Inhibition of vascular smooth muscle G protein-coupled receptor kinase 2 enhances alpha1D-adrenergic receptor constriction

G protein-coupled receptor kinase 2 (GRK2) is a serine/theorinine kinase that phosphorylates and desensitizes agonist-bound G protein-coupled receptors. GRK2 is increased in expression and activity in lymphocytes and vascular smooth muscle (VSM) in human hypertension and animal models of the disease. Inhibition of GRK2 using the carboxyl-terminal portion of the protein (GRK2ct) has been an effective tool to restore compromised beta-adrenergic receptor (AR) function in heart failure and improve outcome. A well-characterized dysfunction in hypertension is attenuation of betaAR-mediated vasodilation. Therefore, we tested the role of inhibition of GRK2 using GRK2ct or VSM-selective GRK2 gene ablation in a renal artery stenosis model of elevated blood pressure (BP) [the two-kidney, one-clip (2K1C) model]. Use of the 2K1C model resulted in a 30% increase in conscious BP, a threefold increase in plasma norepinephrine levels, and a 50% increase in VSM GRK2 mRNA levels. BP remained increased despite VSM-specific GRK2 inhibition by either GRK2 knockout (GRK2KO) or peptide inhibition (GRK2ct). Although betaAR-mediated dilation in vivo and in situ was enhanced, alpha(1)AR-mediated vasoconstriction was also increased. Further pharmacological experiments using alpha(1)AR antagonists revealed that GRK2 inhibition of expression (GRK2KO) or activity (GRK2ct) enhanced alpha(1D)AR vasoconstriction. This is the first study to suggest that VSM alpha(1D)ARs are a GRK2 substrate in vivo.     

Characterization of the indigenous PAH-degrading bacteria of <Emphasis Type="Italic">Spartina</Emphasis> dominated salt marshes in the New York/New Jersey Harbor

The aerobic polyaromatic hydrocarbon (PAH) degrading microbial communities of two petroleum-impacted Spartina-dominated salt marshes in the New York/New Jersey Harbor were examined using a combination of microbiological, molecular and chemical techniques. Microbial isolation studies resulted in the identification of 48 aromatic hydrocarbon-degrading bacterial strains from both vegetated and non-vegetated marsh sediments. The majority of the isolates were from the genera Paenibacillus and Pseudomonas. Radiotracer studies using ¹⁴C-phenanthrene and ¹⁴C-pyrene were used to measure the PAH-mineralization activity in salt marsh sediments. The results suggested a trend towards increased PAH mineralization in vegetated sediments relative to non-vegetated sediments. This trend was supported by the enumeration of PAH-degrading bacteria in non-vegetated and vegetated sediment using a Most Probable Numbers (MPN) technique, which demonstrated that PAH-degrading bacteria existed in non-vegetated and vegetated sediments at levels ranging from 10² to 10⁵ cells/g sediment respectively. No difference between microbial communities present in vegetated versus non-vegetated sediments was found using terminal restriction fragment length polymorphism (of the 16S rRNA gene) or phospholipid fatty acid analysis. These studies provide information on the specific members and activity of the PAH-degrading aerobic bacterial communities present in Spartina-dominated salt marshes in the New York/New Jersey Harbor estuary.     

Targeting erythroblast-specific apoptosis in experimental anemia

Erythrocyte production is regulated by balancing precursor cell apoptosis and survival signaling. Previously, we found that BH3-only proapoptotic factor, Nix, opposed erythroblast-survival signaling by erythropoietin-induced Bcl-xl during normal erythrocyte formation. Since erythropoietin treatment of human anemia has limitations, we explored the therapeutic potential of abrogating Nix-mediated erythroblast apoptosis to enhance erythrocyte production. Nix gene ablation blunted the phenylhydrazine-induced fall in blood count, enhanced hematocrit recovery, and reduced erythroblast apoptosis, despite lower endogenous erythropoietin levels. Similar to erythropoietin, Nix ablation increased early splenic erythroblasts and circulating reticulocytes, while maintaining a pool of mature erythroblasts as erythropoietic reserve. Erythrocytes in Nix-deficient mice showed morphological abnormalities, suggesting that apoptosis during erythropoiesis not only controls red blood cell number, but also serves a “triage” function, preferentially eliminating abnormal erythrocytes. These results support the concept of targeting erythroblast apoptosis to maximize erythrocyte production in acute anemia, which may be of value in erythropoietin resistance. Abhinav Diwan and Andrew G. Koesters contributed equally to this work.     

Induced emissions of apple fruit volatiles by the codling moth: changing patterns with different time periods after infestation and different larval instars

The changes in the emission of volatiles from mature apple fruits in response to larval feeding by the codling moth (Cydia pomonella) under laboratory conditions are reported. A time course experiment investigated the emission of volatiles throughout the period of larval development following infestation. The volatiles consisted mainly of esters, a few aldehydes, and the terpene alpha-farnesene. Infested apples emitted the same compounds as healthy apples. The quantities of volatiles released were much higher for infested as compared to healthy fruits for an initial three day period. Following this period there was a decrease in volatile emissions (days 6-9), eventually declining back to the levels emitted from healthy apples or below by 9-21 days after infestation. In a separate experiment, the volatile emissions from healthy and artificially damaged fruits were compared to those from herbivore damaged fruits for each of the five larval instars of C. pommonella. The results from the discriminant analysis indicate that the most effective induction of volatiles occurred when fruits were infested with first instar larvae. Induction by first instar larvae was generally higher than after infestation by later instars, and for most compounds it also exceeded the emission from artificially damaged fruits.     

Selection on olfactory response to semiochemicals from a plant-host complex in a parasitic wasp

Parasitic wasps (parasitoids) use volatiles from the plants infested by phytophagous insects to locate host herbivores, but their behavioural response to such semiochemicals is highly variable. Bi-directional selection on Cotesia glomerata (Hymenoptera: Braconidae) was conducted to investigate the importance of genetic variation in the olfactory response of parasitoids. Female wasps were assessed for flight orientation and landing success in response to the hexane extract from a plant-host complex in a wind tunnel. After the first generation of selection, two strains significantly differentiated in both flight orientation and landing success, and their divergence continued with further selection. The two selected strains genetically differentiated in olfactory perception rather than upwind flight ability. The realized heritability was estimated as 0.248 for flight orientation and 0.216 for landing success. The selection experiment further demonstrated that a prior exposure to the semiochemicals significantly enhanced the subsequent response of female wasps, independent of genetic differences. These results suggest that both genetic component and environmental conditioning have played an important role in the evolution of host selection and utilization by the parasitoid in a tritrophic system.     

GENETICS OF BRASSICA CAMPESTRIS. 1. GENETIC CONSTRAINTS ON EVOLUTION OF LIFE-HISTORY CHARACTERS

Energy allocation arguments suggest a possible tradeoff between timing and magnitude of reproduction: plants that postpone reproduction may accumulate greater resources and consequently produce more offspring. However, early reproduction may be favored when adult mortality is high. Tradeoffs among life-history characters may be a consequence of constraints imposed by genetic and environmental covariation among traits. In this paper we examine the genetic basis of the relationship between timing and magnitude of reproduction in an annual plant, Brassica campestris, by selecting to change flowering date and plant size in each of four directions (early and large, late and large, early and small, or late and small). There is a strong positive relationship between flowering date and flowering height. The response to selection was greatest along the axis of positive genetic covariation. Populations may evolve to become early flowering and small or late flowering and tall, but there is little response for the alternative combinations of characters. In this instance, the constraints imposed by quantitative genetics are in striking accord with predictions that might be made on physiological, energetic, or ecological grounds.     

Environmental and genetic influences on the germination of Arabidopsis thaliana in the field

Seasonal germination timing strongly influences lifetime fitness and can affect the ability of plant populations to colonize and persist in new environments. To quantify the influence of seasonal environmental factors on germination and to test whether pleiotropy or close linkage are significant constraints on the evolution of germination in different seasonal conditions, we dispersed novel recombinant genotypes of Arabidopsis thaliana into two geographic locations. To decouple the photoperiod during seed maturation from the postdispersal season that maternal photoperiod predicts, replicates of recombinant inbred lines were grown under short days and long days under controlled conditions, and their seeds were dispersed during June in Kentucky (KY) and during June and November in Rhode Island (RI). We found that postdispersal seasonal conditions influenced germination more strongly than did the photoperiod during seed maturation. Genetic variation was detected for germination responses to all environmental factors. Transgressive segregation created novel germination phenotypes, revealing a potential contribution of hybridization of ecotypes to the evolution of germination. A genetic trade-off in germination percentage across sites indicated that determinants of fitness at or before the germination stage may constrain the geographic range that a given genotype can inhabit. However, germination timing exhibited only weak pleiotropy across treatments, suggesting that different sets of genes contribute to variation in germination behavior in different seasonal conditions and geographic locations. Thus, the genetic potential exists for rapid evolution of appropriate germination responses in novel environments, facilitating colonization across a broad geographic range.     

The serosa of Manduca sexta (Insecta, Lepidoptera): ontogeny, secretory activity, structural changes, and functional considerations

In Manduca sexta, the blastoderm forms successively and becomes immediately cellularized as the cleavage energids reach the surface of the oocyte. Presumptive serosal cells are large and contain 2 or 4 large polyploid nuclei; presumptive embryonic cells are small and mononuclear. All parts of the blastoderm participate in the uptake and digestion of yolk material. About 10 h post-oviposition, the blastoderm breaks at the amnioserosal fold and the extraembryonic part closes above the germ band and constitutes the serosa (12 h post-oviposition, i.e. 10% development completed). At once, the serosa starts to secrete a cuticle consisting of an epi- and a lamellated endocuticle. Detachment of the serosal cuticle, 22h post-oviposition, is reminiscent of apolysis of larval cuticle. Thereafter, the serosa deposits a membranous structure, the serosal membrane. The sercretory process lasts from 23h to 44h post-oviposition. At first a fine granular layer, then an amorphous, spongy-like, fibrillar layer is secreted via microvilli. This persisting membrane is tough, rubbery and very elastic. It may serve to bolster the serosa during katatrepsis (48h post-oviposition) and later embryonic movements. After detachment of the serosal membrane, 44h post-oviposition, a distinct subcellular reorganization of the serosa takes place. The nuclei become still larger and more irregular. Uptake of yolk granules, but not of lipid droplets, ceases, although interaction of serosa and yolk cells are intense. Serosal cells include many mitochondria, large areas of rER, besides some sER, increasing amounts of lysosomal bodies and prominent Golgi complexes. Most conspicuous is the assembly of spindle-shaped, electron-lucent vesicles below the apical surface. These vesicles may contain metabolic products which are released into the peripheral space. The studies show that the serosa assumes changing functions during embryogenesis: digestion of yolk substances, synthesis of a serosal cuticle and a serosal membrane, which may have a protective function, and excretion.     

New racing equation for championship performance

A separate investigation of swimming endurance has led to a family of expressions relating maximal human locomotor activity to time and/or distance having such wide application to medicine, physiology, and sports as to merit independent publication. Specifically, variations of a single racing equation with one variable parameter suffice to predict record times, speeds, energy, power, and endurance accurately for cycling, running, and swimming, provided only that performance be categorized as steady-state, aerobic effort.