Pepsin fragmentation of botulinum type E neurotoxin: Isolation and characterization of 112, 48, 46, and 16 kD fragments

Controlled digestion of 150 kD single chain botulinum type E neurotoxin with pepsin atpH 6.0 produced 112, 48, 46, and 16 kD fragments. These were chromatographically purified; their locations in the 1300 amino acid residue long neurotoxin were determined by identifying the amino terminal 10 residues of 112 and 48 kD fragments, 50 residues of 46 kD fragment, and 59 residues of 16 kD fragment. The 48 and 112 kD fragments contain the N-terminal segment of the neurotoxin (i.e., residue no. 1 to 425 and 1 to 990, respectively), the 46 kD fragment corresponds to 407 residues of the C-terminal region, and the 16 kD fragment contains the 140 residues from a segment nearer to the C-terminus. The 48 kD fragment is similar to the 50 kD N-terminal light chain of the 150 kD dichain neurotoxin, which is generated by tryptic cleavage of the 150 kD single chain neurotoxin, and is separated from the 100 kD C-terminal heavy chain by dithiothreitol (DTT) reduction of an intrachain disulfide bond in the presence of 2 M urea (Sathyamoorthy and DasGupta,J. Biol. Chem. 260, 10461, 1985). The pepsin-generated 48 kD fragment, unlike the light chain, was isolated without exposure to DTT and urea. The single chain 112 kD fragment following trypsin digestion yielded 48 and 60 kD fragments that were separable after DTT reduction of the intrachain disulfide which links them. The N-terminal residues of the smaller fragment were identical to that of the single chain 150 kD neurotoxin; the single chain 112 kD fragment is therefore the neurotoxin minus the 50 kD C-terminal half of the heavy chain. The biological activities of the 48 and 112 kD fragments can be demonstrated in permeabilized PC12 cells (Lomnethet al., J. Neurochem. 57, 1413, 1991); they inhibit norepinephrine release.     

Release of carboxylic anions and protons by tomato roots in response to ammonium nitrate ratio and pH in nutrient solution

The exudation of certain organic anions and protons by roots which may affect solubility of metals and P and uptake by plants, is affected by nitrogen form and pH. The objective of this work was to study exudation of carboxylates and H⁺/OH^– by tomato plants in response to NH₄/NO₃ ratio and pH in nutrient solution. Four NH₄/(NH₄+NO₃) ratios (R= 0, 0.33, 0.67 and 1) and constant vs. variable solution pH treatments were investigated. The sum of the exudation rates of all carboxylates tended to decline with increasing R, particularly tri- and dicarboxylates. The molar fraction of the exuded tri- and dicarboxylates, averaged over all treatments and plant ages, increased in the order tartarate 2%), malate (6%), succinate (15%), citrate (26%) and fumarate (46%). At R=1 the solution pH dropped from 5.2 to 3 and at R=0 increased to 8. The R corresponding to the pH stat of tomato plant was 0.3. For the constant solution pH treatment, the effect of solution pH on carboxylate exudation rate was small as compared to the effect of R. The exudation of citrate and H⁺ efflux which were initiated when NO₃ and NH₄ uptake rates per plant exceeded certain threshold values, increased with plant age.     

Characteristics of sarcoplasmic reticulum membrane preparations isolated from skeletal muscles of active and hibernating ground squirrel Spermophilus undulatus

The total Ca-ATPase activity in the sarcoplasmic reticulum (SR) membrane fraction isolated from skeletal muscles of winter hibernating ground squirrel Spermophilus undulatus is 2.2-fold lower than in preparations obtained from summer active animals. This is connected in part with 10% decrease of the content of Ca-ATPase protein in SR membranes. However, the enzyme specific activity calculated with correction for its content in SR preparations is still 2-fold lower in hibernating animals. Analysis of the protein composition of SR membranes has shown that in addition to the decrease in Ca-ATPase content in hibernating animals, the amount of SR Ca-release channel (ryanodine receptor) is decreased 2-fold, content of Ca-binding proteins calsequestrin, sarcalumenin, and histidine-rich Ca-binding protein is decreased 3-4-fold, and the amount of proteins with molecular masses 55, 30, and 22 kD is significantly increased. Using the cross-linking agent cupric–phenanthroline, it was shown that in SR membranes of hibernating ground squirrels Ca-ATPase is present in a more aggregated state. The affinity of SR membranes to the hydrophilic fluorescent probe ANS is higher and the degree of excimerization of the hydrophobic probe pyrene is lower (especially for annular lipids) in preparations from hibernating than from summer active animals. The latter indicates an increase in the microviscosity of the lipid environment of Ca-ATPase during hibernation. We suggest that protein aggregation as well as the changes in protein composition and/or in properties of lipid bilayer SR membranes can result in the decrease of enzyme activity during hibernation.     

Freeze-fracture studies on the cockroach hemocyte membrane complex: Symmetric and asymmetric membrane particle distributions

Summary Freeze-fracture studies were conducted on the membranes of normal cockroach hemocytes. The plasmalemma is asymmetric with the A fracture face containing 80–100 Å membrane intercalated particles at a concentration of 2500/². The B fracture face contains 120–150 Å particles with a relatively low density (800/²). The nuclear envelope displays an asymmetry with the A fracture face containing 1500 particles/² and the B face containing 300/ ². No significant particle size differences were observed in nuclear envelope fracture faces. Two types of symmetric membranes were also found in these cells. Both A and B fracture faces of the membrane surrounding the numerous cytoplasmic inclusion bodies contain particle sizes and concentrations similar to the B face of the plasmalemma. A second type of symmetry was observed in cells apparently engaged in exocytosis. Vesicles (0.1 D) from this process were completely particle free on both fracture faces. Such particle free vesicles could be found in the cytoplasm, attached to the plasmalemma, or completely separated from the cell.Supported by a Pharmaceutical Manufacturers Association Foundation Fellowship.The author wishes to thank Ms. Annalena K. Charla for assistance in plate preparation, Dr. Julius Schultz and the Papanicolaou Cancer Research Institute for use of the freeze-etch device, and Dr. David Smith for the electron microscope facilities.     

Temporal changes of mangrove vegetation boundaries in Amazônia: Application of GIS and remote sensing techniques

The present work analyses a 25-year timeseries of radar and satellite images, andidentifies areas with losses and gains ofvegetation coverage along 166 km of Para'scoastline (North Brazil) and in the innerparts of the Bragança peninsula. Thegeomorphology of this region hassignificantly changed in recent years. Aresult of these changes has been a retreatof the mangrove vegetation along thecoastline, mainly due to landward sandmigration, which covers the mudflat andasphyxiates the vegetation. Image analysissuggests that the loss of vegetationcoverage has been the dominating process inthe last 25 years, occurring on 42% ofthe Bragança coastline and adjacentareas. Vegetation has remained stable along 39% of the coastline, while mudsedimentation has allowed mangrovedevelopment along 19% of it. On the otherhand, during this period of time, mangroveshave invaded 3.4 km² ( 38%) of theelevated herbaceous flats in the highestsectors of the Bragança peninsula.Despite other possible causes for mangrovedeath along the coastline, such as littoraldrift currents or cyclical changes incoastal sediment dynamics, the invasion ofmangrove into herbaceous elevated flats inthe central peninsula cannot be attributedto these factors. The current dynamics ofvegetation coverage change seem to becompatible with a long-term trend relatedto the predicted rates of sea-level rise.     

Identification of a specific protein in the mitochondrial fraction of the rat heart whose phosphorylation is inhibited by taurine

Summary It was previously reported that the mitochondrial fraction of the rat heart contained a specific protein with a molecular weight of approximately 44kDa whose phosphorylation was inhibited by taurine (Lombardini,1994a). Isolation of the 44kDa phosphoprotein on a 1-dimensional polyacrylamide gel using traditional glycine buffers followed by re-electrophoresing the cut out proportion of the gel which corresponds to the 44kDa protein on a tricine-buffered gel resulted in sufficient pure protein for sequence analysis. The results indicate that the 44kDa phosphoprotein is pyruvate dehydrogenase.     

Semper's (zoanthid) larvae: pelagic life,parentage and other problems

Semper's larvae were obtained from <300 out of 1800 plankton tows taken in the world's oceans (1964–1993). Zoanthellae (larvae of Sphenopidae) occurred at 217 stations and zoanthinae (larvae of Zoanthidae) at 86, the two larval types showing distributions clearly delimited by a minimum sea temperature (22 °C for zoanthellae, 18 °C for zoanthinae; a statistically significant difference, P<0.001). Length of formalin-fixed zoanthellae was 2–8.6 mm and of zoanthinae 1.5–5.9 mm. Endodermal zooxanthellae were present in 9/24 zoanthinae but in no zoanthellae (of 19). Three larvae contained an endo-commensal/parasitic amphipod. Septa were externally visible in larger zoanthinae and were counted in transverse sections of other larvae, a majority of which (both kinds) had 12 septa, the normal maximum. The pattern was brachycnemic in 40/43 larvae and anomalous (but non-macrocnemic) in three. If macrocnemic genera reproduce by Semper's larvae, they should have been represented in such a large sample. The distribution of adult Epizoanthus was examined: many species are deep sea (recorded down to 5000 m) but shallow-water species are relatively plentiful in, for example, the Adriatic and North Seas. No Semper's larva has ever been recorded from either. Some Parazoanthus species also occur in shallow water, especially associated with western Atlantic reef sponges. If they produce Semper's larvae, these have never been found. It is probable that macrocnemic zoanthids settle from planulae that do not develop into recognizable zoanthellae or zoanthinae.     

Subunit distribution of calcium-binding sites in Lumbricus terrestris hemoglobin

The giant, 3.6-MDa hexagonal bilayer hemoglobin (Hb) of Lumbricus terrestris consist of twelve 213-kDa globin subassemblies, each comprised of three disulfide-bonded trimers and three monomer globin chains, tethered to a central scaffolding of 36–42 linkers L1–L4 (24–32 kDa). It is known to contain 50–80 Ca and 2–4 Cu and Zn; the latter are thought to be responsible for the superoxide dismutase activity of the Hb. Total reflection X-ray fluorescence spectrometry was used to determine the Ca, Cu, and Zn contents of the Hb dissociated at pH 2.2, the globin dodecamer subassembly, and linker subunits L2 and L4. Although the dissociated Hb retained 20 Ca²⁺ and all the Cu and Zn, the globin subassembly had 0.4 to 3 Ca²⁺, depending on the method of isolation, and only traces of Cu and Zn. The linkers L2 and L4, isolated by reversed-phase high-pressure liquid chromatography at pH 2.2, had 1 Ca per mole and very little Cu and Zn. Electrospray ionization mass spectrometry of linker L3 at pH 2.2 and at neutral pH demonstrated avid binding of 1 Ca²⁺ and additional weaker binding of 7 Ca²⁺ in the presence of added Ca²⁺. Based on these and previous results which document the heterogeneous nature of the Ca²⁺-binding sites in Lumbricus Hb, we propose three classes of Ca²⁺-binding sites with affinities increasing in the following order: (i) a large number of sites (>100) with affinities lower than EDTA associated with linker L3 and dodecamer subassembly, (ii) 30 sites with affinities higher than EDTA occurring within the cysteine-rich domains of linker L3 and dodecamer subassembly, and (iii) 25 very high affinity sites associated with the linker subunits L1, L2, and L4. It is likely that the low-affinity type (i) sites are the ones involved in the effects of 1–100 mM Group IIA cations on Lumbricus Hb structure and function, namely increased stability of its quaternary structure and increased affinity and cooperativity of its oxygen binding.     

Membrane differentiation in the cytoplasmic-tubule system of the intestinal absorptive cells of the lamprey,Lampetra japonica

Summary Specific membrane differentiation occurs in the cytoplasmic-tubule system of the absorptive cells lining the mucosa of the lamprey anterior intestine. The absorptive cells are characterized by the presence of abundant mitochondria and a system of well-developed cytoplasmic tubules (120 nm in diameter). The cytoplasmic tubules open on to the basolateral cell surface and contain numerous lipoprotein particles (50–100 nm diam.) in their lumina. Lipoprotein particles are also observed in the endoplasmic reticulum and the Golgi complex, and they are transfered to the lateral intercellular space and lamina propria by way of the cytoplasmic tubules. Spirally-wound parallel rows of particles are found in the luminal surface of the cytoplasmic tubules. The rows are 17 nm apart and are wound spirally at a pitch of 210 nm. Freeze-fracture images of the tubule membranes also show spiral arrays of particles (9 nm in diameter) on the P-face, and complementary shallow grooves on the E-face. From these observations, it is suggested that the cytoplasmic-tubule system of the intestinal absorptive cells serves as a channel for the transport of synthesized lipoprotein into the interstitium, and is also the site of the ion and water exchange essential for the maintenance of ionic homeostasis.     

A novel cilia-based feature within the food grooves of the ctenophore Mnemiopsis mccradyi Mayer

We describe a novel compound ciliary structure (g-cilium) from the food groove of the lobate ctenophore Mnemiopsis mccradyi. G-cilia are small, flat compound ciliary organelles that are oriented with their tips pointing toward the mouth. Typically three to four rows of g-cilia line the inner surface of the tentacular groove, which together with the transport groove, make up the food groove. G-cilium cells are 11.4 m long and 4.2 m wide at the g-cilium base. The g-cilium itself is 3.4 m long and tapers to a flat, sharp tip. G-cilia are not motile but are surrounded by many hundreds of smaller, actively motile cilia that beat with orally-directed effective strokes. G-cilia contain 50 conventional `9+2' cilia embedded in a fibrous core that arises from the cell body. In addition, g-cilia contain mitochondria, thousands of small membrane-bounded vesicles and rod bacteria. G-cilia basal bodies are anchored by large, strongly-banded rootlets that extend approximately the entire length of the cell. G-cilia may have organizational, sensory and/or secretory function within the feeding apparatus. Their placement strongly suggests that they play critical roles in feeding. They may enhance the efficiency of prey capture and so contribute to M. mccradyi's well-known voracious appetite. By enhancing prey capture they probably play a critical role in the capacity of this organism to follow prey dynamics, so contributing to dense blooms in mid-late summer in coastal regions.     

Heterogeneity of calcium compartmentation: Electron probe analysis of renal tubules

Summary The objective of this study has been to determine the intracellular localization of calcium in cryofixed, cryosectioned suspensions of kidney proximal tubules using quantitative electron probe X-ray microanalysis. Two populations of cells have been identified: 1) Viable cells, representing the majority of cells probed, are defined by their relatively normal K/Na concentration ratio of 41. Their measured Ca content is 4.1±1.4 (sem) mmol/kg dry wt in the cytoplasm and 3.1 ± 1.1 mmol/kg dry wt in the mitochondria, or an average cell calcium content of 3.8 mmol/kg dry wt. 2) Nonviable cells, defined by the presence of dense inclusions in their mitochondria and a K/Na concentration ratio of 1. The Ca content is 15±2 mmol/kg dry wt in the cytoplasm and 685±139 mmol/kg dry wt in the mitochondria of such cells. Assuming 25 to 30% of the cell volume is mitochondrial, the overall calcium content of such nonviable cells is 210 mmol/kg dry wt. The presence of these inclusions in 4 to 5% of the cells would account for the average total Ca content measured in perchloric acid extracts of isolated proximal tubule suspensions ( 18 nmol/mg protein or 12.6 mmol/kg dry wt). Whole kidney tissues display a large variability in toal Ca content (4.5 to 18 nmol/mg protein, or 3.4 to 13.5 mmol/kg dry wt), which could be accounted for by inclusion in 0 to 4% of the cells. The electron probe X-ray microanalysis (EPXMA) data conclusively demonstrate that thein situ mitochondrial Ca content of viable cells from the kidney, proximal tubule is low and support the idea that mitochondrial Ca may regulate dehydrogenase activity but probably does not normally control cytosolic free Ca.     

Photosystem-II repair and chloroplast recovery from irradiance stress: relationship between chronic photoinhibition,light-harvesting chlorophyll antenna size and photosynthetic productivity in Dunaliella salina (green algae)

High-light (HL) grown Dunaliella salina cells exhibit lower pigment content, a highly truncated chlorophyll (Chl) antenna size, and accumulation of photodamaged PS II centers in the chloroplast thylakoids (chronic photoinhibition). In HL-grown cells, the rate of photosynthesis saturated at higher irradiances and the quantum yield was lower compared to that of normally-pigmented low-light (LL) grown cells. In spite of these deficiencies, the light-saturated rate of photosynthesis for the HL-cells, when measured on a per chlorophyll basis, was 3 times greater than that of the LL-grown cells. To delineate the effect of photoinhibition from the Chl antenna size on quantum yield and rate of photosynthesis, HL-acclimated cells were switched to LL-conditions. Repair of photodamaged PS II, estimated from the recovery of functional PS II centers and from the increase in the quantum yield of photosynthesis, occurred with a half-time of 1 h. Chlorophyll accumulation in the cells occurred with a half-time of 4 h. The differential kinetics in repair versus Chl accumulation provided a window of opportunity, within about 2–3 h after the HLLL shift, when cells exhibited a high quantum yield of photosynthesis, a small Chl antenna size and a light-saturated rate that was 6–9 times greater than that of the normally pigmented LL-grown cells. The work provides insight on the temporal sequence of events at the chloroplast and thylakoid membrane levels, leading from a chronic photoinhibition of PS II to repair and recovery. It is suggested that it is possible to maximize photosynthetic productivity and light utilization in mass microalgal cultures by minimizing the light-harvesting Chl antenna size of the photosystems.     

The Effect of Salicylic Acid on Peroxidase Activity in Wheat Calli Cocultured with the Bunt Pathogen Tilletia caries

The effect of salicylic acid (SA) on peroxidase activity in wheat (Triticum aestivum L.) calli cocultured with the bunt pathogen Tilletia caries was studied. Fungal infection was shown to activate cytoplasmic peroxidase. SA suppressed total peroxidase activity but did not inhibit the peroxidase with pI 9.8. A novel chitin-specific peroxidase with pI 3.5 appeared after the SA treatment. The infection of SA-treated cells with Tilletia caries activated the isoenzymes with pI 3.5, 4.8, and 7.5 and stimulated their secretion into the culture medium. The ability of SA to control wheat peroxidase activity during pathogenesis is discussed. The important role of this control in plant defense responses to the bunt pathogen is emphasized.     

Comparative studies on cellular resistance of bivalves from marine and brackish waters

Summary 1. The cellular resistance of representatives from four euryhaline poikilosmotic bivalves caught in Büsum (North Sea, 30 S), Kieler Förde (Belt Sea, 15 S), and Tvärminne (Gulf of Finland, 6 S) has been tested. All experiments were performed in October.2. According to their cellular salinity rangesMacoma baltica andMytilus edulis are more euryhaline thanMya arenaria andCardium edule.3. The cellular heat resistance decreases in specimens from the North Sea in the following order:Cardium, Mya, Macoma, Mytilus. Among the representatives collected near Tvärminne,Macoma has the highest heat resistance.4. The cellular freezing resistance in specimens from the North Sea is highest inMytilus. The bivalves of the Gulf of Finland practically lack resistance to freezing.5. In localities of decreased salinity the cellular resistance of these bivalves to a dilution of the external medium is somewhat increased, but simultaneously the cellular resistance to other environmental factors, such as heat, freezing and high salinity, decreases unspecifically.

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Vergleichende Untersuchungen über zelluläre Resistenz bei Muscheln aus Meer- und Brackwasser

Kurzfassung An vier euryhalinen poikilosmotischen Muschelarten aus Büsum (Nord-seeküste, 30 S), der Kieler Förde Beltsee, 15 S) und Tvärminne (Finnischer Meerbusen, 6 S), wurden zur gleichen Jahreszeit (Herbst) vergleichende Untersuchungen über die zellulären Resistenzgrenzen durchgeführt. Auf Grund ihrer zellulären Salzgehaltsbereiche sindMacoma baltica undMytilus edulis euryhaliner alsMya arenaria undCardium edule. Die zelluläre Hitzeresistenz nimmt bei den Individuen aus der Nordsee in der Reihenfolge:Cardium, Mya, Macoma, Mytilus ab. Bei Exemplaren aus Tvärminne ist die zelluläre Hitzeresistenz vonMacoma am größten. Die zelluläre Gefrierresistenz ist unter den Muscheln aus der Nordsee am größten, fehlt dagegen praktisch bei den Tieren aus dem Finnischen Meerbusen. Bei verringertem Salzgehalt des Fundortwassers ist die zelluläre Resistenz der untersuchten Muscheln gegenüber Verdünnung des Außenmediums jeweils etwas erhöht, gleichzeitig nimmt die zelluläre Resistenz gegenüuber anderen Außenfaktoren wie Hitze, Frost und hohen Salzkonzentrationen unspezifisch ab.

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Dedicated to Professor Dr.Friedrich Krüger on his 65^th birthday, August 18, 1967.     

Effects of taurine on the phosphorylation of specific proteins in subcellular fractions of the rat retina

The effects of 20 mM taurine on the phosphorylation of specific proteins in mitochondrial and rod outer segment subcellular fractions of the rat retina were measured. A band of protein with an apparent molecular wieght of 20K was consistently inhibited by taurine. Densitometry measurements performed on gel electrophoresis autoradiograms from the mitochondrial fraction demonstrated a 42.7±8.3% decrease due to taurine (20 mM) in the area corresponding to radioactivity from the 20K phosphoprotein. However, only a 21.2±9.0% decrease was observed due to taurine in the rod outer segment preparation. These data suggest that taurine is exerting its primary effect on the phosphorylation of the 20K molecular weight protein in the mitochondria of the retina. In addition, calmodulin and phorbol ester had no effect on the phosphorylation of the 20K molecular weight protein.     

Absolute measurement of cell expansion in plant cell suspension cultures

A method is described for the measurement of intracellular volume (V_i) in cell cultures. In principle, any stable compound that neither penetrates the plasma membrane nor binds to the cells can be used to trace the total extracellular (apoplastic) volume and hence to estimate the intracellular volume. No suitable coloured or UV-absorbing compound could be found among those tested; the main problems were binding to the cell surface and/or instability in the medium. However, [¹⁴C]mannitol was an acceptable apoplastic marker, by use of which we showed that 21–47% of total packed cell volume (PCV) was intracellular, and 14–33% of total settled cell volume (SCV) was intracellular. Therefore, measurements of PCV and SCV misrepresent cell expansion to a variable extent. Cultures of Acer, Rosa, Spinacia and Zea achieved final symplastic volumes of only 9, 14, 6 and 6%, respectively, of the total suspension culture volume.     

Quantitation of movement of the phosphoryl group during catalytic transfer in the arginine kinase reaction: 31P relaxation measurements on enzyme-bound equilibrium mixtures

³¹P nuclear spin relaxation measurements have been made on enzyme-bound equilibrium mixtures of lobster-muscle arginine kinase in the presence of substituent activating paramagnetic cation Co(II) (in place of Mg(II)), i.e., on samples in which the reaction, ECoATParginine ECoADPP-arginine, is in progress. The results have been analyzed on the basis of a previously published theory (Nageswara Rao, B.D. (1995) J. Magn. Reson., B108, 289–293) to determine the structural changes in the reaction complex accompanying phosphoryl transfer. The analysis enables the determination of the change in the Co(II)-³¹P (-P(ATP)) vector as the transferable phosphoryl group moves over and attaches to arginine to form P-arginine. It is shown that the Co(II)-³¹P distance of 3.0 Å, representing direct coordination of Co(II) to -P(ATP), changes to 4.0 Å when P-arginine is formed in the enzyme-bound reaction complex. This elongation of the Co(II)-³¹P vector implies an excursion of at least 1.0 Å for the itinerant phosphoryl group on the surface of the enzyme.     

Endothelin-1 and insulin induce cellular inactivation of protein kinase FA/glycogen synthase kinase-3α in a common signaling pathway

In this study, we investigate the effects of endothelin-1 (ET-1) and insulin on the cellular activity of protein kinase F_A/glycogen synthase kinase-3 (kinase F_A/GSK-3) in rat adipocytes. The cellular activity of kinase F_A/GSK-3 is inhibited to 50% of control within 30 min when cells are treated with 1 nM ET-1 at 37°C; in addition, significant inhibition to 60% of control is observed at as low as 1 pM ET-1. Conversely, ET-1 at concentrations up to 1 nM has no direct effect on purified kinase F_A/GSK-3 in vitro. Immunoblotting analysis further reveals that the protein level of this kinase is not significantly changed when treated with 1 nM ET-1 for 30 min. Similar to ET-1, insulin as low as 10 nM can also induce inactivation of kinase F_A/GSK-3 to 50% of control in adipocytes when processed under identical conditions. Most importantly, when treated with both insulin and ET-1, the activity of kinase F_A/GSK-3 can be decreased only to 50% of control. Taken together, the results provide initial evidence that ET-1 and insulin may regulate this important multisubstrate/multifunctional protein kinase in a common signaling pathway in cells.     

Presynaptic Modulation of K+-Evoked [3H]Dopamine Release in Striatal and Frontal Cortical Synaptosomes of Normotensive and Spontaneous-Hypertensive Rats

Regional differences in presynaptic [³H]dopamine ([³H]DA) release and its modulation by D₂ DA-receptors between the frontal cortex and striatum obtained from Wystar-Kyoto (WKY) and spontaneous-hypertensive rats (SHR) have been evaluated using superfused synaptosomes. Synaptosomal tritium content was significantly lower in the frontal cortex than in the striatum in both SHR and WKY (45% and 48%, respectively), but no differences in tritium content were obtained between strains. However, the 15 mM K⁺-evoked [³H]DA overflow was lower in the SHR as compared to WKY rats in both brain regions (striatum 23%, frontal cortex 21). Concentration-response curves for quinpirole (1nM-10 M)-mediated inhibition of 15mM K⁺-evoked [³H]DA release showed no differences between SHR and WKY. These results suggest that SHR has less ability to release [³H]DA as compared to WKY rats, but SHR did not show differences in the autoregulation of such release in both the frontal cortex and striatum.     

Variation with depth and season in metal sulfides in salt marsh soils

Seasonal monitoring of metal sulfides was carried out in four soils ofthe Ría de Ortigueira salt marshes. Soils from the high salt marsh (withsuboxic redox conditions at the surface), had low concentrations of ironsulfides (AVS and pyrite fraction) and thus a low degree of trace metalpyritization (DTMP) in surface layers (0–10 cm), butconcentrations of metals associated with the pyrite fraction increasedconsiderably at depth (27.5 cm). In the low salt marsh soils (withanoxic conditions at the surface) maximum concentrations of metal sulfides werefound in the surface layers of soils colonized by Spartina maritima. These results are explained by the double effectexerted by roots in strongly reduced soils. On the one hand, they stimulate theactivity of sulfate-reducing bacteria and on the other, they favour the partialoxidation of the soil, thus generating polysulfides with which Fe²⁺immediately precipitates as pyrite, whereas in the deepest, permanently anoxiclayers, pyrite must be formed in a reaction in which FeS is an intermediate, asfollows: FeS + H₂S FeS₂ + H₂.Concentrations of metal sulfides also varied greatly with the season, with twopatterns being distinguished. In soils colonized by S. maritima in both high and low salt marshes, the lowestconcentrations were found in summer. At this time of the year there is a netloss of metal sulfides throughout the profile, presumably due to physiologicalactivity of plants (evapotranspiration and release of oxygen from roots). Incontrast, maximum concentrations of AVS and pyritic metals were found in thesummer in the low salt marsh soils not colonized by vascular plants (creekbottom). In this case, the higher temperatures increased the activity ofsulfur-reducing bacteria leading to synthesis and accumulation of metalsulfidesin the soil.