HS Rats Research Articles

Low prepulse inhibition predicts lower social interaction, impaired spatial working memory, reference memory and cognitive flexibility in genetically heterogeneous rats

The “Genetically Heterogeneous National Institutes of Health (NIHHS)” stock rat (hereafter HS) shows a wide phenotypic variation, as a result of having been derived from eight inbred rat strains. Thus, these rats may be a conceivable parallel model of a healthy human sample. In order to evaluate whether HS rats have face validity as an animal model of schizophrenia-relevant features, it should be demonstrated that they present behavioural traits that may model negative and cognitive symptoms of the disorder. Previous studies on HS rats have shown that prepulse inhibition (PPI, a measure of sensorimotor gating processes), which is impaired in schizophrenic patients, is correlated with their working memory performance. In this study, we evaluated whether low PPI in the HS stock rat predicts impairments of spatial working memory (SWM), spatial reference memory and cognitive flexibility in the Morris water maze (MWM) test, and we evaluated HS rats for social interaction (SI) in a social investigation task. HS rats were stratified into 2 different groups according to their PPI scores, i.e. low- and high-PPI. In the SI task, low-PPI rats showed decreased social behaviour compared to high-PPI rats. In addition, relative to high-PPI HS rats, the low-PPI group displayed poorer SWM performance, impaired cognitive flexibility (in a reversal task) and worsened long-term spatial memory. Such differential behaviours in social and cognitive paradigms provide evidence on the face validity of low-PPI HS rats as a model of negative-like and cognitive schizophrenia-relevant traits.

AMPK Phosphorylation Impacts Apoptosis in Differentiating Myoblasts Isolated from Atrophied Rat Soleus Muscle

Regrowth of atrophied myofibers depends on muscle satellite cells (SCs) that exist outside the plasma membrane. Muscle atrophy appears to result in reduced number of SCs due to apoptosis. Given reduced AMP-activated protein kinase (AMPK) activity during differentiation of primary myoblasts derived from atrophic muscle, we hypothesized that there may be a potential link between AMPK and susceptibility of differentiating myoblasts to apoptosis. The aim of this study was to estimate the effect of AMPK activation (via AICAR treatment) on apoptosis in differentiating myoblasts derived from atrophied rat soleus muscle. Thirty rats were randomly assigned to the following two groups: control (C, n = 10) and 7-day hindlimb suspension (HS, n = 20). Myoblasts derived from the soleus muscles of HS rats were divided into two parts: AICAR-treated cells and non-treated cells. Apoptotic processes were evaluated by using TUNEL assay, RT-PCR and WB. In differentiating myoblasts derived from the atrophied soleus, there was a significant decrease (p < 0.05) in AMPK and ACC phosphorylation in parallel with increased number of apoptotic nuclei and a significant upregulation of pro-apoptotic markers (caspase-3, -9, BAX, p53) compared to the cells derived from control muscles. AICAR treatment of atrophic muscle-derived myoblasts during differentiation prevented reductions in AMPK and ACC phosphorylation as well as maintained the number of apoptotic nuclei and the expression of pro-apoptotic markers at the control levels. Thus, the maintenance of AMPK activity can suppress enhanced apoptosis in differentiating myoblasts derived from atrophied rat soleus muscle.

The Chinese Herbal Formula Huoxiang Zhengqi Dropping Pills Prevents Acute Intestinal Injury Induced by Heatstroke by Increasing the Expression of Claudin-3 in Rats.

Intestinal injury has been regarded as an important causative factor for systemic inflammation during heatstroke, and maintaining intestinal integrity has been a potential target for the prevention of HS. Huoxiang Zhengqi Dropping Pills (HZPD) is a modern preparation of Huoxiang Zhengqi and widely used to prevent HS. The present study aims to explore the protective effect of HZDP on intestinal injury during heatstroke and analyze its potential pharmacodynamic basis. Male rats in the control and HS groups were given normal saline, and those in the HZDP groups were given HZDP (0.23, 0.46, and 0.92 g/kg) before induction of HS. Serum contents of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), intestinal fatty acid-binding protein (iFABP), and diamine oxidase (DAO) were determined using ELISA. Histopathology of intestinal injury was observed following H&E staining. The expression of claudin-3 was determined using western blot, immunohistochemistry, and immunofluorescence techniques. Moreover, network pharmacological tools were used to analyze the potential pharmacodynamic basis and the mechanism of HZDP. Treatment with HZDP significantly prolonged the time to reach Tc. Compared with the control group, the contents of TNF-α, IL-6, iFABP, and DAO in HS rats increased markedly. HZDP treatments reduced these levels significantly, and the effects in the middle dose group (0.46 g/kg) were most obvious. HZDP also attenuated intestinal injury and significantly reversed the decrease in claudin-3 expression. Bioinformatics analysis suggested that 35 active ingredients and 128 target genes of HZDP were screened from TCMSP and 93 target genes intersected with heatstroke target genes, which were considered potential therapeutic targets. TNF-α and IL-6 were the main inflammatory target genes of HZDP correlated with HS. These results indicated that HZDP effectively protected intestinal barrier function and prevented acute intestinal injury by increasing the expression of claudin-3 in rats, eventually improving heat resistance.

Effect of enhanced muscle tone on the expression of atrogenes and cytoskeletal proteins during postural muscle unloading

Skeletal muscle unloading leads to the decreased electrical activity and decline of muscle tone. Aims: Current study evaluated the effect of muscle tone preservation achieved by tetanus toxin (TeNT) treatment on signaling pathways regulating atrophic processes during unloading. Main methods: Four groups of rats were used: non-treated control (C), control rats with TeNT administration (CT), 7 days of unloading/hindlimb suspension with placebo (HS), and 7 days of unloading with TeNT administration (HST). Key findings: Absolute and relative force of tetanic contractions was decreased by 65% in soleus muscle of HS rats when compared with C. Treatment with TeNT significantly lessened force decline in soleus muscle of HST rats when compared with HS. TeNT administration increased myosin heavy chain I beta (MyHC Iβ) expression in CT rats and prevented MyHC Iβ loss in HST group when compared with C rats. Desmin content was lower by 31.4% (p < 0.05) in HS group when compared with HST. Calpain-1 expression was increased in HS group when compared with C, CT and HST. There was a decrease in p-p70S6K content (41%, p < 0,05) and an increase in p-eEF2 content (77%, p < 0,05) in HS group when compared with C, while there were no significant differences in the content of these proteins between HST, CT and C groups. Significance: Treatment with TeNT significantly diminished unloading-induced decline of soleus muscle mass and mechanical properties and affected the regulation of MyHC Iβ expression. These effects are mediated by signaling pathways regulating protein synthesis and degradation.

SK Channel Dysfunction in the Hypothalamic Paraventricular Nucleus Contributes to Sympathoexcitation in Dahl Salt‐Sensitive Rats

Salt‐sensitive hypertension is characterized by marked activation of the sympathetic nervous system. Previous studies have consistently demonstrated that increased excitability or activity of pre‐sympathetic neurons in the hypothalamic paraventricular nucleus (PVN) contribute to the increased blood pressure and sympathetic nerve activity (SNA) in salt‐sensitive hypertension; however, the underlying mechanisms have not been fully elucidated. Small conductance calcium‐activated potassium (SK) channels act as negative feedback regulators of neuronal excitability in PVN neurons, and inhibition of SK channel function in the PVN is known to augment SNA and arterial blood pressure. In the present study, we tested the hypothesis that reduced SK channel function in the PVN contributes to sympathoexcitation in Dahl salt‐sensitive rats fed a high salt (HS, 4%) diet compared to a normal salt (NS, 0.4%) diet. Sympathoexcitatory responses to bilateral microinjection of the SK channel blocker apamin into the PVN were assessed in anesthetized 12–14‐week‐old male Dahl salt‐sensitive rats fed a high salt (n=7) or a normal salt (n=6) diet for 4 weeks. Baseline mean arterial pressure (MAP) was significantly elevated in rats fed a HS diet compared to NS (HS 129±4 vs. NS 114±4 mmHg, p&lt;0.05), however baseline heart rate (HR) was not different between groups (HS 411±10 vs. NS 385±13 beats/min, p=0.07). Bilateral microinjection of apamin (12.5pmol, 50nL) into the PVN significantly increased splanchnic SNA in both HS and NS rats compared to baseline, however responses were significantly attenuated in HS rats compared to NS (HS 97±22% vs. NS 249±57%, p&lt;0.01). In contrast, increases in MAP in response to PVN microinjection of apamin were similar between HS and NS rats (HS 24±4 vs. NS 29±4 mmHg, p=0.25), whereas HR responses were variable but not significantly different between groups (HS ‐8.5±9 vs. NS 7.5±7 beats/min, p=0.11). Next, we utilized qPCR to assess mRNA expression of SK1, SK2, and SK3 channels in PVN tissue from HS (n=5) and NS (n=6) rats. Interestingly, there was no significant difference in mRNA levels of either SK1, SK2, or SK3 channels between HS and NS fed rats. In summary, increases in splanchnic SNA due to PVN SK channel blockade with apamin were significantly attenuated in Dahl salt‐sensitive rats fed a HS diet, whereas there was no difference in SK channel expression between rats fed a HS vs. NS diet. These results suggest that dysfunction of SK channels in the PVN, but not decreased expression, contributes to sympathoexcitation Dahl salt‐sensitive rats fed a HS diet.

Low level transcutaneous vagus nerve stimulation reverses cardiac dysfunction and changes left ventricular gene expression in a rat model of heart failure with preserved ejection fraction

Abstract Introduction Heart failure with preserved ejection fraction (HFpEF) has become a major public health concern, accounting for approximately half of all heart failure patients. Despite normal or near normal left ventricular ejection fraction, HFpEF is associated with increased morbidity and mortality. The complex pathophysiology of HFpEF is not yet fully understood. Inflammation and fibrosis play a central role in the development of HFpEF. We have previously shown that low-level transcutaneous vagus nerve stimulation (LLTS) is anti-inflammatory. The goal of this study was to determine the effect of chronic intermittent LLTS on cardiac fibrosis, diastolic dysfunction, and left ventricular (LV) gene expression in a rat model of HFpEF. Methods Dahl salt-sensitive (DSS) rats were randomized into high salt (HS, 8% NaCl) or low salt (LS) diet (0.3% NaCl) at 7 weeks of age. The animals were monitored for the development of HFpEF on a daily basis. After 6 weeks of LS or HS diets, HS rats were randomized into 4 groups: HS active LLTS (n=50), HS sham LLTS (n=48), HS plus olmesartan (n=14) and HS active LLTS plus methyllycaconitine (MLA) (n=36), a specific blocker of the α7-nicotinic acetylcholine receptor (a7nAChR), which mediates the anti-inflammatory effects of LLTS. Stimulation was delivered for 30 min daily (20Hz, 3mA) for 4 weeks. Echocardiography was performed at 13 weeks (baseline) and 17 weeks (endpoint). At endpoint, LV histology and gene expression were examined. Results After 6 weeks of diets, HS rats developed hypertension and LV hypertrophy compared to LS rats. At endpoint, LLTS significantly attenuated blood pressure elevation, prevented the deterioration of diastolic function and improved LV circumferential strain, compared to the HS sham LLTS group. LV fibrosis was decreased in the HS active LLTS compared to HS sham LLTS group. MLA attenuated this effect, suggesting that the anti-inflammatory effect of LLTS is necessary to prevent fibrosis in this model (Figure 1). RNA-seq analysis revealed that LLTS significantly changed the expression of genes involved in mitochondrial dysfunction, sitruin signaling pathway, and oxidative phosphorylation. Conclusion Autonomic modulation with LLTS attenuates the unfavorable changes of echocardiographic parameters and LV fibrosis induced by HS diet through its anti-inflammatory effects. The results support our hypothesis that inhibiting the anti-inflammatory effect of LLTS attenuates the antifibrotic effect. The obtained data suggest that LLTS may be used clinically as a novel non-invasive neuromodulation therapy for the treatment of HFpEF. Funding Acknowledgement Type of funding sources: Public grant(s) – National budget only. Main funding source(s): National Institutes of Health (NIH)

Individual differences in addiction-like behaviors and choice between cocaine versus food in Heterogeneous Stock rats.

Recent studies reported that when given a mutually exclusive choice between cocaine and palatable food, most rats prefer the non-drug reward over cocaine. However, these studies used rat strains with limited genetic and behavioral diversity. Here, we used a unique outbred strain of rats (Heterogeneous Stock, HS) that mimic the genetic variability of humans. We first identified individual differences in addiction-like behaviors (low and high). Next, we tested choice between cocaine and palatable food using a discrete choice procedure. We characterized the individual differences using an addiction score that incorporates key features of addiction: escalated intake, highly motivated responding (progressive ratio), and responding despite adverse consequences (footshock punishment). We assessed food versus cocaine choice at different drug-free days (without pre-choice cocaine self-administration) during acquisition of cocaine self-administration or after escalation of cocaine self-administration. We also assessed drug versus food choice immediately after 1-, 2-, or 6-h cocaine self-administration. Independent of the addiction score, without pre-choice cocaine (1 or more abstinence days), HS rats strongly preferred the palatable food over cocaine, even if the food reward was delayed or its size was reduced. However, rats with high but not low addiction score modestly increased cocaine choice immediately after 1-, 2-, or 6-h cocaine self-administration. Like other strains, HS rats strongly prefer palatable food over cocaine. Individual differences in addiction score were associated with increased drug choice in the presence but not absence (abstinence) of cocaine. The HS strain may be useful in studies on mechanisms of addiction vulnerability.

Ovariectomy and high salt increase blood pressure and alter sodium transport proteins in peripheral blood mononuclear cells of adult Wistar rats.

What is the central question of this study? In a model of salt-sensitive hypertension in ovariectomized (oVx) adult Wistar rats, what is the expression of proteins related to sodium transport in peripheral blood mononuclear cells (PBMCs), and how does the response of proteins to high sodium intake compare with changes in blood pressure in intact female rats? What is the main finding and its importance? Sodium transport proteins in PBMCs react to high sodium and blood pressure markedly differently in oVx versus intact female rats. Protein expression shows sodium and pressure sensitivity. Renal immune cells increase in oVx under high salt. Hypertension is a worldwide public health problem. High sodium consumption is associated with hypertension, and hypertensive mechanisms involve immunity cells. Peripheral blood mononuclear cells (PBMCs) are endowed with proteins related to sodium transport. We studied their abundance in PBMCs from intact (IF) or ovariectomized (oVx) adult Wistar rats under normal (NS) or high (HS) salt intake. Ovariectomy was performed at 60days of life. At 145days, one group of IF and oVx rats received NS or HS intake for 5days. Another group of IF HS and oVx HS rats received hydralazine (HDZ) to reduce blood pressure (BP). Sodium balance and BP were recorded. Expression of Na+ ,K+ -ATPase (NKA), Na+ -K+ -2Cl- cotransporter 1 (NKCC1), serum/glucocorticoid-regulated kinase 1 (SGK1), dopamine D1 like receptor (D1DR), CD4+ and CD8+ were determined in PBMCs and CD45+ leukocytes in renal tissue. IF HS rats showed increased natriuresis and normal BP. NKA and CD4+ expression diminished in IF HS. Instead, oVx HS rats had sodium retention and high BP and increased the expression of NKA, NKCC1, D1DR, CD4+ and CD8+ in PBMCs. Renal CD45+ leukocytes increased in oVx HS rats. HDZ decreased BP in all rats. Upon HDZ treatment, NKA did not change, NKCC1 decreased in oVx HS rats, while SGK1 increased in both IF HS and oVx HS rats. Hormonal background determines BP response and the expression of proteins related to sodium transport in PBMCs and renal immune cells at HS intake. The analysis of NKA, NKCC1 and SGK1 expression in PBMCs differentiated salt-sensitivity from BP variations.

Brown Seaweed Sargassum siliquosum as an Intervention for Diet-Induced Obesity in Male Wistar Rats.

The therapeutic potential of Sargassum siliquosum grown in Australian tropical waters was tested in a rat model of metabolic syndrome. Forty-eight male Wistar rats were divided into four groups of 12 rats and each group was fed a different diet for 16 weeks: corn starch diet (C); high-carbohydrate, high-fat diet (H) containing fructose, sucrose, saturated and trans fats; and C or H diets with 5% S. siliquosum mixed into the food from weeks 9 to 16 (CS and HS). Obesity, hypertension, dyslipidaemia, impaired glucose tolerance, fatty liver and left ventricular fibrosis developed in H rats. In HS rats, S. siliquosum decreased body weight (H, 547 ± 14; HS, 490 ± 16 g), fat mass (H, 248 ± 27; HS, 193 ± 19 g), abdominal fat deposition and liver fat vacuole size but did not reverse cardiovascular and liver effects. H rats showed marked changes in gut microbiota compared to C rats, while S. siliquosum supplementation increased gut microbiota belonging to the family Muribaculaceae. This selective increase in gut microbiota likely complements the prebiotic actions of the alginates. Thus, S. siliquosum may be a useful dietary additive to decrease abdominal and liver fat deposition.

HDAC1 Mediates High Salt Impaired Endothelial NO Signaling

High salt diet (HS) impairment of endothelial nitric oxide (NO) signaling has been described in human and rodents. The mechanisms which mediate this dysfunction remain unknown. We and others have previously shown histone deacetylase 1 (HDAC1) decreases endothelial NO production. We therefore sought to investigate the mechanisms by which HS initiates endothelial dysfunction and if it is HDAC1-dependent. We first investigated if HS induced endothelial dysfunction is acutely regulated by a circulating factor. Male Sprague Dawley rats were fed normal salt diet (NS; 0.49% NaCl) or 2 weeks of HS (4.0% NaCl). Afferent arteriole vasoconstrictor responses to the NOS inhibitor, L-NAME, were monitored using the in vitro, blood perfused juxtamedullary nephron preparation. Arterioles from HS fed rats perfused with blood from HS fed rats demonstrated blunted constriction to L-NAME (96 ± 2% of Con) compared to arterioles from NS fed rats perfused with blood from NS fed rats (76.2 ± 2% of Con; P=0.0017). Perfusing arterioles from NS fed rats with blood HS fed rats blunted constriction to L-NAME (88 ± 1% of Con) compared to perfusing with blood from NS fed rats (P=0.0003), but not to the same extent as arterioles from HS fed rats perfused with blood from HS fed rats (P=0.0095). HS arterioles perfused with blood from NS fed rats had similar L-NAME-induced constriction (75 ± 2% of Con) compared to arterioles from NS fed rats perfused with blood from NS fed rats (P=0.9107). In short, the impaired constriction to L-NAME is dependent up on the diet of the blood donor and not the diet of the arteriole donor suggesting HS impairs endothelial function through an acute-acting circulating factor. This was not due to increased plasma arginase activity (NS: 7.96 ± 3.73 U/L vs HS: 5.42 ± 1.83 U/L P=0.5813), changes in plasma tetrahydrobiopterin (NS: 65.09 ± 11.57 pmol/ml vs HS: 53.54 ± 5.59 pmol/ml, P=0.311) or changes in plasma dihydrobiopterin (NS: 98.18 ± 7.84 pmol/ml vs HS: 87.45 ± 13.28 pmol/ml, P=0.530). We then investigated if HS increased endothelial HDAC1 activity. Renal endothelial cells were isolated from NS and HS fed rats and incubated in plasma from the same rat. HDAC1 activity was defined as the MS-275 (HDAC1 specific inhibitor) inhibitable portion of total HDAC activity. We found HS significantly increased renal endothelial HDAC1 activity (NS: 0.38 ± 0.03 pmol/min vs HS:0.94 ± 0.19 pmol/min, P=0.01). Furthermore, in vitro HDAC1 inhibition with 300 nM MS-275 restored the arteriolar vasoconstrictor responses in HS rats (79 ± 3% of Con) to values similar to NS fed rats (P=0.969) and to NS rats + MS-275 (79 ± 1% of Con; P>0.998). When plasma from HS fed rats was combined with erythrocytes from NS fed rats and perfused through arterioles from NS fed rats, constriction to L-NAME was blunted (85 ± 3% of Con; P=0.0129). Under these same conditions, HDAC1 inhibition restored constriction (75.7 ± 0.95 % of Con) similar to controls (P=0.979). We conclude that HS disruption of renal microvascular NO signaling is initiated by a circulating factor(s) that increases endothelial HDAC1 activity.

Регуляция тонуса артерий у крыс, длительно получавших высокосолевую диету

A high salt diet leads to a decrease in vascular dilatation to agonists, but the vascular mechanisms involved in this process are not extensively studied. A group of male Wistar rats at the age of 3 months was transferred to a diet containing 8% NaCl (HS) for 3 months, while the second group received a diet with a standard salt content (0.34%) (NS). At the end of the experiment, the rats were euthanized and the abdominal aorta and superior mesenteric artery (SMA) were extracted. The vascular segments were placed in a myograph and the acetylcholine (ACh) -induced relaxation of the vascular segments previously contracted with phenylephrine - was measured. A high salt diet led to a weakening of the relaxation of SMA in a calcium-free solution. In response to ACh and sodium nitroprusside, a pronounced relaxation of the vascular segments was observed, while the ACh-induced vascular relaxation of HS rats had a lower amplitude. K+ -channel blockers (TEA, TRAM-34, and apamine) weakened ACh-induced relaxation of the SMA, but not the aorta. In the SMA of HS rats the decrease in relaxation under the action of K+ -channel blockers was more significant. Inhibition of production of endogenous H2S also led to a weakening of the relaxation of the SMA segments on ACh. In SMA of HS rats, the degree of weakening of ACh-induced relaxation against the background of propargylglycine was greater than in NS rats. The data obtained in the study shows that a long-term high salt diet leads to a decrease in agonist-induced relaxation of the aorta segments and SMA. The relaxation weakening of the aorta segments and SMA occurs due to a decrease in the production of NO by the endothelium. In the SMA of HS rats, the decrease in NO-mediated relaxation is partially compensated by an increased role of EDHF in ACh-induced relaxation. The results of the study also show that one of the EDHFs in rat SMA is H2S, the role of which in SMA relaxation increases in HS rats.

Changes Depression- and Anxiety- like Behaviors following Selective Breeding for Cocaine Reinforcement

The LS and HS are rat lines selectively bred for altered cocaine self-administration. Given the importance of mental health in substance use, these lines were evaluated for putative depression- and anxiety- like behaviors through forced swimming and exploration of a plus maze. We found increases of struggling in LS males, climbing in LS females, and swimming in HS males; with biphasic effects on immobility in the HS strain. HS rats had fewer entries into and less time spent in open arms of the plus maze, consistent with greater anxiety-like behavior, which may contribute to enhanced drug taking.

Consuming sucrose solution promotes leptin resistance and site specifically modifies hypothalamic leptin signaling in rats.

Rats consuming 30% sucrose solution and a sucrose-free diet (LiqS) become leptin resistant, whereas rats consuming sucrose from a formulated diet (HS) remain leptin responsive. This study tested whether leptin resistance in LiqS rats extended beyond a failure to inhibit food intake and examined leptin responsiveness in the hypothalamus and hindbrain of rats offered HS, LiqS, or a sucrose-free diet (NS). Female LiqS Sprague-Dawley rats initially only partially compensated for the calories consumed as sucrose, but energy intake matched that of HS and NS rats when they were transferred to calorimetry cages. There was no effect of diet on energy expenditure, intrascapular brown fat tissue (IBAT) temperature, or fat pad weight. A peripheral injection of 2 mg of leptin/kg on day 23 or day 26 inhibited energy intake of HS and NS but not LiqS rats. Inhibition occurred earlier in HS rats than in NS rats and was associated with a smaller meal size. Leptin had no effect on energy expenditure but caused a transient rise in IBAT temperature of HS rats. Leptin increased the phosphorylation of signal transducer and activator of transcription 3 (pSTAT3) in the hindbrain and ventromedial hypothalamus of all rats. There was a minimal effect of leptin in the arcuate nucleus, and only the dorsomedial hypothalamus showed a correlation between pSTAT3 and leptin responsiveness. These data suggest that the primary response to leptin is inhibition of food intake and the pattern of sucrose consumption, rather than calories consumed as sucrose, causes leptin resistance associated with site-specific differences in hypothalamic leptin signaling.

Aortic Stiffness and Diastolic Dysfunction in Sprague Dawley Rats Consuming Short-Term Fructose Plus High Salt Diet.

IntroductionHigh fructose and salt consumption continues to be prevalent in western society. Existing studies show that a rat model reflecting a diet of fructose and salt consumed by the upper 20th percentile of the human population results in salt-sensitive hypertension mitigated by treatment with an antioxidant. We hypothesized that dietary fructose, rather than glucose, combined with high salt leads to aortic stiffening and decreased renal artery compliance. We also expect that daily supplementation with the antioxidant, 4-hydroxy-2,2,6,6-tetramethylpiperidine-1-oxyl (+T; Tempol), will ameliorate the increase in mean arterial pressure (MAP) and vascular changes.MethodsMale Sprague Dawley rats were studied with either 20% fructose or 20% glucose in the drinking water and normal salt (0.4%) or high salt (4%) in the chow resulting in four dietary groups: fructose normal Fru+NS or high salt (Fru+HS) or glucose with normal (Glu+NS) or high salt (Glu+HS). Tempol (+T) was added to the drinking water in half of the rats in each group for 3 weeks.ResultsMAP was significantly elevated and the glucose:insulin ratio was depressed in the Fru+HS. Both parameters were normalized in Fru+HS+T. Plasma renin activity (PRA) and kidney tissue angiotensin II (Ang II) were not suppressed in the high salt groups. Pulse wave velocity (PWV), radial ascending strain, and distensibility coefficient of the ascending aorta were significantly decreased in Fru+HS rats and improved in the Fru+HS+T rats. No differences occurred in left ventricular systolic function, but the ratio of early (E) to late (A) transmitral filling velocities was decreased and renal resistive index (RRI) was higher in Fru+HS rats; antioxidant treatment did not change these indices.DiscussionThus, short-term consumption of high fructose plus high salt diet by rats results in modest hypertension, insulin resistance, diminished aortic and renal artery compliance, and left ventricular diastolic dysfunction. Antioxidant treatment ameliorates the blood pressure, insulin resistance and aortic stiffness, but not renal artery stiffness and left ventricular diastolic dysfunction.

Macrophage Depletion Improves Endothelial Insulin Resistance and Protects against Cardiovascular Injury in Salt-Sensitive Hypertension.

Vascular endothelial insulin signaling is critical for the maintenance of vascular and metabolic homeostasis. We have previously shown that in hypertensive Dahl rats, impaired vascular insulin action is linked to angiotensin II activation of the NFκB inflammatory pathway. Macrophage polarization (M1) has implicated in hypertensive and metabolic diseases. Here, we investigated the effect of macrophage depletion using liposome-encapsulated clodronate (LEC) on endothelial insulin resistance and cardiovascular remodeling in Dahl salt-sensitive (DS) rats. High salt intake (HS) for 5 weeks increased systolic blood pressure (SBP: 192 ± 5 vs. 144 ± 4 mmHg in NS, p < 0.05), aortic and cardiac hypertrophy, cardiac fibrosis, and impaired acetylcholine- and insulin-induced vasorelaxation, accompanied by impaired insulin activation of endothelial nitric oxide synthases (eNOS)/NO signaling. HS rats had a significant increase in CD68 (a monocyte/macrophage marker) expression in the aorta and the heart. LEC reduced SBP (168 ± 5 mmHg, p < 0.05) and cardiovascular injury and improved acetylcholine- and insulin-mediated vasorelaxation and insulin signaling molecules with a reduction in the macrophage infiltration in the aorta and the heart. HS rats also manifested an increase in the aortic expressions of inflammatory cytokines, including the ratio of phosphorylated inhibitory kappa B (Iκb)/Iκb, tumor necrosis factor α, and phosphorylated c-Jun N-terminal kinase (JNK) and oxidative stress, which were reduced in HS/LEC rats. Our results suggest that in salt-sensitive hypertension, macrophage may importantly contribute to endothelial insulin resistance, vascular inflammation, and injury. These findings support the idea that macrophages may be a new target for immunotherapy of vasculopathy in hypertensive and metabolic disorders.

High Salt Intake Impairs Erectile Function in Salt-Sensitive Rats Through Mineralocorticoid Receptor Pathway Beyond Its Effect on Blood Pressure

BackgroundHigh salt intake is a risk factor for hypertension, which can potentially lead to erectile dysfunction (ED); however, the underlying pathological mechanisms remain unclear. AimTo investigate whether erectile function is directly impaired by high salt intake and whether selective inhibition of mineralocorticoid receptor (MR) could provide protection from ED. Methods6-week-old male Dahl salt-sensitive rats were randomly divided into 3 groups: normal diet (0.3% NaCl; control, n = 8), high-salt diet (8% NaCl; HS, n = 8), and high-salt diet plus eplerenone (HS + EPL, n = 11). HS + EPL rats were orally administered daily doses of EPL (75 mg/kg) for 6 weeks; control and HS rats received purified water on the same schedule. OutcomesAt the end of the study period, erectile function was evaluated by measuring intracavernosal pressure and mean arterial pressure after cavernous nerve stimulation. Serum levels of asymmetric dimethylarginine and L-arginine were determined using ultraperformance liquid chromatography–tandem mass spectrometry. Quantitative PCR was used to assess the expression of MR, inflammation, and oxidative stress markers (nicotinamide adenine dinucleotide phosphate oxidase-1/4, p22phox, interleukin-6, and superoxide dismutase-1), and protein arginine N-methyltransferase-1. ResultsThe intracavernosal pressure/mean arterial pressure ratio was significantly lower, whereas systolic blood pressure, MR expression, serum asymmetric dimethylarginine levels, oxidative stress, and levels of inflammatory biomarkers were significantly higher in HS rats than in control rats (P < .05). EPL administration significantly improved each of these parameters except systolic blood pressure and MR expression. No significant intergroup differences were observed for L-arginine and superoxide dismutase-1 levels. Clinical TranslationOur results provide a rationale for the need of salt restriction and the use of selective MR inhibitors in prophylaxis or treatment of ED in men consuming a high-salt diet. Strengths & LimitationsWe are the first to report that the adverse impact of high salt intake on erectile function is mediated via MR activation, independent of its effect on blood pressure. A major limitation of this study is that responses of salt-resistant rats were not studied. ConclusionsHigh salt intake directly impaired erectile function in Dahl salt-sensitive rats, whereas selective MR inhibition ameliorated this effect.Kishimoto T, Kataoka T, Yamamoto Y, et al. High Salt Intake Impairs Erectile Function in Salt-Sensitive Rats Through Mineralocorticoid Receptor Pathway Beyond Its Effect on Blood Pressure. J Sex Med 2020;17:1280–1287.

Can the frequency of defensive posture adoption influence the expression of active behavior of adult rats?

During the youth, rats spend a long time in play fighting (PF), and asymmetries are observed in terms of the frequency of adoption of the supine posture. This study investigated the effects of asymmetry in adopting supine posture during PF on the time spent in locomotor activity-related contextual behaviors during youth and adulthood. Male Wistar rats were divided into three groups according to the number of times of their adoption of a defensive supine (S) posture: low (LS, n = 10), intermediary (IS, n = 10) and high (HS, n = 10). Rats aged 21–62 days were placed in voluntary physical active cages (VPAC) with running wheels. The time spent in social play (SP), play fighting (PF), voluntary physical activity (VPA), spontaneous physical activity (SPA) and resting (Rest) were recorded daily. During adulthood (63–92 days of age), rats were placed individually in the VPAC, with daily quantification of VPA. The average time spent in PF was not different among the groups, but was age-dependent. LS and IS rats spent more time in VPA than the HS rats, between days 49–62 (intragroup analyses) and 72–78 (individual analyses) days of age. However, LS and IS animals spent less time in SPA than HS rats from 42–48 days of age. In conclusion, the expressions of VPA and SPA are influenced by the asymmetries in adoption of the supine posture of members within a social group, remaining even after the separation of the social groups.

The Role of Proteoglycans in Glomerular Physiology and Pathophysiology

BackgroundDiabetic kidney disease (DKD) is the leading cause of renal failure in the world. Diabetes is associated with damage to the endothelial glycocalyx (eGCX), the layer of proteoglycans (PGs) and other negatively charged molecules covering the glomerular endothelial capillaries. The eGCX restrain the flow of charged molecules, such as albumin, over the filtration barrier in the kidney. Loss of the glomerular eGCX leads to proteinuria without other visible damage to the filtration barrier, but the detailed structure and composition is still largely unknown. The aim of this study was to gain new knowledge about the composition and role of the EGCX in health and DKD.MethodsThe negatively charged PGs in the eGCX was eluted from rat kidneys using 1 M NaCl solution (high salt, HS). In addition 1 M mannitol was used as osmotic control (HO) and 0.15 M NaCl to mimic physiological salt concentrations (NS). Solutions were introduced intra‐arterially to rat kidneys under anesthesia in vivo. Venous effluent was analyzed using mass spectrometry. Fractional clearance of albumin and GFR was measured. Electron microscopy (EM) was used to investigate morphological changes. Expression of PGs and PG related genes in glomeruli from patients with DKD was investigated using deep sequencing data from a Swedish DKD cohort (DKD, n=19, controls; pre‐transplant biopsies, n=20).ResultsWe identified 15 PGs in the eluate from rats. PGs were found in highest yields in the HS samples. EM demonstrated that the eGCX thickness was significantly reduced in the HS rats −28% (p&lt;0.05) compared to NS. Rats perfused with HS had increased fractional clearance of albumin and reduced GFR, compared to NS and HO, 10 minutes after perfusion. In glomeruli from patients with DKD 12 PGs were found to be significantly regulated, and 4 of these PGs were also identified in the eGCX eluates from rats. There was an overall decrease in expression of enzymes responsible for building up PG side chains, as well as an increase in proteins involved in PG degradation.ConclusionThe eGCX itself represents a dynamic structure with significant molecular turnover. In our study, we identified several PGs novel to the glomerular eGCX. We show that loss of eGCX leads to proteinuria and reduced GFR, strongly supporting that the eGCX is important for preventing proteinuria. In glomeruli from patients with DKD we found significant alterations in the gene expression of PGs and the enzymes regulating their composition. Further investigation is needed to clarify in detail when and how these alterations appear in DKD.

Chronic exposure to liquid sucrose and dry sucrose diet have differential effects on peripheral taste responses in female rats

Sugar-sweetened beverages are the major source of added calories in the Western diet and their prevalence is associated with obesity and metabolic disruption. Despite the critical role of the taste system in determining food selection and consumption, the effects of chronic sucrose consumption on the peripheral taste system in mammals have received limited attention. We offered female Sprague Dawley rats free access to water and one of three diets for up to 40 days: (1) sucrose-free chow or “NS” diet; (2) a high-sucrose dry diet or “HS”; or (3) 30% sucrose solution and the NS diet, designated “LiqS” diet. Sucrose consumption by LiqS rats gradually increased and by day 14 was equal to that of HS rats. Food intake decreased in LiqS rats, but their energy intake remained higher than for NS or HS rats. There was no significant difference in weight gain of the groups during the study. Recordings from the chorda tympani nerve (CT), which innervates taste buds on the anterior tongue, revealed decreased responses to 1 M sucrose in both LiqS and HS rats and to acesulfame K and salt tastants in LiqS rats after 40 days on diet. Umami, bitter, and acid response magnitudes were unchanged in both groups. These results demonstrate that chronic sucrose exposure inhibits taste responses to higher concentrations of sweet stimuli. More surprisingly, CT responses to NaCl and 0.5M NaAc were significantly reduced in rats on the LiqS diet. Thus, the physical form of the diet influences taste responsiveness to salt and sweet taste function. These data suggest that taste buds are previously unappreciated targets of chronic sucrose consumption.

P3512Sex differences in rats with heart failure with preserved ejection fraction and the effect of autonomic modulation

Abstract Background Heart failure (HF) with preserved ejection fraction (HFpEF) accounts for 50% of HF and sudden death is the leading cause of mortality. There are considerable sex differences in cardiac structure and function, which may be related to outcomes in HFpEF. Transcutaneous vagus nerve stimulation (tVNS) is antiarrhythmic. Purpose To describe sex differences in mortality, autonomic tone and ECG parameters in rats with HFpEF and examine the effect of tVNS on these outcomes. Methods Dahl salt sensitive (DS) rats of either sex were randomized into high salt (HS, 8% NaCl) or low salt (LS) diet (0.3% NaCl) at 7 weeks of age. After 6 weeks of LS or HS diets, HS rats were randomized to receive active or sham tVNS, 30min daily (20Hz, 3mA) for 4 weeks. The rats were monitored daily for 4 weeks for the development of HFpEF. ECG and echocardiogram were performed at 13 weeks (baseline) and 17 weeks (endpoint). Heart rate variability (HRV) was calculated at the respective time points. ECG and HRV parameters were analyzed in a blinded fashion. Logistic regression analysis was performed to identify independent predictors of mortality. Results A total of 58 rats were included (5 male LS, 6 female LS, 22 male HS and 25 female HS). HS rats developed significant hypertension and signs of HFpEF, while 24% of females and 53% of males died (P=0.004). There were 4 sudden cardiac deaths in males (with ventricular tachycardia documented in 1 rat), whereas all the females died of HF or stroke. Corrected QT (QTc) at baseline significantly prolonged in HS compared to LS rats (250.5±14.4ms vs. 226.8±13.9ms, respectively, p=0.0007), while all other ECG parameters did not differ significantly between groups. In HS rats, QTc prolongation was significantly more pronounced in males compared to females (259.4±20.6ms vs. 243.8±14.5ms, respectively, P=0.002). In univariate analysis, prolonged baseline QTc (OR=1.04; 95% CI 1.01–1.06, p=0.003) and male sex (OR=3.21, 95% CI 1.19–8.66, p=0.016) predicted mortality. However, in multivariate analysis, QTc was the only significant predictor of mortality (OR=1.04; 95% CI 1.01–1.06, p=0.003). After 4 weeks of treatment, active tVNS significantly decreased QTc compared to sham (244.6±13.8ms vs. 255.8±14.0ms, respectively, p=0.017) in both male and female rats in a similar manner. The low frequency to high frequency ratio (LF/HF) of HRV, which reflects sympathovagal balance, was significantly decreased in active tVNS rats compared to sham (0.21±0.13 vs. 0.54±0.14, respectively; p=0.001) in both male and female rats in a similar manner. Conclusions Male rats with HFpEF exhibit worse survival compared to females and are at higher risk for sudden death. QTc prolongation accounts for the increased risk of sudden death in males compared to females. Autonomic modulation with tVNS attenuates the unfavorable changes in QTc and HRV induced by HS diet and may be used to prevent ventricular arrhythmias in patients with HFpEF.