Avidin- or streptavidin-biotin as a highly sensitive method to stain total protein on membranes

A sensitive method for staining proteins after transfer from polyacrylamide gels to nitrocellulose paper is described. Transferred proteins are first derivatized by reaction of the nitrocellulose replica with sulfosuccinimidobiotin and are then reacted sequentially with streptavidin, rabbit anti-streptavidin, and horseradish peroxidase-conjugated goat anti-rabbit IgG antibody. Incubation with the enzyme substrate α-chloronaphthol, produces dark protein bands against a white background. The binding of streptavidin to the proteins is dependent on biotin derivatization as demonstrated by competition with biotinylated bovine serum albumin or 10 nM biotin. The procedure detects less than 5ng of transferred protein in a single band and is thus 5–10 times more sensitive than horseradish peroxidase-conjugated avidin alone. For bovine serum albumin, the method is comparable in sensitivity to silver staining of protein in polyacrylamide gels.     

A new family of H3 receptor antagonists based on the natural product Conessine

A new family of Histamine H(3) receptor antagonists (5a-t) has been prepared based on the structure of the natural product Conessine, a known H(3) antagonist. Several members of the new series are highly potent and selective binders of rat and human H(3) receptors and display inverse agonism at the human H(3) receptor. Compound 5n exhibited promising rat pharmacokinetic properties and demonstrated functional antagonism of the H(3) receptor in an in-vivo pharmacological model.     

Enteral arginine modulates inhibition of AP-1/c-Jun by SP600125 in the postischemic gut

We previously demonstrated that enteral arginine increased c-Jun/activator protein-1 (AP-1) DNA-binding activity and iNOS expression in a rodent model of mesenteric ischemia/reperfusion (I/R). The objective of this study was to specifically investigate the role of AP-1 in arginine's deleterious effect on the postischemic gut. We hypothesized that AP-1 inhibition would mitigate the effects of arginine. Using a rodent model of mesenteric I/R we demonstrated that gut neutrophil infiltration, activity of c-Jun/AP-1, as well as iNOS expression were increased by I/R and further increased by arginine while lessened by inhibition of c-Jun using the pharmacologic c-Jun N-terminal kinase inhibitor, SP600125. Similar results were demonstrated using a cell culture model of oxidant stress in IEC-6 cells. Importantly, effects of SP600125 were comparable to those of c-Jun silencing. Lastly, the specific iNOS inhibitor, 1400W, had no effect on either AP-1 or c-Jun. In conclusion, SP600125 attenuated the activity of c-Jun/AP-1, iNOS expression, and neutrophil infiltration induced by arginine following mesenteric I/R. Our data suggest that AP-1 inhibition mitigates the injurious inflammatory effects of arginine in the postischemic gut. Further investigation into the pathologic role of enteral arginine in the postischemic gut is warranted.     

Increasing variance in North Pacific climate relates to unprecedented ecosystem variability off California

Changes in variance are infrequently examined in climate change ecology. We tested the hypothesis that recent high variability in demographic attributes of salmon and seabirds off California is related to increasing variability in remote, large‐scale forcing in the North Pacific operating through changes in local food webs. Linear, indirect numerical responses between krill (primarily Thysanoessa spinifera) and juvenile rockfish abundance (catch per unit effort (CPUE)) explained >80% of the recent variability in the demography of these pelagic predators. We found no relationships between krill and regional upwelling, though a strong connection to the North Pacific Gyre Oscillation (NPGO) index was established. Variance in NPGO and related central Pacific warming index increased after 1985, whereas variance in the canonical ENSO and Pacific Decadal Oscillation did not change. Anthropogenic global warming or natural climate variability may explain recent intensification of the NPGO and its increasing ecological significance. Assessing non‐stationarity in atmospheric‐environmental interactions and placing greater emphasis on documenting changes in variance of bio‐physical systems will enable insight into complex climate‐marine ecosystem dynamics.     

Seabird species assemblages reflect hydrographic and biogeographic zones within Drake Passage

Polar Biology - Drake Passage, extending from the southern tip of South America to the northern Antarctic Peninsula, is a dynamic oceanographic region with well-defined habitats delineated by the...     

Essential krill species habitat resolved by seasonal upwelling and ocean circulation models within the large marine ecosystem of the California Current System

Due to their global distribution, high biomass and energy content, euphausiids (krill) are important prey for many mid and upper trophic level marine organisms. Understanding drivers of krill habitat is essential for forecasting range shifts, and to better understand the response of krill predators to climate change. We hypothesized that the distribution and abundance of krill species derived from ecosystem surveys in spring/summer relates to geomorphic features, coastal upwelling during the preceding winter, and spring mesoscale oceanographic conditions. To test this hypothesis, we used boosted regression trees with environmental data and ocean model output to quantify the habitat associations of two primary krill species (Euphausia pacifica and Thysanoessa spinifera) in the central California Current Ecosystem from 2002 to 2018. Models confirmed the neritic distribution of T. spinifera and pelagic, outer slope association of E. pacifica (deviance explained ~35%). Distribution of these species were influenced by depth and bottom rugosity; chlorophyll-a concentrations and increased winter upwelling conditions; and spring surface currents and wind stress. Thysanoessa spinifera and E. pacifica abundance responded negatively (positively) to warm (cold) climate events, confirming known relationships. As an independent evaluation of krill models, observations of krill predator (seabirds, marine mammals) distribution indicated they were present within habitats of predicted high krill species abundance. Our framework indicates species-specific habitat relationships for these foundational forage species and their negative response to large-scale climate variations, such as El Niño and marine heatwave conditions. The approach can be easily transferred to other ecosystems or krill species that respond to similar ocean and climate forcing.     

Environmental determinants of top predator distribution within the dynamic winter pack ice zone of the northern Antarctic Peninsula

Global warming is predicted to reduce the amount of sea ice concentration in polar environments, thus presenting profound changes for populations of seabirds and marine mammals dependent on sea ice. Using data from a shipboard survey during August 2012, I test the hypothesis that relative abundance of seabird and marine mammals reflects environmental variability associated with the dynamic pack ice zone. Using environmental data and observations of sea ice concentration, I quantified an environmental gradient that describes the spatial organization of the dynamic pack ice zone. The relationship of top predators to this environmental gradient revealed three important aspects: (1) an open water and pack ice community is present with some top predator species exhibiting higher abundance associated with moderate sea ice concentration (40–60 %) as opposed to the pack ice edge (10 %), (2) Antarctic fur seals (Arctocephalus gazella) were the most abundant pinniped and they were observed resting on ice floes and foraging within leads and polynyas, and (3) for the most abundant species, spatial regression models indicate that latitude and sea ice concentration (a principal north/south gradient) are the most important environmental determinants. Winter ocean conditions may strongly influence population dynamics of top predators; therefore, information regarding their habitat use during winter is needed for understanding ecosystem dynamics.     

Developmental control of 2-aminoisobutyric acid transport by 7-and 14-day chick heart cell aggregates. Roles of insulin and amino acids.

Cell aggregates cultured from 7-day embryonic avian heart showed a spontaneous increase in A-system 2-aminoisobutyric acid transport when placed in protein-free and amino acid-free buffer for 3 hr. The apparent V_max increased from 4.0 to 9.9 nmoles/μl of intracellular fluid volume/10 min in 3 hr. l-Proline (5 mM), an amino acid transported primarily by the A system, prevented this rise, but l-phenylalanine, primarily an L-system substrate, had no effect. Actinomycin, puromycin, and cycloheximide (55 μM) also prevented the time-dependent increase in transport. In contrast, cell aggregates cultured from 14-day embryonic heart exhibited a decrease in apparent V_max during the 3-hr incubation, from 8.3 to 3.3 nmoles/μl of intracellular volume/10 min. l-Proline, but not l-phenylalanine, enhanced this decrease in A-system transport. The percentage proline inhibition of transport was reduced by actinomycin or cycloheximide (55 μM) at both ages. Insulin stimulated A-system transport at identical half-maximal concentrations of 18 nM at 7 and 14 days of embryonic development. In the presence of cycloheximide at 7 days of age, insulin prolonged the half-life of transport activity twofold. However, at 14 days, cycloheximide reduced the insulin response by 88% [Elsas, L. J., Wheeler, F. B., Danner, D. J., and DeHaan, R. L. (1975). J. Biol. Chem.250, 9381–9390]. l-Proline or actinomycin reduced both basal and insulin-stimulated transport by 7-day cell aggregates, but neither reduced the percentage insulin stimulation. We conclude that inherent developmental control(s), A-system amino acids, and insulin regulated the maximal velocity of A-system transport by controlling the biological turnover of transport protein(s). l-Proline decreased the existing synthesis of transport protein(s) at both ages. The predominant effect of insulin shifted from a posttranslational level at 7 days to a synthetic level by 14 days of embryonic development. Seven-day cell aggregates spontaneously increased synthesis in the absence of A-system amino acids, but 14-day cell aggregates required hormonal stimulation to shift the balance from degradation to synthesis of transport protein(s).     

Identification of biaryl sulfone derivatives as antagonists of the histamine H₃ receptor: discovery of (R)-1-(2-(4'-(3-methoxypropylsulfonyl)biphenyl-4-yl)ethyl)-2-methylpyrrolidine (APD916)

The design of a new clinical candidate histamine-H(3) receptor antagonist for the potential treatment of excessive daytime sleepiness (EDS) is described. Phenethyl-R-2-methylpyrrolidine containing biphenylsulfonamide compounds were modified by replacement of the sulfonamide linkage with a sulfone. One compound from this series, 2j (APD916) increased wakefulness in rodents as measured by polysomnography with a duration of effect consistent with its pharmacokinetic properties. The identification of a suitable salt form of 2j allowed it to be selected for further development.