Regulation of respiration in brain mitochondria and synaptosomes: restrictions of ADP diffusion in situ, roles of tubulin, and mitochondrial creatine kinase

The role of ubiquitous mitochondrial creatine kinase (uMtCK) reaction in regulation of mitochondrial respiration was studied in purified preparations of rat brain synaptosomes and mitochondria. In permeabilized synaptosomes, apparent Km for exogenous ADP, Km (ADP), in regulation of respiration in situ was rather high (110 +/- 11 microM) in comparison with isolated brain mitochondria (9 +/- 1 microM). This apparent Km for ADP observed in isolated mitochondria in vitro dramatically increased to 169 +/- 52 microM after their incubation with 1 muM of dimeric tubulin showing that in rat brain, particularly in synaptosomes, mitochondrial outer membrane permeability for ADP, and ATP may be restricted by tubulin binding to voltage dependent anion channel (VDAC). On the other hand, in synaptosomes apparent Km (ADP) decreased to 25 +/- 1 microM in the presence of 20 mM creatine. To fully understand this effect of creatine on kinetics of respiration regulation, complete kinetic analysis of uMtCK reaction in isolated brain mitochondria was carried out. This showed that oxidative phosphorylation specifically altered only the dissociation constants for MgATP, by decreasing that from ternary complex MtCK.Cr.MgATP (K (a)) from 0.13 +/- 0.02 to 0.018 +/- 0.007 mM and that from binary complex MtCK.MgATP (K (ia)) from 1.1 +/- 0.29 mM to 0.17 +/- 0.07 mM. Apparent decrease of dissociation constants for MgATP reflects effective cycling of ATP and ADP between uMtCK and adenine nucleotide translocase (ANT). These results emphasize important role and various pathophysiological implications of the phosphocreatine-creatine kinase system in energy transfer in brain cells, including synaptosomes.     

Paralarval development,abundance, and distribution of Pterygioteuthis hoylei (Cephalopoda: Oegopsida: Pyroteuthidae) in the Gulf of California,México

The morphology and ecology of paralarvae (PL) through to the juvenile stages are unknown in most cephalopod species in spite of their importance as indicators of biodiversity, of adult spawning grounds, and as key basis for fisheries management programs. We describe, for the first time, the morphological development of Pterygioteuthis hoylei (Pfeffer 1912 Pfeffer, G. 1912. Die Cephalopoden der Plankton-Expedition. Zugleich eine monographische übersicht der Oegopsiden Cephalopoden. Ergebnisse der Plankton-Expedition der Humboldt-stiftung, 2: 1–815.  [Google Scholar]) and used cytochrome C oxidase for genetic identification of specimens, linking PL with both juveniles and adults. Pterygioteuthis hoylei represents the most abundant species of squid in oblique plankton collections in the Gulf of California, México. PL were collected in plankton tows made in the Gulf of California from 2004 to 2007. Almost all morphological features followed an increasing size/count trend. Most specimens were collected during March–May, accounting for 71.8% of total cephalopod paralarval abundance, and in November (13.5%). Size, abundance and distribution analyses indicate a high abundance of recently hatched PL (1.1–2.0 mm mantle length) off Bahía Concepción to Bahía de La Paz in these months along with the highest abundance of juveniles and adults in the same area. These results indicate that reproductive activity of P. hoylei takes place in the southern Gulf of California in late winter to early spring, and in November, representing an important spawning and hatching area for this species.     

Vitellogenesis and changes in lipid and protein content of oocytes of Trophon geversianus (Neogastropoda: Muricidae) in Golfo San José (Chubut,Argentina)

The reproductive cycle of Trophon geversianus (Pallas 1774) population from Golfo San José (Chubut, Argentina, 42°33′S, 64°33′W) was studied using histochemical methods and digital image analysis. For such purpose, ovary samples were taken monthly between July 2006 and August 2007 and stained with hematoxylin and eosin, Sudan black B (to identify lipids) or Schiff’s ninhydrin (to identify proteins). Four different gametogenic stages were described: oogonias, previtellogenic oocytes, early vitellogenic oocytes, and late vitellogenic oocytes. Two spawning events were registered; one between September and October 2006, and a second one between February and March 2007. Oocyte quality was determined by the changes in lipid and protein composition during gametogenesis. Through digital image analysis, a lipid and a protein indexes (LI and PI) were calculated. Significant differences were observed in LI over month sampled, contrarily to what PI showed. A significant and positive correlation was found between lipid and protein content and oocytes areas, indicating that these nutrients accumulate during the entire vitellogenesis. Also, when dividing the oocytes into two size groups, analysis indicates a positive correlation between LI and oocyte area for smaller area oocytes. This demonstrates that while proteins accumulate linearly throughout vitellogenesis process, lipids accumulate in two steps: first at a growing rate, secondly at constant rate. This information is essential to determine the nutritional requirements of brood stock individuals at hatcheries in this potential fishery resource that inhabits patagonian waters.     

Diversity and distribution of oribatid mites (Acari:Oribatida) in a lowland rain forest in Peru and in several environments of the Brazilians states of Amazonas, Rond?nia, Roraima and Pará.

We are summarizing the current state of knowledge of the diversity and distribution of oribatid mites in 26 environments in northern Brazil and of a rain forest in Peru. The published studies were mostly concentrated in Central Amazon. Only one report is a result from an agricultural polyculture. We are providing the first lists of species for savannas and for the Brazilian states of Roraima and Pará. Up to date, 146 species are definitively identified from a total of 444 taxa with 188 known genera, reinforcing the notion of a rich biodiverse area. The high number of 298 non-described species (morphospecies) clearly shows the inadequacy of the current taxonomic knowledge for the region. Most of the registers are from forest environments. In the soil from primary forests, we registered the highest diversity (54-155 species/morphospecies). Eighty-nine species were unique to primary forests, followed by 34 for savannas, 32 in trees, 10 in "igapó", 4 in caatinga, 3 in secondary forests, two in "várzea" and one in polyculture. Twenty genera were the most speciose. The species with the largest home ranges were Rostrozetes foveolatus, Scheloribates sp. A, and Galumna sp. A. Our numbers reflect the lack of taxonomists and show that the taxonomic knowledge must be improved for the region or we will continue to work with taxonomic resolution of Order or Family and a high percentage of morphospecies, which will probably be appropriate to the question being asked in each study, but not for a comparison among environments.     

Indicators of infestation, colonization and infection of Triatoma dimidiata (Latreille) (Hemiptera: Reduviidae) in Campeche, México

Triatoma dimidiata (Latreille) is considered to be one of the primary vectors of Chagas disease in Southern Mexico and Central America. The objective of the present study was to obtain ecological information on T. dimidiata in two rural communities of Campeche, Mexico, where the vector is poorly studied. Our work consisted of monthly samplings carried out during one-year time at three levels: sylvatic, peridomestic and intradomestic, in order to estimate the population abundance of this species and its rate of infection with Trypanosoma cruzi. Triatoma dimidiata was the unique vector of this disease collected in San Juan Bautista Sakcabchen (SJBS) and Crucero San Luis (CSL). The total of 145 individuals were captured in SJBS; from these, 26.9%, 20% and 53.1% were collected in the sylvatic, peridomestic and intradomestic area, respectively. In CSL captures yielded 108 individuals: 40.7% in the sylvatic area, 20.4% peridomestic and 38.9% intradomestic. We found no correlation between climatic variables and population abundance of T. dimidiata. Dataset obtained suggests that individuals from the sylvatic area exhibit a high rate of natural infection by T. cruzi, with monthly percentages up to 61.5% for SJBS and 50% for CSL. At the peri and intradomestic level, the reservoirs apparently play an important role in the transmission, as the seroprevalence in dogs was 61.5% y 65.4%, for SJBS y CSL, respectively. Based on these findings, it was concluded that inhabitants of both communities are at a high risk of Chagas disease infection.     

Variation in timing and abundance of flowering by Delphinium barbeyi Huth (Ranunculaceae): the roles of snowpack, frost, and La Niña, in the context of climate change

Delphinium barbeyi is a common herbaceous wildflower in montane meadows at 2,900 m near the Rocky Mountain Biological Laboratory, and its flowers are important nectar resources for bumblebees and hummingbirds. During the period 1977-1999 flowering was highly variable in both timing (date of first flower ranged from 5 July to 6 August, mean=17 July) and abundance (maximum open flowers per 2Ƕ-m plot ranged from 11.3 to 197.9, mean=82). Time and abundance of flowering are highly correlated with the previous winter's snowpack, as measured by the amount of snow remaining on the ground on 15 May (range 0-185 cm, mean=67.1). We used structural equation modeling to investigate relationships among snowpack, first date of bare ground, first date of flowering, number of inflorescences produced, and peak number of flowers, all of which are significantly correlated with each other. Snowpack depth on 15 May is a significant predictor of the first date of bare ground (R²=0.872), which in turn is a significant predictor of the first date of flowering (R²=0.858); snowpack depth is also significantly correlated with number of inflorescences produced (R²=0.713). Both the number of inflorescences and mean date of first flowering are significant predictors of flowers produced (but with no residual effect of snowpack). Part of the effect of snowpack on flowering may be mediated through an increased probability of frost damage in years with lower snowpack - the frequency of early-season "frost events" explained a significant proportion of the variance in the number of flowers per stem. There is significant reduction of flower production in La Niña episodes. The variation in number of flowers we have observed is likely to affect the pollination, mating system, and demography of the species. Through its effect on snowpack, frost events, and their interaction, climate change may influence all of these variables.     

Sequence and expression of an α-amylase gene in four related species of prickleback fishes (Teleostei: Stichaeidae): ontogenetic,dietary, and species-level effects

Partial α-amylase gene sequences were determined and α-amylase gene expression was quantified in four species of carnivorous, omnivorous, and herbivorous prickleback fishes (family Stichaeidae) to assess the effects of ontogeny, diet, and species on expression of this gene. Pairwise comparison of α-amylase nucleotide sequences revealed 96–98 % identity, and comparison of amino acid portions revealed 93–95 % similarity among the four prickleback species. Expression was determined using in situ hybridization and intensity of expression quantified using image analysis. Alpha-amylase expression level was compared in three feeding categories of the four species: (1) small, wild-caught carnivorous juveniles; (2) larger, wild-caught juveniles of the carnivorous species and the three that had shifted to herbivory or omnivory; and (3) larger, juveniles produced by feeding a low-starch artificial diet to small juveniles until they reached the size of the larger wild-caught juveniles. The results showed no dietary effect in any species but significant ontogenetic and species-level effects in Cebidichthys violaceus, as well as in the sister species Xiphister mucosus and X. atropurpureus. Based on a phylogeny for the Stichaeidae produced for this study using two mtDNA genes and one nuclear gene, the ontogenetic dietary shifts to herbivory/omnivory evolved independently in C. violaceus and in the clade containing the two species of Xiphister. All three of these species increased α-amylase gene expression with increase in size and had higher expression than Anoplarchus purpurescens, which is a member of a third, stichaeid clade comprising carnivores. These results show the importance of α-amylase in the herbivores and omnivores.     

Streptomycetes in micro-cultures: Growth, production of secondary metabolites, and storage and retrieval in the 96–well format

Mycelium-forming Streptomyces strains were grown in one milliliter liquid micro-cultures in square deep-well microtiter plates. Growth was evaluated with respect to biomass formation and production of secondary metabolites which were found to be very similar in the micro-cultures, bioreactor, and shake flask cultivations, respectively. Despite repetitive sampling and extensive growth on the walls of the wells, no cross contamination occurred. Furthermore, we successfully employed cold storage at –20 °C of spore suspensions (in the 96–well format), directly prepared from cultures grown on agar in the microtitre plate. Cultures were retrieved by replicating aliquots from the frozen spore suspensions.     

Associations of aquatic insects (Ephemeroptera,Plecoptera, and Trichoptera) in a network of subarctic lakes and streams in Québec

As part of an impact assessment of large hydroelectric projects in the James Bay drainage in Northwestern Québec, the aquatic insect communities were studied in a network of rivers, lakes and streams during the summer of 1975. Thirty-eight emergence traps operated over the ice-free season yielded 10 888 insects (5559 Ephemeroptera, 2817 Plecoptera, and 2512 Trichoptera), representing 148 species (respectively 44, 18, and 86), most of temperate and boreal afinities. There was no arctic element.Similarity analyses and clustering procedures on the emergence series revealed the existence of distinct insect communities in the river (fast and slow sections), the streams (fast and slow), the lakes and the bogs, each characterized by a particular assemblage of species. Many of the species were more or less ubiquitous and differences between communities were marked more by changes in the dominance of the species and differences in the frequency distributions, than absolute shifts in the species lists. The yields in the traps set in fast water were much greater than those in slow running water, and these in turn greater than those of standing water.By comparison with more southerly sites, the seasonal succession of species was retarded in the spring and early summer, but was not shortened appreciably in the fall. The usual emergence patterns associated with these taxa was observed, namely those of spring, summer and autumn species.     

Development, longevity and reproduction of Trissolcus basalis (Wollaston) and Telenomus podisi Ashmead (Hymenoptera: Scelionidae) in natural conditions during autumn and winter, in southern Paraná, Brazil

The development, longevity and reproduction of Trissolcus basalis (Wollaston) and Telenomus podisi Ashmead were investigated under natural conditions during autumn and winter from 1999 to 2001, in Southern Paraná State, Brazil, in order to understand how these parasitoids overwinter in subtropical regions. Adults of T. basalis e T. podisi emerged during autumn and winter from eggs parasitized between April and June. Adult longevity in natural conditions was higher than eight and seven months, for T. basalis and T. podisi respectively, showing that both species overwinter in the adult stage. Percentage survival of T. basalis and T. podisi females during autumn and winter was higher than 85% and mortality increased significantly in spring. Females maintained for six months in natural conditions parasitized E. heros eggs after transference to 25 degrees C, and the presence of females in the offspring showed that mating occurred during autumn and winter. Temperature during immature development influenced significantly the adult performance, regulating the longevity and reproductive capacity after dormancy. Female parasitoids that developed in higher temperatures lived longer and parasitized more host eggs after transference to 25 degrees C than females that developed under colder conditions.     

Oxo-bridged isomers of aza-trishomocubane sigma (σ) receptor ligands: Synthesis,in vitro binding,and molecular modeling

Isomeric oxo-bridged analogs of aza-trishomocubane sigma (σ) receptor ligands were synthesized and shown to display a reduced affinity for the σ receptor. In the case of phenethyl derivative 4, there was a concomitant introduction of high-affinity for the α_2C adrenergic receptor, and moderate affinity for the dopamine transporter. Molecular modeling was undertaken to rationalize these results.     

Where and why? Bees,snail shells and climate: Distribution of Rhodanthidium (Hymenoptera: Megachilidae) in the Iberian Peninsula

Species distribution patterns are widely studied through species distribution models (SDMs), focusing mostly on climatic variables. Joint species distribution models (JSDMs) allow inferring if other factors (biotic interactions, shared phylogenetic history or other unmeasured variables) can also have an influence on species distribution. We identified current distributional areas and optimal suitability areas of three species of the solitary snail‐shell bee Rhodanthidium (Hymenoptera: Megachilidae), and their host gastropod species in the Iberian Peninsula. We undertook SDMs using Maxent software, based on presence points and climatic variables. We also undertook JSDMs for the bees and the snails to infer if co‐occurrence could be a result of biotic interactions. We found that the three bee species: (1) use at least five different species of Mediterranean snails; (2) use empty shells not only for nesting but also for sheltering when there is adverse weather and during the night; (3) have their most suitable areas in the eastern and southern Iberian Peninsula, mostly on limestone areas; and (4) have their optimal range under Mediterranean climatic values for the studied variables. There is positive co‐occurrence of Rhodanthidium with the gastropod species, especially with the snail Sphincterochila candidissima. The contribution of the environmental component to the co‐occurrence is less than that of the residual component in those cases, suggesting that: (i) the use of biotic resources (between Rhodanthidium and the gastropod species); (ii) shared phylogenetic history (between R. septemdentatum and R. sticticum); or (iii) unmeasured variables are largely responsible for co‐occurrence.     

Theobromine, theophylline, and caffeine in 42 samples and products of Guaraná (Paullinia cupana, Sapindaceae)

Guaranás, the carbonated beverages (sodas) of choice throughout much of Brazil, are mandated by Brazilian law to contain at least 300 mg guara’a (Paullinia cupana, Sapindaceae) seed per 100 ml soda. Were all the soda manufacturers adhering to the law, they would be consuming almost three times the annual production of seed. Guaraná seeds contain unusually high levels of caffeine, along with smaller amounts of the related purine alkaloids theobromine and theophylline. We investigated the purine alkaloid content of three ethnobotanical guaraná collections and 39 commercial products using HPLC/UV. Many of the products did contain caffeine as the major alkaloid, with traces of theobromine and theophylline. Numerous sodas and syrups contained up to ten times more theobromine than caffeine, and we suspect that these products were adulterated with cacao (Theobroma cacao, Sterculiaceae), the major source of theobromine.     

The role of His-50 of α-synuclein in binding Cu(II): pH dependence, speciation, thermodynamics and structure

Copper interaction with alpha synuclein (αS) has been shown to accelerate aggregation and oligomerization of the protein. Three different αS copper binding domains have been proposed: (i) the N-terminal residues (1-9) that represent the minimal copper binding domain; (ii) the His-50 imidazole and (iii) the Asp and Glu residues within the acidic C-terminal domain. The copper coordination at the N-terminus has been extensively characterized and it is generally accepted that it provides the highest affinity site. The same does not hold for the role played by His-50 in copper binding. In this work Cu(ii) coordination to peptide fragments encompassing residues 45-55 of αS has been exhaustively characterized, including systems containing the inherited mutations E46K and A53T, as model peptides of the His-50 site. Through potentiometric titrations all the speciation profiles have been determined and the stability constants have been used to estimate the dissociation constants of complexes corresponding to the binding modes at pH 6.5 and 7.5. Spectroscopic analyses allowed determination of (i) the copper coordination sphere, (ii) its geometry and (iii) the constraints wherefrom the 3D structural models of the copper complexes could be obtained.     

Global Co‐Distribution of Light at Night (LAN) and Cancers of Prostate,Colon, and Lung in Men

The incidence rates of cancers in men differ by countries of the world. We compared the incidence rates of three of the most common cancers (prostate, lung, and colon) in men residing in 164 different countries with the population‐weighted light at night (LAN) exposure and with several developmental and environmental indicators, including per capita income, percent urban population, and electricity consumption. The estimate of per capita LAN exposure was a novel aspect of this study. Both ordinary least squares (OLS) and spatial error (SE) regression models were used in the analysis. We found a significant positive association between population exposure to LAN and incidence rates of prostate cancer, but no such association with lung cancer or colon cancer. The prostate cancer result is consistent with a biological theory and a limited number of previous studies of circadian disruption and risk. The LAN‐prostate cancer connection is postulated to be due to suppression of melatonin and/or disruption of clock gene function. An analysis holding other variables at average values across the 164 countries yielded a risk of prostate cancer in the highest LAN‐exposed countries 110% higher than in the lowest LAN exposed countries. This observed association is a necessary condition for a potentially large effect of LAN on risk of prostate cancer. However, it is not sufficient due to potential confounding by factors that increase the risk of prostate cancer and are also associated with LAN among the studied countries.     

Rates of entry and oxidation of acetate,glucose, d(?)-β-hydroxybutyrate,palmitate, oleate and stearate,and rates of production and oxidation of propionate and butyrate in fed and starved sheep

1. Rates of entry and oxidation of a range of metabolites have been measured in tracheostomized sheep (diet, 800g. of lucerne chaff and 100g. of maize/day) by combining isotope-dilution techniques with the continuous measurement of total respiratory gas exchange, and ¹⁴CO₂ production during the intravenous or intraruminal infusion of ¹⁴C-labelled substrates. 2. Mean entry rates in fed and starved (24hr.) sheep respectively, expressed as mg./min./kg. body wt.^0·75, were: glucose, 5·0 (range 4·8–5·1, 2 observations) and 3·8 (3·2–4·2, 4); acetate, 10·8 (9·1–13·5, 4) and 5·8 (1); d(−)-β-hydroxybutyrate, 1·4 (1) and 1·5 (0·8–2·4, 4); palmitate, oleate and stearate (starved sheep only) 1·0 (0·6–1·9, 7), 0·9 (0·2–1·6, 10) and 0·9 (0·5–1·1, 11) respectively. 3. Production rates of propionate and butyrate in continuously feeding sheep were 6·4 (4·7–8·3, 4) and 4·3 (3·4–6·1, 4) mg./min./kg.^0·75 respectively, and in starved (24hr.) sheep were 2·5 (2·2–2·9, 2) and 1·0 (0·8–1·2, 2) mg./min./kg.^0·75 respectively. 4. Calculated terminal values for the specific radioactivity of respiratory ¹⁴CO₂ during measurements of entry rates and production rates were used to calculate the contributions of individual substrates to overall oxidative metabolism. Mean values for fed and starved sheep respectively were: glucose, 9·1 (8·6–9·6, 2) and 11·2 (5·9–15·1, 4)%; acetate, 31·6 (26·8–38·1, 4) and 22·1 (1)%; d(−)-β-hydroxybutyrate, 10·4 (1) and 4·8 (1·9–7·7, 4)%; propionate, 23·0 (13·8–29·9, 4) and 7·1 (6·8–7·4, 2)%; butyrate, 16·5 (13·7–20·5, 4) and 5·3 (5·2–5·3, 2)%; palmitate, oleate and stearate (starved sheep only), 4·7 (2·0–7·7, 7), 4·0 (1·2–6·6, 10) and 4·4 (3·8–5·8, 9)% respectively. The sum of these values for individual substrates in fed and starved sheep, excluding that of β-hydroxybutyrate and after correction of the glucose value for the known interrelations of this substrate with propionate, accounted for 76% and 58% respectively of total production of carbon dioxide. 5. Calculations based on the proportion of substrate entry directly oxidized indicated that the substrates studied accounted for 63% (fed sheep) and 43% (starved sheep) of total energy expenditure measured by oxygen uptake. The contribution of β-hydroxybutyrate was excluded, and corrections were made for glucose–propionate interrelations, and for the different rates of oxidation of the methyl and carboxyl fragments of acetate. 6. The present results have been combined with those obtained earlier in this Laboratory to examine the relationships between rates of substrate entry and oxidation, and concentrations of substrate in blood. Rates of entry of acetate, glucose, d(−)-β-hydroxybutyrate, palmitate and oleate (but not stearate) were well correlated with concentration in blood, and substrate contribution to production of carbon dioxide showed a similar correlation to blood concentration, except with glucose. 7. It was concluded that the general technique is of potential value in providing valid quantitative parameters of animal metabolism.