Kathie Wickwire

Direct effects of nutrients, acetylcholine, CCK, and insulin on ghrelin release from the isolated stomachs of rats

Ghrelin is a powerful orexigenic peptide predominantly secreted by the stomach. Blood concentration of ghrelin increases before meals and fall postprandial. Its regulation appears to be influenced by the type of macronutrient ingested, the vagus nerve stimulation and by other post-meal stimulated hormonal factors. However, the direct role of nutrients (amino acids or lipids), neuronal (vagal neurotransmitter acetylcholine) and satiety-inducing factor such as CCK are not known. To study this we applied amino acids, lipids, acetylcholine and CCK via vascular perfusion to the isolated stomachs and found that amino acids significantly reduced ghrelin release from the isolated stomach by approximately approximately 30% vs. the control while lipids (10% intralipid) had no affect. Acetylcholine (1 microM) increased ghrelin release from the stomach by approximately 37% whereas insulin (10nM) decreased it by approximately 30% vs. the control. Interestingly, CCK (100 nM) potently increased ghrelin release by approximately 200% vs. the control. Therefore it appears that ghrelin secretion from the stomach is under direct influence of amino acids, neurotransmitter acetylcholine and hormones such as insulin and CCK.

Effect of reducing hypothalamic ghrelin receptor gene expression on energy balance

Central and peripheral injections of fghrelin potently stimulates food intake via its receptor, GHSR1a expressed in the brain. In this study, we explored the role of GHSR1a in the paraventricular nucleus of the hypothalamus (PVN) by reducing their gene expression using the RNA interference (RNAi). pSUPER plasmids inserted with sh (short hairpin)-GHSR1a were injected into the PVN to reduce its expression. The transfected rats were monitored daily for their food intake and body weight throughout the experimental period lasting 8 days. We found that knockdown of GHSR1a did not affect daily food intake but significantly reduced body weight and blood ghrelin levels. This suggests that the central ghrelin system could selectively regulate body weight without affecting energy intake.

Maternal high fat feeding and gestational dietary restriction: effects on offspring body weight, food intake and hypothalamic gene expression over three generations in mice.

Excessive gestational weight gain and maternal obesity have both been associated with increased incidence of obesity and metabolic disorder in offspring in both humans and animal models. The objectives of this study were to determine (1) whether mild gestational food restriction during the third trimester (GFR) would alter food intake and growth parameters of offspring, (2) whether effects of GFR depended on diet (high fat [HF] vs chow), (3) whether effects of excessive gestational weight gain (WG) would become magnified across generations, and (4) whether diet and GFR would alter hypothalamic gene expression in adult offspring. Three generations of female C57BL/6 mice were fed chow or HF diet, mated at 11 weeks of age and assigned to ad libitum feeding or 25% GFR. Offspring were fed the same diet as their mothers. Results showed (1) maternal gestational WG was positively correlated with offspring WG. (2) HF offspring weighed less (p<0.01) at weaning (WWT) but gained more during the 8 weeks after weaning than chow-fed offspring (p<0.05), resulting in higher final body weights (BW) (p<0.01). (3) HF males from GFR mothers had higher WWT (p<0.05), but subsequent WG and final BW were less (p<0.05) compared to males from ad lib mothers. (4) In the HF group, GFR also resulted in decreased FI (p<0.05) and FE (p<0.07) in offspring, compared to offspring from ad lib mothers. (5) In generation 3, hypothalamic expression of tyrosine hydroxylase was lower in HF males from GFR mothers compared to HF males from ad lib mothers (p<0.05). In conclusion, gender and maternal GFR had independent effects on growth and FI, and hypothalamic gene expression was dependent on both gender and maternal GFR in HF offspring. Even mild food restriction of obese mothers during pregnancy may have beneficial effects in reducing the risk or degree of obesity in offspring.

Genetic analysis of tissue glutathione concentrations and redox balance

Glutathione redox balance-defined as the ratio GSH/GSSG-is a critical regulator of cellular redox state, and declines in this ratio are closely associated with oxidative stress and disease. However, little is known about the impact of genetic variation on this trait. Previous mouse studies suggest that tissue GSH/GSSG is regulated by genetic background and is therefore heritable. In this study, we measured glutathione concentrations and GSH/GSSG in liver and kidney of 30 genetically diverse inbred mouse strains. Genetic background caused an approximately threefold difference in hepatic and renal GSH/GSSG between the most disparate strains. Haplotype association mapping determined the loci associated with hepatic and renal glutathione phenotypes. We narrowed the number of significant loci by focusing on those located within protein-coding genes, which we now consider to be candidate genes for glutathione homeostasis. No candidate genes were associated with both hepatic and renal GSH/GSSG, suggesting that genetic regulation of GSH/GSSG occurs predominantly in a tissue-specific manner. This is the first quantitative trait locus study to examine the genetic regulation of glutathione concentrations and redox balance in mammals. We identified novel candidate genes that have the potential to redefine our knowledge of redox biochemistry and its regulation and inform future therapeutic applications.

Agouti-Related Protein: Appetite or Reward?

Abstract: Agouti‐related protein (AgRP) is an orexigenic peptide that acts as an antagonist of the melanocortin‐3 and ‐4 receptors in the hypothalamus. Studies suggest that the melanocortin and opioid systems interact in the control of ingestive behavior. Also, AgRP has been shown to especially increase intake of a palatable diet. Given these observations, we wished to examine whether the effects of AgRP on ingestive behavior resemble those of opioids. AgRP was injected into the hypothalamic paraventricular nucleus in animals given a choice between a palatable sucrose solution and calorically dense chow. As a result of AgRP injection, animals increased intake of chow but not sucrose relative to controls, in contrast to what has been seen with opioid agonists. These results together with prior findings suggest that the primary effect of AgRP is to cause an increase in food intake to satisfy energy needs, though AgRP also has opioid‐like effects, possibly due to melanocortin‐opioid interactions.

Role of AgRP on Ghrelin-induced feeding in the hypothalamic paraventricular nucleus.

MT II, agonist for MC3/4-Rs, inhibited Ghrelins orexigenic effect in the paraventricular nucleus of the hypothalamus (PVN). To further investigate the role of the melanocortin system as mediator of ghrelins orexigenic actions, we explored the involvement of AgRP in Ghrelins orexigenic effect by testing the effect on food intake after their co-administration in the PVN, during the light and dark phases of feeding in rats. During both the phases of feeding, co-administration of Ghrelin with either AgRP 50 or AgRP 100 pmol into the PVN did not produce a synergistic effect on the food intake, suggesting that ghrelin induction of feeding occurs by recruiting Agrp as one of the obligatory mediators of its orexigenic effect.

WHOLE-EGG DIET DELAYS THE AGE-RELATED IMPAIRED GLUCOSE TOLERANCE OF BHE/CDB RATS

Abstract The effects of dietary egg on the age-related progression of impaired glucose tolerance and glomerulonephropathy in diabetes-prone BHE/Cdb rats were studied. This rat strain mimics the human with NIDDM. The development of impaired glucose tolerance was delayed in rats fed the whole-egg diet, however, feeding this diet resulted in elevated hepatic weight but had no effect on the age-related changes in renal lesions or renal function. We conclude that in this animal model for NIDDM, the development of glomerulonephropathy is independent of the development of impaired glucose tolerance and that diet can affect the time course for impaired glucose tolerance without affecting renal disease development.

Menhaden Oil Feeding Increases Potential for Renal Free Radical Production in BHE/cdb Rats

Abstract The effects of feeding 9% beef tallow (BT) or menhaden oil (MO) in a 10% fat-60% sucrose-20% protein diet on renal cortex fatty acid profile, renal lipid peroxide formation potential, and the blood pressure response to a norepinephrine challenge was studied. Male weanling BHE/cdb prediabetic rats were studied after 8 weeks of diet treatment. Half the rats were subjected to a norepinephrine challenge, and their mean arterial blood pressure was determined. Plasma renin and angiotensin II levels were determined in the presence or absence of the challenge. The source of dietary fat had no effect on these measurements. MO fed rats had a greater potential to form lipid free radicals in the kidney than BT fed rats despite the fact that the renal tissue from both groups had an equivalent number of unsaturations on a mole % basis. From these results we conclude that the accelerated renal disease in menhaden oil fed rats is not due to a diet fat effect on blood pressure regulation but might be due to a diet fat effect on free radical production. These free radicals can be cytotoxic and if produced in large amounts could result in a loss of glomerular cells. Whether this occurs and can be reversed by a change in diet was not determined.

Differential hepatic and renal cholesterol levels in diabetes-prone BHE/cdb rats fed menhaden oil or beef tallow.

Abstract A series of experiments were conducted to determine whether the feeding of beef tallow compared with menhaden oil would affect renal cortex membrane composition, Na, K-ATPase activity, renal cholesterol uptake, and plasma lipoprotein cholesterol profile. BHE/cdb rats were used because they carry a genetic trait for non–insulin-dependent diabetes mellitus and are prone to develop diabetic nephropathy. Beef tallow feeding resulted in an increase in HDL cholesterol and an increase in Na, K-ATPase activity. The different fats also affected the arachidonic acid content of the membrane but not the membrane cholesterol content. These diet effects may explain why the development of renal disease in beef tallow–fed rats is delayed when compared with rats fed an equivalent amount of menhaden oil.

Neither the cholesterol or arginine content of whole egg explains its beneficial effect on glucose homeostasis in BHE/cdb rats 1

Abstract Male weanling BHE/cdb rats were fed either a whole-egg diet, a control diet, or a control diet that matched either the arginine or cholesterol content of the whole-egg diet. Glucose tolerance, renal function, cholesterol status, and urea cycle activity was assessed at 250 days of age. Neither the cholesterol content of the whole-egg diet nor its arginine content could explain the whole-egg diet effect on glucose tolerance, renal metabolism, or renal function. At 250 days of age BHE/cdb rats fed the whole-egg diet had a near normal glucose tolerance, whereas rats fed the other diets had impaired tolerance as well as impaired renal function. This diet difference might be attributable to either the vitamin A or lecithin content of the whole egg, but is not attributable to its arginine or cholesterol content.