Global Down-Regulation of Gene Expression in the Brain Using RNA Interference,with Emphasis on Monoamine Transporters and GPCRs: Implications for Target Characterization in Psychiatric and Neurological Disorders

RNA interference (RNAi) is a natural mechanism for regulating gene expression, which exists in plants, invertebrates, and mammals. We investigated whether non-viral infusion of short interfering RNA (siRNA) by the intracerebroventricular route would enable a sequence-specific gene knockdown in the mouse brain and whether the knockdown translates into disease-relevant behavioral changes. Initially, we targeted enhanced green fluorescent protein (EGFP) in mice overexpressing EGFP. A selective knockdown of both EGFP protein and mRNA was observed throughout the brain, with lesser down-regulation in regions distal to the infusion site. We then targeted endogenous genes, encoding the dopamine (DAT) and serotonin transporters (SERT). DAT-siRNA infusion in adult mice produced a significant down-regulation of DAT mRNA and protein and elicited hyperlocomotion similar, but delayed, to that produced on infusion of GBR-12909, a potent and selective DAT inhibitor. Similarly, SERT-siRNA infusion resulted in significant knockdown of SERT mRNA and protein and elicited reduced immobility in the forced swim test similar to that obtained on infusion of citalopram, a very selective and potent SSRI. Application of this non-viral RNAi approach may accelerate target validation for neuropsychiatric disorders that involve a complex interplay of gene(s) from various brain regions.     

Benthic metabolism in two turbid dryland rivers

1. Australian dryland rivers have among the most variable discharge of any rivers worldwide and are characterized by extended periods of no flow during which aquatic habitat contracts into isolated waterholes. Despite naturally high turbidity, benthic primary production is known to be the main source of carbon to waterhole food webs. The objective of this study was to quantify rates of benthic metabolism and identify factors influencing these rates in two Australian dryland rivers, the Cooper Creek and the Warrego River. 2. Both rivers have similar variable hydrology and high levels of turbidity (photic depths < 0.4 m), but fish abundance in Cooper Creek is 10 times than that of the Warrego River. Therefore, an additional aim of the study was to determine if fish abundances reflected underlying differences in benthic primary production. 3. Benthic gross primary production (GPP), benthic respiration, nutrient concentrations and light penetration were measured immediately after flow had ceased (‘post‐flow’) and after at least 2 months of zero flow (‘no‐flow’) in 15 waterholes from each river. A subset of four waterholes from each river was sampled on two additional occasions to determine if patterns were consistent over time. 4. Cooper Creek generally had higher rates of GPP and a more autotrophic benthic zone than the Warrego River. As a result, the expected positive relationship between fish abundance and GPP was generally observed at a broad catchment scale. 4. Light was the major control in benthic GPP in both rivers, as nutrient concentrations were high on all sampling occasions. However, for similar values of photic depth, GPP was greater in Cooper Creek than in the Warrego River. This suggests that more frequent disturbance of the littoral zone may inhibit biofilm development in waterholes of the Warrego River. 5. Although flow variability in dryland rivers is extreme compared with other rivers worldwide, cycles of expansion and contraction of aquatic habitat in these two rivers were associated with a shift in the dominance of regional scale (subcatchments contributing to river flow) versus local scale (waterhole morphology) influences on ecosystem functioning, similar to floodplain rivers in tropical and temperate regions.     

The Mediterranean Sea as a ‘cul‐de‐sac’ for endemic fishes facing climate change

The Mediterranean Sea is a hotspot of biodiversity, and climate warming is expected to have a significant influence on its endemic fish species. However, no previous studies have predicted whether fish species will experience geographic range extensions or contractions as a consequence of warming. Here, we projected the potential future climatic niches of 75 Mediterranean Sea endemic fish species based on a global warming scenario implemented with the Mediterranean model OPAMED8 and a multimodel inference, which included uncertainty. By 2070–2099, the average surface temperature of the Mediterranean Sea was projected to warm by 3.1 °C. Projections for 2041–2060 are that 25 species would qualify for the International Union for the Conservation of Nature and Natural Resources (IUCN) Red List, and six species would become extinct. By 2070–2099, 45 species were expected to qualify for the IUCN Red List whereas 14 were expected to become extinct. By the middle of the 21st century, the coldest areas of the Mediterranean Sea (Adriatic Sea and Gulf of Lion) would act as a refuge for cold‐water species, but by the end of the century, those areas were projected to become a ‘cul‐de‐sac’ that would drive those species towards extinction. In addition, the range size of endemic species was projected to undergo extensive fragmentation, which is a potentially aggravating factor. Since a majority of endemic fishes are specialists, regarding substratum and diet, we may expect a reduced ability to track projected climatic niches. As a whole, 25% of the Mediterranean Sea continental shelf was predicted to experience a total modification of endemic species assemblages by the end of the 21st century. This expected turnover rate could be mitigated by marine protected areas or accelerated by fishing pressure or competition from exotic fishes. It remains a challenge to predict how these assemblage modifications might affect ecosystem function.     

Mites as biological tags of their hosts

Movements and spatial distribution of host populations are expected to shape the genetic structure of their parasite populations. Comparing the genetic patterns of both interacting species may improve our understanding of their evolutionary history. Moreover, genetic analyses of parasites with horizontal transmission may serve as indicators of historical events or current demographic processes that are not apparent in the genetic signature of their hosts. Here, we compared mitochondrial variation in populations of the ectoparasitic mite Spinturnix myoti with the genetic pattern of its host, the Maghrebian bat Myotis punicus in North Africa and in the islands of Corsica and Sardinia. Mite mitochondrial differentiation among populations was correlated with both host mitochondrial and nuclear differentiation, suggesting spatial co‐differentiation of the lineages of the two interacting species. Therefore our results suggest that parasite dispersal is exclusively mediated by host movements, with open water between landmasses as a main barrier for host and parasite dispersal. Surprisingly the unique presence of a continental European mite lineage in Corsica was inconsistent with host phylogeographical history and strongly suggests the former presence of European mouse‐eared bats on this island. Parasites may thus act as biological tags to reveal the presence of their now locally extinct host.     

Thirsty sea snakes forsake refuge during rainfall

Abstract Vertebrates living in dry or salty habitats spend significant amounts of time and energy maintaining their hydro‐mineral balance; any opportunity to drink fresh water should reduce such expenses. However, to find fresh water, individuals are generally forced to leave the safe and buffered conditions that prevail in their refuges. Therefore, they must face harsh environmental conditions and increased predation risks. We examined this trade‐off in the field, using a very large dataset gathered on amphibious sea snakes that confront periodic droughts. The study was carried out on different islets in the lagoon of New Caledonia. As expected, sea kraits remain concealed most of the time under rocks or within burrows during dry weather. They select precise time windows to move rapidly between the sea and the land, thereby minimizing the time spent in the open. Rainfall triggered massive disruptions of this rhythm: many snakes quickly forsook their shelter to drink in the open, and remained almost motionless until satiety. Interestingly, they adopted specific and unusual postures to drink. They perched on the top of rocks that were benefiting from running and less salty water during downpours. They also drank the droplets attached to vegetation during drizzle.     

Differential responses of plasma membrane aquaporins in mediating water transport of cucumber seedlings under osmotic and salt stresses

It has long been recognized that inhibition of plant water transport by either osmotic stress or salinity is mediated by aquaporins (AQPs), but the function and regulation of AQPs are highly variable among distinct isoforms and across different species. In this study, cucumber seedlings were subjected to polyethylene glycol (PEG) or NaCl stress for duration of 2 h or 24 h. The 2 h treatment with PEG or NaCl had non‐significant effect on the expression of plasma membrane AQP (CsPIPs) in roots, indicating the decrease in hydraulic conductivity of roots (Lp_r) and root cells (Lp_rc) measured in these conditions were due to changes in AQP activity. After both 2 h and 24 h PEG or NaCl exposure, the decrease in hydraulic conductivity of leaves (K_leaf) and leaf cells (Lp_lc) could be attributed to a down‐regulation of the two most highly expressed isoforms, CsPIP1;2 and CsPIP2;4. In roots, both Lp_r and Lp_rc were further reduced after 24 h PEG exposure, but partially recovered after 24 h NaCl treatment, which were consistent with changes in the expression of CsPIP genes. Overall, the results demonstrated differential responses of CsPIPs in mediating water transport of cucumber seedlings, and the regulatory mechanisms differed according to applied stresses, stress durations and specific organs.     

The evolution of redd site selection in brook charr in different environments: same cue,same benefit for fitness*

1. Despite the great interest in characterising fish reproductive habitat, the relationship between the selection of a given spawning site and individual fitness has not been experimentally tested. 2. In this study, we used an in situ experimental approach to determine (i) the relative contribution of substrate characteristics as well as hydrological and physicochemical variables to small‐scale redd site selection by brook charr, Salvelinus fontinalis (Mitchill) and (ii) if hatching and emergence success, used as a surrogate of fitness, are improved in selected compared to non‐selected sites in both lake and stream habitats. 3. Our results show that upwelling groundwater flow was always significantly higher in selected than in non‐selected sites in both lake and stream habitats. We found no significant difference in the mean geometric substrate diameter and no consistent differences in substrate composition between selected and non‐selected sites. Oxygen concentration was higher (significantly so in three of four comparisons) and conductivity tended to be lower in selected than in non‐selected sites, while temperature showed no significant or consistent variations. We found a significant positive relationship between the selection of a given spawning habitat and hatching and emergence success in these systems. 4. These results show that the main cue that brook charr use to select their spawning sites is upwelling groundwater in lake and stream habitats, and that active selection of these sites increases individual fitness. This suggests that natural selection acted on the same cues in lentic and lotic environments; this could have been highly adaptive in a species that used both habitats as colonisation routes after the last glaciation.     

Is nitrogen transfer among plants enhanced by contrasting nutrient‐acquisition strategies?

Nitrogen (N) transfer among plants has been found where at least one plant can fix N₂. In nutrient‐poor soils, where plants with contrasting nutrient‐acquisition strategies (without N₂ fixation) co‐occur, it is unclear if N transfer exists and what promotes it. A novel multi‐species microcosm pot experiment was conducted to quantify N transfer between arbuscular mycorrhizal (AM), ectomycorrhizal (EM), dual AM/EM, and non‐mycorrhizal cluster‐rooted plants in nutrient‐poor soils with mycorrhizal mesh barriers. We foliar‐fed plants with a K¹⁵NO₃ solution to quantify one‐way N transfer from ‘donor’ to ‘receiver’ plants. We also quantified mycorrhizal colonization and root intermingling. Transfer of N between plants with contrasting nutrient‐acquisition strategies occurred at both low and high soil nutrient levels with or without root intermingling. The magnitude of N transfer was relatively high (representing 4% of donor plant N) given the lack of N₂ fixation. Receiver plants forming ectomycorrhizas or cluster roots were more enriched compared with AM‐only plants. We demonstrate N transfer between plants of contrasting nutrient‐acquisition strategies, and a preferential enrichment of cluster‐rooted and EM plants compared with AM plants. Nutrient exchanges among plants are potentially important in promoting plant coexistence in nutrient‐poor soils.     

Effects of an experimental drought and recovery on soil emissions of carbon dioxide,methane, nitrous oxide,and nitric oxide in a moist tropical forest

Changes in precipitation in the Amazon Basin resulting from regional deforestation, global warming, and El Niño events may affect emissions of carbon dioxide (CO₂), methane (CH₄), nitrous oxide (N₂O), and nitric oxide (NO) from soils. Changes in soil emissions of radiatively important gases could have feedback implications for regional and global climate. Here, we report the final results of a 5‐year, large‐scale (1 ha) throughfall exclusion experiment, followed by 1 year of recovery with natural throughfall, conducted in a mature evergreen forest near Santarém, Brazil. The exclusion manipulation lowered annual N₂O emissions in four out of five treatment years (a natural drought year being the exception), and then recovered during the first year after the drought treatment stopped. Similarly, consumption of atmospheric CH₄ increased under drought treatment, except during a natural drought year, and it also recovered to pretreatment values during the first year that natural throughfall was permitted back on the plot. No treatment effect was detected for NO emissions during the first 3 treatment years, but NO emissions increased in the fourth year under the extremely dry conditions of the exclusion plot during a natural drought. Surprisingly, there was no treatment effect on soil CO₂ efflux in any year. The drought treatment provoked significant tree mortality and reduced the allocation of C to stems, but allocation of C to foliage and roots were less affected. Taken together, these results suggest that the dominant effect of throughfall exclusion on soil processes during this 6‐year period was on soil aeration conditions that transiently affected CH₄, N₂O, and NO production and consumption.     

Can we predict nutrient limitation in streams and rivers?

1. Anthropogenic impacts on the biogeochemical cycles of nitrogen (N) and phosphorus (P) affect natural ecosystems worldwide. Modelling is required to predict where and when these key nutrients limit primary production in freshwaters. 2. We reviewed 382 nutrient‐enrichment experiments to examine which factors promote limitation of microphytobenthos biomass by N or P in streams and rivers. Using regression models, we examined whether the response of microphytobenthos biomass to N and P additions could be predicted by the absolute N and P concentrations in the water, the water N:P ratio or a combination of the two. 3. The absolute N concentration in the water was the best predictor of the magnitude of the response of microphytobenthos biomass to N additions. In comparison, the N:P ratio was the best predictor of whether or not N was limiting. However, predictions were uncertain except at extreme N:P ratios <1 : 1 and >100 : 1. 4. The absolute P concentration in the water was the best predictor of the magnitude of the response of microphytobenthos biomass to P additions. Neither the absolute nor the relative N and P concentrations predicted whether or not P was limiting. 5. The absolute and the relative N and P water concentrations contribute significant and complementary insights into the responses of microphytobenthos biomass to nutrient enrichment in running waters. However, ability to predict nutrient limitation from these concentrations is constrained by substantial error in the models. In the future, the prediction ability of models of nutrient limitation might be improved by focussing on regional scales and accounting for additional factors such as light and disturbance.