos and larvae may not rely solely upon endogenous energy reserves during development.     

Attraction of zebrafish, Brachydanio rerio, to alanine and its suppression by copper

Preference responses of zebrafish to 10⁻³, 10⁻⁴ and 10⁻⁵M alanine (Ala) were concentration- dependent. Behavioural responses to copper (Cu) and Cu + Ala mixtures were also assessed. Zebrafish avoided 100 and 10 μg Cu l⁻¹, but not 1 μg l⁻¹. Mixtures of 10⁻³ m Ala+ 100 μg Cu l⁻¹ and 10 ⁴ M Ala + 10 μg Cu 1⁻¹ were avoided as intensely as was Cu alone. Responses to 10⁻³ M Ala + 10 or 1 μg Cu l⁻¹ and 10 ⁴ M Ala +1 μg Cu l⁻¹ did not differ statistically from controls (no detectable preference or avoidance). These results demonstrate, firstly, that a concentration of a pollutant avoided by itself (10 μg Cu l⁻¹) may not be avoided when encountered with an attractant chemical stimulus (Ala) and may suppress the preference for an attractant stimulus, and secondly, that a concentration of a pollutant not avoided by itself and not considered deleterious (1 μg Cu l⁻¹) suppresses attraction to Ala (an important constituent of prey odours for many fishes).     

Photosynthesis and carbon metabolism in photoautotrophic cell suspensions of Chenopodium rubrum from different phases of batch growth

Rapidly dividing photoautotrophic cell suspensions from Chenopodium rubrum L. assimilated about 85 μmol CO₂ (mg chlorophyll)⁻¹ h⁻¹. During the late stationary phase of culture growth, CO₂ fixation rate was reduced to about 60 μmol CO₂ (mg chlorophyll)⁻¹ h⁻¹. Actively dividing cells characteristically incorporated a smaller proportion of ¹⁴C into starch than cells from non-dividing stationary phases. In rapidly dividing cells, [¹⁴C]-turnover from free sugars, sugar-phosphates, organic and amino acids was substantially higher compared to non-dividing cells from stationary growth phase. Higher proportions of photosynthetically fixed carbon were channelled into proteins, lipids and structural components in actively dividing cells than in non-dividing cells. In the latter. ¹⁴C was preferentially channeled into starch, and a striking increase in starch accumulation was observed. The transfer of non-dividing, stationary growth-phase cells into fresh culture medium resulted in an increase in the maximum extractable activities of some enzymes involved in the glycolytic and dark respiratory pathways and in the citric acid cycle. In contrast, the maximum extractable activities of the chloroplastic enzymes, ribulose-1,5-bisphosphate carboxylase (EC 4.1.1.38) and NADP⁺-glyceraldehyde-3-phosphate dehydrogenase (EC 1.2.1.13) were highest after the cells had reached the stationary growth phase.