The role of adenosine monophosphate activated protein kinase in luteal progesterone production

Prostaglandin F2 alpha (PGF2&agr;) has in vivo luteolytic actions on the bovine corpus luteum (CL). Although PGF2&agr; is utilized extensively in livestock to synchronize estrus, its actions in vitro are controversial and do not always result in a reduction in progesterone (P4) secretion. The mechanism of action of PGF2&agr; is thought to involve activation of the phospholipase C-Ca2+ pathway. The luteolytic actions of PGF2&agr; are mediated through an elevation of cytosolic Ca2+ involving activation of protein kinase C (PKC). Expression of PKC isozymes and calcium/calmodulin-dependent protein kinase kinase 2 (CAMKK2) change as the CL develops to a mature stage. Either CAMKK2 directly affects steroidogenesis or works indirectly through an intermediary step, such as adenosine- monophosphate kinase (AMPK), which is activated through CAMKK2 targeted phosphorylation. Specifically, the hypothesis that activation of PGF2&agr; receptor (FP) in the mature CL involves AMPK was examined. Expression of mRNA encoding a potential target AMPK for the action of CAMKK2, was increased in the mature versus developing CL. Furthermore, activation of FP induced rapid phosphorylation of AMPK. An AMPK-specific inhibitor, dorsomorphin dichloride (DM), eliminated effects of PGF2&agr; on secretion of P4, supporting the hypothesis that activation of the FP receptor in mature CL involves participation of AMPK. Furthermore, in vitro two AMPK activators, 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR) and metformin, decreased basal P4 secretion in mature but not in developing CL. Effect of FP activation on cholesterol transport in the mature CL was investigated. Both serum and luteal P4 declined as soon as 2 hours after PGF 2&agr; administration, which was paralleled by a decrease in protein concentration of low density lipoprotein receptor (LDLR). Human mural granulosal cells are collected routinely when a patient is undergoing in vitro fertilization (IVF) and are useful for studying granulosal lutein cells of the CL. Altering intracellular calcium in these cells through an ionophore, A23187, increased basal P4 production. However, at high concentrations a calcium chelator, BAPTA, decreased P4 from mural granulosal cells. Infertility is a problem that affects more than 6.1 million women and their partners across the United States. Causes of infertility range from female related tubal factors such as blockage, endometriosis, diminished ovarian reserve, to male related infertility due to abnormal sperm motility or morphology to unexplained infertility. Primary infertility types, age, BMI, and pregnancy outcome influenced ability of granulosal cells to increase P4 production when stimulated with human chorionic gonadotropin (hCG). AICAR decreased basal P4 production in these cells, which is similar to earlier observations in the mature bovine CL. Thus, AMPK appears to be a distal target in the pathway responsible for mediating the actions of PGF2&agr;. Directly activating AMPK could lead to better estrous cycle manipulation in the cow, or overcoming luteal insufficiency in women undergoing IVF

The host immune response to gastrointestinal nematode infection in sheep

non peer reviewedGastrointestinal nematode infection represents a major threat to the health, welfare and productivity of sheep populations worldwide. Infected lambs have a reduced ability to absorb nutrients from the gastrointestinal tract, resulting in morbidity and occasional mortality. The current chemo-dominant approach to nematode control is considered unsustainable due to the increasing incidence of anthelmintic resistance. In addition there is growing consumer demand for food products from animals not subjected to chemical treatment. Future mechanisms of nematode control must rely on alternative, sustainable strategies such as vaccination or selective breeding of resistant animals. Such strategies take advantage of the host's natural immune response to nematodes. The ability to resist gastrointestinal nematode infection is considered to be dependent on the development of a protective acquired immune response; although the precise immune mechanisms involved in initiating this process remain to be fully elucidated. In this paper current knowledge on the innate and acquired host immune response to gastrointestinal nematode infection in sheep and the development of immunity is reviewed.We gratefully acknowledge funding support for the research in our laboratories from the Teagasc Walsh Fellowship Programme, the Allan and Grace Kay Overseas Scholarship and the EC-funded FP7 Programme. We also thank the BBSRC Animal Health Research Club for funding part of this research (grant BB/l004070/1

Outdoor adventure education in schools: curriculum or pedagogy? Considerations for teacher preparation and program implementation

Although outdoor education has a rich history of providing key transformative moments and personal growth for students, its incorporation into public school education has been weak at best. With such an established role of providing effective learning environments, this naturally raises the question: What has prevented the greater inclusion of outdoor education in our public school system? This research addresses this question by demonstrating that the role of outdoor education in public schools can be framed as either pedagogy or curriculum. The relationship between pedagogy and curriculum is revealed to create difficulties for outdoor education’s incorporation into schools because the public education system has the ability to use it selectively in a piecemeal fashion. This research demonstrates that the current lack of articulation of this duality has enabled those opposed to its inclusion in schools to create false arguments against its use, while those in favor of incorporation face challenges by not understanding outdoor education in this way. The importance of framing outdoor education’s role for schools in this manner is shown to affect two areas: developing integrated school programs and its incorporation into teacher training programs. In order to adequately explore the above problem this dissertation divides and separately considers three frames of reference for outdoor education programming relative to schools: supplementary, curricula-based, and integrated. Curricula-based outdoor programs are shown to emphasize outdoor education as curriculum for schools (as a body of knowledge), while integrated outdoor programs largely focus on outdoor education as pedagogy (as experiential education). Primary research investigated Canadian integrated outdoor programs through surveys of veteran outdoor education teachers operating such programs in our schools. This research identified two key roles for outdoor program inclusion: an experiential learning framework, and personal life skills development. By redefining the roles of outdoor education in schools in this way we now understand that previous reference to education about the natural heritage refers to curriculum initiatives while education through the natural heritage refers to pedagogical aims. In addition, the roles of the institution and the educator are examined in relation to the compatibility of outdoor education in public schools

The Association between Microsatellite Marker CSRD 2138 and Parasite Resistance in Katahdin Hair Sheep

Gastrointestinal nematode infection in sheep is currently considered to be a problem of great magnitude in sheep production worldwide. This problem has been compounded by the overuse of anthelmintics used to treat parasite infection. In efforts to reduce chemical dependant production, research has focused on selection for parasite resistance based on worm load. Due to the low heritability of this trait any change effected by selection would take many years to realize. Selection for resistance can be improved by using more specific traits acted upon by fewer genes. Markers that are linked to the immune response to a parasite infection are great candidate genes to evaluate use in a genetic marker selection program. For the past five years, the Northeast Katahdin Hair Sheep Project has been C breeding Katahdins for improved parasite resistance. Lambs from the hair sheep project were used in this study, all of which were 3/4 Katahdin (n=28). To evaluate resistance, lambs were naturally and artificially infected with the nematode Haemonchus contortus in a process called priming. Data from the initial artificial infection were used to determine levels of resistance based upon fecal egg count (FEC) and packed cell volume (PCV). Lambs under artificial infection showed a weak infection and there was no difference in FEC or PCV between the lambs (p \u3e 0.05) and were all considered to be resistant. Lambs were treated with levamisole (8mgIkg) and held kept free of parasite infection for fourteen days post anthelmintic treatment. Lambs were then divided into control and treatment groups, where they received an oral dose of PBS or 10,000 L3 Haemonchus contortus larvae, respectively. Lambs were tested weekly post infection for 6 weeks. On the third week p.i. the lambs showed symptoms of parasite infection. Fecal egg count peaked on 28d p.i. at 4,168 epg and began to recede each following week to the end of the study where the mean FEC was 3,021 epg. The control lambs, grown in the same location, did not become infected. Resistance was determined on the mean fecal egg count, packed cell volume and FAMACHA anemia scores and lambs were grouped into low and high resistance groups. The low resistance group differed from the high group in FEC, PCV, and FAMACHA (p C0.05). There was no difference found in the DNA sequences of the 27 lambs tested as they all had the same allele of CSRD 2138. It is unlikely that a different allele exists in the local population as it is inherited codominantly. This candidate marker needs more research with breeds susceptible to Haemonchus contortus. The response from the lambs in this study shows that breeding for resistance can be improved faster by crossbreeding with breeds that show more parasite resistance

The Association between Microsatellite Marker CSRD 2138 and Parasite Resistance in Katahdin Hair Sheep

Gastrointestinal nematode infection in sheep is currently considered to be a problem of great magnitude in sheep production worldwide. This problem has been compounded by the overuse of anthelmintics used to treat parasite infection. In efforts to reduce chemical dependant production, research has focused on selection for parasite resistance based on worm load. Due to the low heritability of this trait any change effected by selection would take many years to realize. Selection for resistance can be improved by using more specific traits acted upon by fewer genes. Markers that are linked to the immune response to a parasite infection are great candidate genes to evaluate use in a genetic marker selection program. For the past five years, the Northeast Katahdin Hair Sheep Project has been C breeding Katahdins for improved parasite resistance. Lambs from the hair sheep project were used in this study, all of which were 3/4 Katahdin (n=28). To evaluate resistance, lambs were naturally and artificially infected with the nematode Haemonchus contortus in a process called priming. Data from the initial artificial infection were used to determine levels of resistance based upon fecal egg count (FEC) and packed cell volume (PCV). Lambs under artificial infection showed a weak infection and there was no difference in FEC or PCV between the lambs (p \u3e 0.05) and were all considered to be resistant. Lambs were treated with levamisole (8mgIkg) and held kept free of parasite infection for fourteen days post anthelmintic treatment. Lambs were then divided into control and treatment groups, where they received an oral dose of PBS or 10,000 L3 Haemonchus contortus larvae, respectively. Lambs were tested weekly post infection for 6 weeks. On the third week p.i. the lambs showed symptoms of parasite infection. Fecal egg count peaked on 28d p.i. at 4,168 epg and began to recede each following week to the end of the study where the mean FEC was 3,021 epg. The control lambs, grown in the same location, did not become infected. Resistance was determined on the mean fecal egg count, packed cell volume and FAMACHA anemia scores and lambs were grouped into low and high resistance groups. The low resistance group differed from the high group in FEC, PCV, and FAMACHA (p C0.05). There was no difference found in the DNA sequences of the 27 lambs tested as they all had the same allele of CSRD 2138. It is unlikely that a different allele exists in the local population as it is inherited codominantly. This candidate marker needs more research with breeds susceptible to Haemonchus contortus. The response from the lambs in this study shows that breeding for resistance can be improved faster by crossbreeding with breeds that show more parasite resistance

Niobium phosphine macrocyclic complexes

The chemistry of the macrocycle PhP(CH₂SiMe₂NSiMe₂CH₂)₂PPh (abbreviated as [P₂N₂] within this thesis) with niobium(III) is explored. The paramagnetic chloride compound NbCl[P₂N₂] (1) is used as a starting material for other derivatives, such as {[P₂N₂]Nb}₂ (µ-N₂) (2) , Nb(CH₂SiMe₃)[P₂N₂] (6), and Nb(CH₂Ph)[P₂N₂ ] (9). Compound 1 forms adducts with small molecules and solvents, including CO which causes the complex to become diamagnetic. NbCl[P₂N₂] (1) reacts with KC₈ under 1 atm of N₂ to give {[P₂N₂]Nb}₂(µ-N₂) (2). Complex 2, characterized by X-ray crystallography, is found to have an end-on bound dinitrogen ligand with an N-N bond length of 1.272 (5) A . This is only the fourth niobium dinitrogen species to be structurally characterized. Reaction of 2 with 2 equivalents of Me₃N-HCl gives the original chloride 1 and 0.31 equivalents of hydrazine. Nb(CH₂SiMe₃)[P₂N₂] (6) and Nb(CH₂Ph)[P₂N₂] (9) are synthesized by reaction of chloride 1 with LiCH₂SiMe₃ and KCH₂Ph respectively. Reaction of trimethylsilylmethyl 6 with H₂ in benzene results in the formation of a cyclohexadienyl ligand complex, Nb(ƞ⁵-C₆H₇)[P₂N₂] (8). Also, reaction of compound 6 with CO results in the CO insertion and rearrangement with the trimethylsilylmethyl ligand to give an enolate complex. Compound 9 is determined to have an ƞ⁵ bound benzyl ligand. Preliminary investigations in the reaction of compound 1 with LiN(SiMe₃)₂, Mel, and KC₈ while under Ar give Nb{N(SiMe₃)₂}[P₂N₂] (4), Nb(MeI)Cl[P₂N₂] (10), and {[P₂N₂]Nb}₂ (3) respectively. Molecular structure information is given for {[P₂N₂]Nb}₂(µ-N₂)(µ-Cl) (5); the first characterized side-on bridged dinitrogen niobium complex. Studies into the elemental determinations of {[P₂N₂]Nb(µ-N₂) (2) and Nb(CH₂Ph)[P₂N₂] (9) reveal limitations in microanalysis due to the formation of stable niobium nitrides and carbides. ¹H NMR spectroscopy is successfully used for identification of all paramagnetic complexes within this thesis.Science, Faculty ofChemistry, Department ofGraduat