A novel approach to evaluation of tumor response for advanced pulmonary adenocarcinoma using the intertumoral heterogeneity response score.

Treatment response and prognosis estimation in advanced pulmonary adenocarcinoma are challenged by the significant heterogeneity of the disease. The current Response Evaluation Criteria in Solid Tumors (RECIST) criteria, despite providing a basis for solid tumor response evaluation, do not fully encompass this heterogeneity. To better represent these nuances, we introduce the intertumoral heterogeneity response score (THRscore), a measure built upon and expanding the RECIST criteria. This retrospective study included patients with 3-10 measurable advanced lung adenocarcinoma lesions who underwent first-line chemotherapy or targeted therapy. The THRscore, derived from the coefficient of variation in size for each measurable tumor before and 4-6 weeks posttreatment, unveiled a correlation with patient outcomes. Specifically, a high THRscore was associated with shorter progression-free survival, lower tumor response rate, and a higher tumor mutation burden. These associations were further validated in an external cohort, confirming THRscore's effectiveness in stratifying patients based on progression risk and treatment response, and enhancing the utility of RECIST in capturing complex tumor behaviors in lung adenocarcinoma. These findings affirm the promise of THRscore as an enhanced tool for tumor response assessment in advanced lung adenocarcinoma, extending the RECIST criteria's utility.

Changes of insulin receptors in high fat and high glucose diet mice with insulin resistance.

This study aimed to observe the expression of insulin-signaling molecules in different organs of mice with insulin resistance (IR). Firstly, mice were fed a high-fat and high-sugar diet (HF group) to establish an IR model, and the controls (NF group) were fed with a normal diet. Next, the weight, fasting blood glucose (FBG), serum insulin and insulin tolerance were detected. Pathological changes of liver tissues were observed by H&E staining. The expressions of INSR, IRS-1 and IRS-2 in the liver, skeletal muscle and ovary were measured by qRT-PCR and western blotting. As a result, compared with the NF group, the HF group mice had increased weight, FBG, insulin and IR index after 6-week of feeding as well as a worse performance in the insulin tolerance test and H&E staining showed fatty liver-like changes after 12-week of feeding, exhibited lower expression of INSR, IRS-1 and IRS-2 in the liver of mice at 6 and 12 weeks. The expression of INSR and IRS-1 in skeletal muscle tissues exhibited the same trend, while those in ovary organs showed the opposite trend. These results suggested that the insulin signaling alters in the liver, skeletal muscle and ovary organs with the progress of IR.

Both prolonged high-fat diet consumption and calorie restriction boost hepatic NAD+ metabolism in mice.

Hepatic NAD+ homeostasis is essential to metabolic flexibility upon energy balance challenges. The molecular mechanism is unclear. This study aimed to determine how the enzymes involved in NAD+ salvage (Nampt, Nmnat1, Nrk1), clearance (Nnmt, Aox1, Cyp2e1), and consumption pathways (Sirt1, Sirt3, Sirt6, Parp1, Cd38) were regulated in the liver upon energy overload or shortage, as well as their relationships with glucose and lipid metabolism. Male C57BL/6N mice were fed ad libitum with the CHOW diet, high-fat diet (HFD), or subjected to 40% calorie restriction (CR) CHOW diet for 16 weeks respectively. HFD feeding increased hepatic lipids content and inflammatory markers, while lipids accumulation was not changed by CR. Both HFD feeding and CR elevated the hepatic NAD+ levels, as well as gene and protein levels of Nampt and Nmnat1. Furthermore, both HFD feeding and CR lowered acetylation of PGC-1α in parallel with the reduced hepatic lipogenesis and enhanced fatty acid oxidation, while CR enhanced hepatic AMPK activity and gluconeogenesis. Hepatic Nampt and Nnmt gene expression negatively correlated with fasting plasma glucose levels concomitant with positive correlations with Pck1 gene expression. Nrk1 and Cyp2e1 gene expression positively correlated with fat mass and plasma cholesterol levels, as well as Srebf1 gene expression. These data highlight that hepatic NAD+ metabolism will be induced for either the down-regulation of lipogenesis upon over nutrition or up-regulation of gluconeogenesis in response to CR, thus contributing to the hepatic metabolic flexibility upon energy balance challenges.

Decreased taste sensitivity to sucrose in dopamine D3 receptor mutant mice.

Dopamine plays a key role in food rewards and sweet-taste stimulation. We examined the basis for behavioral responses to sweet taste in dopamine D3 receptor-deficient (D3-/-) mice by determining whether the absence of D3 receptors affects the sensitivity to dilute sucrose solutions. In experiment 1, we measured the intensity generalization threshold of conditioned taste aversion (CTA) to a 0.2 M sucrose solution. Results showed that the generalization thresholds were 0.025-0.05 M in D3-/- mice and 0.0025-0.005 M in wild-type (WT) mice. In experiment 2, we found that D3-/- and WT mice had similar capabilities to form and extinguish CTAs. Since the intensity generalization threshold is mainly due to a combination of sweet-taste sensitivity and the robust nature of CTA formation, the results showed that taste sensitivity to sucrose in D3-/- mice was lower than that in WT mice. In experiment 3, to test whether the peripheral sensory signaling may also be affected by the disruption of the dopamine D3 receptors, the mRNA expression levels of sweet-taste-related proteins in taste buds of D3-/- mice were determined. The T1R1 and BDNF mRNA expression levels in D3-/- mice were higher than the controls, whereas T1R2, T1R3, α-gustducin, and TRPM5 mRNA were similar. These findings suggest that disruption of dopamine D3 receptor-mediated signaling decreases the sweet-taste sensitivity and alters the mRNA expression levels of some taste-related molecules.

Maternal exercise and high-fat diet affect hypothalamic neural projections in rat offspring in a sex-specific manner.

This study aimed to investigate the effect of maternal high-fat (HF) diet and exercise during gestation and lactation on hypothalamic neural projection development in the offspring. Pregnant Sprague-Dawley rats were fed a CHOW or HF diet during gestation and lactation, and further divided into two subgroups: voluntary exercised and sedentary. Offspring's brains and tissue were collected at weaning and 16 weeks of age. Maternal exercise downregulated dams' body weight and food intake during lactation, but failed to normalize increased fat weight, plasma and milk leptin levels of HF dams at weaning. Maternal HF diet significantly increased offspring's body weight, adipose depots, plasma insulin, and leptin at weaning and had long-term effect on body weight of male offspring, while maternal exercise decreased offspring's body weight from 3 to 5 weeks of age in both sexes. At weaning, maternal exercise decreased αMSH fiber density and maternal HF diet impaired agouti-related peptide fiber density in the paraventricular nucleus of hypothalamus of male pups, while maternal HF diet disrupted αMSH fiber density in the paraventricular nucleus of hypothalamus of female pups. The impaired αMSH fiber density was consistent with reduced STAT3 signaling in the arcuate nucleus of hypothalamus, while the reduced agouti-related peptide fiber density was consistent with reduced ERK1/2 signaling in the arcuate nucleus of hypothalamus. The impaired hypothalamic projections were compensated in adulthood in both sexes. Our findings suggest that maternal HF diet and exercise exerts different effects on hypothalamic neural projections development through distinct signaling pathways in a sex-specific manner.

NNMT is induced dynamically during beige adipogenesis in adipose tissues depot-specific manner.

Nicotinamide N-methyltransferase (NNMT) is a novel regulator, shown recently to regulate adipose tissue energy expenditure partly through changing NAD + content, which is essential for mitochondrial. We determine whether NNMT plays important role in energy metabolism during the beige adipogenesis in vivo and in vitro. Male C57BL/6 mice at 8 weeks old were exposed to 4 ℃ for 1, 2, 3, 4, and 5 days, respectively. Interscapular brown adipose tissue (iBAT), inguinal subcutaneous WAT (sWAT), and epididymal WAT (eWAT) were harvested for gene and protein expression analysis and the correlation analysis. In addition, cultured primary mice brown adipocyte (BA) and white adipocyte (WA) treated with or without ß3-adrenoceptor agonist (CL316, 243) were also harvested for these analyses. A combination of NNMT and its related genetic (Nmnat1, Nampt, Cyp2e1, Nrk1, Cd38) and proteic analyses and also the NAD + levels demonstrated the dynamical and depot-specific remodeling of NAD metabolism in different adipose tissues in response to cold exposure. While upon CL316, 243 treatment, gene expression of Nnmt, Nampt, Cyp2e1, and Nrk1 was all significantly decreased in WA but not in BA. The increased NAD + amount in BA and WA during the beige adipogenesis was observed. Besides, it is demonstrated that the expression of NNMT both in sWAT and WA showed significant negative correlation with browning markers UCP-1 and PGC-1α at protein levels. Above all, NNMT was induced in WAT during the 'cold remodeling' phase and correlated negatively with the process of browning in sWAT and WA, indicating the specific role of NNMT in the regulation of energy homeostasis during the process of beige adipogenesis.

Melanocortin receptor-4 mediates the anorectic effect induced by the nucleus tractus solitarius injection of glucagon-like Peptide-2 in fasted rats.

Glucagon-like peptide-2 (GLP-2) is secreted from enteroendocrine L-type cells of the gut and also released from preproglucagonergic (PPG) neurons in the nucleus tractus solitarius (NTS) and adjacent medial reticular nucleus of the brain stem. The neurons in the NTS express GLP-2, and the neurons send extensive projections to the hypothalamus. Recent studies show that the intracerebroventricular administration of GLP-2 significantly suppresses food intake in animals and some evidence suggest that the melanocortin receptor-4 (MC4-R) signaling in the hypothalamus is required for intracerebroventricular GLP-2-mediated inhibition of feeding. There is proopiomelanocortin (POMC) positive neurons expressing MC4-R in the NTS. Suppression of MC4-R expressing neurons in the brain stem inhibits gastric emptying. In this study, we tested the effects of NTS GLP-2R activation and blockade on feeding behavior and evaluated the endogenous melanocortin system's role in the NTS in mediating effects of GLP-2 on feeding behavior in fed and fasted rats. Our results demonstrated that microinjection of GLP-2 into the NTS suppressed food intake in fasted-refeeding rats but did not affect food intake in free-feeding rats, and this inhibition was blocked by pretreatment of either Exendin (9-39) or SHU 9119, suggesting the GLP-2 system in the NTS exerts an inhibitory action on food intake. MC4-R mediates this action in the NTS.

GLP-2 decreases food intake in the dorsomedial hypothalamic nucleus (DMH) through Exendin (9-39) in male Sprague-Dawley (SD) rats.

Glucagon-like peptide 2 (GLP-2), a member of Glucagon peptide family involved in regulating energy metabolism, can be produced and secreted by preproglucagonergic (PPG) neurons in the brain. GLP-2 reduces food intake but at which brain sites GLP-2 exerts its feeding-suppress effects are still unclear. In this study, we used the stereological microinjection technique and behavioral test to examine the functions of locally delivered GLP-2 into DMH on feeding behavior. We compared effects of different concentration of GLP-2 on the food intake behavior in free-feeding rats and fasted-refeeding rats. We found that GLP-2 inhibited food intake in fasted rats after a short-term intervention in a dose-dependent manner. Importantly, the effects of locally delivered GLP-2 can be blocked by specific GLP-1 receptor antagonist Exendin(9-39), but not the melanocortin-4 receptor antagonist SHU9119, indicating the involvement of specificity of GLP-2 signaling in regulating the feeding behavior. Taken together, our data revealed that GLP-2 peptide pharmacologically inhibited food intake in DMH and this effect could be blocked functionally by Exendin(9-39).

Maternal exercise during gestation and lactation decreases high-fat diet preference by altering central reward system gene expression in adult female offspring from high-fat fed dams.

Exposure to maternal high-fat (HF) diet during gestation and lactation alters adult offspring's feeding behavior and diet preference. However, the impact of maternal exercise on offspring's diet preference and reward system development is less studied. In this study, we investigate the effect of perinatal maternal exercise on the development of diet preference, dopamine- and opioid-related gene expression in the central reward system in female offspring from HF-fed Sprague-Dawley rat dams. We found maternal HF diet did not alter adult offspring HF preference, but influenced offspring's dopamine and opioid system both at weaning and in adulthood, and these offspring retained higher body weight in adulthood. However, offspring from dams exposed to both HF diet and exercise during gestation and lactation had normalized body weight, decreased fat mass and lower HF-diet preference but increased energy intake in adulthood. The dopamine- and opioid-related gene expression in central reward system and POMC expression in hypothalamus was elevated in these adult offspring. We conclude that maternal exercise during gestation and lactation can potentially overcome the negative effects of perinatal exposure to HF diet in female offspring by altering their diet preference, central reward system signaling and hypothalamus neuropeptide expression.

Depot-specific regulation of NAD+/SIRTs metabolism identified in adipose tissue of mice in response to high-fat diet feeding or calorie restriction.

Deteriorated nicotinamide adenine dinucleotide (NAD+)/sirtuins (SIRTs) metabolism in adipose tissue is implicated in diet-induced obesity, while calorie restriction (CR)-induced beneficial effects require sufficient NAD+ biosynthesis. Mechanistic links have not been defined. This study aims to identify changes of specific components of NAD+/SIRTs system in white adipose tissue (WAT) and brown adipose tissue (BAT) of mice upon energy imbalance, focusing on key enzymes in NAD+ salvage (Nampt, Nmnat1, Nrk1), clearance (Nnmt, Aox1, Cyp2e1) and consumption pathways (Sirt1, Sirt2, Sirt3, Sirt6, Parp1). Male C57BL/6J mice were fed ad libitum with the standard laboratory chow diet, high-fat diet (HFD) or 40% CR diet, respectively. The epididymal and inguinal WAT (eWAT and iWAT) and interscapular BAT (iBAT) were harvested for histological, NAD+ assay, gene and protein expression analysis after 16 weeks of dietary regimen. HFD decreased, while CR increased, the NAD+ and NADH levels in eWAT, iWAT and iBAT. NAD+ content negatively correlated with plasma cholesterol, TNF-α levels and calorie intake, while it positively correlated with plasma adiponectin level. The change trend of SIRT1 is quite the same as that of NAD+/NADH ratio. Nmnat1 gene is sensitive to energy imbalance in WAT but not in BAT. Nrk1 gene expression was decreased in eWAT and iWAT but increased in iBAT of HFD mice. Nnmt mRNA and protein abundance was increased in iWAT of HFD mice. Nampt, Cyp2e1 and Sirt3 were the most robust genes responding to energy imbalance. In summary, adipose tissue responds to long-term energy excess or shortage with depot-specific transcriptional activation or repression of NAD+/SIRTs metabolic components.

NAD+ /sirtuin metabolism is enhanced in response to cold-induced changes in lipid metabolism in mouse liver.

The nicotinamide adenine dinucleotide (NAD+ )/Sirtuin (SIRT) system is linked to metabolic adaptation. This study aimed to determine the temporal profile of metabolic responses of the liver to cold exposure and changes in the hepatic NAD+ /SIRT system. Eight-week-old male C57BL/6 mice were individually housed in conventional cages under cold exposure (4 °C) for up to 5 days. Cold exposure decreased the hepatic triglyceride level and cholesterol level in mice by 1.7- and 1.6-fold, respectively. Lipogenic gene expression was persistently reduced, while gluconeogenic gene expression was transiently increased. Hepatic NAD+ /SIRT metabolism was induced during the 'cold remodeling' phase (days 1-3) and correlated with decreasing lipogenic and increasing gluconeogenic gene expression, contributing to the maintenance of whole-body lipid and glucose homeostasis.

Maternal low protein exposure alters glucose tolerance and intestinal nutrient-responsive receptors and transporters expression of rat offspring.

AIMS: Maternal protein malnutrition during perinatal period has long-term consequences on the offspring's metabolic phenotype. Here we determined the effects of maternal protein-restricted (PR) diet on offspring's metabolism in 3- and 12-week-old. MAIN METHODS: Sprague-Dawley rats were fed with standard chow diet or PR diet during pregnancy and lactation. Food intake and body weight of offspring were measured weekly. The oral glucose tolerance tests were underwent, the pancreases were collected for histochemical staining, and the duodenum, jejunum and ileum were collected for gene and protein expression analysis in 3- and 12-week-old offspring. KEY FINDINGS: PR offspring had significant lower body weight and persisted till 12-week-old. From 3- to 12-week-old, PR offspring presented considerably impaired glucose tolerance, while no marked change was shown in control rats. Additionally, the average islet size of PR offspring decreased significantly in 12-week-old. The mRNA and protein expression of nutrient-responsive receptors and transporters T1R3, SGLT1 and GLUT2 increased significantly in the intestine of 3-week-old PR offspring. And from 3- and 12-week-old, the increase tendency of expression subdued. SIGNIFICANCE: These results suggest that maternal PR diet during critical developmental windows influences offspring metabolism, which may be subdued partially, but not be reversed completely by chow diet after weaning.

Prenatal exercise reverses high-fat-diet-induced placental alterations and alters male fetal hypothalamus during late gestation in rats†.

Maternal high-fat (HF) diet negatively affects maternal metabolism and placental function. This study aimed to determine whether gestational exercise prevents the effect of HF diet on placental amino acid transporter expression and nutrient-sensing signaling and the fetal response. Pregnant Sprague-Dawley rats were either fed with a CHOW (13.5% fat) or HF (60% fat) diet during gestation and further divided into two subgroups: voluntary exercised and sedentary. Placentae were collected on gestational day (GD) 14 and GD20, and male placentae were used in this study. We found that gestational exercise ameliorated the detrimental effects of HF diet on dams' adiposity, plasma leptin, and insulin concentrations. Maternal exercise did not influence fetoplacental growth but affected male fetal hypothalamic Leprb, Stat3, Insr, Agrp, and Pomc expressions on GD20. Maternal HF diet decreased placental labyrinth thickness and increased system A amino acid transporter SNAT2 expression, while these changes were normalized by exercise. The activation of placental mechanistic target of rapamycin complex 1/4E-BP1 and LepRb/STAT3 signaling might contribute to the increased placental SNAT2 expression in HF-fed dams, which were reversed by exercise on GD20. These data highlight that gestational exercise reverses HF-diet-induced placental alterations during late gestation without influencing fetal growth. However, maternal exercise altered fetal hypothalamic gene expression, which may affect long-term offspring health.

Maternal high-fat diet during gestation and lactation increases conditioned aversion threshold for sucrose and alters sweet taste receptors expression in taste buds in rat offspring.

Maternal high-fat (HF) diet affects offspring's metabolic phenotype. Sweet taste is an important factor in promoting appetite. In order to determine the effects of maternal HF diet throughout gestation and lactation on taste sensitivity to sucrose in rat offspring, we measured conditioned aversion threshold for sucrose by conditioned taste aversion (CTA) associated with two-bottle choice tests, and measured mRNA expression of sweet taste receptors in taste buds. In male offspring, conditioned aversion threshold for sucrose lay between 0.007 M and 0.009 M in control group, while in those with HF dams, the threshold significantly increased to be between 0.011 M and 0.02 M. In female offspring, conditioned aversion threshold for sucrose lay between 0.003 M and 0.005 M in control group, whereas maternal HF diet increased it to be between 0.007 M and 0.009 M. Maternal HF diet increased T1R2 and T1R3 mRNA expression in taste buds of male offspring, while only increased T1R2 mRNA expression in female offspring. Both male and female offspring with HF dams had lower α-gustducin mRNA expression, whereas only male offspring with HF dams had lower OB-Rb mRNA expression in taste buds. Our data suggest that maternal HF diet decreased taste sensitivity to sucrose in both male and female offspring, which may be partly due to altered expression of sweet taste receptors and related downstream pathways in taste buds.

Targeting Trim69 alleviates high fat diet (HFD)-induced hippocampal injury in mice by inhibiting apoptosis and inflammation through ASK1 inactivation.

The prevalence of obesity is growing, and high fat diet (HFD)-induced obesity can alter the brain and cognition. However, the link between HFD, hippocampal function, and inflammation is still not fully understood. Tripartite motif (TRIM) family has been implicated in various cellular processes, such as apoptosis, neurogenesis, and innate immune responses. Trim69, a member of TRIM family, was investigated in the present study to determine its role in HFD-induced hippocampal damage. Here, we first found that hippocampal Trim69 expression was markedly down-regulated in wild-type (WT) mice challenged with HFD. Trim69 knockout (KO) mice exhibited an exaggerated version of the metabolic disorder after HFD challenge, as evidenced by their increased body weight and elevated insulin resistance. HFD-induced hippocampal injury was further aggravated by Trim69 deletion, as confirmed by the reduced survival of neurons and increased level of apoptotic cell death. In addition, the inflammatory response triggered by HFD was more pronounced in the hippocampi of Trim69-KO mice after blockage of the activation of the nuclear factor kappa B (NF-κB) signaling pathway. Phosphorylation of mitogen-activated protein kinase (MAPK) kinase 4 (MKK4), MKK7, and c-Jun N-terminal kinase (JNK) in the hippocampi of HFD-challenged mice was intensified by the loss of Trim69. Hippocampal-apoptosis-signal-regulating kinase 1 (ASK1) phosphorylation was also found to be up-regulated by HFD, especially in mice with Trim69 deletion. Of note, we found that Trim69 directly interacted with and deubiquitinated ASK1 in microglial cells. Microglial cell-specific suppression of Trim69 exacerbated inflammation and apoptosis in response to lipopolysaccharide (LPS). Trim69 over-expression markedly alleviated LPS-induced inflammatory response and apoptotic cell death in microglial cells. Together, these results indicated that Trim69 might be a functionally essential inhibitor of ASK1 activation during the pathogenesis of hippocampal inflammation and apoptosis, and it could serve as a novel molecular target for obesity-associated brain damage.

[Nutritional status alters the mRNA expressions of galanin and its receptors in taste buds of rats].

The nutritional and metabolic status alters the peripheral taste perception and food intake by participating in the modulation of taste information integration. The taste receptors and neuropeptides in the taste buds are the important targets of this modulation process. To explore the effects of nutritional status on the expressions of galanin and its receptors in the taste buds, we compared the mRNA levels of galanin and its specific receptor GalR2 in the taste buds among the high-fat diet induced obese rats (HF), chronically restricted diet rats (CR) and control rats. The high-fat diet, half of chow diet, and normal chow diet were given to HF, CR and control groups for 6 weeks, respectively. The body weight and some metabolic indexes, including blood glucose, triglyceride and cholesterol levels were detected. The mRNA expressions of galanin and its receptors in taste buds were determined using real-time PCR. Results showed that compared with control rats, the body weights, levels of blood glucose and triglyceride were significantly elevated in HF rats; while the mRNA expressions of galanin and GalR2 were dramatically decreased. However, galanin mRNA expression in CR rats was increased to 2.3 times of that in control group. Considering the results obtained from our previous studies, we conclude that the behavioral changes in tasting choice of HF rats may be related to the expressions of galanin and GalR2 in the taste buds. The changes of galanin and GalR2 in taste buds are involved in the peripheral mechanism of nutritional status regulating taste perception and feeding behavior in rats.

[Role of the central nucleus of the amygdala in regulating the nongenomic effect of aldosterone on sodium intake in rat nucleus tractus solitarius].

OBJECTIVE: To reveal the nongenomic effect of aldosterone on the regulation of sodium intake in the nucleus tractus solitarius (NTS) and the role of central nucleus of the amygdala (CeA) in regulating this effect. METHODS: Adult male SD rats were divided into four groups and underwent operations to induce bilateral CeA electrolytic lesions (400 µA, 25 s; n=28), bilateral sham CeA lesions (n=28), unilateral CeA lesions (n=28), or unilateral sham CeA lesions (n=26). After 3 days of recovery, the rats received implantation of a stainless steel 23-gauge cannula wih two tubes into the NTS followed by a recovery period of 7 days. The rats in each group were then divided into two subgroups for microinjection of aldosterone (50 ng/µL) or control solution in the NTS, and the cumulative intake within 30 min of 0.3 mol/L NaCl solution was recorded for each rat. RESULTS: Bilateral CeA lesions (3 days) eliminated the increased 0.3 mol/L NaCl intake induced by aldosterone microinjected into the NTS (0.3±0.04 mL in CeA lesion group vs 1.3±0.3 mL in sham lesion group). Unilateral CeA lesion (3 days) reduced aldosterone-induced increase of NaCl intake in the first 15 min (P < 0.05) but not in 15-30 min (P > 0.05). In rats with sham lesions, aldosterone (50 ng/µL) still induced a significant increase in NaCl intake[1.3±0.3 mL vs 0.25±0.02 mL in the control group; F (3, 224)=24.0, P < 0.05]. CONCLUSIONS: The regulation of sodium intake by aldosterone is subjected to descending facilitatory modulation by the bilateral CeA, and CeA integrity is essential for aldosterone to execute the nongenomic effect in regulating rapid sodium intake.

The purinergic mechanism of the central nucleus of amygdala is involved in the modulation of salt intake in sodium-depleted rats.

The central nucleus of the amygdala (CeA) is a critical region in regulating sodium intake, and interestingly, purinergic receptors reportedly related to fluid balance, are also expressed in CeA. In this study, we investigated whether the purinergic mechanisms of CeA were involved in regulating sodium intake. Male Sprague-Dawley rats had cannulas implanted bilaterally into the CeA and were sodium depleted with furosemide (FURO 20 mg/kg) plus 24 h-sodium deficient food fed. Bilateral injections of the P2X purinergic agonist, α,ß-methyleneadenosine 5'-triphosphate (α,ß-methylene ATP 1.0, 2.0, 4.0 nmol, respectively) into the CeA region induced dose-related reductions in sodium intake without affecting water intake. Injection of P2X purinergic antagonist, pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid (PPADS 4.0 nmol/0.5 µl) into the CeA region did not alter sodium and water intake, however, prior injection of PPADS into the CeA area abolished the inhibitory effects on sodium intake by α,ß-methylene ATP. Interestingly, prior injection of γ-aminobutyric acid type A (GABAA) receptor antagonist, bicuculline (4.0 nmol/0.5 µl) into the CeA region partially reversed the deficit of sodium intake induced by α,ß-methylene ATP. These results suggest that purinergic receptors in the CeA are involved in the control of sodium intake in the sodium-depleted rats and this negative modulation may be, at least partly, mediated by the GABAA receptor.

Effects of saccharin supplementation on body weight, sweet receptor mRNA expression and appetite signals regulation in post-weanling rats.

Non-nutritive sweeteners have been considered to promote diet healthfulness by delivering a pleasant sweet taste without calories. We investigated the effects of long term supplementation with drinks containing saccharin on body weight and possible mechanisms of the effects in post-weanling rats. Our results showed that saccharin solution intake increased food intake and energy intake in male rats. In males, saccharin solution intake increased TIR3 mRNA expression in the taste buds and ghrelin receptor mRNA expression both in the taste buds and hypothalamus, whereas no effects were observed in females. These results suggest the effects of saccharin solution exposure on food intake and body weight gain may be different in developmental males and females. In males, peripheral sweet taste receptors and both peripheral and central ghrelin receptors may be involved in the effect of saccharin solution intake to promote food intake and weight gain.

Expression of Proinflammatory Cytokines Is Upregulated in the Hypothalamic Paraventricular Nucleus of Dahl Salt-Sensitive Hypertensive Rats.

Accumulating evidence indicates that inflammation is implicated in hypertension. However, the role of brain proinflammatory cytokines (PICs) in salt sensitive hypertension remains to be determined. Thus, the objective of this study was to test the hypothesis that high salt (HS) diet increases PICs expression in the paraventricular nucleus (PVN) and leads to PVN neuronal activation. Eight-week-old male Dahl salt sensitive (Dahl S) rats, and age and sex matched normal Sprague Dawley (SD) rats were divided into two groups and fed with either a HS (4% NaCl) or normal salt (NS, 0.4% NaCl) diet for 5 consecutive weeks. HS diet induced hypertension and significantly increased cerebrospinal fluid (CSF) sodium concentration ([Na+]) in Dahl S rats, but not in normal SD rats. In addition, HS diet intake triggered increases in mRNA levels and immunoreactivities of PVN PICs including TNF-α, IL-6, and IL-1ß, as well as Fra1, a chronic marker of neuronal activation, in Dahl S rats, but not in SD rats. Next, we investigated whether this increase in the expression of PVN PICs and Fra1 was induced by increased CSF [Na+]. Adult male SD rats were intracerebroventricular (ICV) infused with 8 µl of either hypertonic salt (4 µmol NaCl), mannitol (8 µmol, as osmolarity control), or isotonic salt (0.9% NaCl as vehicle control). Three hours following the ICV infusion, rats were euthanized and their PVN PICs expression was measured. The results showed that central administration of hypertonic saline in SD rats significantly increased the expression of PICs including TNF-α, IL-6, and IL-1ß, as well as neuronal activation marker Fra1, compared to isotonic NaCl controls and osmolarity controls. Finally, we tested whether the increase in PICs expression occurred in neurons. Incubation of hypothalamic neurons with 10 mM NaCl in a culture medium for 6 h elicited significant increases in TNF-α, IL-6, and Fra1 mRNA levels. These observations, coupled with the important role of PICs in modulating neuronal activity and stimulating vasopressin release, suggest that HS intake induces an inflammatory state in the PVN, which, may in turn, augments sympathetic nerve activity and vasopressin secretion, contributing to the development of salt sensitive hypertension.