Role and Mechanism of Chronic Restraint Stress in Regulating Energy Metabolism and Reproductive Function Through Hypothalamic Kisspeptin Neurons

Abstract Background: Stress activates the hypothalamic-pituitary-adrenal (HPA) axis, affecting energy homeostasis and reproductive function. The hypothalamic Kisspeptin neurons might be a new important central target in stress affecting energy metabolism and reproductive function.The aim of this study is to investigate whether stress affected energy metabolism and reproductive function through the glucocorticoid receptor on Kisspeptin neurons in the hypothalamus. Methods: There were four groups, that were control group, chronic restraint stress group (stress group), Kisspeptin specific glucocorticoid receptor knock out group (KGRKO group) and KGRKO+stress group. Body weight, food intake, the estrous cycle of female mice, serum sex hormone levels, serum corticosterone and prolactin, Kisspeptin expression in the hypothalamus were measured. Results: The restraint stress group showed a significant weight loss compared with the control group. Compared with the restraint stress group, the KGRKO+restraint stress group had a reduced weight loss, suggesting that restraint stress might partially affect the energy metabolism through GR on Kisspeptin neurons. In terms of reproductive function, the restraint stress group and the KGRKO+restraint stress group showed missing pre-estrus period or prolonged estrous cycles. Serum LH and FSH in KGRKO + restraint stress group decreased significantly compared with KGRKO group. However, no significant difference in the level of serum testosterone was observed. After restraint stress, the levels of serum cortisol and prolactin in male and female mice were significantly higher than the control group, and the hypothalamus Kiss1 gene mRNA expression and Kisspeptin protein expression were significantly decreased. Conclusion: Chronic restraint stress induced weight loss in mice. Chronic restraint stress played a negative role in regulating reproductive function. The effects of chronic restraint stress on energy metabolism and reproduction were partially mediated by glucocorticoid receptor on Kisspeptin neurons in the hypothalamus.

Download Full-text

Role and Mechanism of Chronic Restraint Stress in Regulating Energy Metabolism and Reproductive Function Through Hypothalamic Kisspeptin Neurons

Journal of the Endocrine Society ◽

10.1210/jendso/bvab048.1124 ◽

2021 ◽

Vol 5 (Supplement_1) ◽

pp. A551-A552

Author(s):

Yinqiong Huang ◽

Junping Wen ◽

Gang Chen

Keyword(s):

Weight Loss ◽

Body Weight ◽

Energy Metabolism ◽

Restraint Stress ◽

Reproductive Function ◽

Serum Cortisol ◽

Control Group ◽

Chronic Restraint Stress ◽

Stress Group ◽

Female Mice

Abstract Objectives: The purpose of this study was to investigate whether stress affected energy metabolism and reproductive function through the glucocorticoid receptor (GR) on Kisspeptin neurons in the hypothalamus. Methods: 1. The first part of this study focused on the effects of chronic restraint stress on energy metabolism and reproductive function in mice. 2. The second part of this study focused on the effects of chronic restraint stress on the expression of serum cortisol and prolactin and the expression of GR and Kiss1 genes in the hypothalamus.3. Based on the above research, we constructed Kisspeptin specific glucocorticoid receptor knock out (KGRKO) mice. The mice were subjected to restraint stress intervention, the body weight of the mice was monitored every day, and the vaginal smear was performed on the female mice to observe the estrous cycle. After 28 days of restraint stress, serum was collected to detect serum sex hormone levels, including LH, FSH and estrogen levels in female mice and testosterone level in male mice by ELISA. Results: 1.The body weight and energy intake of the restraint stress group were lower than those of the control. 2.After restraint stress, the female mice showed irregular estrous cycle. LH and FSH levels in the stress group were significantly decreased compared with the control group.3. After restraint stress, the levels of serum cortisol and prolactin in male and female mice were significantly higher than the control group, and the hypothalamus Kiss1 gene mRNA expression and Kisspeptin protein expression were significantly decreased.4. In female mice, compared with GRflox/flox/Kiss1Cre-control mice, the nadir serum cortisol increased significantly in the morning in GRflox/flox/Kiss1Cre+ KGRKO mice, however, there was no significant difference in the serum peak cortisol in the afternoon. Compared with the control group, the restraint stress group and the KGRKO + restraint stress group had a significant weight loss compared with the KGRKO group. Compared with the restraint stress group, the KGRKO+restraint stress group had a reduced weight loss, suggesting that restraint stress might partially affect the energy metabolism through GR on Kisspeptin neurons. Conclusions: Chronic restraint stress induced weight loss in mice and reversed body weight gain induced by high-fat diet. Chronic restraint stress played a negative role in regulating reproductive function. The effects of chronic restraint stress on energy metabolism and reproduction were partially mediated by GR on Kisspeptin neurons in the hypothalamus. This study provided a central mechanism for chronic restraint stress in affecting energy metabolism and reproductive function, and was of great significance in revealing the relationship between the HPA axis with energy metabolism and reproduction.

Download Full-text

The effect of ethanol extract of pegagan (Centella asiatica) on the performance of Wistar rats after restraint stress

Biofarmasi Journal of Natural Product Biochemistry ◽

10.13057/biofar/f080102 ◽

2010 ◽

Vol 8 (1) ◽

pp. 11-16

Author(s):

BAARID LUQMAN HAMIDI ◽

SAMIGUN SAMIGUN ◽

ANIK LESTARI

Keyword(s):

Restraint Stress ◽

Wistar Rats ◽

Radial Maze ◽

Centella Asiatica ◽

Ethanol Extract ◽

The Body ◽

Control Group ◽

Error Type ◽

Stress Group ◽

Significant Difference

Hamidi BL, Samigun, Lestari A. 2010. The effect of extract ethanol of pegagan (Centella asiatica) on the performance of Wistar rats after restraint stress. Biofarmasi 8: 11-16. The aim of this research was to investigate the effects of extract ethanol of pegagan (Centella asiatica) after treated with restraint stress by measuring the eight arms radial maze performance of rats. Pre-test and post-test controlled groups design was applied in this research. Male Wistar rats (Rattus novergicus) with the mean age of 8 weeks and the body weight of 150-200 grams which used for sample were divided randomly into 4 groups, each group consisted of 6 rats, i.e. (i) control group (without any treatment), (ii) stress group (it was given by restraint stress for 2 hours/day for each rat), (iii) pegagan group (it was given by 0.3 mg/g BW/day/rat extract ethanol of pegagan), and (iv) pegagan and stress group (it was given by 0.3 mg/g BW/day/rat extract ethanol of pegagan and restraint stress for 2 hours/day for each rat). The treatments were given for 21 days. Within 12 days for each pre-treatment and post-treatment, a test on the eight arm radial maze was conducted on individual rat to observe its performance. The assessment of rat performance in the eight arms radial maze test was conducted based on error type B. Kruskal-Wallis and Mann-Whitney tests with SPSS for Windows 16 version were applied to analyze statistically the difference between four groups. Kruskal-Wallis test was used to show the significant performance level difference between four groups of rats with p=0.001, while Mann-Whitney test was used to determine the significant difference between stress group and pegagan group (p=0.001), also stress group and pegagan and stress group (p=0.001). The result of research showed that there was no significant difference between control group and stress group (p=0.051), control group and pegagan group (p=0.143), control group and pegagan and stress group (p=0.143), also pegagan group and pegagan and stress group (p=0.952). It was concluded that extract ethanol of pegagan improved the performances of rats on the eight arms radial maze after treated with restraint stress.

Download Full-text

Decreased luteinizing hormone pulse frequency is associated with elevated 24-hour ghrelin after calorie restriction and exercise in premenopausal women

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00360.2012 ◽

2013 ◽

Vol 304 (1) ◽

pp. E109-E116 ◽

Cited By ~ 13

Author(s):

Jennifer L. Scheid ◽

Mary Jane De Souza ◽

Brenna R. Hill ◽

Heather J. Leidy ◽

Nancy I. Williams

Keyword(s):

Weight Loss ◽

Luteinizing Hormone ◽

Premenopausal Women ◽

Reproductive Function ◽

Pulse Frequency ◽

Energy Deficit ◽

Control Group ◽

Supervised Exercise ◽

Lh Pulsatility ◽

Change In Mean

Elevated ghrelin has been shown to be associated with reduced luteinizing hormone (LH) pulsatility in Rhesus monkeys, rats, men, and recently women. We previously reported that 24-h ghrelin concentrations are elevated in women following a 3-mo exercise and diet program leading to weight loss. We investigated whether the elevations in ghrelin following an ∼3-mo exercise and diet program leading to weight loss are associated with a decrease in LH pulsatility. The nonexercising control group (Control, n = 5) consumed a controlled diet that matched energy needs, whereas energy intake in the exercise group (Energy Deficit, n = 16) was reduced from baseline energy requirements and supervised exercise training occurred five times per a week. Significant decreases in body weight (−3.0 ± 0.6 kg), body fat (−2.9 ± 0.4 kg) and 24-h LH pulse frequency (−0.18 ± 0.08 pulses/h), and a significant increase in 24-h mean ghrelin were observed in only the Energy Deficit group. The pre-post change in LH pulse frequency was negatively correlated with the change in mean 24-h ghrelin ( R = −0.485, P = 0.030) and the change in peak ghrelin at lunch ( R = −0.518, P = 0.019). Interestingly, pre-post change in night LH pulse frequency was negatively correlated with the change in mean day ghrelin ( R = −0.704, P = 0.001). Elevated total ghrelin concentrations are associated with the suppression of LH pulsatility in premenopausal women and may play a role in the suppression of reproductive function following weight loss.

Download Full-text

Hyperthermia Affects Immune and Oxidative Stress Indices of Immune Organs of Broilers by Changing the Expressions of ABCG2, SVCT-2 and MCU

10.21203/rs.3.rs-936596/v1 ◽

2021 ◽

Author(s):

Xiuheng Xue ◽

Chunhuan Ren ◽

Luping Wang ◽

Mengzhu Xu Xu ◽

Caiyun Fan ◽

...

Keyword(s):

Oxidative Stress ◽

Heat Stress ◽

Immune Function ◽

Immunoglobulin M ◽

Control Group ◽

Rt Pcr ◽

Stress Group ◽

Immune Organs ◽

Significant Difference ◽

And Oxidative Stress

Abstract Background: As global temperatures rise, heat stress has become one of the major environmental stressors in the poultry industry. The purpose of the study was to investigate the effects of heat stress on immune function and oxidative stress, and further reveal the possible mechanisms of oxidative stress induced by heat stress for thymus and spleen of broilers. Methods: At the age of 28 days, thirty broilers were randomly divided into the control group (25 ± 2°C; 24 h/day) and the heat stress group (36 ± 2°C; 8 h/day); the experience was lasted for 1 week. At the end of the experience, the broilers per group were respectively euthanized and collected some samples, then to be analyzed. Results: The results showed that the levels of heat shock proteins 70 (HSP70,P＜ 0.01), corticosterone (CORT,P＜ 0.01), the contents of malondialdehyde (MDA, P＜ 0.05), interleukin-6 (IL-6, P＜ 0.01) and tumor necrosis factor-alpha (TNF-α, P＜ 0.01) in serum were significantly higher in heat stress group than that in the control group; The activities of total antioxidant capacity (T-AOC), glutathione peroxidase (GSH-Px) and superoxide dismutase (SOD) and contents of glutathione (GSH) in heat stress group significantly reduced (P＜ 0.05) in serum. Compared with the control group, the birds subjected to heat stress reduced the weight (P＜ 0.01) and the indices of thymus (P＜ 0.01), the activities of T-AOC (P＜ 0.01) and SOD (P＜ 0.05) of spleen, and levels of IL-10 (P＜ 0.05) and the GSH-PX (P＜ 0.05) in thymus and spleen, and increased the IL-6 content of thymus (P＜ 0.05), the MDA content (P＜ 0.01), and the reactive oxygen species (ROS) levels (P＜ 0.01) in thymus and spleen. Moreover, the expression of immunoglobulin G (IgG) gene in thymus and spleen of heat stressed broiler significantly increased by reverse transcription-polymerase chain reaction (RT-PCR) and real time RT-PCR (qRT-PCR; P＜ 0.05); However, the expression of immunoglobulin M (IgM) gene in spleen significantly increased (P＜ 0.05), and had no significant difference (P＞ 0.05) in thymus of heat-stressed broiler. Furthermore, the relative expression of ATP binding cassette subfamily G member 2 (ABCG2) in thymus and spleen (P＜ 0.05), sodium dependent vitamin C transporter-2 (SVCT-2, P＜ 0.01) and mitochondria calcium uniporter (MCU, P＜ 0.01) mRNA in thymus of heat stressed broilers significantly increased; and the expression of ABCG2 (P＜ 0.05), SVCT-2 (P＜ 0.01) and MCU (P＜ 0.01) protein of thymus and spleen in the heat-stressed broiler increased significantly compared with the control group. Conclusions: In summary, the study confirmed that heat stress caused oxidative stress to immune organs of broilers, further reduced immune function. Moreover, the potential mechanisms of heat stress-induced oxidative stress for thymus and spleen was further reveal in broilers.

Download Full-text

Weight and Glycemic Control Outcomes of Bariatric Surgery and Pharmacotherapy in Patients With Melanocortin-4 Receptor Deficiency

Frontiers in Endocrinology ◽

10.3389/fendo.2021.792354 ◽

2022 ◽

Vol 12 ◽

Author(s):

Esphie Grace Fodra Fojas ◽

Saradalekshmi Koramannil Radha ◽

Tomader Ali ◽

Evan P. Nadler ◽

Nader Lessan

Keyword(s):

Bariatric Surgery ◽

Weight Loss ◽

Glycemic Control ◽

Therapeutic Interventions ◽

Control Group ◽

Short Term ◽

Melanocortin 4 Receptor ◽

Significant Difference

BackgroundMelanocortin-4 receptor (MC4R) mutations are the most common of the rare monogenic forms of obesity. However, the efficacy of bariatric surgery (BS) and pharmacotherapy on weight and glycemic control in individuals with MC4R deficiency (MC4R-d) is not well-established. We investigated and compared the outcomes of BS and pharmacotherapy in patients with and without MC4R-d.MethodsPertinent details were derived from the electronic database among identified patients who had BS with MC4R-d (study group, SG) and wild-type controls (age- and sex-matched control group, CG). Short- and long-term outcomes were reported for the SG. Short-term outcomes were compared between the two groups.ResultsSeventy patients were screened for MC4R-d. The SG [six individuals (four females, two males); 18 (10–27) years old at BS; 50.3 (41.8–61.9) kg/m2 at BS, three patients with homozygous T162I mutations, two patients with heterozygous T162I mutations, and one patient with heterozygous I170V mutation] had a follow-up duration of up to 10 years. Weight loss, which varied depending on mutation type [17.99 (6.10–22.54) %] was stable for 6 months; heterogeneity of results was observed thereafter. BS was found superior to liraglutide on weight and glycemic control outcomes. At a median follow-up of 6 months, no significant difference was observed on weight loss (20.8% vs. 23.0%, p = 0.65) between the SG and the CG [eight individuals (four females, four males); 19.0 (17.8–36.8) years old at BS, 46.2 (42.0–48.3) kg/m2 at BS or phamacotherapeutic intervention]. Glycemic control in patients with MC4R-d and Type 2 diabetes improved post-BS.ConclusionOur data indicate efficacious short-term but varied long-term weight loss and glycemic control outcomes of BS on patients with MC4R-d, suggesting the importance of ongoing monitoring and complementary therapeutic interventions.

Download Full-text

The effects of kumiss supplementation on body weight, food intake, nesfatin-1 and irisin levels in BALB/C mice

Veterinarski arhiv ◽

10.24099/vet.arhiv.0928 ◽

2020 ◽

Vol 90 (6) ◽

pp. 627-636

Author(s):

Canan Gulmez ◽

◽

Onur Atakisi

Keyword(s):

Weight Loss ◽

Food Intake ◽

Live Weight ◽

Feed Consumption ◽

Control Group ◽

Male Mice ◽

Tissue Samples ◽

Bacterial Colony ◽

Metabolic Functions ◽

Significant Difference

The aim of this study was to investigate the plasma and tissue levels of nesfatin-1 and irisin hormones, which were discovered in recent years and are associated with endocrine and metabolic functions, in kumiss-supplemented mice. Sixteen BALB/C male mice were divided into two groups as control and kumiss groups. During the experiment, the kumiss was added to the drinking water of mice at a ratio of 1:1 to obtain a daily 2×108 cfu/mL bacterial colony, and was given once a day orally for 20 weeks. The weights and food intake of the animals were monitored during the experiment. The nesfatin-1 and irisin levels in plasma and tissue samples were determined using ELISA kits. Kumiss supplementation reduced the live weight for 2-12 weeks (P<0.05). However, no significant difference was observed after the 12th week. The feed consumption of the kumiss group was lower at the beginning and the 10th week, and at the end, compared to the control group (P<0.05). The plasma levels of nesfatin-1 and irisin (P<0.001) decreased while the liver levels increased (P<0.05 and P<0.001, respectively). The results indicate that plasma and liver levels of nesfatin-1 and irisin are regulated by diet and are effective in weight loss and food intake.

Download Full-text

The Effect of Choline Alphoscerate on Non spatial memory and Neurogenesis in a Rat Model of Dual Stress

10.1101/2020.06.16.154310 ◽

2020 ◽

Author(s):

Hyo Jeong Yu ◽

Min Jung Kim ◽

Jung Mee Park ◽

So Young Park ◽

Shi Nae Park ◽

...

Keyword(s):

Cognitive Function ◽

Restraint Stress ◽

Memory Function ◽

Treated Group ◽

Control Group ◽

Bdnf Expression ◽

Object Recognition Test ◽

Stress Group ◽

White Band ◽

Immature Cells

AbstractCholine alphoscerate (α-GPC) is a choline-based compound and acetylcholine precursor commonly found in the brain; it has been known to be effective in treating neuronal injury and increasing the levels of acetylcholine (Ach) and brain-derived neurotrophic factor (BDNF) which in turn enhances memory and cognitive function. This study was designed to establish rat models of dual stress using noise and restraint in order to investigate the effect of α-GPC on cognitive function and neurogenesis after dual stress. The rats were randomly divided into four groups as follows: a control group (CG), a control with α-GPC group (CDG), a noise-restraint stress group (NRSG), and a noise-restraint stress with α-GPC group (NRSDG). Two experimental groups were exposed to the double stress stimuli of noise and restraint, which involved 110dB sound pressure level (SPL) white band noise and restraint at the same time for 3 hours/day for 7 days. While the CG and NRSG received saline, the CDG and NRSDG received α-GPC (400mg/kg) orally after stress exposure. The α-GPC–treated group showed increased memory function compared to the dual stress group in the novel object recognition test. In analysis of the hippocampus, the α-GPC–treated group showed greater Choline acetyltransferase (ChAT) and BDNF expression compared to the dual stress group. The α-GPC–treated group showed significantly increased neuroblast expression compared to the dual stress group, which suggests that α-GPC enhances BDNF expression and protects the activity of the immature cells at the dentate gyrus. Our results suggest that α-GPC treatment can protect cognitive function and neurogenesis in a dual stress model.

Download Full-text

Intraventricular T3 reverses chronic restraint stress-induced depressive-like behaviors: Inhibition of NF-κB/ NLRP3 inflammasome pathway in the hippocampus

10.21203/rs.3.rs-226593/v1 ◽

2021 ◽

Author(s):

Tahmineh Mokhtari ◽

AymanEl-Meghawry El-Kenawy ◽

Li Hu

Keyword(s):

Nlrp3 Inflammasome ◽

Restraint Stress ◽

Male Rats ◽

Control Group ◽

Therapeutic Effects ◽

Chronic Restraint Stress ◽

Model Group ◽

Nissl Staining ◽

Caspase 1 ◽

Ca1 Region

Abstract In this study, the effects of triiodothyronine (T3) were evaluated on the NLR family pyrin domain containing 3 (NLRP3) inflammasome complex formation in the rat's hippocampus with restraint stress-induced depressive-like behaviors.Thirty-six Wistar male rats were randomly allocated to following groups: Control, Model, and Model + T3. In the Model or Model+T3 group, a single dose of PBS or T3 was administered into the lateral ventricle. Depressive-like behaviors were induced by chronic restraint stress. The forced swimming (FST), tail suspension (TST), and open field (OFT) tests were used to investigate the depression. The rats were sacrificed, and brain tissues were stored for molecular and pathological evaluations. Chronic stress increased the immobility of rats in the Model group according to FST, TST, and OFT (P < 0.05). T3 significantly improved depressive-like behaviors (P < 0.05). The gene expression and protein level of hippocampal nuclear factor kappa B (NF-κB), NLRP3, apoptosis-associated speck-like protein (ASC), and Caspase-1 significantly increased in the Model group compared to the control group (P < 0.05). The reduced hippocampal levels of NF-κB, NLRP3, ASC, and Caspase-1 were observed in the T3 group compared to the Model group (P < 0.05). The Nissl staining of the CA1 region showed an increased number of dark neurons (P < 0.05) and reduced pyramidal layer thickness (P < 0.05) in the Model group. These histopathological alterations were changed by T3 administration compared to the Model group (P < 0.05). The findings confirmed the therapeutic effects of intraventricularly T3 on depressive-like behaviors induced by restraint stress via surviving pyramidal neurons of the CA1 region and inhibition of NF-κB/NLRP3 inflammasome pathway.

Download Full-text

Identification of Plant Soot as Novel Safe Feed Additive: Evaluation of 90-Day Oral Toxicity and Prenatal Developmental Toxicity in Rats

Frontiers in Veterinary Science ◽

10.3389/fvets.2020.610627 ◽

2020 ◽

Vol 7 ◽

Author(s):

Xingyao Pei ◽

Xilong Xiao ◽

Jianling Liao ◽

Linfu Ke ◽

Daowen Li ◽

...

Keyword(s):

Bacterial Infections ◽

Developmental Toxicity ◽

Reproductive Function ◽

Feed Additive ◽

Control Group ◽

Oral Toxicity ◽

Subchronic Toxicity ◽

Pregnant Rats ◽

Significant Difference ◽

Effect Level

Plant soot, as a novel feed additive, could not only improve digestive function but also adsorb mycotoxins and inhibit bacterial infections. The subchronic toxicity and prenatal developmental effects of plant soot were studied for the first time. Our results indicated that there was no subchronic toxicity in the range of 2,000–50,000 mg/kg plant soot added in the feed, and there was no significant difference in reproductive function, embryo development, and teratogenicity between the pregnant rats exposed to 312.5, 1,250, and 5,000 mg/kg plant soot and the control group. The maximum no-observed effect level (NOEL) of supplemental dosage in feed could be set to 50,000 mg/kg, and the maximum intragastric NOEL could be set to 5,000 mg/kg, which preliminarily provided guidance on daily additive amount or clinical protocols for plant soot, as well as promoting the development and application of this harmless antibiotic substitutes.

Download Full-text

Prenatal Dexamethasone Exposure Increases the Susceptibility to Autoimmunity in Offspring Rats by Epigenetic Programing of Glucocorticoid Receptor

BioMed Research International ◽

10.1155/2016/9409452 ◽

2016 ◽

Vol 2016 ◽

pp. 1-9 ◽

Cited By ~ 6

Author(s):

Yanhong Sun ◽

Xiaoyan Wan ◽

Juan Ouyang ◽

Renfeng Xie ◽

Xueping Wang ◽

...

Keyword(s):

Glucocorticoid Receptor ◽

Mononuclear Cells ◽

Methylation Status ◽

Saline Control ◽

Control Group ◽

Long Term Effects ◽

Peripheral Blood Mononuclear ◽

Glucocorticoid Response ◽

Significant Difference ◽

Exon 1

Objective. Prenatal glucocorticoids (GC) can induce long term effects on offspring health. However, reports and related studies regarding the prolonged effects of prenatal GC on the development of autoimmunity are limited. Here, we aimed to explore the immunological effects of dexamethasone (DEX) exposure on young adults and whether glucocorticoid receptor (GR) is involved in this process. Methods. Wistar rats were given DEX during pregnancy. Susceptibility to autoimmunity in offspring was assessed using experimental autoimmune encephalomyelitis (EAE) and adjuvant-induced arthritis (AIA) animal models. To reveal the possible mechanism, glucocorticoid response, GR expression, and methylation status were measured in peripheral blood mononuclear cells (PBMCs). Results. Our results showed that the DEX-treated rats had greater susceptibility to EAE (100% versus 62.5%, P<0.05) and AIA (63.6% versus 0%, P<0.05) than saline control group. Glucocorticoid response and GR expression were decreased in DEX rats. Significant difference was also found in the methylation levels of GR exon 1-10 to exon 1-11 region. Conclusions. Prenatal DEX administration increases the susceptibility to autoimmune diseases, which is potentially mediated by programming GR methylation status and glucocorticoid sensitivity.

Download Full-text