Utveckling av metod för utvärdering av synnervens funktion hos häst

VEP, visual evoked potential is an electrophysiologic test that can be used for evaluating the higher visual pathways. The aim of this study was to investigate if it is possible to adapt this method to horses. This method could have a clinical use in addition to the existing diagnostic methods that are available to evaluate vision in horses. Firstly, this study contains a summary of relevant studies done on animals and, secondly, an experimental part where the possibility of recording VEP in nonsedated and sedated horses is investigated and where to position the recording electrode to obtain the best results. Our results show that it is possible to record VEP in horses in a clinical setting. VEP was best recorded in the sedated horse to avoid movement artifacts and other disturbances. The best signal was obtained when the active electrode was positioned on top of the nuchal crest. When the active electrodes were positioned lateral to the midline, isolation of signals from the right and left hemispheres was possible. The electrodes were easy to apply, but the impedance had to be checked to evaluate the connection between the electrodes and the skin. Akinesia of the eyelids had to be induced to reduce blink artifacts. Because this study just contains results from VEP-recordings from two horses, further studies are needed that will provide normative data and assessment of its clinical use.Visual evoked potentials, VEP, är en elektrofysiologisk undersökningsmetod som kan användas för att objektivt utvärdera de högre synvägarnas funktion. Syftet med denna studie var att undersöka om det var möjligt att anpassa denna metod för häst. Metoden skulle kunna vara ett viktigt komplement till de diagnostiska hjälpmedel som finns idag för att utvärdera hästens synförmåga och även följa upp behandlingar och ställa prognos. Studien består dels av en litteraturstudie, där en sammanställning över relevanta studier på ett flertal djurslag är gjord, samt dels en experimentell studie, där möjligheten att registrera VEP på osederad respektive sederad häst undersökts, samt även vilken elektrodplacering som är optimal. Resultaten visar att det är möjligt att registrera VEP på häst utan större besvär under kliniska förhållanden. Bäst registrering fås vid sedering av hästen eftersom man annars får mycket störningar. Optimal placering av aktiv elektrod är mitt på crista nuchae. En referenselektrod placerad kaudalt om ögonen minskar sannolikt ERG-kontamination. Placering av aktiva elektroder lateralt på crista nuchae möjliggör viss separation av signaler från vänster och höger synkortex. Kutana knappelektroder är lätta att applicera, men impedansen måste kontrolleras för att se till att det är bra kontakt mellan elektroder och hud. Akinesi av ögonlocken krävs för att reducera blinkartefakterna. Eftersom det är så litet försöksmaterial i detta examensarbete bör ytterligare studier göras för att kunna dra mer långtgående slutsatser, dokumentera normalvärden samt bedöma eventuell klinisk betydelse

Plant species from mesotrophic wetlands cause relatively high methane emissions from peat soil

Plants can influence methane emissions from wetland ecosystems by altering its production, consumption and transport in the soil. The aim of this study was to investigate how eight vascular plant species from mesotrophic to eutrophic wetlands vary in their influence on CH4 emissions from peat cores, under low and high N supply. Additionally, we measured the production of low-molecular-weight organic acids (LOA) by the same species (also at low and high N supply), because LOA form a substrate for methanogenesis. There were considerable differences among species in their effects upon rates of CH4 emission. Six of the species (Eriophorum latifolium Hoppe, Potentilla palustris (L.) Scop., Anthoxanthum odoratum (L.) s. str., Carex rostrata Stokes, Carex elata All., Carex acutiformis Ehrh.) increased CH4 emissions up to five times compared to control peat cores without plants, whereas two species (Phalaris arundinacea L., Phragmites australis (Cav.) Trin. ex Steud.) had no effect. There was a weak negative correlation between plant biomass and CH4 emission. N addition had no significant general effect upon CH4 emission. LOA production varied considerably among species, and tended to be highest for species from mesotrophic habitats. LOA production was stimulated by N addition. We conclude that some species from mesotrophic wetlands tend to cause higher CH4 emissions than species from eutrophic wetlands. This pattern, which contradicts what is often mentioned in literature, may be explained by the higher LOA production rates of species adapted to less productive habitat

Site-dependent N uptake from N-form mixtures by arctic plants, soil microbes and ectomycorrhizal fungi

Abstract in Undetermined Soil microbes constitute an important control on nitrogen (N) turnover and retention in arctic ecosystems where N availability is the main constraint on primary production. Ectomycorrhizal (ECM) symbioses may facilitate plant competition for the specific N pools available in various arctic ecosystems. We report here our study on the N uptake patterns of coexisting plants and microbes at two tundra sites with contrasting dominance of the circumpolar ECM shrub Betula nana. We added equimolar mixtures of glycine-N, NH4+-N and NO3--N, with one N form labelled with N-15 at a time, and in the case of glycine, also labelled with C-13, either directly to the soil or to ECM fungal ingrowth bags. After 2 days, the vegetation contained 5.6, 7.7 and 9.1% (heath tundra) and 7.1, 14.3 and 12.5% (shrub tundra) of the glycine-, NH4+- and NO3--N-15, respectively, recovered in the plant-soil system, and the major part of N-15 in the soil was immobilized by microbes (chloroform fumigation-extraction). In the subsequent 24 days, microbial N turnover transferred about half of the immobilized N-15 to the non-extractable soil organic N pool, demonstrating that soil microbes played a major role in N turnover and retention in both tundra types. The ECM mycelial communities at the two tundras differed in N-form preferences, with a higher contribution of glycine to total N uptake at the heath tundra; however, the ECM mycelial communities at both sites strongly discriminated against NO3-. Betula nana did not directly reflect ECM mycelial N uptake, and we conclude that N uptake by ECM plants is modulated by the N uptake patterns of both fungal and plant components of the symbiosis and by competitive interactions in the soil. Our field study furthermore showed that intact free amino acids are potentially important N sources for arctic ECM fungi and plants as well as for soil microorganisms

Plant and microbial uptake and allocation of organic and inorganic nitrogen related to plant growth forms and soil conditions at two subarctic tundra sites in Sweden

In order to follow the uptake and allocation of N in different plant functional types and microbes in two tundra ecosystems differing in nutrient availability, we performed a 15N-labeling experiment with three N forms and followed the partitioning of 15N label among plants, microorganisms and soil organic matter. At both sites the deciduous dwarf shrub Betula nana and the evergreen Empetrum hermaphroditum absorbed added 15N at rates in the order: NH4+ > NO3− > glycine, in contrast to the graminoid Carex species which took up added 15N at rates in the order NO3− > NH4+ > glycine. Carex transported a high proportion of 15N to aboveground parts, whereas the dwarf shrubs allocated most 15N to underground storage. Enhanced 13C in Betula nana roots represents the first field evidence of uptake of intact glycine by this important circumpolar plant. Plant and microbial uptake of label was complementary as plants took up more inorganic than organic N, while microbes preferred organic N. Microbes initially took up a large part of the added label, but over the following four weeks microbial 15N decreased by 50% and most 15N was recovered in soil organic matter, while a smaller but slowly increasing proportion was retained in plant biomass

Plasma atropine concentrations associated with decreased intestinal motility in horses

IntroductionAtropine is an essential part of the treatment protocol for equine uveitis. Topical atropine administration has been associated with decreased intestinal motility and abdominal pain in horses. Experimental studies have indicated that frequent dosing is associated with a higher risk than dosing every 6 h. Unfortunately, no quantitative pharmacodynamic data for inhibition of the equine gut are published. Materials and methodsEight standardbred horses were assigned to receive either atropine or saline (control) to be infused over 30 min in a two-treatment cross-over design. Atropine concentrations in plasma were measured using ultra-high-performance liquid chromatography-tandem mass spectrometry. Intestinal motility was measured using borborygmi frequency and electrointestinography (EIG). Experimental data were analyzed using a non-linear mixed effects model. The model was then used to simulate different dosing regimens. ResultsAtropine significantly decreased borborygmi response and EIG response. Six horses developed clinical signs of abdominal pain. The pharmacokinetic typical values were 0.31, 1.38, 0.69, and 1.95 L/kg center dot h for the volumes of the central, the highly perfused, the scarcely perfused compartments, and the total body clearance, respectively. The pharmacodynamic typical values were 0.31 mu g/L and 0.6 and 207 nV(2)7 cpm for the plasma concentration at 50% of the maximum response and the maximum response and the baseline of cecal EIG response, respectively. Six different dosing regimens of topical atropine sulfate to the eye (0.4 and 1 mg every hour, every 3 h, and every 6 h) were simulated. ConclusionThe IV PK/PD data coupled with simulations predict that administration of 1 mg of topical atropine sulfate administered to the eye every hour or every 3 h will lead to atropine accumulation in plasma and decreased intestinal myoelectric activity. Administration every 6 h predicted a safe dosing regimen in full-sized horses. Clinical studies would be valuable to confirm the conclusions. For smaller equids and horses put at risk for colic due to othercauses, droplet bottles that deliver 40 mu l of 1% atropine sulfate per drop or less may be used to lower the risk further

N-1-methylnicotinamide is a signalling molecule produced in skeletal muscle coordinating energy metabolism

Obesity is a major health problem, and although caloric restriction and exercise are successful strategies to lose adipose tissue in obese individuals, a simultaneous decrease in skeletal muscle mass, negatively effects metabolism and muscle function. To deeper understand molecular events occurring in muscle during weight-loss, we measured the expressional change in human skeletal muscle following a combination of severe caloric restriction and exercise over 4 days in 15 Swedish men. Key metabolic genes were regulated after the intervention, indicating a shift from carbohydrate to fat metabolism. Nicotinamide N-methyltransferase (NNMT) was the most consistently upregulated gene following the energy-deficit exercise. Circulating levels of N-1-methylnicotinamide (MNA), the product of NNMT activity, were doubled after the intervention. The fasting-fed state was an important determinant of plasma MNA levels, peaking at similar to 18 h of fasting and being lowest similar to 3 h after a meal. In culture, MNA was secreted by isolated human myotubes and stimulated lipolysis directly, with no effect on glucagon or insulin secretion. We propose that MNA is a novel myokine that enhances the utilization of energy stores in response to low muscle energy availability. Future research should focus on applying MNA as a biomarker to identify individuals with metabolic disturbances at an early stage.Peer reviewe

Organic acids in root exudates and soil solutions. Importance to calcicole and calcifuge behaviour of plants.

Many vascular plant species are unable to colonise calcareous sites, the floristic composition of adjacent limestone and acid silicate soils differing markedly. The inability of these 'calcifuge' species to establish themselves and grow on limestone soil appears to mainly be related to their low capacity for solubilising and absorbing phosphate and/or iron from such soils. Differences between calcifuge and calcicole species (the latter normally growing on calcareous soil) in their root exudation of low-molecular organic acids (LOAs) can lead to their ability to solubilise mineral nutrients from the soil differing. In the studies presented in this thesis I examined exudation by many calcicole and calcifuge species. Exudation was studied for plants grown hydroponically, for germinating seeds and seedlings and for plants grown in a pH intermediate soil. I found that calcicole species generally have a higher, often much higher, exudation rate of dicarboxylic oxalate and tricarboxylic citrate than calcifuge species do. These two LOAs were also shown to have a strong solubilising effect on soil phosphate and iron, both in calcareous and silicate soil. I also studied the calcifuge behaviour of Rumex acetosella in greater detail. R. acetosella was found to be a highly phosphate-limited species, its growth being closely correlated to the amount of easily exchangeable phosphate in the soil. This species, which contain large amounts of oxalate in its leaves, appears in any case to lack the ability to increase the exudation rate of LOAs in response to phosphate limitation. In my thesis I propose that high exudation rates of dicarboxylic and tricarboxylic organic acids is one of the mechanisms explaining calcifuge behaviour in plants. I also propose that high root exudation rates of LOAs from plants can be an adaptation to nutrient limiting conditions in soils

Visual evoked potentials in the horse

Vision is an important sense for horses, both for survival in the wild and when horses are used for work, sport or recreation. However, it is often difficult to diagnose visual impairment in this species. Traditional techniques available in clinical equine practice, are based on subjective evaluations, and their results are many times difficult to interpret. Electrodiagnostic methods, flash electroretinography (FERG) and recording of flash visual evoked potentials (FVEP) are used to objectively evaluate the function of the retinal and post-retinal visual pathways. The electrical potentials generated in response to brief visual stimuli are measured non-invasively. Abnormal function in visual pathways can affect the FERG and FVEP waveforms, peak times and amplitudes. Lesions can thereby be detected, and their approximate localization evaluated. FVEPs are used in human medicine, and occasionally in animal species, but have not been described in the horse. The general aim of this thesis was to establish a technique for recording of FVEPs in horses in clinical practice. The results showed that FVEPs can be readily recorded in sedated horses in a clinical setting. The recorded waveform consisted of a series of positive (P1-P5) and negative (N1-N2) wavelets. The overall appearance of the waveform was shown to be similar in foals, young horses and adult horses. An age-related effect on peak times and amplitudes was observed, but most of the changes occurred early in life. Important data on FVEP variability and repeatability was reported, and it was concluded that P2, N2 and P4 peak times should be included in the evaluation of equine, clinical FVEPs. The large inherent variability of FVEP amplitudes made them less useful, but they occasionally provided support to a clinical diagnosis. In clinical patients, electrodiagnostic testing helped assessing functional impact of potentially visual-threatening diseases. By recording FERGs and FVEPs simultaneously, a subdivision into retinal vs post-retinal dysfunction could be made in many patients, such as horses with optic neuropathies and cortical visual impairment. FVEPs may also be of prognostic value in horses with traumatic optic neuropathy and possibly in cases with cortical visual impairment. The results from this thesis, opens up for the use of the FVEP as an adjunctive, objective method in the evaluation of equine patients with suspected visual impairment and neurological disease, but also for studies of development and function of the visual pathways in this species

Regulation of the murine immunoglobulin GL gamma-1 and epsilon promoters

The immune system takes advantage of different genetic alterations to amplify its diversity. First at early stages of B cell differentiation when the antigen (ag) receptors are assembled from different gene segments. Secondly during the proliferative stage induced by ag challenge, when the lg genes undergo two types of modifications: somatic hypermutation and class switch recombination (CSR). During CSR the effector function of an lg molecule is altered, while the antigen specificity is remained, resulting in expression of IgG, lgA or lgE instead of the initial IgM. Switching to a given isotype requires induction of transcription over the targeted CH-gene, while it is still in germline (GL) configuration. However, the specific function of GL transcripts in CSR is still unknown and regulation of GL transcription over the C- gamma-1 and C-epsilon regions has been the main focus of this thesis. Initially, we explored whether the CD40/CD4OL interaction was sufficient for B cells to undergo CSR. In vivo distribution of an agonistic monoclonal rat anti-mouse CD40-antibody (ab) induced up- regulation of all lg isotypes in mice completely devoid of T cells. This induction was to some extent independent of IL-4, since lgE synthesis was only partially inhibited by co-injection of anti-IL-4. Anti- CD40 also induced GL c transcripts and expression of lgE in B cells from IL-4-/- mice in vitro. We then investigated the molecular mechanisms of CD40 signalling in induction of GL transcripts and CSR. We found that signalling through CD40 stimulated GL gamma-1, gamma-2-b and low levels of E transcripts. In B cells activated by LPS or anti-CD40, we could detect CSR to C-gamma-2-b. In addition, antiCD40 together with IL-4 or IL-5 induced CSR to the gamma-1 region. We showed that NF-kappaB activated both by LPS and anti-CD40 bound to the gamma-1 promoter. However the total concentration of bound NF-kappaB varied, in addition to the subunit composition induced by the two stimuli. Furthermore, we have investigated the dependency of proliferation for CSR and synthesis of GL transcripts. We examined what effect inhibition of DNA synthesis had on CSR. Cell cycle arrest was induced in cells stimulated with LPS+IL-4 by addition of the DNA synthesis inhibitors Hydroxyurea (HU) and Aphidicholin (AC). We found that CSR was completely abrogated. Addition of HU also reduced GL gamma-1 and c transcripts in LPS plus IL-4 activated B cells. To enrich for B cell blasts in different cell cycle phases we used elutriator centrifugation and found that GL gamma-1 transcripts were expressed in G1, and S phases, at lower levels in G2/M but not in G.. When investigating binding patterns of nuclear proteins to the GL gamma-1 promoter, we found two major LPS-induced DNA binding protein complexes. These complexes were shown by elutriation experiments to be expressed in G, and presumably S, but not G0. or G2/M. Furthermore, the complexes bound to an Ets consensus element. We concluded that there exists a relation between proliferation, Ig class switching and GL transcription. The Ets consensus binding site in the gamma-1 promoter overlaps with a binding site for Ikaros, a protein important for development of all lymphoid cells, which was initially defined as a transcriptional activator but later has been suggested to act as a repressor. Ikaros forms spectacular clusters in the nuclei of activated B cells at loci of heterochromatin. There are a number of possible Ikaros binding sites in the gamma-1 and epsilon GL promoters. We showed that Ikaros could bind to several of the possible binding sites in the GL gamma-1 and epsilon promoters, presumably in a cooperative manner. Co-transfection of dominant negative Ikaros, unable to bind DNA, with gamma-1 and epsilon promoter-luciferase reporter constructs to a B cell line, led to modestly increased basal transcription levels from these promoters. However, retroviral transduction of wildtype or a dominant negative mutation of Ikaros into primary LPS activated B cells, had no effect on lg class switching or GL transcription, even though the endogenous clusters were disrupted by introduction of the mutated Ikaros. Thus, Ikaros might not be directly involved in transcriptional regulation of specific genes but might exert its function at other levels. Colocalisation of Ikaros with methyltransferase, which serves to direct replication-coupled DNA methylation, might suggest a role in inheritable silencing of genetic material somehow selected for methylation