bluuebunny/crossref_metadata_2025 huggingface dataset:
Molecular mechanisms involved in the constitutive nuclear localization of the mineralocorticoid receptor in cardiac myocytes
Authors: de la Rosa Diego Alvarez , Hernandez‐Diaz Ivan , Giraldez Teresa
Published: June 2021

The mineralocorticoid receptor (MR) is a member of the nuclear receptor family. Aldosterone activates MR in target epithelia, modulating the expression of genes that enhance sodium reabsorption. In addition, MR may have other physiological functions. Altered expression/activation of cardiac MR leads to the development of cardiac fibrosis and MR blockade is beneficial for the treatment of heart failure. However, the physiological role and activation status of MR in the heart are poorly known. Since steroid receptors are typically cytoplasmic and translocate to the nucleus upon ligand binding, we examined the subcellular localization of MR. Our results show that MR is a constitutive chromatin‐bound factor in mouse left ventricle and in HL‐1 cardiomyocytes, regardless of corticosteroid levels. MR constitutive nuclear localization depends on two nuclear localization signals (NLS0 and NLS1), which are also essential for MR activity. Overexpression of hsp90 prevents the constitutive nuclear localization of MR, suggesting that heat shock protein expression levels could be a limiting factor in mouse cardiac myocytes, shifting the equilibrium of MR localization to the nucleus. The nuclear localization of MR in cardiac myocytes may have important consequences for its function and pharmacological regulation. This work was supported by grant BFU2007‐61148 from Ministerio de Ciencia e Innovación (Spain) to DAR.
Reduced Uterine Perfusion Pressure in Pregnant Rats Impairs Cerebral Vascular Myogenic Responses
Authors: Ryan Michael J , Gilbert Emily L , Glover Porter H , LaMarca Babbette D , Granger Joey P
Published: June 2021

Women with preeclampsia are at increased risk for encephalopathies; however, the mechanisms for this are not clear. One proposed mechanism is that cerebral vascular autoregulation is impaired. Therefore, we tested whether reduced uterine perfusion pressure in pregnant rats (RUPP, an established model that closely mimics preeclampsia) impairs middle cerebral artery (MCA) responses to changes in pressure. Mean pressure was lower in normal pregnant rats (NP, 103±2 mmHg) and increased in RUPP rats (135±3 mmHg) compared with non‐pregnant controls (CTRL, 116±1 mmHg). Rats were sacrificed at day 19 of gestation and MCA function was assessed in a pressure myograph. Luminal pressure was servo‐controlled between 25 and 150 mmHg under active (1.8mMCa2+) and passive (0mMCa2+) conditions and vessel tone was calculated. The slope of the relationship between tone and pressure was 0.08±0.01 in CTRL rats. This was blunted in NP rats (0.05±0.01) and ameliorated in RUPP rats (0.01±0.00, p<0.05). Endothelial dependent and independent dilation was not different between groups. A trend towards increased brain water content (wet:dry ratio) was observed in NP (4.3±0.1) and RUPP rats (4.9±0.3) when compared to CTRL (3.9±0.2) but this was not statistically different. These data suggest that RUPP in rats impairs cerebral vascular myogenic responses and could contribute to an increased risk for encephalopathies during preeclampsia.
Regulation of sFlt‐1 and VEGF secretion by adenosine receptor signaling in rat placental villous explants
Authors: George Eric Matthew , Granger Joey P , Adair Thomas H
Published: June 2021

Recent studies have suggested that an imbalance of pro‐ and anti‐ angiogenic factors in response to placental ischemia plays an important role in the pathogenesis of preeclampsia. Adenosine signaling is a well‐established factor in the regulation of cardiovascular function, but has only recently been recognized as a regulator of the pro‐angiogenic protein VEGF. Because VEGF and the anti‐angiogenic protein sFlt‐1 are known to be secreted in response to placental hypoxia, we determined whether adenosine plays a role in the regulation of sFlt‐1 and VEGF in the placenta. Treatment of cultured rat placental villous explants with the adenosine receptor antagonist 8‐SPT had no effect on VEGF (p=0.38) or sFlt‐1 (p=0.056) secretion under normoxic (6% oxygen) conditions. However, the hypoxia (1% oxygen) induced increases in both sFlt‐1 and VEGF secretion was significantly (p<0.005 and p<0.05 respectively) attenuated by 8‐SPT. Administration of DPP, an adenosine transport inhibitor that increases extracellular adenosine, caused a significant increase in sFlt‐1 (p<0.005) under normoxic conditions, but had no effect under hypoxic conditions. These results suggest that adenosine could be an important regulator of sFlt‐1 and VEGF in the hypoxic placenta, possibly affecting the balance of these two opposing factors in response to placental ischemia.
ACUTE ANDROGEN INFUSIONS REDUCE ARTERIAL BLOOD PRESSURE BY A NONGENOMIC MECHANISM IN CONSCIOUS MALE RATS
Authors: Perusquia Mercedes , Sellers Minga M. , Greenway Clayton D. , Guedes Alonso G.P. , Stallone John N
Published: June 2021

Previously we reported that androgens produce rapid nongenomic vasorelaxation that is structurally‐specific and independent of cytosolic androgen receptor (AR). Thus, the effects of i.v. androgen infusions on mean arterial blood pressure (MAP) were studied in conscious, unrestrained, chronically‐catheterized, ganglionic‐blocked (hexamethonium, 30 mg/kg, i.p.) male Sprague‐Dawley (SD) and Testicular‐Feminized Rats (Tfm, AR‐deficient), 16–20 wks. age. MAP and heart rate (HR) were recorded at baseline and with increasing doses of androgen (0.375–6.00 μM/kg‐min, i.v.). Control rats received vehicle (10% EtOH/saline, 0.15 ml/kg‐min, i.v.). Data are means ± S.E. (n = 4–8 rats/group). In SD, baseline MAP and HR averaged 104 ± 4 mmHg and 353 ± 12 beats/min. Testosterone (TES) produced a dose‐dependent reduction in BP to a low of 87 ± 4 (Δ16%), HR was unchanged (354 ± 14). In Tfm, TES produced a similar reduction in BP (99 ± 4 to 84 ± 3, Δ15%); HR was unchanged (406 ± 15). Neither BP (109 ± 3) nor HR (395 ± 13) were altered by vehicle. In SD, 5β‐dihydrotestosterone (genomically‐inactive metabolite) produced a greater reduction in BP than TES (104 ± 2 to 80 ± 1, Δ23%); HR was unchanged (371 ± 15). These data suggest the hypotensive effect of androgens involves a direct vasodilatory action on the peripheral vasculature which, like effects observed in isolated arteries, is structurally‐specific and AR‐independent. (NIH HL‐080402)
VARIATIONS IN THE DEFICIENCY LEVELS OF ENDOGENOUS ESTRADIOL ON THE RATE OF POSTMENOPAUSAL BONE RESORPTION
Authors: Malepati Dhananjaya Naidu , Suresh M
Published: June 2021

Over a period of three years, 257 healthy postmenopausal women (mean age of 51.7 years) and 47 premenopausal women (control group, mean age of 34.23 years) were selected into the study. The postmenopausal women were divided into three estradiol‐deficient (study) groups based on the endogenous estradiol levels such as Group – I (10–25 pg/ml), Group ‐ II (>25–50 pg/ml) and Group ‐ III (>50 pg/ml). Bone formation marker (total alkaline phosphatase), bone resorption markers (tartrate‐resistant acid phosphatase, calcium, free hydroxyproline, free deoxypyridinoline and calcium/creatinine) and intact paratharmone levels were estimated in every subject. The bone formation marker and bone resorption markers were found to be elevated in all the three study groups over controls (P<0.0010) When the study groups compared among one another, the bone resorption markers were found elevated in Group‐I over Group ‐ II and III and in Group ‐ II over Group ‐ III. But, the intact paratharmone levels were found to be normal in all the groups.

This study concludes that the degree of bone resorption was high in Group ‐ III (>50 pg/ml, higher in Group – II (> 25–50 pg/ml) and highest in Group ‐ I (10–25 pg m/l) over controls. This view portends that the endogenous estradiol levels are the important determinants of bone resorption risk and also the variations in deficient endogenous estradiol levels in postmenopausal women might contribute to differences in the rate of postmenopausal bone resorption.
Ovariectomy induces an enhanced sensitivity to angiotensin II in female growth restricted rats
Authors: Ojeda Norma B , Royals Thomas P , Alexander Barbara T
Published: June 2021

Placental insufficiency in the rat results in intrauterine growth restriction (IUGR) and sex differences in adult hypertension. Male IUGR are hypertensive; ovariectomy (OVX) induces hypertension in female IUGR suggesting estradiol (E) may provide a protective status. Enhanced sensitivity to acute angiotensin II (ANG II) is observed in male IUGR, but not in intact normotensive female IUGR. Thus, the purpose of this study was to determine if OVX enhances sensitivity to ANG II in female IUGR. Female rats were subjected to OVX or sham OVX at 10 weeks of age. Cardio‐renal physiological parameters were determined before and after acute infusion of ANG II (100 ng/kg/min for 30 minutes) at 16 weeks of age in rats pretreated with enalapril (250 mg/L for 1 week). Mean arterial pressure (MAP) and renal vascular resistance (RVR) were increased to a greater extent in OVX IUGR; a greater decrease in glomerular filtration rate (GFR) was observed in OVX IUGR. Thus, these data suggest that estradiol modulates renal responsiveness to ANG II in female IUGR and may contribute to sex differences in IUGR hypertension.

Response after acute ANG II
Increase in MAP (mmHg)
Increase in RVR (mmHg/ml/min)
Decrease in GFR (ml/min)

Control female intact
21±0.5
8±3
2.3±0.3

IUGR female intact
20±9
16±7
2.3±0.3

Control female OVX
18±2
12±6
3.5±0.9

IUGR female OVX
49±4*†
17±1.5*†
5.1±0.1*†

P < 0.05 vs. control OVX

P < 0.05 vs. intact IUGR
L‐Arginine supplementation abolishes the blood pressure and endothelin response to chronic increases in plasma sFlt‐1 in pregnant rats
Authors: Murphy Sydney , Cockrell Kathy , Granger Joey P.
Published: June 2021

While soluble fms‐like tyrosine kinase (sFlt‐1) and endothelin (ET‐1) have been implicated in the pathogenesis of preeclampsia (PE), the mechanisms whereby increased sFlt‐1 leads to enhanced ET‐1 production and hypertension remain unclear. NO production is reduced in PE, however, whether a reduction in NO synthesis plays a role in increasing ET‐1 and blood pressure in response to chronic increases in plasma sFlt‐1 remains unclear. The purpose of this study was to determine the role of reduced NO synthesis is mediating the increase in blood pressure and ET‐1 in response to sFlt‐1 in pregnant rats. To achieve this goal, sFlt‐1 was infused into NP rats (3.7μg/kg/d for 6 days) and pregnant rats supplemented with 2% L‐arginine (in drinking water for 6 days). Infusion of sFlt‐1 into NP rats significantly elevated MAP (117±2 mmHg, p<0.05) versus NP (103±1 mmHg). sFlt‐1 hypertensive pregnant rats had ~3.5 fold increase in preproendothelin‐1 (PPET‐1) in the renal cortex when compared to NP rats. Supplementation with L‐arginine decreased the sFlt‐1 hypertension (108±3 mmHg, p<0.05) while having no effect on the blood pressure response in NP rats (108±2 mmHg), and abolished the enhanced sFlt‐1 induced PPET‐1 expression. In conclusion, a reduction in nitric oxide synthesis may play an important role in the enhanced ET‐1 production in response to sFlt‐1 hypertension in pregnant rats. NIH grant HL51971
Combined effects of hypoxia and endurance training: role of tissue diffusion evaluated by NIRS.
Authors: Richalet Jean‐Paul , Bourdillon Nicolas , Beaudry Michèle
Published: June 2021

Exposure to hypoxia provokes a decrease in peak oxygen consumption that is greater than expected from the decrease in arterial oxygen content (CaO2), suggesting the participation of other factors. We hypothetized that O2 transfer within the active muscle may play a role. Therefore we used Near Infra Red Spectroscopy (NIRS) to assess oxy (O2Hb) and deoxyhemoglobin (HHb) concentration in the vastus lateralis of trained athletes (TA) and untrained subjects (US) exercising at various inspired oxygen pressure (PiO2 = 149.4, 131.4, 107.3 and 87.0 mmHg). A mathematical model has been developed to compute i) the pulmonary (Kp) and muscular (Ktm) O2 diffusion coefficients and ii) the proportion of arteriolar:capillary:venous blood participating in the NIRS signal at every exercise intensity from rest to peak exercise in the normoxic and various hypoxic conditions. In TA, O2Hb decreased near maximal exercise at 2500 and 4000 m, while in US, altitude had no effect. In normoxia O2Hb was higher in TA than in US, the difference disappearing in hypoxia. Ktm increased linearly with workload and altitude and was higher in TA than US while Kp plateaued near maximal exercise, which was consistent with athletes’ greater decrease in CaO2. The greater participation of arterial blood in the NIRS signal in TA at altitudes account for their higher O2Hb values as well as the greater decrease they underwent in hypoxia. At 4000 m, athletes loose their advantages of adaptation to training due to a reduced arterial content, and both from NIRS variables and model output, characteristics of O2 transfer of TA converge toward those of US. The model was also used to predict the decrease in peak O2 consumption and the various factors involved in this decrease both in trained and untrained subjects.
The Gene Profile of the Skeletal Muscle in Response to Burn and Hindlimb Unloading
Authors: Wu Xiaowu , Xiao Yufei , Baer Lisa A , Chen Yidong , Wolf Steven E , Walters Thomas j , Wade Charles E
Published: June 2021

Hindlimb unloading (HU) and Burn (B) both cause muscle atrophy. In this study, we analyzed the gene profiles in the muscles of plantaris (PL) and soleus (SL) in rats with combined B and HU (B/HU) or B with ambulation (B/A) to determine the impact of HU on burn induced muscle atrophy. The muscles of PL and SL were collected and weighed from B/A or B/HU (40% TBSA of burn) at days (D) 1, 3 and 7 (n=5/group), and from controls (n=4). The extracted total RNA of PL and SL was reverse transcribed and hybridized to Affymetrix rat 230 2.0 chips. Significant genes were selected by the fold changes (+/−2), and adjusted p value (¡Ü0.05) versus control. We found that the weight of PL and SL from B/HU was significantly less than B/A on D3 and D7. Genes were uniquely expressed in PL and SL in response to B/A and B/HU. B/HU induced more profound and sustained gene responses than B/A (Number of genes at D1, B/A: 2277 (PL) and 410 (SL) with 258 genes overlapped; B/HU: 3151 (PL) and 1750 (SL) with 1140 genes overlapped; at D7, B/A: none; B/HU: 259 (PL) and 710 (SL) with 101 genes overlapped). The significantly enriched functional clusters were: cell cycle, signaling, metabolism, apoptosis, protein catabolism, and extracelluar matrix. The genes regulating muscle protein catabolic processes (e.g. Fbxo 32) were enhanced and anabolic processes (e.g. IGF‐1) were further down‐regulated in both PL and SL of B/HU compared to B/A.
FOXO3a mediates signaling cross‐talk that coordinates ubiquitin and atrogin‐1/MAFbx expression during muscle atrophy
Authors: Price Russ , Zheng Bin , Li Haiyan , Roberts‐Wilson Tiffany K.
Published: June 2021

Muscle atrophy due to chronic diseases (e.g., diabetes) or glucocorticoid‐induced insulin resistance results from programmed changes in ubiquitin‐proteasome system (UPS) gene expression (i.e., atrogenes). Different signaling pathways regulate various atrogenes like ubiquitin and the muscle‐specific E3 ligase atrogin‐1/MAFbx. A novel signaling model was tested in which FOXO3a mediates cross‐talk between the PI3‐kinase (PI3K)/Akt and MEK/ERK pathways to coordinate the expression of atrogin‐1 (AT‐1) and ubiquitin C (UbC), respectively. In L6 myotubes, dexamethasone reduced IRS‐1 protein and PI3K/Akt signaling while increasing AT‐1 expression; IRS‐2 protein, MEK/ERK/Sp1 signaling and UbC transcription were simultaneously increased. Expression of constitutively‐activated FOXO3a or knockdown of IRS‐1 increased IRS‐2 protein, MEK/ERK signaling and UbC expression. In rat muscles undergoing atrophy due to streptozotocin‐induced insulin deficiency, Akt signaling was decreased whereas MEK/ERK signaling and expression of UbC and AT‐1 were increased. These results demonstrate that FOXO3a mediates a reciprocal communication between the PI3K/Akt and MEK/ERK pathways that coordinates AT‐1/MAFbx and ubiquitin expression, thereby supporting muscle atrophy. Supported by NIH DK50740 and DK61521.
Protein S‐nitrosylation prevented by resistive vibration exercise (RVE) in human skeletal muscle following 60‐days of bed rest immobilization (BBR2‐2 Study)
Authors: Salanova Michele , Schiffl Gudrun , Gutsmann Martina , Felsenberg Dieter , Belavý Daniel , Blottner Dieter
Published: June 2021

Reactive Nitrogen Species (RNS) are important regulators of signalling pathways in muscle atrophy during prolonged disuse. We used the bed rest analogue (2nd Berlin Bed Rest Study, BBR2‐2) as a model of muscle inactivity, to study the pattern of S‐nitrosylated proteins in healthy human subjects following prolonged immobilization with and without exercise countermeasures. Twenty‐four healthy male subjects underwent 60‐days of 6° head down tilt bed‐rest (BR) and were randomized to an inactive control group (BR‐CTR, n=8), a high‐load resistive exercise group (BR‐RE, n=8) and a high‐load resistive vibration exercise (BR‐RVE, n=8) group. Muscle biopsy from m. soleus (SOL) and m. vastus lateralis (VL) were performed before and after BR and the pattern of S‐nitrosylated proteins were investigated by the “biotin switch assay”. Preliminary results showed that the level of S‐nitrosylated muscular proteins was altered after BR in all groups (i.e., BR‐CTR group: +10% SOL; +15% VL; BR‐RE group: no change in SOL, +18% in VL; BR‐RVE group: −19% in SOL, −15% in VL). More detailed proteome analysis of s‐nitrosylated proteins is presently underway. Vibration exercise thus serves as an effective exercise countermeasure protocol to prevent oxidative stress during disuse‐induced skeletal muscle atrophy or exposure to microgravity. Sponsor: Deutsches Zentrum für Luft‐und Raumfahrt (DLR), Germany (# 50WB0821 to D.B.)
Complexity in intracellular regulation of protein balance following unilateral diaphragm denervation
Authors: Argadine Heather M. , Mantilla Carlos B. , Sieck Gary C.
Published: June 2021

The balance between protein synthesis and degradation determines if there is a net loss (atrophy) or net gain (hypertrophy) in muscle mass. Recent studies suggest that PI3K‐dependent activation of Akt is an important regulator of both protein synthesis (via Erk and mTOR) and degradation (via the transcription factor FoxO1). Unilateral denervation (DNV) of the diaphragm muscle (DIAm) increases protein synthesis at 3d and degradation by 5d post‐DNV such that protein breakdown becomes evident by 5d after DNV. We hypothesized that DIAm DNV results in time‐dependent changes in Akt activation, leading to initial mTOR and subsequent FoxO1 activation. Akt, Erk, and mTOR phosphorylation and FoxO1 nuclear translocation were measured by Western blot analyses. At 1h and 6h post‐DNV, pAkt/Akt and pErk/Erk decreased. pErk/Erk increased at 1d and 3d, and pAkt/Akt increased at 5d post‐DNV. Although mTOR phosphorylation and expression increased 5d post‐DNV, the ratio of phospho‐mTOR/mTOR did not change at any time point post‐DNV. Nuclear translocation of FoxO1 increased 1d post‐DNV. Taken together, these results indicate that concomitant changes in PI3K/Akt, Erk, mTOR, and FoxO1 activity all contribute to, but do not completely account for, the time‐dependent changes in protein balance observed after DNV. Regulation of other signaling pathways, e.g., AMPK, and/or trophic influences, e.g., neuregulin, must be considered.
Intercellular adhesion molecule‐1 is expressed by skeletal muscle cells in hypertrophying muscle of mice
Authors: Dearth Christopher L. , Marino Joseph S. , Pizza Francis X.
Published: June 2021

Our laboratory recently demonstrated that β2 integrins contribute to overload‐induced skeletal muscle hypertrophy. One potential mechanism by which β2 integrin expressing leukocytes influence the hypertrophic response of skeletal muscle involves intercellular adhesion molecule‐1 (ICAM‐1), the major ligand for β2 integrins. The objective of our current work is to examine ICAM‐1 expression and localization to skeletal muscle cells in hypertrophying muscle and to determine if β2 integrins influence ICAM‐1 expression and/or its localization.

Plantaris muscles of wild type and β2 integrin deficient mice were exposed to 3, 7, or 14 days of mechanical overload via synergist ablation. Real‐time PCR and western blotting analysis revealed that ICAM‐1 gene and protein expression was significantly elevated in overloaded muscle. Experiments using flow cytometry and multi‐photon laser scanning confocal microscopy revealed that ICAM‐1 was expressed by mononuclear cells (satellite cells/myoblasts and leukocytes) as well as mature muscle fibers following mechanical loading.

We speculate that expression of ICAM‐1 by skeletal muscle cells after mechanical loading serves as a mechanism by which neutrophils and macrophages can bind to and directly communicate with skeletal muscle cells to promote hypertrophy.

This work was partially supported by The University of Toledo Interdisciplinary Grant Program.
HSP70 response to aerobic training in aging human skeletal muscle
Authors: Konopka Adam , Harber Matthew
Published: June 2021

The response of heat shock proteins (HSP) to aerobic training has been investigated in animal models, but understudied in human skeletal muscle. Furthermore, very little is known regarding HSP in aging skeletal muscle. The purpose of this investigation was to determine basal mRNA and protein content of HSP70 before and after 12‐weeks of aerobic training on a cycle ergometer in nine elderly women (70±2y). Post training muscle biopsy samples were taken ~48 hours after the last exercise session in order to assess steady state levels of HSP70 mRNA and protein. Basal HSP70 mRNA was reduced (P<0.05) by 23±8% while HSP70 protein content was unchanged after aerobic training. The reductions in HSP70 mRNA may represent a reduction in age‐related basal stress after the training program. This investigation provides the first assessment of HSP70 regulation in aging human skeletal muscle with aerobic exercise training.

Supported by NIH Grant AG032127
Parathyroid Hormone, Platelet Free Cytosolic Calcium and Platelet Indices among Asian Indian and Qatari Subjects
Authors: Afifi Nahla M. , Ewis Soliman A. , Benslimane Fatiha M. , Bejaoui Yosra R. , Alghafer Aisha
Published: June 2021

Asian Indians (AI) are more prone to coronary artery disease (CAD) than other ethnic groups. These variations in events are not fully explained only by differences in conventional cardiovascular risk factors. Thus, other factors must be account for such pattern. Disturbed some of the platelet activation indices including mean platelet volume (MPV) & platelet distribution width (PDW) as well as increased platelet free cytosolic calcium (Cai) play critical roles in platelet activation and thrombus formation. Parathyroid hormone (PTH) may increase risk for ischemic arrhythmias and its effects on MPV, PDW and Cai is of growing interest. There are limited data concerning these parameters in AI. Therefore we aimed to study the PTH and platelet Cai levels as well as MPV and PDW in healthy AI versus Qatari (Qt) subjects. Twelve healthy, nonsmoking AI were age matched with 11 healthy nonsmoking Qt subjects. PTH levels, as measured by ELISA, demonstrated a significant higher levels in AI (32.7±10.3 vs 23.5±9.95, P<0.05). Platelet Cai, as measured by X ray microanalysis, exhibited a non‐significant increase among AI subjects compared with Qt ones (0.117±0.0854 vs 0.097±0.034, P=0.366). MPV and PDW were elevated among AI versus Qt subjects (MPV, 7.865±0.57 vs 7.7±0.294, P=0.284)) & (PDW, 15.929±0.369 vs 15.4±0.698, P<0.05). These differences may clarify the reported discrepancy in CAD rates between the two ethnic groups.
Effect of diamide‐induced oxidative stress on Ca2+ signaling in cultured bovine aortic endothelial cells (BAEC)
Authors: Lock Jeffrey T. , Schilling William P.
Published: June 2021

Diamide is a thiol‐oxidizing agent commonly used to rapidly and reversibly oxidize intracellular glutathione (GSH) to its disulfide (GSSG) and to increase the formation of protein‐glutathione mixed disulfides (P‐SSG). In the present study we examined the acute effects of diamide on free cytosolic Ca2+ ([Ca2+]i) of cultured BAECs loaded with fura‐2. Diamide increased spontaneous Ca2+ oscillations and elevated basal [Ca2+]i when compared to vehicle‐treated controls. Low concentrations (50, 100 μM) progressively increased the number of cells spontaneously oscillating as well as the oscillation frequency of individual cells, whereas high concentrations (250, 500 μM) caused a synchronized oscillation of almost all cells immediately upon diamide exposure followed by a significant rise in basal [Ca2+]i. Similar dose‐ and time‐dependent effects of diamide were observed in the absence of extracellular Ca2+. Pretreatment with the phospholipase C inhibitor U73122, but not the inactive analog U73343, prevented the observed changes in [Ca2+]i. These results indicate that diamide promotes Ca2+ release from the inositol 1,4,5‐trisphosphate (IP3)‐sensitive internal Ca2+ store and is capable of elevating basal [Ca2+]i in the absence of extracellular Ca2+, effects which may be related to a diamide‐induced glutathionylation of the IP3 receptor and the plasmalemmal Ca2+ pump, respectively. Supported in part by HL097355
ACE2 over‐expression decreases the development of neurogenic hypertension and is associated with activation of nitric oxide synthase and nitric oxide release in human ACE2 transgenic mice
Authors: Feng Yumei , Cai Yanhui , Lazartigues Eric
Published: June 2021

Central ACE2 prevents the development of hypertension induced by chronic AngII infusion in transgenic mice (SA) overexpressing ACE2 in the brain. To elucidate the mechanisms involved, we investigated the expression of NOS isoforms (immunohistochemistry) and NO release (free radical measurement) in the brain of these mice. At baseline, ACE2 over‐expression was associated with increased (Relative Intensity, RI) nNOS (1.9 ±0.4), eNOS (1.5 ±0.1), phosphorylated‐eNOS‐Ser1177/Thr495 ratio (3.4 ±0.2), an index of phosphorylation vs. dephosphorylation and NO release (3.57 ±0.02 μM) vs. non‐transgenic (NT) littermates (1.0 ±0.1 RI and 2.95 ±0.11 μM, respectively, P<0.05). AngII infusion dramatically reduced nNOS (1.4 ±0.3) and eNOS (0.8 ±0.1) expression; (P<0.05) and NO release (2.68 ±0.19 μM) in SA but it remained significantly higher than in NT (nNOS: 0.4 ±0.03; eNOS: 0.4 ±0.1; NO: 1.82 ±0.22 μM, P<0.05) mice. Interestingly, Ang II+Mas blocker dramatically decreased the Phospho‐eNOS‐Ser1177/Thr495 ratio (0.7 ±0.02) compared to SA mice infused with AngII alone (3.6 ±0.5, P<0.05) and further reduced NO levels (1.75 ±0.08 μM). These data suggest that ACE2 exerts its modulatory effects partly through up‐regulation of NOS expression and phosphorylation, reinforcing Ang‐(1–7)‐mediated NO release in the brain, thus preventing the development of neurogenic hypertension. (AHA 0825459E, NIH RR018766 and HL093178)
Insulin‐mediated increases in arterial baroreflex control of muscle sympathetic nerve activity following meal intake in humans
Authors: Young Colin N. , Mikus Catherine R. , Thyfault John P. , Fadel Paul J.
Published: June 2021

Animal studies indicate that insulin enhances arterial baroreflex (ABR) control of sympathetic nerve activity (SNA); however, the extent to which these findings can be extrapolated to humans is unknown. To begin to address this, we utilized a mixed meal as a physiological method to evoke sustained increases in plasma insulin. Muscle SNA and arterial blood pressure (BP) were measured in 10 subjects (25 ± 2 yr) before and for 120 minutes following mixed meal intake. Weighted linear regression analysis between muscle SNA and diastolic BP was used to determine the gain (i.e. sensitivity) of ABR control of SNA. Plasma insulin was significantly elevated within 30 minutes following meal intake (Δ29 ± 7 uIU; P<0.05) and remained above baseline for up to 120 minutes. Similarly, 30 minutes after meal intake, ABR gain of muscle SNA burst incidence (−3.31 ± 0.64 vs. −5.13 ± 0.95 bursts/100 heart beats/mmHg; baseline vs. 30 min, P<0.05) and total muscle SNA (−3.22 ± 0.60 vs. −5.89 ± 1.30 arbitrary units/beat/mmHg; baseline vs. 30 min, P<0.05) was increased and remained elevated for the duration of the protocol. BP was unchanged by meal intake. In a subset of subjects the ABR muscle SNA gain remained unchanged during time control experiments. These preliminary findings suggest that increases in plasma insulin following meal intake enhance the gain of ABR control of muscle SNA in healthy humans.

Support: NIH RO1HL093167
Hyperinsulinemia and ectopic fat deposition develop in the face of hyperadiponectinemia in young obese rats
Authors: Ronis Martin J , Marecki John C , Shankar Kartik , Badger Thomas M
Published: June 2021

The role of reduced adiponenctin signaling in childhood obesity is unclear. Weanling male Sprague‐Dawley rats were overfed a high fat diet via total enteral nutrition. Excessive caloric intake led to increased weight and fat mass; dyslipidemia; ectopic fat deposition; and hyperinsulinemia (P<0.05). Expression of fatty acid transporter CD36 was elevated in both liver and muscle (P<0.05). Akt phosphorylation was elevated in liver but not muscle and hepatic nuclear FoxO1 protein reduced (P<0.05). Overfeeding increased serum adiponectin from 24.6±1.9 μg/ml to 46.3±5.9 μg/ml (P<0.05) correlating with increased fat mass. TNF‐α expression in fat was unchanged. AMP kinase phosphorylation, PPAR‐α expression and fatty acid oxidation was elevated in both liver and muscle (P<0.05). Thus excessive intake of high fat diet in young rats resulted in “adiponectin‐independent” increases in serum insulin and elevated ectopic fat deposition. Fatty acid transport appears to be the major mechanism underlying ectopic fat deposition. These data suggest age‐related differences in the role of adiponectin in pathological responses associated with obesity. USDA CRIS‐6251‐51000‐007‐03S.
Role of adrenergic receptors beta1 and beta2 in adaptive thermogenesis
Authors: Ribeiro Miriam Oliveira , Ueta Cintia Bagne , Capelo Luciane , Fonseca Tatiane , Chistoffolete Marcelo A , Aoki Marcelo S , Kim Brian
Published: June 2021

Brown adipose tissue (BAT) is the main site of adaptive thermogenesis, and its function is mostly regulated by the sympathetic nervous system and the β‐adrenergic receptor signaling pathway. To evaluate the independent roles played by β1 and β2 isoforms in energy homeostasis, mice with targeted disruption of either one of these genes were studied. While the KOβ1 mice failed to increase resting energy expenditure (REE) and developed hypothermia during cold exposure, they also exhibited decreased interscapular BAT thermal response during norepinephrine or dobutaine infusion. In contrast, KOβ2 mice behaved normally under these conditions. When placed on a high fat diet (24% fat) for 21 weeks, both KOβ1 and KOβ2 mice doubled their caloric intake, body weight gain and body fat content, not different from wild type animals, but failed to increase REE. In conclusion, our data indicate that the β1 signaling pathway is critical in cold‐induced thermogenesis whereas both β1 and β2 pathways play a role in the thermogenesis induced by high‐fat feeding. Supported by FAPESP.
Peripheral Chemoreceptors Enhance Renal Sympathetic Nerve Activity and Hypertension in SHR
Authors: Tan Zhiyong , Morgan Donald A. , Whiteis Carol A. , Rahmouni Kamal , Abboud Francois M.
Published: June 2021

In previous studies we reported that glomus cells of SHR are hypersensitive to low pH and overexpress ASIC (acid sensing ion channels) and TASK (two‐pore domain acid‐sensing potassium channels) that are pH‐sensitive. In this study, we tested the hypothesis that glomus cell hypersensitivity in SHR results in exaggerated chemoreceptor reflex increases in renal sympathetic nerve activity (RNA) and blood pressure (BP). Rats were anesthetized (ketamine and xylazine) and artificially ventilated. Changes in RNA and in mean arterial BP were recorded in response to intracarotid injections of 0.01, 0.02 and 0.05 mmol lactic acid. There were significant ventilatory oscillations in mean arterial pressure (BP) and nerve activity (RNA), which were greater in SHR (P<0.05). Control values of maximum and minimum BP and RNA (integrated voltage) averaged 133±7.8/109±8.0 mmHg and 0.59±0.06/0.35±0.03 Volts/sec respectively in SHR (n=9) vs. 87.5±7.3/73.5±6.4 mmHg and 0.43±0.04/0.24±0.02 Volts/sec in WKY (n=9). Following lactic acid injections, the maximal peak values of RNA and their corresponding mean BP are shown below.

These values of RNA were significantly greater in SHR (ANOVA‐Newman‐Keuls: F ratio 6.8, P=0.018) despite significantly higher levels of BP. Thus, the enhanced chemoreceptor sensitivity in SHR overrides the inhibitory influence of baroreceptors and may contribute to the excessive sympathetic nerve activity and hypertension in SHR.

This work was supported by NIH grant HL14388.
Sympathetic nervous system interacts with renin‐angiotensin system to control renal sodium and water handling in SHR rat
Authors: Salman Ibrahim M , Ameer Omar Z , Sattar Munavvar A , Abdullah Nor A , Yam Mun Fei , Khan Md. Abdul Hye , Johns Edward J
Published: June 2021

This study investigated the impact of acute renal denervation (ARD) on sodium and water excretory responses to i.v. infusion of norepinephrine (NE) and angiotensin II (Ang II) in anesthetized spontaneously hypertensive rats (SHR). Anesthetized SHR were acutely denervated and NE or Ang II was continuously infused for 1hr. A screw‐controlled snare was placed around the aorta above the renal arteries and systemic blood pressure was regulated to avoid any increments in the renal perfusion pressure (RPP) in response to the vasoactive stimuli. Three 20min urine clearances were collected to measure urine flow rate (V) and sodium excretion (UNaV). The administration of NE or Ang II to innervated SHR produced marked (P<0.05) antidiuresis/antinatriuresis response compared to innervated saline treated SHR. Lower (P<0.05) diuresis/natriuresis response was observed in denervated NE treated SHR compared to denervated saline treated rats. There was a larger (P<0.05) diuresis/natriuresis response to ARD in Ang II treated SHR compared to denervated saline treated counterparts. NE retains its typical antidiuretic/antinatriuretic action in denervated SHR. Data demonstrated that the antidiuretic/antinatriuretic action of Ang II depends on the presence of intact renal innervation. It can also be suggested that Ang II facilitates the NE release from renal sympathetic nerve by a presynaptic action.
Lowered activation of voltage‐gated sodium channels contributes to blunted baroreflex sensitivity in heart failure rats
Authors: Tu Huiyin , Zhang Libin , Tran Thai P. , Muelleman Robert L. , Li Yu‐Long
Published: June 2021

Voltage‐gated sodium (Nav) channels are responsible for initiation and propagation of action potential in the neurons. Here we measured the role of Nav channels on aortic baroreceptor neuron excitability and baroreflex sensitivity in sham and CHF rats. CHF was induced by left coronary artery ligation. Immunofluorescent data indicated that Nav1.7 was expressed in A‐type (myelinated) and C‐type (unmylinated) nodose neurons but Nav1.8 and Nav1.9 were expressed only in C‐type nodose neurons. Real‐time RT‐PCR and western blot data showed that CHF reduced mRNA and protein expression levels of Nav channels in nodose neurons. In addition, using the whole cell patch‐clamp technique, we found that Nav current density and cell excitability of the aortic baroreceptor neurons were lower in CHF rats than that in sham rats. Aortic baroreflex sensitivity was blunted in anesthetized CHF rats, compared with that in sham rats. Furthermore, Nav channel activator (rATX II, 100 nM) significantly enhanced Nav current density and cell excitability of aortic baroreceptor neurons and improved aortic baroreflex sensitivity in CHF rats. These results suggest that reduced Nav channel activation is involved in the attenuation of baroreceptor neuron excitability, which subsequently contributes to the impairment of baroreflex in CHF state.
Exposure to chronic intermittent hypoxia in rats leads to exaggerated sympatho‐excitatory responses that are buffered by reduced adrenergic vasoconstriction in the mesentery
Authors: Silva Ana Quenia , Schreihofer Ann M
Published: June 2021

Chronic intermittent hypoxia (CIH) enhances sympatho‐excitatory reflexes in rats with blunted rises in arterial pressure (AP). Phenylephrine (PE; iv) evokes smaller rises in AP, suggesting adrenergic vascular reactivity is reduced. We examined if CIH reduces sympathetically‐mediated vasoconstriction in mesenteric (MES) vascular bed. Male Sprague‐Dawley rats were exposed to CIH (6% O2 for 40s, every 9 min, 8h/day, 2 wks). Under isoflurane rats were ventilated, paralyzed, prepared to record AP, splanchnic SNA, MES artery blood flow and then anesthesia was changed to urethane (750mg/kg)/chloralose (60mg/kg, iv). Compared to controls (n=9), rats with CIH (n=11) had higher AP (127±3 vs 107±7 mmHg, P<0.05) and SNA (85±11 vs 47±5 mV, P<0.05). After CIH, stimulation of brachial nerve evoked larger rises in SNA (137±21 vs 52± 11%, P<0.01) with blunted rises in AP and MES resistance (RES; 110±1 vs 117± 2%, P<0.05). Stimulation of brachial nerve after propranolol induced similar responses, suggesting beta adrenergic receptors were not mediating the blunted effects on AP and MES RES. After ganglionic block to eliminate SNA, PE evoked smaller rises in AP (n=5; 55±9 vs 83±7 mmHg, P<0.05) and MES RES (566±14 vs 971±234 %, P<0.05) in CIH rats. These data suggest CIH reduces sympathetically‐mediated alpha adrenergic vasoconstriction in the mesentery, perhaps as a buffer for chronically elevated SNA. AHA0755292, NIH‐HL075174
Effects of voluntary exercise on levodopa‐induced motor response complications in hemiparkinsonian rats
Authors: Corpt Briana Jean , Oh‐Lee Justin D.
Published: June 2021

Research evidence suggests that voluntary exercise may be beneficial in restoring motor skills traditionally lost in Parkinson's disease (PD). Levodopa pharmacotherapy remains the most effective treatment for PD, but is often associated with the genesis of disabling motor response complications (MRCs). This study was designed to investigate the effects of voluntary exercise on these motor complications in 6‐hydroxydopamine (6‐OHDA) hemiparkinsonian rats. Adult male Sprague‐Dawley rats were rendered parkinsonian by 6‐OHDA injections into the left medial forebrain bundle. After two weeks, half of these animals received twice‐daily levodopa while the other half received twice‐daily vehicle. An equal number of animals in each drug treatment group underwent exercise (2 hrs twice daily, 14 days) after drug injection, and the other half served as controls. Voluntary exercise therapy when combined with levodopa acted to provide a substantially greater attenuating effect on the forelimb deficits (p < 0.05) and the shortened motor response (p < 0.05) produced by chronic levodopa therapy. These results suggest that mechanisms mediated by voluntary exercise could be crucial for an accurate understanding of the pathogenesis of motor complications and further suggest novel approaches to more effective and safer therapies of PD and other dopamine disease states.

ORSP PRIF Grant, CMU, to JD Oh‐Lee
Frequency dependent effects of vibration on nerve function, oxidative stress and inflammation
Authors: Pacurari Maricica , Johnson Claud , Krajnak Kristine
Published: June 2021

Prolonged occupational exposure to hand‐held vibrating tools leads to pain and reductions in tactile sensitivity, grip strength and manual dexterity. The goal of the current study was to use a rat‐tail model to determine if vibration frequency influences factors related to sensorineural dysfunction. We tested the hypothesis that exposure to tail vibration (4 hr/day for 3 days) at a high frequency (250 Hz) would result in sensorineural dysfunction and greater increases in oxidative stress and inflammation in tail nerves than exposure to vibration at a lower frequency (62.5 Hz). Nerve function was assessed using transcutaneous electrical stimulation. Exposure to 250‐Hz, but not 62.5‐Hz vibration, resulted in an increased sensitivity of Aβ‐ and Aδ‐nerve fibers to electrical stimulation. These changes in nerve fiber sensitivity were associated with increased expression of interleukin (IL)‐1β and tumor necrosis factor ‐α in tail nerves, and a moderate increase in cyclooxygenase‐2 gene expression. Circulating concentrations of IL‐1 β were also increased. Vibration exposure at both frequencies resulted in reductions in glutathione and glutathione peroxidase 3 levels in tail nerves. However gene expression for other oxidative factors and nitrotyrosine protein levels were significantly increased after exposure to vibration at 62.5 Hz, but not 250 Hz. These results demonstrate that the effects of vibration on sensorineural function and physiology are frequency dependent, and that changes in nerve function may be related to increases in inflammation and oxidative activity.
Relationship between apolipoprotein E genotype and cerebral palsy in Chinese Han children
Authors: Li Xiao‐Jie , Wang Li‐Ping , Lu Zhi‐Hai , Wang Shu‐Qiu , Kang Yu‐Ming
Published: June 2021

The present study was undertaken to investigate the relationship between apolipoprotein E genotype and cerebral palsy in the Chinese Han children. 120 Chinese Han children with cerebral palsy age range from 1 to 12 years old were selected as CP group including 86 spastic and 34 dyskinetic children, 120 normal Chinese Han children age range from 1 to 12 years old as control group. Apolipoprotein E DNA was obtained from blood cell. Polymerase chain reaction‐restricted fragment length polymorphism (PCR‐RFLP) was used to determine allele and apolipoprotein E genotype in CP group and control group. The severity of impairment was assessed by gross motor function classification system (GMFCS). The results showed that gene frequency of the ε4 allele in CP group was higher than control group. Gene frequency of the ε4 allele in GMFCS level IV and level V children was higher than in GMFCS level I, level II, level III ones in CP group. The difference of gene frequency of the ε4 allele between spastic children and in dyskinetic ones was not significant.

Conclusion

Gene frequency of the ε4 allele was associated with cerebral palsy in the Chinese Han children, particularly with GMFCS level IV and level V ones, but it was no association with CP children who was the spastic or the dyskinetic.
Effects of chronic iron deficiency anemia on the protein expression of rat thalamus
Authors: Kawano Fuminori , Oke Yoshihiko , Nakai Naoya , Ohira Takashi , Nomura Sachiko , Fujita Ryo , Ohira Yoshinobu
Published: June 2021

The present study was carried out to investigate the effects of anemia on the protein expression in rat thalamus. The powdered diet containing 100 or 3 ppm iron was fed in rats with the age of 3‐week. After 20 weeks of supplementation, blood and thalamus including hypothalamus were sampled from both groups. The hematocrit and hemoglobin content were 44 and 15 g/dl in the 100 ppm group, respectively. However, they were less (17 and 4 g/dl) in the 3 ppm group. The two‐dimensional electrophoresis was performed using the homogenate of thalamus. As the result, 73 kDa heat shock protein (HSP73) was found in the spot which expression level was greater in 3 ppm group. Further, western blot was performed to check the quantitative expression level. Although the level of total HSP73 was greater (+89%) in the 3 ppm group, no differences were seen if they were compared among spots with the different isoelectric points. The iron deficiency‐induced anemia may cause some changes in thalamus function via HSP73 expression. This study was supported by Grant‐in‐Aid for Scientific Research S (19100009) and Exploratory Research (20650104) from JSPS, and Young Scientists B (21700656) from MEXT.
Plasticity of gastrointestinal tract structure and function in the invasive fish Pterygoplichthys disjunctivus (Teleostei: Loricariidae)
Authors: German Donovan Parks , Evans David H
Published: June 2021

Pterygoplichthys disjunctivus, which is native to tropical South America, is an invasive fish species found in at least nine countries outside of the neotropics, including the United States. We tested the hypothesis that P. disjunctivus has a broad diet and exhibits plasticity of gastrointestinal (GI) tract structure and function in relation to food quality and intake, thereby contributing to its success. Stable isotope analyses (δ13C and δ15N) of several tissues of wild‐caught fish confirmed that P. disjunctivus has a variable diet over time. Individuals of this species were raised in the laboratory on a high‐quality algal diet, a low‐quality wood diet, or no food at all (starvation) for 150 days and the effects of diet, intake, and starvation on GI tract structure and function were examined. Individuals that were starved or consumed large amounts of a low‐quality diet showed little mortality but reduced their intestinal surface areas by 70 and 78%, respectively, and reduced the microvilli surface area by 52 and 27%, respectively, in comparison to wild‐caught fish. Intake and dietary quality did not affect the patterns of 10 digestive enzymes active along the fish's intestine, but the algae‐fed fish had higher activities of most enzymes and higher levels of GI fermentation. Overall, our results support our hypothesis and may provide insight into the success of P. disjunctivus as an invasive species across the globe.
Skeletal Muscle and Hepatic Glycogen Content in Birds
Authors: Ferguson Donna , Braun Eldon J.
Published: June 2021

Plasma glucose levels (Pglu) in birds are relatively elevated (10–15 mM/L) compared to mammals of similar body mass (5 mM/L). In mammals, insulin stimulated glucose uptake into myocytes via GLUT4, and diffusive glucose uptake into hepatocytes via GLUT2 removes glucose from circulation to maintain a consistent Pglu and the glucose can be subsequently converted to glycogen for storage. In birds, there is evidence that GLUT4 is absent from myocytes (Sweazea and Braun, 2005), which would potentially limit the amount of glucose available to skeletal muscle tissue for metabolism and glycogen storage. In our study, we attempt to quantify the glycogen content in liver and muscle tissue of birds with rat tissue as a control. Using a phenol‐sulfuric acid assay (Siu Lo, et al, 1970), we are able to measure the amount of glycogen stored by hydrolyzing the glycogen to glucose monomers and use colorimetric spectroscopy to measure absorbance versus micrograms of glycogen. Preliminary data show that although fasted and fed rats have a large deviation in hepatic glycogen stores, bird hepatic glycogen stores are relatively stable, suggesting that glycogenolysis is not a primary metabolic process. The lower glycogen stores in bird skeletal muscle also suggests a lack of insulin stimulated, post‐prandial glucose uptake into muscle and therefore a limited use of this substrate for metabolism, despite the markedly elevated Pglu.