Permanent Cerebral Vessel Occlusion via Double Ligature and Transection

Stroke is a leading cause of death, disability, and socioeconomic loss worldwide. The majority of all strokes result from an interruption in blood flow (ischemia) ¹. Middle cerebral artery (MCA) delivers a great majority of blood to the lateral surface of the cortex ², is the most common site of human stroke ³, and ischemia within its territory can result in extensive dysfunction or death ^1,4,5. Survivors of ischemic stroke often suffer loss or disruption of motor capabilities, sensory deficits, and infarct. In an effort to capture these key characteristics of stroke, and thereby develop effective treatment, a great deal of emphasis is placed upon animal models of ischemia in MCA.Here we present a method of permanently occluding a cortical surface blood vessel. We will present this method using an example of a relevant vessel occlusion that models the most common type, location, and outcome of human stroke, permanent middle cerebral artery occlusion (pMCAO). In this model, we surgically expose MCA in the adult rat and subsequently occlude via double ligature and transection of the vessel. This pMCAO blocks the proximal cortical branch of MCA, causing ischemia in all of MCA cortical territory, a large portion of the cortex. This method of occlusion can also be used to occlude more distal portions of cortical vessels in order to achieve more focal ischemia targeting a smaller region of cortex. The primary disadvantages of pMCAO are that the surgical procedure is somewhat invasive as a small craniotomy is required to access MCA, though this results in minimal tissue damage. The primary advantages of this model, however, are: the site of occlusion is well defined, the degree of blood flow reduction is consistent, functional and neurological impairment occurs rapidly, infarct size is consistent, and the high rate of survival allows for long-term chronic assessment.     

Mother to Offspring Transmission of Chronic Wasting Disease in Reeves’ Muntjac Deer

The horizontal transmission of prion diseases has been well characterized in bovine spongiform encephalopathy (BSE), chronic wasting disease (CWD) of deer and elk and scrapie of sheep, and has been regarded as the primary mode of transmission. Few studies have monitored the possibility of vertical transmission occurring within an infected mother during pregnancy. To study the potential for and pathway of vertical transmission of CWD in the native cervid species, we used a small cervid model–the polyestrous breeding, indoor maintainable, Reeves’ muntjac deer–and determined that the susceptibility and pathogenesis of CWD in these deer reproduce that in native mule and white-tailed deer. Moreover, we demonstrate here that CWD prions are transmitted from doe to fawn. Maternal CWD infection also appears to result in lower percentage of live birth offspring. In addition, evolving evidence from protein misfolding cyclic amplification (PMCA) assays on fetal tissues suggest that covert prion infection occurs in utero. Overall, our findings demonstrate that transmission of prions from mother to offspring can occur, and may be underestimated for all prion diseases.     

Arabidopsis phosphatidylinositol phosphate kinase 1 binds F-actin and recruits phosphatidylinositol 4-kinase beta1 to the actin cytoskeleton

The actin cytoskeleton can be influenced by phospholipids and lipid-modifying enzymes. In animals the phosphatidylinositol phosphate kinases (PIPKs) are associated with the cytoskeleton through a scaffold of proteins; however, in plants such an interaction was not clear. Our approach was to determine which of the plant PIPKs interact with actin and determine whether the PIPK-actin interaction is direct. Our results indicate that AtPIPK1 interacts directly with actin and that the binding is mediated through a predicted linker region in the lipid kinase. AtPIPK1 also recruits AtPI4Kbeta1 to the cytoskeleton. Recruitment of AtPI4Kbeta1 to F-actin was dependent on the C-terminal catalytic domain of phosphatidylinositol-4-phosphate 5-kinase but did not require the presence of the N-terminal 251 amino acids, which includes 7 putative membrane occupation and recognition nexus motifs. In vivo studies confirm the interaction of plant lipid kinases with the cytoskeleton and suggest a role for actin in targeting PIPKs to the membrane.     

Birthdating studies reshape models for pituitary gland cell specification

The intermediate and anterior lobes of the pituitary gland are derived from an invagination of oral ectoderm that forms Rathke's pouch. During gestation proliferating cells are enriched around the pouch lumen, and they appear to delaminate as they exit the cell cycle and differentiate. During late mouse gestation and the postnatal period, anterior lobe progenitors re-enter the cell cycle and expand the populations of specialized, hormone-producing cells. At birth, all cell types are present, and their localization appears stratified based on cell type. We conducted a birth dating study of Rathke's pouch derivatives to determine whether the location of specialized cells at birth is correlated with the timing of cell cycle exit. We find that all of the anterior lobe cell types initiate differentiation concurrently with a peak between e11.5 and e13.5. Differentiation of intermediate lobe melanotropes is delayed relative to anterior lobe cell types. We discovered that specialized cell types are not grouped together based on birth date and are dispersed throughout the anterior lobe. Thus, the apparent stratification of specialized cells at birth is not correlated with cell cycle exit. Thus, the currently popular model of cell specification, dependent upon timing of extrinsic, directional gradients of signaling molecules, needs revision. We propose that signals intrinsic to Rathke's pouch are necessary for cell specification between e11.5 and e13.5 and that cell–cell communication likely plays an important role in regulating this process.     

Variations in the coordination environment of Co, Cu and Zn complexes prepared from a tridentate (imino)pyridine ligand and their structural comparisons

The variations in the coordination environment of Co(II), Cu(II) and Zn(II) complexes with the neutral, tridentate ligand bis[1-(cyclohexylimino)ethyl]pyridine (BCIP) are reported. Analogous syntheses were carried out utilizing either the M(BF₄)₂ · xH₂O or MCl₂ · xH₂O metal salts (where M = Co(II), Cu(II) or Zn(II)) with one equivalent of BCIP. When the hydrated metal starting material was used, cationic, octahedral complexes of the type [M(BCIP)₂]²⁺ were isolated as the tetrafluoroborate salt (4, 5). Conversely, when the hydrated chloride metal salt was used as the starting material, only neutral, pentacoordinate [M(BCIP)Cl₂] complexes (1-3) formed. All complexes were characterized by X-ray diffraction studies. The three complexes that are five coordinate have distortions due mainly to the pyridine di-imine bite angle. The [Cu(BCIP)Cl₂] (2) also exhibits deviations in the Cu(II)-Cl bond distances with values of 2.4242(9) and 2.2505(9) Å, which are not seen in the analogous Zn(II) and Co(II) structures. Similarly, the two six coordinate complexes (5, 6) are also altered by the ligand frame bite angle giving rise to distorted octahedral geometries in each complex. The [Cu(BCIP)₂](BF₄)₂ (6) also exhibits Cu(II)-N_imine bond lengths that are on average 0.14 Å longer than those found in the analogous 5 coordinate complex, [Cu(BCIP)Cl₂]. In addition to X-ray analysis, all complexes were also characterized by UV/Vis and IR spectroscopy with ¹H NMR spectroscopy being used for the analysis of the Zn(II) analogue (3).     

Development of a pharmacodynamic model of murine malaria and antimalarial treatment with dihydroartemisinin

Antimalarial treatment strategies based on in vitro studies are limited by the paucity of pharmacodynamic information for dosage regimen design. We postulated that a murine model could be used for pre-clinical stages of drug development, especially in dose–response studies and evaluation of combination therapies. Swiss mice infected with Plasmodium berghei parasites (2–5% starting parasitaemia) were given dihydroartemisinin (0–100 mg/kg single dose). Parasite density was regularly determined from thin blood films. A parasite population growth model comprising parasite multiplication, decline in erythrocyte count with increasing parasitaemia and parasite clearance after drug administration was developed. This model described the rise in parasitaemia following inoculation, the nadir following dihydroartemisinin administration, and the subsequent resurgence of parasitaemia (analogous to ‘recrudescence’). At doses of 10, 30 and 100 mg/kg dihydroartemisinin, there was a graded response with 2.5 ± 1, 5 ± 1 and 12 ± 4-fold decreases in parasitaemia, respectively. The nadir parasitaemia (at 21–27 h) was also dose-dependent. This study demonstrates that a murine malaria pharmacodynamic model is a valuable tool for understanding how single drugs and their dosing schedules alter the time course and level of infection.     

GOAT induced ghrelin acylation regulates hedonic feeding

Ghrelin is an orexigenic hormone that regulates homeostatic and reward-related feeding behavior. Recent evidence indicates that acylation of ghrelin by the gut enzyme ghrelin O-acyl transferase (GOAT) is necessary to render ghrelin maximally active within its target tissues. Here we tested the hypothesis that GOAT activity modulates food motivation and food hedonics using behavioral pharmacology and mutant mice deficient for GOAT and the ghrelin receptor (GHSR). We evaluated operant responding following pharmacological administration of acyl-ghrelin and assessed the necessity of endogenous GOAT activity for operant responding in GOAT and GHSR-null mice. Hedonic-based feeding behavior also was examined in GOAT-KO and GHSR-null mice using a “Dessert Effect” protocol in which the intake of a palatable high fat diet “dessert” was assessed in calorically-sated mice. Pharmacological administration of acyl-ghrelin augmented operant responding; notably, this effect was dependent on intact GHSR signaling. GOAT-KO mice displayed attenuated operant responding and decreased hedonic feeding relative to controls. These behavioral results correlated with decreased expression of the orexin-1 receptor in reward-related brain regions in GOAT-KO mice. In summary, the ability of ghrelin to stimulate food motivation is dependent on intact GHSR signaling and modified by endogenous GOAT activity. Furthermore, GOAT activity is required for hedonic feeding behavior, an effect potentially mediated by forebrain orexin signaling. These data highlight the significance of the GOAT–ghrelin system for the mediation of food motivation and hedonic feeding.     

Genetic Variation for the Sex Ratio in NASONIA VITRIPENNIS

We detected genetic variation for the sex ratio in the parasitoid wasp Nasonia vitripennis by analysis of inbred lines and with an artificial selection experiment. Sex ratios differed significantly among five independently isolated lines. Furthermore, sex ratio in broods produced by single females in single hosts shifted from 80-90% female to 50-55% female in 13 to 15 generations in each of two replicate selection lines. The final sex ratios of both selection lines were significantly lower than any of the inbred line sex ratios. Backcrosses revealed that the selection response was due to nuclear genes acting through the female parent. In light of known facultative sex ratio behavior and major genes affecting sex ratio in Nasonia, our results suggest that population and individual sex ratios in this species are molded by processes at both genetic and behavioral levels.     

Precipitation of the warmest quarter and temperature of the warmest month are key to understanding the effect of climate change on plant species diversity in Southern African savannah

In this study, we test for the key bioclimatic variables that significantly explain the current distribution of plant species richness in a southern African ecosystem as a preamble to predicting plant species richness under a changed climate. We used 54,000 records of georeferenced plant species data to calculate species richness and spatially interpolated climate data to derive nineteen bioclimatic variables. Next, we determined the key bioclimatic variables explaining variation in species richness across Zimbabwe using regression analysis. Our results show that two bioclimatic variables, that is, precipitation of the warmest quarter (R² = 0.92, P < 0.001) and temperature of the warmest month (R² = 0.67, P < 0.001) significantly explain variation in plant species richness. In addition, results of bioclimatic modelling using future climate change projections show a reduction in the current bio‐climatically suitable area that supports high plant species richness. However, in high‐altitude areas, plant richness is less sensitive to climate change while low‐altitude areas show high sensitivity. Our results have important implications to biodiversity conservation in areas sensitive to climate change; for example, high‐altitude areas are likely to continue being biodiversity hotspots, as such future conservation efforts should be concentrated in these areas.     

The Role of Estrogen Signaling in a Mouse Model of Inflammatory Bowel Disease: A Helicobacter Hepaticus Model

The pathogenesis of inflammatory bowel diseases (IBD), Crohn''s disease and ulcerative colitis, is due in part to interactions between the immune system, genetics, the environment, and endogenous microbiota. Gonadal sex hormones (GSH), such as estrogen, are thought to be involved in the development of IBD as variations in disease severity occur during pregnancy, menopause, or oral contraceptives use. In certain strains of mice, infection with Helicobacter hepaticus triggers IBD-like mucosal inflammation that is more severe in female mice than in males, suggesting a role for GSH in this model. To determine the role of estrogen signaling in microbiota-induced intestinal inflammation, estrogen receptor (ER) α and β knock-out (KO) mice, ER agonists, and adoptive transfers were utilized. We demonstrate that, when signaling is limited to ERβ on a non-CD4⁺ cell subset, disease is less severe and this correlates with decreased expression of pro-inflammatory mediators.