bluuebunny/crossref_metadata_2025_split huggingface dataset:
Molecular Characterization of the Ca2+ Release Activated Ca2+ Current in Native Brain Capillary Endothelial Cells
Authors: Sanchez Alfredo , Lavanderos Boris , Trebak Mohamed , Earley Scott 
Published: May 2024

Brain capillary endothelial cells (cECs) form the blood-brain barrier orchestrate on-demand increases in blood flow to serve the brain’s unique metabolic needs. An in vivo imaging study showed that brain capillaries generate an array of dynamic Ca2+ signals primarily dependent on the release of Ca2+ from the endoplasmic reticulum (ER). Little is known about the physiological functions and origins of these signals. Store-operated Ca2+ entry (SOCE) is a fundamental Ca2+ influx mechanism activated by emptying ER Ca2+ stores. Following store depletion, interactions between the ER Ca2+-sensor stromal interaction molecule 1 (STIM1) and Orai1, 2, and/or 3 ion channel subunits on the plasma membrane initiate the Ca2+-release activated Ca2+ current (ICRAC) and SOCE. ICRAC and SOCE in brain cECs are uncharacterized. Therefore, we investigated STIM and Orai expression and the biophysical properties of ICRAC in primary brain cECs. Using droplet digital PCR, we found that native brain cECs express Stim1, Stim2, Orai1, and Orai3 but not Orai2. Conventional whole-cell patch-clamp electrophysiology was used to record ICRAC from freshly isolated brain cECs. Currents were induced by including the slow Ca2+ buffer EGTA in the intracellular solution. In contrast to the strong inward rectification of canonical ICRAC, currents recorded from native brain cECs were dually rectifying. The inward and outward components were blocked by La3+, enhanced with 2-APB, and inhibited with the selective Orai1 channel antagonist Synta66. Both ICRAC components were increased when a divalent-free solution (Ca2+ replaced with Cs+) was used. To gain insight into the molecular composition of CRAC channels in brain cECs, we generated EC-specific knockout (ecKO) Stim1, Orai1, and Orai3 mice. ICRAC was absent in cECs obtained from Stim1-ecKO mice. In contrast, ICRAC recorded from Orai1-ecKO mice displayed strong inward rectification with no outward current and was insensitive to Synta66. ICRAC recorded from Orai3-ecKO mice were also inwardly rectifying but were highly sensitive to Synta66. ICRAC was undetectable in cells from Orai1-Orai3-ecKO mice. These data suggest that native ICRAC in brain cECs requires the expression of Stim1, Orai1, and Orai3. The outward component of native ICRAC in these cells requires the expression of Orai1 and Orai3, but the expression of either Orai1 or Orai3 alone is suffcient to maintain the inward component. R35HL155008, R33NS115132, and P20GM130459 (to SE). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Early life stress-induced aortic endothelial dysfunction is mediated by endothelium-specific HDAC9
Authors: Kelly Gillian , Kellum Cailin , Ho Dao , Colson Jackson , Walker Savannah , Weintraub Neal L. , Kovacic Jason 
Published: May 2024

Early life stress (ELS) is highly prevalent worldwide and has been associated with an elevated risk of developing cardiovascular disease (CVD) in adulthood. We previously showed that maternal separation with early weaning (MSEW), an established ELS model of neglect in mice, leads to blunted endothelium-dependent relaxation as well as increased histone deacetylase (HDAC) activity. We hypothesized that specific HDAC isoform(s) mediate the ELS-induced endothelial dysfunction. In a screen of all HDAC isoforms, HDAC1, 6, and 9 gene expression were significantly elevated in aortas from male adult MSEW and normally reared (NR) C57BL6/J mice (N=3/group, p<0.05). Western blot analysis found that only HDAC9 protein abundance was significantly elevated in aortas from male MSEW as compared to NR mice (N=6/group, p=0.01). Detection of HDAC9 via immunohistochemistry was greater in the aortic endothelium of MSEW mice compared to NR (N=5/group). Subsequently, vascular reactivity was measured with wire myography on tamoxifen inducible endothelium-specific HDAC9KO (Cdh5CreERT2-HDAC9) MSEW and NR adult mice to determine the role of HDAC9 in MSEW-induced endothelial dysfunction. Male mice exposed to MSEW displayed blunted aortic endothelial-dependent relaxation to acetylcholine compared to NR, whereas endothelial-specific knockout of HDAC9 (HDAC9KO) restored endothelial-dependent relaxation (NR: Emax=89.27%±4.15, N=5; MSEW: Emax=48.12%±22.12, N=2; NR HDAC9 KO: Emax=68.98%±4.73, N=5, MSEW HDAC9KO: Emax=70.50%±1.85, N=3). Females demonstrated no difference between MSEW and NR endothelial dependent relaxation, however, HDAC9KO MSEW displayed blunted relaxation compared to all other groups (NR: Emax= 77.14%±4.86, N=5; MSEW: Emax=84.30%±8.71, N=2; NR HDAC9 KO: Emax=70.80%±5.87, N=5, MSEW HDAC9KO: Emax=69.76%±9.10, N=4). Endothelial-independent relaxation to sodium nitroprusside (SNP) was similar in all groups in both sexes (Male: NR: Emax=96.92%±1.26, N=5; MSEW: Emax=100%±0, N=2; NR HDAC9 KO: Emax=96.52%±3.484, N=5; MSEW HDAC9KO: Emax=100%±0, N=3; Female: NR: Emax=98.58%±0.42, N=5; MSEW: Emax=100%±0, N=2; NR HDAC9 KO: Emax=98.48%±1.08, N=5; MSEW HDAC9KO: Emax=99.15%±0.85, N=4). These findings suggest that HDAC9 mediates ELS-induced endothelial dysfunction in males, however, HDAC9 may play a protective role in females. Future studies aim to further investigate sex-dependent differences in the specific role of HDAC9 in ELS related CVD. NIH P01 HL158500, F31HL165863-01, T32GM135028. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Effects of Heavy Alcohol Use on Nighttime Blood Pressure, Sleep Quality, and Urine Catecholamines
Authors: Haun Tabitha , Chang Keng-Yu , Phan Kelly , Gil Alfredo , Pham Jonathan , Truong Preston , Liu Zhaoli 
Published: May 2024

Heavy alcohol drinking habits contribute to the development of hypertension; this is partly due to overactivation of the sympathetic nervous system and the increased release of catecholamines. Recently, heavy alcohol drinking has been linked to poor sleep quality, but the underlying mechanisms remain unclear. We postulated that long-term frequent heavy alcohol use could increase catecholamine levels and cause blood pressure dysregulation at night, altogether influencing sleep quality. Therefore, the purpose of this study is to test the following hypothesis that compared to non-heavy drinkers, heavy drinkers have a higher level of nighttime catecholamines, a higher levels of nighttime blood pressure, and poorer sleep quality. A total of 44 men and postmenopausal women non-smokers, free of major clinical diseases, were included in this study. Heavy drinkers were defined as those having a dried blood spot phosphatidylethanol (PEth, a biomarker to detect heavy alcohol drinking) level ≥ 20 ng/mL. Participants wore an ambulatory blood pressure monitor for 24 hours and catecholamines were measured from urine samples collected in the same 24-hour period. Sleep quality was measured by using the Pittsburgh Sleep Quality Index (PSQI) questionnaire. There were 21 heavy drinkers (mean±SEM for age: 58±1 and BMI: 26.2±0.7 kg/m2) and 23 non-heavy drinkers (age: 58±1 and BMI: 27.5±0.8 kg/m2; P≥0.3 vs. heavy drinkers). Compared to non-heavy drinkers, heavy drinkers had a higher nighttime systolic blood pressure (SBP), higher diastolic blood pressure (DBP), and a lower SBP dipping ratio (heavy drinkers vs. non-heavy drinkers for nighttime SBP: 118±3 vs. 109±2 mmHg, P=0.02; nighttime DBP: 70±2 vs. 65±2 mmHg, P=0.04; and SBP dipping ratio: 10.6±1.4% vs. 14.5±1.6%, P=0.04). No differences were found in nighttime urine catecholamines between heavy drinkers and non-heavy drinkers (P≥0.3). A total of 13 participants had a PSQI score higher than 5, indicating poor sleep quality; however, PSQI scores were similar between heavy and non-heavy drinkers (P=0.8). These findings indicate that heavy drinkers had a higher nighttime blood pressure and are consistent with the observation that heavy drinking is associated with an increased risk of hypertension in mid-life adults. Interestingly, the increase in nighttime blood pressure does not appear to be related to catecholamines and sleep quality in apparently healthy mid-life adults does not appear to be influenced by heavy drinking habits. This work was supported by NIAAA AA028537 to CLH and AHA 1266805160 to TH. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Functional sympatholysis of Neuropeptide Y-mediated vasoconstriction in humans
Authors: Wakeham Denis , MacNamara James , Hissen Sarah , Levine Benjamin , Hearon Christopher 
Published: May 2024

Metabolic inhibition of sympathetic vasoconstriction (functional sympatholysis) is essential for adequate perfusion of skeletal muscle during exercise. Functional sympatholysis occurs primarily through post-junctional attenuation of norepinephrine and α-adrenergic-mediated vasoconstriction, though other non-adrenergic neurotransmitters are co-released with norepinephrine. Neuropeptide Y (NPY) is a neurotransmitter that elicits robust vasoconstriction and is co-released with norepinephrine during exercise in an intensity dependent manner. Evidence from animal models indicate that NPY is sensitive to metabolic inhibition, however metabolic inhibition of NPY-mediated vasoconstriction has not been tested in humans. We tested the hypothesis that NPY-mediated vasoconstriction would be attenuated during handgrip exercise in humans. In 10 healthy adults (7M:3F, age: 30±7 years, BMI: 24±5) we measured forearm blood flow (Doppler ultrasound), blood pressure (brachial artery catheter), and calculated changes in forearm vascular conductance (FVC) to local intra-arterial infusions of phenylephrine (PE; α1-agonist) or NPY (Y1R-agonist) during 1) intra-arterial infusion of sodium nitroprusside (SNP; nitric oxide donor) to serve as a non-metabolic vasodilatory control or 2) dynamic rhythmic handgrip exercise (EX; 15% maximal voluntary contraction). The vasoconstrictor responses to PE and NPY were quantified as a percent ΔFVC during each condition (SNP vs. EX) and were compared by two-way RM-ANOVA. The magnitude of sympatholysis (ΔFVC during exercise expressed as a percent of control) for PE and NPY was compared by a paired t-test. As expected, the vasoconstrictor response to PE was attenuated during handgrip exercise compared to SNP (ΔFVC: SNP: -50±24 vs. EX: -17±9 %; p=0.002). Similarly, NPY-mediated vasoconstriction was blunted during handgrip exercise compared to SNP (ΔFVC: SNP: -32±22 vs. EX: -11±7 %; p=0.029). There was no difference in the magnitude of sympatholysis between PE and NPY (PE: 68±18, NPY: 52±34 %; p=0.28). These data indicate that NPY-mediated vasoconstriction is sensitive to metabolic inhibition in humans, and the magnitude of sympatholysis is not different from α1-adrenergic vasoconstriction. Funding: NIH R00HL153777. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Human Adenovirus Serotype 5 (HAdV-5) Infection of Kidney Fibroblast Cells Disrupts Cysteine, Purine, and Unsaturated Fatty Acid Metabolism in a Dose- and Time-Dependent Manner
Authors: Sanchez Bailey-J C. , Grasis Juris A. , Ortiz Rudy M. 
Published: May 2024

Kidneys partake in numerous metabolic responsibilities, notably clearance of insulin from circulation, gluconeogenesis, and reabsorption of amino acids. Viral-induced kidney damage, either acutely or chronically, can occur from direct impact on kidney cells and from systemic and local immunological responses to counter the viral infection. Thus, it is likely that metabolic dysfunction is a consequence of infections. The detriments of viral infection on kidney cell metabolism remains an unelucidated area. We used human adenovirus serotype 5 (HAdV-5) to infect human embryonic kidney cells (HEK293T) in a dose- and time-dependent manner. We hypothesized that optimal metabolic dysfunction results after 24 hours of infection and at the highest viral dose. Cells were exposed to three viral dosages (0.5,1.0,2.0 multiplicity of infection [MOI]) and metabolism assessed by untargeted metabolomics at four time points (6-,12-,24-, 36-hours post infection [HPI]). A total of 182 metabolites were identified in our study. As early as 6 hrs, the metabolic profile of infected cells exhibited downregulation of cystathionine, cysteine, homocysteine, hypoxanthine, linoleic acid, and oleic acid with upregulation of 5'-Deoxy-5'-methylthioadenosine (MTA) and adenine. Peak dysregulation for carbohydrate, cysteine, organic acid, purine, and unsaturated fatty acid-related metabolites was observed at 12 hrs. At 24 hours, tapering of the overall metabolite response was observed with further dysregulation seen at 36 hours. Across all three dosages, responses to infection were similar. Perturbation of the cysteine, purine, and unsaturated fatty acid metabolic pathways has not previously been considered with HAdV-5 infection and a commonly used cell line exhibiting properties of kidney, and historically neuronal components. Although not investigated, uncovering adenoviral impact on enzymatic activity in these pathways could elucidate more understanding of renal impairment. These data altogether suggest that substantial DNA viral infection disrupts cellular metabolic pathways that may impair renal functions but need to be evaluated in an in vivo model. USDA HSI Educational Grant 2021-03397. Bailey-J Sanchez was supported by USDA HSI Educational Grant 2021-03397. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Captopril treatment does not affect inflammatory and autophagy signaling in skeletal muscle of Zucker diabetic fatty rat
Authors: Lee Ji Heun , Reed Carter H , Arriaga Claudia , Carrillo Claudia , Rowling Matthew J. , Valentine Rudy J. , Selsby Joshua T. 
Published: May 2024

Skeletal muscle plays a key role in the development of Type 2 Diabetes Mellitus (T2DM) as its large capacity to sequester glucose and contribute to the regulation of energy metabolism is compromised. Previous studies indicate that in skeletal muscle, T2DM drives inflammatory signaling and impairs autophagy. Captopril (Cap), an angiotensin-converting enzyme (ACE) inhibitor, is a common treatment for hypertension in T2DM patients; however, the impact of Cap on inflammatory signaling and autophagy in insulin resistant skeletal muscle remains ill-defined. We hypothesized that inflammatory signaling would be elevated and degradation of autophagosomes would be impaired in obese Zucker diabetic fatty (ZDF) rats, which are hyperphagic as they lack the leptin receptor. We further hypothesized that daily Cap treatment would attenuate these disturbances. At 14 weeks of age, the absolute weights of gastrocnemius (p<0.0001), tibialis anterior (p<0.0001), soleus (p=0.002), and extensor digitorum longus (EDL, p<0.0001) muscles were decreased by 23-34% in ZDF compared to lean rats but were not altered by 8 weeks of Cap treatment (0.5g/kg/day). Immunoblotting revealed that in soleus and EDL, inflammatory signaling proteins TLR4, phosphorylated (p)IKK α/β (ser176/180), pNFkB p65 (Ser 536), IkBα, TNFα, and AP1 were similar amongst all groups, whereas MyD88 was elevated in ZDF soleus (p=0.03) but was not corrected by Cap. The autophagy protein, LC3A/B I was increased 32% (p=0.01) in EDL from ZDF rats compared to Con; however, PI3K III, pBeclin1 (ser 93), pULK1 (ser555), LAMP2, LC3A/B II, and the LC3A/B II/I ratio were similar amongst groups in the soleus and EDL and were not altered by Cap. Compared to control, P62, a key autophagy adapter protein, was similar in soleus and decreased 27% in EDL (p<0.01) from ZDF rats and was not altered by Cap. Counter to our expectations, although muscle mass was lower in ZDF groups, obesity and insulin resistance in young ZDF rats were not suffcient to drive inflammatory signaling or disrupt autophagy in glycolytic or oxidative skeletal muscle, and these processes appeared to be resistant to Cap treatment in ZDF rats. USDA-NIFA awarded to MJR (2021-67017-33937). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Compromised Plasma Membrane Repair Contributes to Neuronal Cell Death and Neurodegeneration During Alzheimer’s Disease
Authors: Bulgart Hannah , Lopez Perez Miguel , Banford Kassidy , Tucker Alexis , Bonser Sidney , Wold Loren , Weisleder Noah 
Published: May 2024

Alzheimer’s Disease (AD) is the most common neurodegenerative disease and involves accumulation of intracellular amyloid beta (Aβ) in the early stages of the disease. Aβ has been shown to compromise neuronal cell membrane integrity by increasing membrane permeability, elevating intracellular calcium concentrations, and Aβ directly penetrating and damaging the neuronal cell membrane. Damage to the cell membrane requires a rapid and effcient repair mechanism to restore the barrier function of the membrane and avoid cell death. Here, we aimed to study the effects of intracellular Aβ on neuronal cell membrane repair. To determine if membrane permeability is compromised in vivo, we stained 6-month FVB and APP/PS1 brain sections with anti-IgG. We observed a significant increase in IgG positivity in the APP/PS1 tissue as compared to the FVB brains. We repeated this with post-mortem frontal lobe tissue and observed a significant increase in IgG-positivity in the AD samples as compared to the non-AD. To assess the effect of neuronal membrane repair in AD patients, we treated mouse neuroblastomas (N2A) and primary mouse hippocampal neurons with non-AD and AD patient cerebrospinal fluid (CSF) at a 1:100 dilution or 1μM recombinant human Aβ42. We conducted laser damage assays and observed a membrane repair defect in these neuronal cell models. We explored the biochemical components of the CSF that may be contributing to this induced repair defect, we measured the Aβ42 by ELISA and stratified Aβ42 concentration by the corresponding CSF AUC. We observed a positive correlation between Aβ42 concentrations and reduced membrane repair capacity. We sought to determine the mechanism of the reduced membrane repair capacity by assessing the expression levels of membrane repair proteins and observed decreased dysferlin in cell models, 6-month APP/PS1 mouse models, and human postmortem tissues with high levels of Aβ. Overexpression of dysferlin in the AD cell models can increase dysferlin expression and restore membrane repair capacity in neurons treated with AD CSF samples or recombinant Aβ42. We have determined this reduction in repair capacity is due to reduced dysferlin expression, and future studies will determine the mechanism leading to dysferlin downregulation. This work was supported by The Ohio State University College of Medicine Dean’s Discovery Grants Program. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Myotube growth is associated with cancer-like metabolic reprogramming and is limited by phosphoglycerate dehydrogenase and pyruvate kinase M2
Authors: Stadhouders Lian , Smith Jonathon , Gabriel Brendan , Verbrugge Sander , Hammersen Tim , Kolijn Detmar , Vogel Ilse 
Published: May 2024

Skeletal myofiber size and oxidative metabolism are inversely related. Glycolytic fast type myofibers are larger than high high oxidative, slow type myofibers. Paradoxically, despite their lower capacity for protein synthesis (lower myonucluar density and RNA content), glycolytic, low oxidative are more responsive to an anabolic stimulus. We hypothesized that glycolytic myofibers may make use of a Warburg-like metabolic reprogramming in response to an anabolic stimulus. The Warburg effect links growth and glycolysis in tumor cells to generate glycolytic intermediates for synthesis of nucleotides and amino. The aim of this study was to test whether a similar ‘glycolysis-for-anabolism’ metabolic reprogramming also occurs in post-mitotic, hypertrophying myofibers. For this purpose, we first induced C2C12 myotube hypertrophy with IGF-1. We then added 14C glucose to the differentiation medium and measured radioactivity in isolated protein and RNA to establish whether 14C had entered anabolism. We found that especially protein became radioactive, suggesting a glucose → glycolytic intermediates → non-essential amino acid(s) → protein series of reactions, the rate of which was increased by IGF-1. Next, to investigate the importance of glycolytic flux and non-essential amino acid synthesis for myotube hypertrophy, we exposed C2C12 and primary mouse myotubes to the glycolysis inhibitor 2-Deoxy-D-glucose (2DG). We found that inhibiting glycolysis lowered C2C12 and primary myotube size. Similarly, siRNA silencing of PHGDH and PKM2, key enzymes of the Warburg-like metabolic reprogramming, reduced C2C12 and primary myotube growth; whereas retroviral PHGDH overexpression increased C2C12 myotube size. Together these results suggest that glycolysis is important for hypertrophying myotubes, which reprogramme their metabolism to facilitate anabolism, similar to cancer cells. Brendan M. Gabriel was supported by fellowships from the Novo Nordisk Foundation (NNF19OC0055072) & the Wenner-Gren Foundation, an Albert Renold Travel Fellowship from the European Foundation for the Study of Diabetes, and an Eric Reid Fund for Methodology from the Biochemical Society. Abdalla D. Mohamed was funded initially by Sarcoma UK (grant number SUK09.2015), then supported by funding from Postdoctoral Fellowship Program (Helmholtz Zentrum München, Germany), and currently by Cancer Research UK. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.