The US dairy industry loses approximately $1.2 billion due to heat stress related production losses annually. It was formerly believed that heat-stressed lactating dairy cattle produce less milk because they consume less feed. It has since been established that the reduction of feed intake is only responsible for about 50% of the reduced milk yield in HS cows. It is believed that HS increases gastrointestinal permeability (GIP), resulting in microbial components leaking from the lumen of the gastrointestinal tract into underlying tissue and stimulating an immune response. The immune response is suspected to alter overall metabolism, and milk production specifically, by diverting nutrients away from the mammary gland and other non-essential processes to support immune system activation. Topics examined herein focus on identifying markers to assess gastrointestinal permeability and the influence of heat stress on GIP and nutrient metabolism. The first study utilized an in vitro rumen fermentation system to determine if lactulose, sucralose, and D-mannitol could persist in an in vitro rumen culture. Lactulose could not be quantified in the rumen fluid matrix, D-mannitol was rapidly degraded, and sucralose concentrations did not change after 48 h of incubation, establishing sucralose as an indigestible marker in mature ruminants. The second study utilized a pair feeding design to directly assess the effect of HS on GIP, milk yield, and immune activation by lipopolysaccharide (LPS). HS cows (n=7) were exposed to a temperature-humidity index (THI) value of 74-80 for 4 d. The pair-fed thermoneutral cows (PFTN, n=8) were exposed to a constant THI of 64 with their intake matched to the HS cows. HS lowered milk yield without altering GIP, measured using orally dosed sucralose as a permeability marker, or eliciting an LPS related immune response. Jejunal mucosal scrapings were harvested from each cow, tight junction proteins were quantified, and no differences were detected. Lack of treatment responses in GIP marker recovery and tight junction protein abundance indicate that increased GIP may not be a driving force behind production losses in HS dairy cows. The third study focused on energy substrate utilization during HS with the objective of determining if tissue-level energy substrate metabolism could be influencing glucose sparing mechanisms. Metabolic flexibility of skeletal muscle, liver, and mammary tissue was assessed after 4 d of HS. It was determined that HS reduced skeletal muscle metabolic flexibility and did not alter liver and mammary metabolic flexibility. This indicates that skeletal muscle has a greater dependency on glucose as an energy substrate, which may decrease the pool of glucose available for lactose synthesis in lactating cows. Finally, the last study had the objective of assessing branched-chain amino acid (BCAA) requirements during HS. BCAA are oxidized for ATP synthesis in extrahepatic tissues and provide precursors for the biosynthesis of non-essential amino acids. They are also taken up by the mammary gland at a rate greater than what they are used in milk protein. Taken together, it was hypothesized that BCAA requirements may be increased during HS. BCAA entry rates into blood were assessed using a stable isotope approach and a 4-pool model. No differences were detected in daily entry rates or flux rates between pools indicating no change in requirements. When considering the results of all studies, reductions in milk yield are likely a result of altered macronutrient metabolism but further work is needed to confirm that hypothesis. Understanding the physiology behind HS related production losses is the first step in developing mitigation strategies. / Doctor of Philosophy / Heat stress (HS) is a global issue that compromises dairy cattle welfare and reduces milk production. On average the US dairy industry loses approximately $1.2 billion due to heat stress related production losses annually. With the global population expected to exceed 9 billion by 2050, strategies to mitigate HS related production losses are needed. Although cows exposed to HS conditions eat less food, that only explains about 50% of the production losses. It is hypothesized that the other 50% of milk yield losses is at least partially caused by increased gastrointestinal permeability (GIP), which elicits an immune response. Questions examined herein primarily focus on quantification of physiological and metabolic responses to HS. The objective of the first study was to identify a marker to assess GIP we could give orally to cows and detect in their urine. Commonly used GIP markers used in monogastric are carbohydrates, which have the potential to be fermented in the rumen. Sucralose was identified as a suitable marker due to its resistance to degradation in the rumen. The next study focused on measuring physiological responses of lactating dairy cows when exposed to HS conditions. Under the conditions of our experiment, HS decreased dry matter intake and milk yield without increasing GIP or inducing an immune response. It was determined that the reduction in dry matter intake was responsible for 66% of the reduced milk yield with the other 34% being associated with physiological changes other than increased GIP. The next study focused on how HS impacts the ability of skeletal muscle, mammary, and liver tissue to utilize glucose or palmitic acid as an energy substrate. The ability to switch between energy substrates is called metabolic flexibility. It was found that HS lowered the ability of skeletal muscle because it was unable to utilize fat as an energy source. Mammary and liver tissue exhibited no change in metabolic flexibility. The final study focused on how HS changed branched-chain amino acid (BCAA) plasma entry rates into plasma. An in vivo stable isotope method and a 4-pool mathematical model was used to predict how BCAA moved between pools which corresponds to the rate of protein turnover. Under the conditions of this experiment, no differences in BCAA entry rates were observed. Overall, results indicate altered energy substrate metabolism independent of immune activation stemming from altered GIP may be a driving factor in HS related production losses. Overall, this work contributes to understanding of HS biology and questions the established belief that increased GIP resulting in immune activation is responsible for about 50% of production losses.

Univ., Diss.--Passau, 2004.

This thesis presents the first randomised controlled trial (RCT) to investigate whether supplemented probiotic bacteria-"live microorganisms that, when administered in adequate amounts, confer a health benefit on the host" (Sanders, 2008)-affect mood and other psychological outcomes in people presenting with low mood. Seventy-nine participants with at least moderate symptoms of depression were randomised in a double-blind manner to receive either a probiotic preparation containing Lactobacillus helveticus and Bifidobacterium longum or a matched placebo for eight weeks. The RCT phase was followed by an open label extension in which all participants were offered the active study product for a further 8 weeks. Participants were followed up at 6 months post-study. Based on the existing evidence from gut-brain axis research, and on models linking depression with inflammation, immune activation, low vitamin D levels, and the gut microbiota (outlined in Chapters 1 and 2), it was hypothesised that: the overall sample would have elevated levels of inflammatory biomarkers and low levels of vitamin D at baseline, and that this would be associated with scores on psychological and irritable bowel syndrome (IBS) outcome measures; that group differences (active treatment versus placebo) would be observed in scores on psychological outcome measures after eight weeks of probiotic intervention; that group differences would also be observed in blood levels of proinflammatory cytokines, hsCRP, vitamin D and BDNF, and scores on a measure of gut function/IBS, and that levels of these variables may predict or impact on treatment response; and that group differences would be observed on outcome measures at the point of the 6-month follow-up between those who continued to take the probiotic and those who discontinued probiotic use. In total, 58 of the 77 participants who provided baseline blood samples (75%) had at least one marker of inflammation elevated outside the normal reference range at baseline. Baseline vitamin D was approaching the deficient level, displayed a seasonal pattern, and was associated with severity on one measure of cognition. No significant differences were found between the active treatment and placebo groups on any psychological outcome measure, the measure of gut function or in the level of any blood-based biomarker in the randomised phase. Baseline vitamin D level was found to moderate treatment effect on several outcome measures. The results of the open label extension supported the lack of efficacy observed in the randomised phase, and also allowed for the comparison of efficacy over intervention periods of varying durations. The results of the follow-up at 6 months post-trial indicated that, while mean scores on psychological outcome measures remained lower than baseline, there was regression on some outcome measures after the study. When the participants who replied to the 6 month follow-up questionnaire were divided into groups based on their self-reported dominant treatment since the trial (probiotics/nutrition, standard treatment or no treatment) there was no difference in mood or other psychological outcomes among the groups at 6 months. The current trial found no evidence that this probiotic formulation is effective in treating the symptoms of depression or IBS, or in moderating the levels of inflammatory and other biomarkers in a sample recruited with moderate depression. This finding does not support the theory proposed in several narrative reviews which suggests probiotics as a possible intervention for depression and other mental health outcomes, but is supported by the systematic review of human probiotics studies presented in Chapter 3 which found overall limited evidence of probiotic efficacy for psychological outcomes. Future studies in the area should attempt to further broaden this field, in particular by recruiting samples with mild and/or non-chronic depression for interventional studies, or by approaching probiotics as a preventative or adjuvant treatment strategy for depression.

<p>The formation of a single lumen during tubulogenesis is crucial for the development and function of many organs. Although 3D cell culture models have identified molecular mechanisms controlling lumen formation in vitro, their function during vertebrate organogenesis is poorly understood. In this work we used the zebrafish gut as a model to investigate single lumen formation during tubulogenesis. Previous work has shown that multiple small lumens enlarge through fluid accumulation and coalesce into a single lumen. However, since lumen formation occurs in the absence of apoptosis, other cellular processes are necessary to facilitate single lumen formation. </p><p>Using light sheet microscopy and genetic approaches we identified a distinct intermediate stage in lumen formation, characterized by two adjacent un-fused lumens. These lumens are separated by cell contacts that contain basolateral adhesion proteins. We observed that lumens arise independently from each other along the length of the gut and do not share a continuous apical surface. Resolution of this intermediate phenotype into a single, continuous lumen requires the remodeling of basolateral contacts between adjacent lumens and subsequent lumen fusion. </p><p>Furthermore, we provide insight into the genetic mechanisms regulating lumen formation through the analysis of the Hedgehog pathway. We show that lumen resolution, but not lumen opening, is impaired in <italic>smoothened (smo)</italic> mutants, indicating that fluid-driven lumen enlargement and resolution are two distinct processes. We also show that <italic>smo</italic> mutants exhibit perturbations in the Rab11 trafficking pathway, which led us to demonstrate that Rab11-mediated recycling, but not degradation, is necessary for single lumen formation. Taken together, this work demonstrates that lumen resolution is a distinct genetically-controlled process, requiring cellular rearrangement and lumen fusion events, to create a single, continuous lumen in the zebrafish gut.</p> / Dissertation

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9 pp. / When we go to the grocery store it is a fairly easy task to select and purchase nutritious meals. A readily available, predictable food supply is conveniently organized and displayed in the aisles. The nutritional composition of most foods is clearly labeled so you can immediately know what nutrients (and perhaps, toxins) you will be consuming. In contrast, rangeland animals live in a world where nutrients and toxins are constantly changing across space and time. For example, there may be 10s to 100s of plant species growing on a single acre, and each plant can differ widely in the kinds and amounts of nutrients and toxins it offers to free-ranging herbivores. Even at the level of the individual plant, plant parts vary in their concentration of nutrients and toxins; leaves, stems, and flowers, all differ in the kinds and amounts of nutrients and toxins they contain. Nutrient and toxin content of the same plant species can also vary depending on where it grows (in the sun vs. shade, on a wet vs. dry site, on a fertile vs. infertile site, etc.). Mother Nature can also drastically alter foraging environments as a result of natural disasters like floods, fires, or droughts. Wild animals may find themselves in unfamiliar environments during their natural migration patterns. Range and wildlife management practices can also place wild and domestic herbivores in unfamiliar environments via relocation and reintroduction programs or via grazing management practices. Despite all these challenges, rangeland herbivores are remarkably adept at selecting plants that meet their nutritional needs while largely avoiding plants that do not. The fact that animals preferentially select plant species that are more nutritious than what is available, on average, is strong evidence that animals are able to somehow detect nutrient and toxin levels in plants as they change across space and time. In this paper, we examine recent important discoveries that underscore the importance of learning as a critical mechanism which allows rangeland herbivores to survive in a world where the only constant is change (Provenza, 2003; www.behave.net).

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No description available.

This study was directed at characterising the cannabinoid receptor activity modulating the electrical field stimulation (EFS) evoked contractions of the rat isolated ileum myenteric plexus longitudinal muscle (MPLM) preparation, and the capsaicin, nicotine and veratridine evoked secretory responses of the rat isolated colonic submucosal plexus-mucosal (SPM) preparation. EFS of the MPLM preparation with single pulses at a repetition frequency of 0.05 Hz elicited a transient twitch contraction immediately in response to each electrical pulse. In contrast, stimulation of the MPLM preparation with 2 second trains of pulses every minute at a frequency of 30 Hz elicited a rapid transient rebound contraction on termination of each train of EFS. The non-selective cannabinoid receptor agonists AEA, CP 55,940, D9-THC and WIN 55,212-2 inhibited both EFS-evoked twitch and rebound contractions of the rat ileum MPLM elicited by 0.05 Hz and 30 Hz EFS respectively. The inhibition of the twitch contractions was competitively antagonised by the cannabinoid CB1 receptor antagonist / inverse agonist SR 141716 with pKB values of 8.60. In contrast, SR 141716 only antagonised the ability of AEA, D9-THC and WIN 55,212-2 but not CP 55,940 to inhibit the rebound contractions with pA2 values of 6.60. These observations extended to the inhibitory effect of WIN 55,212-2 on the twitch and rebound contractions of the guinea-pig ileum MPLM. The CB2 antagonist / inverse agonist SR 144528 did not alter the effects of the agonists. Additionally, the inhibitory effect of AEA was refractory to the vanilloid TRPV1 receptor antagonist capsazepine. WIN 55,212-3 a stereoisomer of WIN 55,212-2 was without effect on the rat MPLM. SR 141716 alone concentration-dependently increased the twitch contractions but inhibited the rebound contractions. Both types of the EFS-evoked contractions were abolished by the Na+ channel blocker tetrodotoxin or the muscarinic acetylcholine (ACh) receptor antagonist atropine but not the nicotinic ACh receptor antagonist hexamethonium. None of the cannabinoids altered the contractions to exogenously applied ACh. These data suggested that the cannabinoid agonists inhibited the twitch contractions through a stereospecific presynaptic CB1 receptor-mediated reduction in the release of ACh. Additionally, the inhibition of the rebound contractions occurred because of an inhibition of ACh release by a novel stereospecific presynaptic non-CB1 -non CB2 -non -TRPV1 site. The ability of SR 141716 to inhibit the rebound contractions and antagonise AEA, D9-THC and WIN 55,212-2 may be though partial agonism at the non-CB1-non CB2-non-TRPV1 site. The ability of SR 141716 to potentiate the twitch contractions by increasing the release of ACh suggested that the CB1 receptor was constitutively active or was subjected to a tonic activation by endocannabinoid agonists. A comparison between the maximal enhancement of the twitch contractions of the rat and the guinea-pig ileum MPLM caused by three CB1 receptor antagonists/inverse agonists AM 251, SR 141716 and O-2050 showed that each cannabinoid had a different maximum. This suggested inverse agonism. These data were supported with studies showing the lack of effect of three fatty acid amide hydrolase (FAAH) inhibitors AA-5HT, PMSF, URB–597 and VDM-11, an inhibitor of the AEA uptake transporter on EFS-evoked contractions. These studies showed that all three FAAH inhibitors increased the potency of exogenously applied AEA but not WIN 55,212-2, and that VDM-11 had no effect on the potency of exogenously applied AEA. This data suggested that a functional endocannabinoid tone and the uptake transporter were not present in the MPLM, but FAAH was present. These data provide supporting evidence that SR 141716 behaved as an inverse agonist in the MPLM to augment twitch contractions. The interaction between CP 55,940 or WIN 55,212-2 with SR 141716 was investigated using the rat colonic SPM sheet. Both CP 55,940 and WIN 55,212-2 attenuated the secretory responses to capsaicin and nicotine in a SR 141716 sensitive manner. SR 140333, a neurokinin 1 receptor antagonist, abolished the capsaicin and nicotine. This suggested that CP 55,940 and WIN 55,212-2 inhibited the capsaicin and nicotine response through a CB1 receptor-mediated inhibition of the release of substance P or neurokinin A. The sensitivity of the veratridine response to TTX and a-chymotrypsin and the failure of the cannabinoids to attenuate the response suggested the absence of the CB1 receptor on the neurones releasing the undetermined neuropeptide. Together, these data suggest that both the CB1 receptor and non-CB1-non-CB2 -non-TRPV1 receptor can mediate the inhibitory effects of cannabinoid agonists in the rat ileum MPLM depending on the frequency of EFS. These data also show that SR 141716 is an inverse agonist in the MPLM. In the SPM preparation, the CB1 receptor appears to be involved in the modulation of some forms of peptidergic transmission.

The gut and the brain are in constant communication through pathways that include the immune system, the nervous system, neurotransmitters, and hormones. Modifications in the gut, especially the gut microbiome, have the potential to cause changes in the brain resulting in behavioral and cognitive changes. A healthy and diverse microbiome, which may be achieved by a high fiber diet or probiotic or prebiotic treatments, is associated with improvements in cognition. Gut dysbiosis and a decrease in diversity of the microbiota, which may be caused by a western diet or antibiotic treatments, is associated with cognitive decline and decreased memory. There are many possible pathways through which these changes in the gut act to change cognition, including the immune system, the expression of brain derived neurotropic factor, metabolites such as short chain fatty acids, gut hormones, and neurotransmitters. If researchers can decipher which pathways are involved in modifying cognition, they may be able to identify treatments that can help improve memory and specifically decrease age-related cognitive decline.