The field of food intake and satiety has received increasing interest from the research community in recent years. The mechanisms and factors that regulate satiety gains their importance from the crucial role they play in food consumption and consequently control of body weight. Leukotrienes are mediators that are released in inflammatory conditions. One of the receptors on which Leukotrienes perform their actions is Cysteinyl Leukotriene Receptor Type 2 (CysLT2Rs). Recently, our colleagues made the observation that CysLT2Rs are expressed in vagal afferent neurons. In addition, CysLT2R-/- mice appeared to be heavier than WT (Moos and Funk, unpublished observations). Based on these findings, I hypothesized that CysLT2Rs play a role in regulating food intake via vagal afferent activity. In-vivo studies were performed to characterize body weight gain and investigate whether weight gain was associated with increased food intake. I found that CysLT2R-/- mice not only have significantly higher body weight, but also eat significantly more than CysLT2R+/+ mice. Using calcium imaging techniques, I demonstrated that LTD4 and LTC4 increased calcium Ca2+ influx in nodose ganglion neuron. Moreover, the level of neuronal activation in the brainstem (NTS area) was measured in both groups of mice using immunohistochemical techniques, which suggested less postprandial neuronal activity in KO mice. These data suggest that CysLT2Rs take part in regulating body weight and food intake. In addition these results implicate vagal afferents as a possible pathway. These findings may have implications for the control of food intake in both health and disease and may lead to novel insights in the causes and treatment of disordered weight such as overweight and obesity, or even anorexia. / Thesis (Master, Physiology) -- Queen's University, 2011-02-04 10:48:47.018

This project aimed to investigate the role of the γ-lactone of 3,4-dihydroxybutanoic acid (3,4DB) which was claimed by Japanese investigators to be one of several endogenous γ-lactones involved in the control of food intake. Tissue, plasma and urine organic acid profiles were screened for the γ-lactone of 3,4DB using both GC and GCMS. Careful mass spectral analysis and <I>in vitro</I> acid-γ-lactone exchange analyses with structural validation studies showed that the γ-lactone of 3,4DB did not occur <I>in vivo</I> but that the free acid of 3,4DB did. This rejects previous claims to the contrary. Fasting increased rat plasma 3,4DB with urinary output significantly elevated for the first 24h of fasting. A metabolic route from glutamate to acetate is proposed with 3,4DB as an intermediate. Peripheral administration of glutamate, γ-aminobutyric acid (GABA) and 4-hydroxybutyric acid (4HB) to rats reduced food intake with increasing effects at each stage of the pathway. The γ-lactone 2-buten-4-olide is toxic and other γ-lactones had more potent effects than their free acids on rats. The intragastric administration of GABA stimulated 4HB production <I>in vivo</I> and GABA or 4HB increased plasma 3,4DB levels, thus implying that 3,4DB occurs at an intermediate in our proposed pathway. A likely endogenous source of 3,4DB is as a minor GABA metabolite. The response of the free acid of 3,4DB to fasting followed a similar trend as that reported for its γ-lactone and related γ-lactones are unlikely to occur <I>in vivo</I>. Given the similarities of 4HB (a precursor of 3,4DB) and 3,4DB in structure to the ketone body 3-hydroxybutyrate (3HB) and the similarity of 3,4DB in fasting response to 3HB, both 4HB and 3,4DB may initiate anorexia in fasting.

No description available.

Appetite is a complex behavior which can be influenced by factors within the animal's body as well as the environment around it. Internal factors include hormonal and nutrient concentrations found in the blood stream and subsequent neuropeptide and neurotransmitter signaling in the hypothalamus. External factors, such as high ambient temperature (HAT), can indirectly affect appetite regulation through other neuroendocrine systems such as the hypothalamo-pituitary-adrenal (HPA) axis. Understanding the physiological responses to endogenous factors and HAT exposure in birds will have implications in both the agricultural and biomedical fields. Thus, the purpose of this dissertation research was to explore the hypothalamic molecular mechanisms associated with food intake in broiler type chickens and Japanese quail, and the effect of HAT exposure on food intake in broiler type chickens. Broiler type chickens have undergone intense artificial selection for traits that promote rapid growth, consequently driving them to consume feed incessantly. It is hypothesized that broiler type chickens lack a mechanism that signals satiety, causing them to eat significantly more than layer type chickens. Selection for rapid growth of meat (muscle tissue) has made the broiler more susceptible to the deficits associated with HAT exposure, as animals composed primarily of muscle dissipate less heat while also producing more heat than those composed of fat. The Japanese quail have undergone relatively minor artificial selection compared to the chicken, suggesting that use of this model may provide insight into the mechanisms of appetite regulation in wild-type bird species. This research involved administrating appetite associated factors into the avian brain via an intracerebroventricular (ICV) injection including gastrin releasing peptide, vasoactive intestinal polypeptide, neuropeptide AF, and prostaglandin D2. Additionally, I explored the effects of HAT on food intake, and on the efficacy of several ICV administered appetite associated factors including neuropeptide Y, corticotropin releasing factor and α-melanocyte stimulating hormone. After treatment administration, I measured changes in food intake and behavior, activation of hypothalamic nuclei including the arcuate nucleus, dorsomedial nucleus, lateral hypothalamus, paraventricular nucleus, and the ventromedial nucleus, and the nucleus of the hippocampal commissure. I then measured changes in gene expression in both whole hypothalamic samples and specific hypothalamic nuclei. The data from non-HAT associated studies provided information on the hypothalamic nuclei which respond to the various appetite associated factors and the molecular mechanisms mediating changes in appetite. The data from the HAT study provided information on the hypothalamic nuclei involved in the avian response to HAT exposure, and the molecular mechanisms involved in the effect on food intake. Overall, these data provide insight on the mechanisms associated with short-term regulation of appetite, and pathways associated with stress and food intake. / Doctor of Philosophy / Appetite regulation can be affected by factors both in the body and out in the environment. Understanding how both internal and external factors affect appetite regulation can have positive implications in both the agriculture industry as well as the biomedical field. In agriculture, animals exposed to high ambient temperatures often exhibit several deficits including immunosuppression, decreased body weight, and ultimately an increased risk of mortality. It is hypothesized that the factor linking negative wellbeing to heat exposure is a reduction in food intake. Animals aside, the prevalence of eating disorders has doubled worldwide every 6 years since the year 2000. These numbers have increased even more during the recent COVID-19 pandemic. In order to improve the wellbeing of both humans and animals exposed to stressing stimuli, it is imperative we understand how individual appetite associated factors affect food intake, and how external stressors can impact the normal physiology of the hypothalamus. Thus, the purpose of this dissertation was to elucidate the hypothalamic mechanisms mediating appetite regulation using broiler type chickens and Japanese quail as models. Related pathways and molecular mechanisms were explored for several appetite associated factors including gastrin releasing peptide, vasoactive intestinal polypeptide, neuropeptide AF, and prostaglandin D2. Additionally, the effect of high ambient temperature on food intake, on the efficacy of several appetite associated factors including neuropeptide Y, corticotropin releasing factor and α-melanocyte stimulating hormone, and the hypothalamic pathways and molecular mechanisms mediating heat-induced anorexia were assessed.

This study was to determine (1) if genetic selection for high (HWS) or low (LWS) body weight in chickens has altered the hypothalamic AMP-activated protein kinase (AMPK) system and (2) if this alteration contributes to the dissimilar feeding response to various appetite modulators between HWS and LWS lines. Compared to HWS, LWS chickens had higher levels of AMPK α and acetyl-CoA carboxylase (ACC) phosphorylation, which was caused by upregulation of the upstream factor calcium/calmodulin-dependent protein kinase kinase 2 (CAMKK β). There was greater mRNA expression of carnitine palmitoyltransferase I (CPT1), leptin receptor (LEPR) and neuropeptide Y (NPY) and less mRNA expression of ACC α, fatty acid synthase (FAS), fat mass and obesity associated gene (FTO), pro-opiomelanocortin (POMC) and orexin in LWS than HWS chickens. At 5 days of age, intracerebroventricular (ICV) injection of AICAR, 5-amino- 4-imidazolecarboxamide riboside, caused a quadratic dose-dependent decrease in food intake in LWS but not HWS chicks. Compound C, (6-(4-(2-piperidin-1-yl-ethoxy)-phenyl))-3-pyridin-4-yl-pyrazolo(1,5-a)-pyrimidine, caused a quadratic dose-dependent increase in food intake in HWS but not LWS chicks. The anorexigenic effect of AICAR in LWS chicks and orexigenic effect of Compound C in HWS chicks resulted from either activation or inhibition of other kinase pathways separate from AMPK. There is a lower threshold for the anorexigenic effect of ghrelin in LWS than HWS chicks, which was associated with differential hypothalamic AMPK signaling. ICV injection of ghrelin inhibited corticotrophin-releasing hormone (CRH), 20-hydroxysteroid dehydrogenase (20HSD), glucocorticoid receptor (GR), CPT1 and FTO expression in LWS but not HWS chicks. Additionally, the hypothalamic mRNA level of ghrelin was significantly higher in LWS than HWS chicks, which may also contribute to the differential threshold response to ghrelin in these two lines. Obestatin caused a linear dose-dependent increase in food intake in HWS but not LWS chicks. The orexigenic effect of obestatin in HWS chicks was not associated with altered AMPK. Obestatin inhibited LEPR and FTO expression in HWS but not LWS chicks. Thus, selection for body weight may alter the hypothalamic response to ghrelin by the AMPK pathway, CRH pathway, CPT1 and FTO, and to obestatin by LEPR and FTO. / Ph. D.

Experiments were conducted to evaluate nitric oxide (NO) involvement in the leptin effect on food intake in both broiler and Leghorn chickens. The first experiment studied the effect of leptin combined with L-arginine on the food intake in broilers. Intracerebroventricular (ICV) administration of human recombinant leptin injection decreased (P=.01) food intake from 15 to 150 minutes compared to the control group treated with artificial cerebrospinal fluid ( aCSF) while food intake was increased by L-arginine. Food intake between the group receiving leptin and L-arginine was similar to the control group. Therefore, broilers were sensitive to the anoregenic effects of leptin, while L-arginine, a NO precursor appeared to attenuate the leptin effect on food intake. The effect of leptin and L-NNA on food intake in broilers was measured in the second experiment. Lepin, L-NNA and leptin plus L-NNA decreased food intake. The NO inhibitor L-NNA tended to enhance the suppression of leptin on food intake. In the third experiment, using Leghorns instead of broilers, the ICV injection of leptin decreased food intake from 15 to 60 minutes postinjection (P=.05). However, food intake was not affected by injection of L-arginine plus leptin. Therefore, L-arginine appeared to antagonize the leptin inhibitory effect on food intake. A small increase food intake induced by L-arginine was also observed (P=.09). The change of food intake in Leghorns administered leptin and L-NNA were measured in Experiment 4. Food intake was decreased by L-NNA and leptin with the effects lasting 60 minutes, similar to that observed in broilers (P<0.0l). For group B (leptin treatment), there was decreased food intake within 45 minutes (P=.04) and the effect disappeared 60 minutes, post injection. Also, the results along with Experiment 2 demonstrated that NO mediated the effect of leptin in Leghorns. The fifth experiment investigated the change in concentration of metabolites of nitric oxide after injection of leptin within 30 minutes. The group treated with the leptin had a lower level of metabolites of nitric oxide in the hypothalamus than the control group (P=.004). This effect further demonstrates that leptin modulated feeding activity through its inhibition on nNOS activity in the hypothalamus. These results showed that both leptin and NO participated in the regulation of food intake in broiler and Leghorn chickens, and the effect of hypothalamic neuropeptidergic circuitry leptin on food intake was mediated by NO. / Master of Science

The present set of experiments were designed to examine the role of histamine, and its interaction with corticotropin releasing factor (CRF) and bombesin (BM) in food intake regulation of chickens. The hypothesis being tested was as follows: One component of the neuroregulation of food intake involves histaminergic activity in the hypothalamus, acting on either H1 or H2 receptors, how these receptors interact with CRF neurons and if BM elicits its effects on feeding through CRF release. Single Comb White Leghorn (SCWL) and broiler cockerels were utilized for these experiments. Birds were stereotaxically implanted with a 23-gauge thin-walled stainless steel guide cannula, and were provided a mash diet and water for ad libitum consumption. All compounds were infused into the right lateral ventricle. Effects were monitored at 15-minute intervals through three hours postinjection. Experiment 1 examined the effects of intracereboventricular (ICV) injections of histamine (HA) and two HA antagonists, the H1 receptor antagonist chloropheneramine maleate (CM) and H2 receptor antagonist cimetidine (CIM), on food and water consumption and body temperature. Histamine was infused using 0, 25, 50, and 100 µg per 10 µl of artificial cerebrospinal fluid (aCSF). Histamine significantly decreased food and water consumption (P< 0.05) over the three hour observation period in a dose-dependent manner. Histamine was then infused to observe if the decrease in water intake was dependent upon the decrease in food intake. Birds were not allowed access to feed during this experiment. Water intake was not affected by HA in either SCWL or broilers when food was not available. To observe the effects of HA on thermoregulation, HA was infused using the same dosages and body temperature recorded for three hours. Histamine produced hypothermia at a dose of 25 µg in SCWL cockerels, with a quadratic trend at 165 and 180 min. Broiler cockerels did not show hypothermia, but rather a constant hyperthermia compared to the control with a quadratic trend throughout the latter part of the experiment. The last phase of the first set of experiments, birds were pretreated with either CM or CIM (100 µg/10 µl aCSF) followed by HA. When the birds were pretreated with either CM or CIM, the hypophagic responses to HA were attenuated. The pair of experiments that utilized H1 and H2 receptors demonstrated that these receptors are involved in the neural regulation of food intake. These experiments also demonstrated that the aphagic effects of HA on food intake can be blocked with the pretreatment of antihistaminics affecting both H1 and H2 receptors. In Experiment 2, studies were conducted to determine if neuronal CRF elicited its effects on feeding through the release of HA. Birds were infused with 0 or 20 µg CRF and either 0 or 100 µg of CM or CIM. CRF decreased food and water intake in both SCWL and broiler cockerels. When birds were pretreated with CM, the hypophagic responses to CRF were attenuated. When birds were pretreated with CIM, the hypophagic responses of CRF were attenuated in broiler cockerels; this response was not seen in SCWL cockerels. Water intake followed a similar pattern. It was concluded that, contrary to studies showing that HA causes the release of CRF in other species, CRF may cause the release of HA in chickens. Experiment 3 was designed to investigate whether bombesin (BM) elicited its effects on feeding through the release of CRF. Birds were infused with either, 0 or 0.5 µg BM, 0 or 5 µg aCRF (9-41) (CRF antagonist), or a combination of both. These compounds were infused to test whether the effects of BM could be blocked with the pretreatment of anticorticotropics. Food and water consumption were significantly decreased (P< 0.05) with the infusion of BM in both SCWL and broiler cockerels. Food intake was not affected with the infusion of aCRF in SCWL or broilers cockerels. However, water consumption was increased when birds were given ICV injections of aCRF. When birds were pretreated with aCRF, the anorexigenic and adipsic effects of BM were attenuated. It was concluded that BM elicits its effects on feeding through the release of CRF. These results also demonstrate that the aphagic effects of BM could be blocked with the pretreatment of anticorticotropics. / Master of Science

Marijuana has been used for medicinal and recreational purposes for thousands of years. Many people think of marijuana in the context of an illegal drug. Because of the antimarijuana attitude, research with cannabinoids was neglected for a long time. Although this substance is related to social problems, scientists are interested in its action and possible medicinal properties. Since the identification of the structure of Î 9--tetrahydrocannabinol, the main psychoactive ingredient of marijuana, there has been increased interest in this compound. Following the discovery of two cannabinoid receptors, CB1 and CB2 receptors, it was determined that CB1 receptors are in high density in the central nervous system while CB2 receptors are found primarily in the immune system. The endogenous cannabinoid ligands, anandamide and 2-arachidonoylglycerol, were observed in the central nervous system and peripheral tissues. Endocannabinoids differ from other "classical" neurotransmitters because they do not appear to be stored in synaptic vesicles, and they act as retrograde messengers within the brain. The endogenous cannabinoid signaling system includes cannabinoid receptors, their endogenous ligands called endocannabinoids, and the proteins for their synthesis and inactivation. The cannabinoid system appears to act as a neuromodulatory system. During the past ten years, the endogenous cannabinoid system has been implicated in a variety of physiological functions including pain reduction, motor regulation, learning, memory, and reward. Because obesity and eating disorders are prevalent, scientists are working at the molecular level to study the mechanisms controlling body weight and regulation of food intake. Several of the neuropeptides present in hypothalamic nuclei contribute to energy balance and food intake regulation. Endogenous cannabinoid and cannobinoid receptors are found in the hypothalamus and are associated with the regulation of food intake. Although the mechanisms whereby cannabinoids influence food intake remain unclear, results suggest that the cannabinoid system will be an important target in future studies in obesity. Most research on cannabinoids has focused on their role in food intake regulation in mammalian species. It is important to determine the role of endocannabinoids in other species. The effect of intracerebroventricular injection of agonists and antagonists of both CB1 and CB2 receptors in 8 to 11 week-old male Single Comb White Leghorn and 3 to 6 weeks old male broilers was investigated. It was found that agonists of both the CB1 and CB2 receptor increased food intake significantly; however, the CB2 receptor agonist had a stronger and longer lasting effect. Antagonists of both receptors decreased food intake significantly. The CB1 receptor antagonist appeared to block both cannabinoid receptors in birds, whereas the CB2 receptor antagonist did not block both receptors. Previous studies have indicated that the CB2 receptor is found only outside the brain and spinal cord, and is involved with the immune system. From the present results, it appears that both cannabinoid receptors are present in the chicken brain. Furthermore, the CB2 receptor may also be localize in the chicken brain. There are also differences in cannabinoid system between Leghorn and broilers. / Master of Science

Prolactin-releasing peptide (PrRP) is an endogenous hypothalamic neuropeptide that when exogenously injected increases food intake in chickens, but decreases it in rodents and goldfish. We designed three sets of experiments to elucidate the mechanisms of PrRP's orexigenic effect in chicks. In experiment one, food and water intake were evaluated in chicks after receiving intracerebroventricular (ICV) injection of the vehicle, 0.75, 3, 12, 47 or 188 pmol PrRP. The administration of 12 and 47 pmol PrRP increased food intake for up to 120 min after injection, and 188 pmol increased it for up to 180 min. The lowest effective dose was 3 pmol, which increased food intake for up to 60 min after injection. Water intake was not affected. To investigate the molecular mechanisms, c-Fos immunohistochemistry was performed and mRNA expression of some appetite-associated neurotransmitters was measured in chicks that received either vehicle or 188 pmol of PrRP. The rostral paraventricular nucleus (PVN) was activated which coincided with increased neuropeptide Y (NPY) mRNA expression in the whole hypothalamus. In experiment two, food and water intake were evaluated in chicks fed a high carbohydrate (HC), high fat (HF) or high protein (HP) diet after ICV injection of vehicle, 3 or 188 pmol PrRP. Chicks fed the HP diet increased food intake at a lower dose than chicks fed HF and HP diets after ICV PrRP injection. In addition, ICV injection of vehicle, 3 and 188 pmol PrRP were performed in chicks fed all three diets, and ICV PrRP injection induced preferential intake of the HP diet over HC and HF diets. The expression of some appetite-associated neuropeptides in the hypothalamus was also measured in chicks fed the HC, HF or HP diet after ICV injection of vehicle or 188 pmol PrRP. There was a diet effect on mRNA abundance of all appetite-associated genes measured (P < 0.05), with greater expression in chicks fed the HF or HP than HC diet. While neuropeptide Y (NPY) mRNA abundance was similar between vehicle and PrRP-injected chicks that consumed HP or HF diets, expression was greater (P < 0.05) in PrRP- than vehicle-injected chicks that consumed the HC. In experiment three, the orexigenic effect of PrRP was tested in chicks selected for low (LWS) and high (HWS) body weight after central administration of vehicle, 24, 94 and 375 pmol PrRP. The LWS chicks had a lower threshold and higher magnitude of food intake increase in response to PrRP injection. Results demonstrate that PrRP is a potent orexigenic factor in chickens and that effects are likely mediated through the hypothalamus. The orexigenic effect of PrRP was influenced by dietary macronutrient composition, and diet in turn influenced the food intake response to PrRP. These results may contribute to a novel understanding of appetite regulation. / Master of Science

Feeding behavior is a complex trait that is regulated by various hypothalamic neuropeptides and neuronal populations (nuclei). Understanding the physiological regulation of food intake is important for improving nutrient utilization efficiency in agricultural species and for understanding and treating eating disorders. Knowledge about appetite in birds has agricultural and biomedical relevance and provides evolutionary perspective. I thus investigated hypothalamic molecular mechanisms associated with appetite in broilers, layers, chicken lines selected for low (LWS) or high (HWS) body weight, and Japanese quail, which provide a unique perspective to understanding appetite. Broiler-type chicks have been genetically selected for rapid growth and consume much more feed than do layer-type chicks which have been selected for egg production. Long-term selection has caused the LWS chicks to have different severities of anorexia while the HWS chicks become obese, thus making these lines a valuable model for metabolic disorders. Lastly, the Japanese quail have not undergone as extensive artificial selection as the chicken, thus this model may provide insights on how human intervention has changed the mechanisms that regulate feeding behavior in birds. This research involved applying a variety of different treatments including fasting and refeeding, diets differing in macronutrient composition, and/or central administration of neuropeptide Y, xenopsin, neuropeptide K, oxytocin, mesotocin, gonadotropin-inhibitory hormone, and prolactin-releasing peptide, after which I measured feeding behavior and various aspects of hypothalamic physiology. I measured nuclei activation in hypothalamic appetite-associated regions including the lateral hypothalamus, paraventricular nucleus, ventromedial hypothalamus, dorsomedial nucleus, and arcuate nucleus and I measured gene expression of various appetite-associated factors in the whole hypothalamus and individual nuclei. These data provided information about the regions of the brain involved in mediating effects on appetite and the molecular pathways involved in the effect on appetite. There were differences in dose threshold sensitivity to various injected factors in the different stocks, differential responses to fasting and refeeding, and differences in nuclei and genes that were activated in response to the various treatments. These data provide valuable insights on the molecular mechanisms that are associated with the short-term regulation of feeding behavior and pathways that may be genetically stock-dependent. / PHD