bluuebunny/crossref_metadata_2025_split huggingface dataset:
Angiotensin AT1A Receptor Signaling in AgRP Neurons in Metabolism Control
Authors: Balapattabi Kirthikaa , Ritter McKenzie , Mouradian Gary , Hodges Matthew , Sigmund Curt , Grobe Justin 
Published: May 2023

Resting metabolic rate (RMR) accounts for ≈70% of energy expenditure in humans. Obesity induced by high fat diet (HFD) feeding is associated with “RMR adaptation” (i.e., reduced RMR relative to body size). We previously reported angiotensin II (ANG) receptors (AT1A) in a subset of neurons of the arcuate nucleus of the hypothalamus (ARC) that express Agouti-related peptide (AgRP) are critically required for the integrative control of RMR. Understanding signal mechanisms in this unique subset of RMR-controlling AgRP neurons is the goal of this study, and is critical to decipher the pathogenesis of obesity-associated hypertension. Cell-attached voltage clamp recording of AgRP neurons in ARC sections of Ai9AgRP mice shows that ANG application results in three distinct electrical responses in lean mice. In particular, ANG causes inhibition in ≈1/3 of AgRP neurons, and this effect is abolished by losartan or genetic deletion of AT1A, supporting involvement of AT1A receptors (“Type 1i” responses; Firing rate; aCSF:0.85 ± 0.03 vs ANG:0.20 ± 0.04 vs ANG+LOS:0.91 ± 0.11 Hz, p<0.05). After HFD-induced obesity (DIO), roughly half of the Type 1i cells begin to exhibit AT1A-dependent stimulatory instead of inhibitory responses to ANG (“Type 1s”; Firing rate; aCSF: 0.86 ± 0.03 vs ANG:1.10 ± 0.12 vs ANG+LOS:0.82 ± 0.03 Hz, p<0.05). This implies a change in AT1A second-messenger coupling within the Type 1 neurons. ANG inhibition of Type 1i cells was blocked by pre-treatment with the Gαi inhibitor, pertussis toxin (PTX) but not by Gαq inhibitor, BIM-46187 (Δ Firing rate: PTX: -0.39 ± 0.09 Hz, p<0.05 vs BIM: 0.10 ± 0.09 Hz). In contrast, ANG stimulation of Type 1s cells was blocked by BIM-46187 and not by PTX (Δ Firing rate: PTX: 0.07 ± 0.06 Hz vs BIM: 0.49 ± 0.06 Hz, p<0.05). Stereotaxic delivery of AAV constructs encoding Cre-dependent DREADDs into the ARC of AT1A-Cre mice and specific activation of hM4Di (Gαi) within AT1A-expressing (Type 1i) AgRP neurons significantly increased RMR (n= 8-10, p<0.05) in both lean and obese mice. Activation of hM3Dq (Gαq) in Type 1i AgRP neurons did not alter RMR in obese mice.These findings establish a specific ANG-inhibited subtype of AgRP neuron and implicate the AT1A/Gαi signaling cascade ultimately controls RMR. AT1A “signal switching” from Gαi to Gαq cascade within these cells may underlie the pathogenesis of RMR adaptation. These results indicate that RMR adaptation is likely secondary to the loss of AT1R—Gαi coupling, and not the gain of AT1R—Gαq coupling. American Physiological Society and American Heart Association postdoctoral fellowship(KB) & R01 HL134850 (JLG) This is the full abstract presented at the American Physiology Summit 2023 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.
Bioengineering approaches to promote maturity in human skeletal muscle constructs
Authors: Tollitt Benjamin , Jones Samantha , Henstock James , McArdle Christopher , Hoettges Kai , Jackson Malcolm , McArdle Anne 
Published: May 2023

In 2021/22, the MicroAge project, a UK Space Agency-funded mission, sent 3D muscle constructs to the International Space Station to study microgravity as a model of accelerated musculoskeletal ageing on Earth. The aim of this work is to further develop the muscle constructs, compromised of human skeletal muscle cells encapsulated in fibrin hydrogels, induced to differentiate into myotubes and grown in 3D. The study uses several bioengineering approaches to determine the extent of maturity achievable in these muscle constructs in vitro, with the aim of producing a pharmacological or physiological screening platform of mature muscle, targeting muscle disorders such as sarcopenia.To enable measurement of muscle functional properties, it was also necessary to develop a suitable surrogate for force generation due to constraints of strain gauge use, and so this study also determined whether electrochemical impedance spectroscopy (EIS) was appropriate in numerous instances, opening up possibilities for rapid and automated measurement of contractile properties of in vitro skeletal muscle models. This was undertaken by the development of the MicroAge culture system to allow simultaneous measurement of force characteristic using a micro stain gauge and EIS measurements via the stimulating electrodes.The MicroAge muscle constructs have been characterised using a panel of maturation indices for skeletal muscle, including measuring contractile properties, and myosin heavy chain isoform expression using immunocytochemistry with spinning disc microscopy, RT-qPCR and Western blotting and these data will be presented.Of particular interest are the data that demonstrated a linear relationship between force generation measured using the ‘gold-standard’ approach of using a strain gauge and EIS (R2 = 0.9), validating the use of EIS as a proxy for force generation in a range of conditions such as in muscle constructs at rest and during contraction.This allows for rapid and automated measurement of contractile properties of in vitro models of skeletal muscle, providing an exciting new platform to rapidly test pharmacological interventions that may result in improved muscle function in 3D models of disease. The study is also currently implementing bioprinting of muscle constructs to streamline construct formation and investigating varying patterns of electrical stimulation on construct maturity throughout differentiation and the results of this approach will also be presented. Supported by a PhD studentship from MRC DiMeN Doctoral Training Programme and CIMA. This is the full abstract presented at the American Physiology Summit 2023 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.
Degeneration specific NF-L immunoreactivity in spinal cord and brainstem 10 days post-cervical spinal cord contusion
Authors: Fusco Anna , Rana Sabhya , Bindi Victoria , Sunshine Michael , Shaw Gerry , Fuller David 
Published: May 2023

We recently described a panel of novel antibodies to epitopes on the neurofilament light chain (NF-L) which are hidden in healthy neurons and their processes but specifically exposed during neurodegeneration ( doi.org/10.1101/2022.08.27.504533 ). We saw widespread immunopositivity for such antibodies in degenerating and degenerated processes in the rat spinal cord 1-5 days following cervical spinal cord injury (cSCI). Here we aimed to determine if such immunopositivity would persist at 10 days following cSCI. Male Sprague Dawley rats received laminectomy (n=3) or right unilateral contusion at C4 (n=3; 150kdyn; Infinite Horizon Impactor). Tissue was harvested 10 days post-SCI and stained with either Cresyl Violet for routine histology or for immunohistochemistry using mouse monoclonal antibody MCA-6H63, one of several novel antibodies binding to degeneration specific NF-L epitopes (EnCor Biotechnology Inc.). Spinal cord MCA-6H63 immunopositivity was not detected in spinal cords from the laminectomy group. Contusion produced the expected histopathology with ipsilateral grey and white matter disruption and some damage to the contralateral dorsal columns. Punctate labeling for MCA-6H63 was prominent at the lesion epicenter, as well as in the ipsilateral cord rostral and caudal to the injury, consistent with axonal degeneration. Sparse, punctate MCA-6H63 positivity was observed in the contralateral cervical white matter. Qualitatively, MCA-6H63 staining was most robust in the ventral and lateral white matter ipsilateral and caudal to the lesion. This location was consistent with ongoing Wallerian degeneration in prominent spinal tracts including spinothalamic, spinocerebellar, reticulospinal, vestibulospinal and raphespinal. Bilateral MCA-6H63 staining in the dorsal columns was present mostly rostral to the injury. Some MCA-6H63 staining was noted in the medulla, most prominently ipsilateral to the lesion. We conclude that at 10 days post-cSCI this novel NF-L antibody specifically identifies degenerating and degenerated axonal profiles. This is a promising technique in assessing neurodegeneration following SCI and other neurodegenerative diseases. R01HL139708-01A1 (DDF), R01 HL153140-01 (DDF), and EnCor Biotechnology Inc. This is the full abstract presented at the American Physiology Summit 2023 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.
Physiological effects of fentanyl in goats: acute withdrawal and known and novel reversal agents
Authors: Hodges Matthew , Neumueller Suzanne , Buiter Nicole , Hilbert Grace , Pan Lawrence , Lewis Stephen , Forster Hubert 
Published: May 2023

Synthetic opioids alter several vital physiological functions and discontinuation of its use can cause acute withdrawal symptoms. In this and three adjoining posters we present data on the dose-dependent physiologic and behavioral effects of intravenous (IV) fentanyl in adult awake goats. Here, we tested two hypotheses: 1) that fentanyl will lead to acute, dose-dependent withdrawal symptoms, and 2) that co-administration of a known reversal agent (naloxone (NAL)) or novel respiratory stimulant (D-cysteine ethyl ester; D-CYSee) can mitigate the acute physiological and behavioral effects of fentanyl. Acute withdrawal was measured for 4 hrs. beginning 90 min post-injection of IV vehicle or fentanyl (n=5-7) using a wearable activity tracking device (Biopac) and subjective videographic observations. Animal movement (Vector Magnitude; g) was lowest with vehicle across all time points but was increased (p<0.05) in a dose-dependent manner from 90-210 min after fentanyl administration. Additional behaviors such as pawing, vocalization, hyperphagia, itching and rearing also appeared dose-dependent and were increased from 90-210 min post-injection but waned across the 4h observation period (n=4). Based on the demonstrated efficacy of D-CYSee in reversing OIRD in rats (Getsy et al., 2022), we next tested if fentanyl-induced physiological dysfunction or withdrawal symptoms were mitigated by co-administration of either D-CYSee (500 μmol/kg) or NAL (0.06 mg/kg). Transient suppression of minute ventilation (VI) and breathing frequency was followed by a secondary increase >10 min after 50 μg/kg fentanyl, where co-administration of D-CYSee stimulated VI and NAL prevented the secondary increase in VI (n=5-6). Fentanyl alone moderately decreased HR where D-CYSee prevented this effect, whereas NAL exacerbated the fentanyl-induced bradycardia for up to 45 min. Mean, systolic and diastolic blood pressures were increased with fentanyl for >90 min post-injection, where D-CYSee had no additional effect but NAL prevented the hypertension. Finally, NAL but not D-CYSee reversed sedation (n=7), but neither NAL or D-CYSee appeared to mitigate withdrawal behaviors. These preliminary data suggest that: 1) fentanyl causes withdrawal behaviors in goats, and 2) NAL and D-CYSee have differential effects on fentanyl-induced physiological dysfunction suggesting D-CYSee has a distinctly different mechanism of action as a potential OIRD countermeasure. Supported by NIH DA050571. This is the full abstract presented at the American Physiology Summit 2023 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.
Caveolin-1 facilitates lipid uptake into endothelium by stabilizing Cd36 at the plasma membrane
Authors: Luse Melissa , Dunaway Luke , Pavelic Caitlin , Tessema Rachael , Juskiewicz Zuzanna , Minshall Richard , Leitnger Norbert , Isakson Brant 
Published: May 2023

Endothelial cells (ECs) can take up lipids from the circulation through Caveolin-1 (Cav1)-mediated endocytosis. In metabolic disease, fatty acids accumulate in blood plasma, creating a pathological role for ECs as lipid metabolizing cells. ScRNAseq performed on adipose endothelium shows an inverse relationship between Cav1 expression and dietary fat percentage; with increased dietary fat consumption, there is a decrease in Cav1 mRNA. To investigate the link between EC Cav1 and lipids we employed the use of an inducible EC-specific Cav1 knockout mouse (Cav1fl/fl/Cdh5-CreERT2+). Cav1 genetic deletion from endothelium was induced at 6 weeks, and mice were fed either a normal chow (NC; 5% fat) or high fat diet (HFD; 60% fat) with all mice examined at 20 weeks. Within diet groups, all mice regardless of genotype had comparable weight gain, gonadal fat pad mass, and food consumption. Following induction, electron microscopy was used to confirm presence of caveolae in adipocytes, but not endothelium. The gross vascular network of adipose, from all experimental mice, was visualized using fluorescence microscopy following tissue clearing of whole epididymal fat pads. Deletion of EC Cav1 resulted in increased blood lipids (triglycerides, cholesterol, LDL) independent of diet, suggesting Cav1-mediated endocytosis as a major mechanism of endothelial lipid uptake under normal and pathophysiological conditions. Further investigation determined that intact endothelium of adipose arteries from mice lacking EC Cav1 had decreased cellular lipid content when compared to mice with functional Cav1. Caveolae serve as signaling microdomains in ECs, leading us to question whether the lipid uptake impairments could be due to the failure to enrich proper fatty acid uptake machinery to caveolae decreasing circulating fatty acid uptake. We found Cd36, a known fatty acid translocase, to be located within these Cav1-mediated signaling microdomains via immune staining of an adipose artery en face and cellular membrane fractionation. EC Cav1 stabilizes Cd36 at the plasma membrane and with loss of Cav1, Cd36 gets recycled into the endosomal network. The decreased localization of Cd36 at the plasma membrane impairs endothelial lipid uptake and increases plasma lipids as seen in the EC Cav1 deficient mice. This work provides initial evidence for the unique metabolic roles of endothelial Research Support: T32 HL007284, AHA predoctoral fellowship, HL120840 (BEI), and Lipidema Foundation (BEI). This is the full abstract presented at the American Physiology Summit 2023 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.
HOX/PBX Inhibition Attenuates Tsc2-Null Cell Viability and Lung Metastasis in Lymphangioleiomyomatosis
Authors: Olatoke Tasnim , Astrinidis Aristotelis , Guo Minzhe , Zhang Erik , Zhang Alan , McCormack Francis , Xu Yan , Yu Jane , Wagner Andrew 
Published: May 2023

Lymphangioleiomyomatosis (LAM) is a rare lung disease that exclusively affects premenopausal women. LAM is caused by infiltration of metastasizing atypical smooth muscle-like cells into the lungs leading to progressive respiratory failure. Though the origin of these cells remains elusive, LAM cells carry loss-of-function mutations in the Tuberous Sclerosis Complex (TSC1/TSC2) genes, leading to hyperactivation of the mechanistic target of rapamycin complex 1 (mTORC1) and aberrant cell growth. There is no cure for LAM. The greatest barriers to finding a cure for LAM include its undetermined origin and unclear underlying pathogenesis. Our integrative single cell omics analyses of LAM lung identified the activation of uterine-similar HOX/PBX transcriptional network in LAM cells compared to other cells, suggesting that the uterus is the primary origin of LAM. HOX/PBX signaling is critical for female reproductive tract development and maintenance, and their overexpression is implicated in some female reproductive cancers. The objective of this study is to elucidate cellular and molecular mechanisms by which HOX/PBX signaling may regulate LAM pathogenesis, with hopes to uncover novel therapeutic strategies to cure LAM. We hypothesize that blockade of HOX/PBX attenuates LAM patient-derived (TSC2-null) cell viability and lung metastasis in LAM. We first confirmed HOX/PBX dimerization in LAM cells using immunofluorescence and immunoprecipitation assays. Using cell viability assays including crystal violet assay and MTT, we found that treatment with a cell-permeable HOX/PBX antagonist (HXR9) decreased TSC2-null cell growth and viability in vitro (*p<0.05, n=3/treatment group). We also showed that inhibiting HOX/PBX dimerization enhanced apoptosis and necroptosis in TSC2-null cells compared to controls using immunoblotting. Finally, using a xenograft mouse model of LAM, we tracked lung metastasis in vivo (Xenogen IVIS Spectrum System) and quantified total photon flux in the chest region. We found that pharmacologic and molecular suppression of HOX/PBX attenuates lung metastasis of LAM cells (*p<0.05, n=4/group). Our findings suggest that HOX/PBX signaling is critical for LAM progression and should be explored as a potential therapeutic target for LAM. Funding Sources: Public: National Heart Lung and Blood Institute (R01HL153045) and Department of Defense (W81XWH-19-1-0474). Private: LAM Foundation (LAM0141E01-20). This is the full abstract presented at the American Physiology Summit 2023 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.
Sex-specific effects of chronic unpredictable stress on mitochondrial function in the HPA axis
Authors: Crockett Alexia , Frambes Noelle , Stegmann Kyle , Churillo Amelia , Priviero Fernanda , Webb Clinton , Wood Susan , Spinale Francis , Ryan Michael , Hollis Fiona 
Published: May 2023

Background: Stress is a real or perceived threat that elicits physiological and behavioral responses via activation of innate response mechanisms like the hypothalamic-pituitary-adrenal (HPA) axis and sympathetic nervous system (SNS). Mitochondria are dynamic organelles responsible for providing cellular energy and the synthesis and release of glucocorticoids from the adrenal glands in response to stress. Thus, mitochondria represent the first line of response to environmental challenges, however, the effects of chronic stress on mitochondrial function in the HPA axis, particularly with respect to sex differences, are relatively unexplored. Hypothesis: Chronic unpredictable stress (CUS) induces mitochondrial dysfunction in key brain regions within the HPA axis.Methods: Adult male and female C57Bl6/J mice were exposed to 28 consecutive days of CUS or handling (CON) with weekly weighing. During the last week of stress, anxiety-like behavior was evaluated using the elevated plus maze and open field tests and coping behavior was assessed using the forced swim test. Mice were euthanized and the brain and adrenal glands dissected, weighed, and mitochondrial function was assessed using high resolution respirometry in hypothalamus, amygdala, and adrenal tissues. Results: CUS exposure decreased total body weight gain by 53% (p=0.04) and increased anxiety-like behavior (p=0.01) in stressed males compared to controls but had no effect on female weight or anxiety-like behavior. Stressed males and females both exhibited 46% decreased latency to immobility in the forced swim test (p=0.0001). CUS induced significant sex-dependent decreases in mitochondrial respiration within key stress centers, with stressed males exhibiting decreased respiration in the amygdala (20%, p=0.05), and stressed females exhibiting decreased respiration in the hypothalamus (20%, p=0.004) and adrenal glands (17%, p=0.06). Co nclusions: The unique findings of this study identified sex dependent CUS effects on HPA axis mitochondrial function. These observations suggest targeting mitochondria as a therapeutic strategy against stress-related pathologies. This work was supported by a NIH grants P20GM109091 (F.H.), R01HL130972-01A1 (F.G.S.), R01HL5949 (F.G.S.), RO1DK132948 (C.W. & F.P.) and R01 MH129798 (SKW); VA grants: VISN7 RDA (F.H.), Merit awards: BX000168-10A1 (F.G.S.), BX005320 (F.G.S.), & BX002604 (M.J.R.). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the abstract. This is the full abstract presented at the American Physiology Summit 2023 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.
Deletion of the stress response protein REDD1 prevents impaired cardiac function in diabetic mice
Authors: Stevens Shaunaci , Cavus Omer , Subrahmanian Sandeep , Sunilkumar Siddharth , Toro Allyson , Pfleger Jessica , Bradley Elisa , Dennis Michael 
Published: May 2023

The overwhelming impact of heart failure on morbidity and mortality in obesity and type 2 diabetes requires a deeper understanding of the molecular events that mediate cardiac dysfunction. We recently demonstrated using in vivo and in vitro models of diabetes that upregulation of endoplasmic reticulum stress in cardiomyocytes promoted expression of the stress response protein regulated in development and DNA damage 1 (REDD1). REDD1 has been implicated in diabetes-induced functional deficits in both the retina and kidney; however, a role for REDD1 in diabetes-induced cardiac dysfunction has not been investigated. Thus, we explored the hypothesis that REDD1 contributes to the development of diabetes-induced cardiac dysfunction. To induce diabetes, 6-week old male wild-type and REDD1 knock-out (KO) B6;129 mice were fed a high-fat high-sugar diet (42% kcal fat, 34% sucrose by weight) for 16 weeks; 2 weeks after initiation of the diet, mice were administered low-dose streptozotocin (STZ) for five consecutive days. Non-diabetic control mice were fed a standard rodent diet and received vehicle injections. Mice with a fasting blood glucose level >250mg/dL were defined as diabetic. At 10 weeks of age, transthoracic echocardiogram was evaluated every 2 weeks using a GE Logique small animal echocardiography machine. After 12 weeks of diabetes, left ventricular ejection fraction (EF) was reduced in diabetic wild-type mice as compared to non-diabetic wild-type mice (51.0 +/-9.8% versus 39.5 +/- 5.5%, p = 0.015). Meanwhile, EF in diabetic REDD1 KO mice (47.4 +/- 6.6%) was similar to that observed in non-diabetic REDD1 KO mice (49.6 +/- 6.7%). To investigate a potential cause of the preserved EF in diabetic REDD1 KO mice, tissue homogenates from the left ventricle were examined by PCR analysis. REDD1 mRNA expression in diabetic wild-type mice was enhanced as compared to non-diabetic controls in coordination with increased expression of Tgfb1, as well as expression of the pro-fibrotic genes Fn1, Col1a1, and Col3a1. By contrast, there was no increase in pro-fibrotic mRNA expression in cardiac tissue homogenates from diabetic REDD1 KO mice, as compared to non-diabetic REDD1 KO mice. Overall, the data support a role for REDD1 expression in the development of impaired cardiac function and the pro-fibrotic response of the heart to diabetes. This is the full abstract presented at the American Physiology Summit 2023 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.