bluuebunny/crossref_metadata_2025 huggingface dataset:
Using small group presentation of primary research literature to facilitate understanding and discussion in a graduate neurophysiology class
Authors: Knight Marvar Sarah , Knight Marvar Sarah
Published: May 2023

In order to stimulate a deeper understanding of physiology concepts in a graduate neurophysiology course, students analyzed and discussed primary research literature in the form of small group presentations. Our aim was to use original research articles to enhance understanding of major concepts and for students to gain historical context of seminal findings in neurophysiology. We hypothesized that giving students the opportunity to discuss the article in small groups, focusing on a specific section then presenting to the class before a large-group discussion would deepen their understanding of the material and build confidence in discussing scientific literature. Students were required to read and submit a short synopsis of a primary research article provided by the class instructor before class. In class, students were separated into groups of between 4-8 and allowed 1 hour to prepare a 5 - 10 minute presentation summarizing a section of the research article, such as the methods section. Following the small group presentations students were encouraged to discuss the findings and implications further. Students reflected on the use of primary research literature in the course and its impact on their learning.In a student survey 76% of students reported feeling more confident in reading, discussing and presenting primary research articles because of this course, whereas just 41% believed that reading primary research articles were a valuable part of the course and improved their understanding of the material. The inclusion of a group presentation of the material to their peers appeared to encourage deeper analysis, discussion and therefore better understanding of the literature. Whether that translated to deeper understanding of the physiological concepts of the class requires further study.

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.
Carbon dioxide attenuates PKC signaling and is sensed in monocytes and macrophages by a CA2-dependent, coupled intracellular pH-response
Authors: Strowitzki Moritz J. , Bleul Marc , Tuffs Christopher , Dupovac Mareen , Blanco Alfonso , Cummins Eoin P. , Schneider Martin , Taylor Cormac
Published: May 2023

Background: Increased levels of CO2 reduce macrophage function during inflammatory processes and wound healing. However, little is known how macrophages sense CO2 and adapt upon high levels of CO2 during differentiation and activation. We, therefore, elucidated the effects of CO2 on gene and protein expression during basic inflammatory processes, such as monocyte differentiation and macrophage activation. Methods: Monocyte differentiation was induced by phorbol 12-myristate 13-acetate (PMA). Primary macrophages (BMDMs) were polarized using different cytokines to induce proinflammatory (M1) macrophages (lipopolysaccharides) or immunomodulatory (M2) macrophages (interleukin-4). Cells were simultaneously subjected to different levels of CO2. Morphological changes, mRNA and protein expression of markers for cell differentiation and macrophage polarization were determined. Cell culture medium was buffered and intracellular pH (pHi) was monitored to determine pH-dependent effects of CO2. Direct effects of CO2 on PKC protein function were analyzed. Results: High levels of CO2 attenuated PMA-induced cell differentiation of human monocytes. In BMDMs, CO2 significantly reduced transcript levels of M1-marker. Mechanistically, CO2 mitigated PMA-stimulated PKC activity and function. Experiments with buffered medium revealed that changes in pH were responsible for most, but not all, of the CO2-mediated effects on monocyte differentiation and macrophage polarization. Pharmacological and genetic inhibition of carbonic anhydrases (CA) uncoupled the CO2-elicited intracellular pH-response, thereby preventing CO2-sensitivity during monocyte differentiation, activation, and migration, which could be reversed by CRISPR-mediated upregulation of CA2. Conclusion: Carbon dioxide is highly immunomodulatory and might thus affect wound healing properties in vivo. In M1 (but not M2) macrophages, CO2 is sensed by CA2-driven changes in pHi.

MJS received funding from the German Research Foundation (DFG; STR 1570/1-1) and the Braun Foundation (Braun®; BBST-D-18-00018).

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.
Chronic alcohol-induced mesenteric lymphatic leakage is associated with local and systemic metabolic dysregulation
Authors: Weaver Kourtney , Simon Liz , Molina Patricia , Souza-Smith Flavia
Published: May 2023

Introduction: Chronic alcohol administration produces significant metabolic dysfunction as determined in pre-clinical and clinical studies. Our studies using a rodent model of chronic alcohol feeding showed that alcohol induces mesenteric lymphatic leakage and perilymphatic adipose tissue (PLAT) immunometabolic dysregulation. Specifically, chronic alcohol induced localized PLAT inflammation and decreased glucose uptake. Whether chronic alcohol-induced lymphatic leakage is also associated with altered systemic glucose/insulin dynamics remained to be determined. We hypothesized that chronic alcohol feeding impairs the endocrine response to a glucose tolerance test. Methods: Male Fischer 344 rats were randomized to a Lieber-DeCarli liquid diet containing 36% of calories from alcohol or pair-fed an isocaloric diet for 10 weeks. Animals consumed approximately 75 mL of diet per day. Body weights were not significantly different between groups; the average weights were 275.7 grams for control animals and 275.9 grams for alcohol-fed animals. Weights in both groups remained stable for the duration of the study. At 10 weeks, animals were fasted for 16 hours prior to intraperitoneal (IP) injections of 20% glucose solution (2g/kg). Blood samples were collected from the rat tail prior to and at 15-minute intervals following initial glucose injection for 2 hours. Blood glucose was tested at each interval using the AlphaTRAK glucometer. 2-way ANOVA and paired T-test were used to assess significant differences between groups. Results and Conclusion: Prior to IP glucose injection, blood glucose levels averaged 77 mg/dL for control animals and 71.5 mg/dL for alcohol-fed animals. Glucose levels in alcohol-fed animals averaged 38% higher at 15 minutes and 37% higher 30 minutes after IP glucose bolus than that of control animals. Total area under the curve (AUC) calculated from the glucose concentrations measured following IP glucose injection was significantly higher in alcohol-fed animals compared to control animals; average AUC for alcohol-fed animals was 26.8% higher than AUC for control animals. These results suggest that chronic alcohol-induced mesenteric lymphatic leakage and localized PLAT inflammation and altered glucose metabolism are also associated with disturbances in systemic glucose dysregulation. Future studies will examine the potential link between alcohol-induced PLAT immunometabolic dysregulation and alterations in systemic glucose metabolism.

Supported by T32 (PM), K01 (FS).

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.
Contribution of local metabolic control to coronary pressure-flow autoregulation
Authors: Warne Cooper , Essajee Salman , Tucker Selina , Figueroa Alberto , Beard Daniel , Dick Gregory , Tune Johnathan
Published: May 2023

The coronary circulation has an innate ability to maintain constant blood flow over a wide range of perfusion pressures. This study interrogated the local metabolic hypothesis, which proposes that myocardial oxygen tension, indexed by coronary venous PO2 (CvPO2), determines the degree of coronary pressure-flow autoregulation by increasing the production of vasodilator metabolites as coronary perfusion pressure (CPP) is reduced. We tested this hypothesis by examining the extent to which dobutamine-induced increases in myocardial oxygen consumption (MVO2) under both normoxic and hypoxemic conditions, influences coronary autoregulatory capability. Experiments were performed on open-chest anesthetized swine during 20 mmHg stepwise changes in coronary perfusion pressure (CPP) from 140 to 40 mmHg, via servo-controlled roller pump. Measurements were made in 3 conditions: a) normoxia (n = 7); b) normoxia with dobutamine (10 μg/kg/min, iv; n = 7); and c) hypoxemia (PaO2 36-38 mmHg) plus dobutamine (n = 5). Under control-normoxic conditions, CvPO2 decreased from 38 ± 2 to 22 ± 1 mmHg and coronary blood flow fell from 0.93 ± 0.15 to 0.32 ± 0.03 ml/min/g as CPP was reduced from 140 to 40 mmHg. Administration of dobutamine significantly increased heart rate (~60%; P < 0.001), but the combination of hypoxemia plus dobutamine did not further influence heart rate. Further, dobutamine with and without hypoxemia significantly increased MVO2 (P < 0.001); however, CvPO2 was reduced (P < 0.001) only in the presence of hypoxemia. Calculation of closed-loop autoregulatory gain (Gc) over a CPP range of 120 to 60 mmHg (value of 1 represents perfect autoregulation) was unaffected by dobutamine alone but tended to increase when dobutamine infusion was combined with reductions in PaO2 ≤ 40 mmHg (P = 0.069). Additional analysis of the autoregulatory slope within this same CPP range revealed no differences between groups. However, there was a significant inverse association between Gc and CvPO2, measured at CPP of 100 mmHg, in the absence and presence of dobutamine-induced increases in MVO2 ± hypoxemic conditions (P < 0.001, r = -0.722). The relationship of coronary resistance (CPP range of 120 to 60 mmHg) relative to respective CvPO2 revealed that decreases in CvPO2 were not associated with changes in coronary resistance until CvPO2 fell below a relative threshold of ~ 25 mmHg (r = 0.88). These findings support the interpretation that the sensitivity of coronary autoregulatory behavior is directly coupled with the activation of local metabolic vasodilator pathways below a critical level of myocardial oxygenation.

This work was supported by National Institutes of Health grant R01 HL158723.

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.
Alpha-glucosidase inhibitor mitigates bone loss in type-1 diabetes
Authors: Juin Subir Kumar , Pushpakumar Sathnur , Sen Utpal
Published: May 2023

Diabetic nephropathy is a hallmark of diabetes mellitus (DM) and is characterized by kidney dysfunction. The kidney plays an important role in bone health by preserving the equilibrium of minerals viz., calcium and phosphate in the blood and 1,25-Dihydoxyvitamin D3 (Calcitriol) production. Previous studies have suggested that renal dysfunction is associated with dysregulated mineral reabsorption due to mineral-hormone imbalance, leading to bone loss and increased fracture risk in DM. Alpha-glucosidase inhibitor (AGI) is known to decelerate carbohydrate catabolism and delay glucose production to improve overall diabetic health. Nimbidiol is an AGI derived from the medicinal plant, Azadirachta indica and is considered a potential anti-diabetic natural compound. The purpose of our present study was to investigate whether AGI mitigates the imbalance of mineral homeostasis and eventual bone loss in type-1 diabetes. Twelve - fourteen weeks old wild-type, C57BL/6J (WT) and type-1 diabetic, C57BL6/‐ Ins2Akita /J (Akita) mice were either treated with saline or AGI (0.40 mg kg-1 d-1) by subcutaneous implantation of micro-osmotic pump for eight weeks. Diabetic Akita mice showed a distinct downregulation of sodium-phosphate co-transporter, Npt2a expression in the renal tubules compared to the WT mice. In addition, Akita mice exhibited a significant increase in alkaline phosphatase activity and decrease in Calcitriol and bone minerals such as calcium and phosphate levels in the blood. The changes were associated with a significant bone loss as evidenced by the increased thinning and porosity of cortical and trabecular bone and reduction of osteoblasts and osteocytes in diabetic femur. AGI treatment mitigated the pathological changes in Akita mice. Taken together, our results suggest that AGI preserves mineral homeostasis and thereby protects from bone loss in type-1 diabetes.

DK116591

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.
Endothelial Cell-Selective Adhesion Molecule Enhances Tumor Growth in a HFpEF Mouse Model
Authors: Lang Liwei , Ajith Ashwin , Horuzsko Anatolij , Bagi Zsolt
Published: May 2023

Accumulating epidemiological evidence shows that the incidence of multiple types of cancer, such as melanoma and GI cancers, is significantly higher in patients with heart failure with preserved ejection fraction (HFpEF). However, the mechanistic relationships between HFpEF and cancer development are poorly understood. Impaired angiogenesis, upregulation of endothelial adhesion molecules and chronic low-grade inflammation are common in HFpEF. Here we examined the role of endothelial cell-selective adhesion molecule (ESAM) in the progression of malignant melanoma in a HFpEF mouse model. Orthotopic melanoma xenografts (B16F10-Luc2) were used and implanted subcutaneously in mice underwent uninephrectomy and aldosterone infusion (UNX-Aldo with 1% high salt drinking water for 4 weeks) and tumor growth was monitored with bioluminescent imaging. We found that ESAM expression and serum levels were elevated in the UNX-Aldo mice, which was accompanied by enhanced melanoma growth and metastasis rate. ESAM promoted tube formation and increased phospho-Akt, phospho-PKCα and phospho-ERK1/2 in cultured endothelial cells. Flow cytometry analysis showed elevated number of tumor-associated neutrophils (CD11b+Ly6G+Ly6C-) and myeloid-derived suppressor cells (MDSCs, CD11b+F4/80-Gr-1+) in the UNX-Aldo mice. UNX-Aldo mice with genetic deletion of ESAM (ESAM KO) showed a reduced tumor growth and reduced number of tumor-associated neutrophils and MDSCs. Collectively, our results indicate that in HFpEF, a proinflammatory ESAM activation may act to promote tumor angiogenesis, neutrophil and MDSCs tumor infiltration, thereby mitigating a proper immune response and facilitate tumor growth.

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.
A circadian oscillator regulates a food anticipatory increase in heart rate and body temperature
Authors: Prabhat Abhilash , Schroder Elizabeth , Stumpf Isabel , Stalcup Rebecca , Seward Tanya , Pendergast Julie , Delisle Brian
Published: May 2023

Objective: The daily rhythm in heart rate is classically considered to be regulated by the circadian clock in the suprachiasmatic nucleus (SCN). The circadian clock in the SCN entrains to the light-dark cycle. However, some circadian rhythms can be entrained to meal timing independently of the SCN. In this study, we tested the hypothesis that heart rate is entrained by meal timing. Methodology: Wild-type male and female mice were implanted with telemetry devices to continually record HR and CBT in thermoneutral conditions. The mice were initially housed in a 12 h light: 12 h dark cycle (LD) with ad libitum access to food (ALF) for 7 days. Mice were then subjected to light cycle time-restricted feeding (TRF) between hours 2 and 9 (hour 0: lights on; hour 12: lights off) for 5 days, followed by 3 days in constant darkness (DD). The mice were then fed ALF for 3 days in DD to determine if food anticipatory rhythms persisted in constant conditions. Results: During ALF feeding in LD, HR and CBT increased at hours 11-13 and corresponded with the onset of nocturnal activity. In contrast, after 1 day of light cycle TRF, HR increased during the 2 hours prior to mealtime between hours 0-2. The food anticipatory increase in HR persisted during TRF in DD. The food-entrained anticipatory increase in HR and CBT remained phase-locked to prior mealtime when mice were returned to ALF after TRF. In contrast to the food-entrained anticipatory increase in HR and CBT, the circadian rhythms in HR and CBT dissociated and shifted back to the phase seen in ALF in LD. There were no sex differences. Conclusions: These data demonstrate that male and female mice have food-anticipatory increases in HR and CBT. Entrainment of the food anticipatory increase in HR and CBT to meal timing during TRF persisted in the absence of zeitgebers (in constant feeding and lighting conditions). Together these findings suggest that a food-entertainable oscillator regulates food-anticipatory increase HR and CBT rhythms independently of the SCN.

Funding sources: The present work is supported by the National Heart Lung and Blood Institute grants R01HL153042 and R01HL141343.

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.
Avian mitochondria respiration is not predicated solely on complex abundance and DNA content
Authors: Yen Con-Ning , Bodmer Jocelyn , Wicks Jordan , Shi Tim , Gerrard David
Published: May 2023

Skeletal muscle is a collection of heterogenous muscle fiber types differing in their ability to contract and metabolize energy substrates. The relative proportion of each fiber type in muscle drive function, where oxidative muscles are more fatigue resistant and generally have greater amounts of mitochondria than more glycolytic muscles that have less mitochondria because of their reliance on glycolytic reactions for energy production. However, even highly glycolytic muscles, like the breast muscle of chickens have mitochondria. Yet, differences in mitochondrial characteristics across muscle types remain largely unknown.We compared glycolytic and oxidative muscles of two different species in an attempt to understand better the relationship between mitochondrial DNA (mtDNA) and protein abundance, and mitochondria function. Glycolytic muscles longissimus lumborum (LL) and the pectoralis major (PM) and oxidative muscles masseter (MS) and quadriceps femoris (QF) were collected from adult pigs and chickens, respectively. Greater mtDNA abundance (2-ΔΔCT) was noted in oxidative muscles of both species compared to their glycolytic counterparts ( P < 0.01). However, when absolute mtDNA was estimated, porcine MS had more mtDNA than LL ( P < 0.001), whereas both avian muscles were equivalent. Porcine MS muscle had greater succinate dehydrogenase (SDHA, P < 0.05) and citrate synthase (CS, P < 0.001) than LL, while avian QF muscle had greater CS ( P < 0.001) and voltage dependent anion channel (VDAC, P < 0.05) compared to PM muscle. Isolated mitochondria from the MS of pigs had more electron transport chain complex I (CI, P < 0.010) and complex II (CII, P < 0.05) than those isolated from the LL. Avian mitochondria had no differences in either electron transport chain complexes across muscle, but mitochondria from the PM had greater VDAC than those purified from the QF. Oxygen consumption rates (OCR) followed muscle type, where mitochondria from oxidative muscles consumed more oxygen than those of glycolytic muscle when stimulated with ADP ( P < 0.05). These data show that porcine mtDNA abundance, mitochondrial protein abundance, and mitochondrial function is greater in oxidative muscle compared to glycolytic muscle, and that respiration is greater in mitochondria from more oxidative muscle. However, differences in the relative abundance of mtDNA and electron transport chain complexes were not evident in isolated mitochondria from the chicken, yet respired differently nonetheless suggesting mtDNA and protein may not be directly correlated to the overall function of mitochondria.

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.
A humanized protease-activated receptor-2 mouse as a model for allergic asthma
Authors: Gillman Samuel , Schiff Hillary , Pederson Wil , Sandoval Estevan , Rivera Benjamin , Dussor Gregory , Price Theodore , Vagner Josef , Defea Kathryn , Boitano Scott
Published: May 2023

Asthma is a complex disease characterized by airway hyperresponsiveness (AHR) inflammation, and mucus overproduction, and has differential presentation in males and females. There exists strong demand to improve upon current asthma treatments by targeting upstream signaling pathways, including the G-protein coupled receptor protease-activated receptor-2 (PAR2). We have recently shown effectiveness of full (C391) and ß-arrestin biased (C781) PAR2 antagonism in limiting allergen-induced asthma indicators in mouse models (C57Bl/6 and Balb/C). To better test PAR2 antagonists as asthma drugs, we have developed a transgenic mouse model that expresses human PAR2 without mouse PAR2 expression in a C57Bl/6 background (htgPAR2). The htgPAR2 mice have heightened responses to asthma allergens [house dust mite (HDM) or Alternaria alternata] following acute exposure. Limited sex differences were observed in the HDM challenge model. However, when challenged with A. alternata there was significantly higher AHR in the male mice and significantly higher inflammatory and mucus responses in the female mice. The full and ß-arrestin biased PAR2 antagonists also displayed differential effects with C781 providing broader control of allergen-induced indicators (AHR, inflammation and mucus overproduction). We conclude that the htgPAR2 mouse model is a promising tool for preclinical in vivo screening of potential asthma treatments targeting PAR2.

Grants from the National Institute of Health (NS098826, AI140257, HL16024, HL152942)

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.
Seasonal changes in the metabolomic profile of white adipose tissue in hibernating thirteen-lined ground squirrels
Authors: Heinis Frazer , Alvarez Sophie , Rugira Kevin , McMurchie Ashley , Schuster Olivia , Andrews Matthew
Published: May 2023

Hypothesis: Hibernator white adipose tissue will exhibit season-dependent remodeling of metabolic pathways. Methods: Thirteen-lined ground squirrels were captured near Lincoln, Nebraska, and housed in University of Nebraska-Lincoln veterinary facilities. Seasonal adjustments in environmental chamber lighting and air temperature were used to induce hibernation, and tissues were taken at different timepoints throughout the seasonal activity-torpor cycle. Polar compounds were extracted from white adipose tissue (N=47 across five groups) and analyzed by hydrophilic interaction liquid chromatography-mass spectrometry (HILIC-MS) in both positive and negative ion modes. Spectra were aligned, and peaks detected and identified using MS2 libraries search (mzCloud, MoNA, in-house) with Compound Discoverer (Thermo). Normalized and QC corrected feature intensities were processed using MetaboAnalyst 5.0 for statistical analysis and pathway enrichment. Results: We identified 185 compounds and 85 compounds from positive and negative HILIC-MS, respectively, including 15 compounds identified in both ion modes. Unsupervised clustering using principal component analysis revealed discrete clusters representing the different seasonal timepoints throughout hibernation. One-way analysis of variance and feature intensity heatmaps revealed metabolites that varied in abundance between active and torpid timepoints. Pathway analysis compared against the KEGG database demonstrated enrichment of amino acid metabolism and aminoacyl-tRNA biosynthesis pathways among our identified compounds. Carnitine derivatives and ketones were among molecules found to be elevated during torpor-associated timepoints relative to the active state. Summary and Conclusion: We report here the polar metabolomic profile of ground squirrel white adipose tissue at different points throughout the seasonal hibernation cycle. We identified 255 compounds and showed seasonal differences in white adipose metabolites indicating a shift of hibernator metabolism associated with torpor. Our findings indicate it is possible that white adipose directly produces ketones, rather than solely relying on hepatic ketogenesis. As our results represent steady-state measurements of metabolite abundance, further work is currently underway to identify whether hibernator adipose tissue can directly synthesize these compounds and contribute to the elevated plasma ketone concentrations during torpor observed in previous work.

Funding: University of Nebraska

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.
Chronic RAS Inhibition Prevents Aberrant Irritant-evoked Pulmonary-Cardiac Reflexes in the Spontaneously Hypertensive (SH) Rat
Authors: Hooper Justin , Taylor-Clark Thomas
Published: May 2023

Cardiopulmonary reflexes evoked by activation of airway vagal afferent nerves via irritant/pollution inhalation are remodeled from bradycardia in normal subjects to tachycardia and ventricular arrhythmia in patients with pre-existing cardiovascular disease (CVD). We have shown similar remodeling in the spontaneously hypertensive (SH) rat: selective activation of airway vagal afferent nerves via inhalation of the transient receptor potential ankyrin 1 (TRPA1)-specific agonist allyl isothiocyanate (AITC) evoked parasympathetic-mediated reflex bradycardia in normotensive wistar-kyoto rats but evoked sympathetically-mediated tachycardia and premature ventricular contractions (PVCs) in spontaneously hypertensive (SH) rats. Hypertension in the SH rat develops with age (~7 weeks) and this is accompanied by autonomic dysfunction.The renin-angiotensin system (RAS) is closely linked with blood pressure (BP), cardiac remodeling and autonomic dysfunction, with the main effector being angiotensin II (Ang II), produced downstream of angiotensin converting enzyme (ACE). Acute treatment with ACE inhibitors or antagonists of Ang II-sensitive receptor AT1R causes temporary BP normalization in SH rats. Chronic treatment with ACE inhibitors or AT1R inhibitors prevents the development of hypertension, baroreceptor dysfunction and cardiac remodeling in SH rats. Previously, we have shown that acute normalization of BP with the ACE inhibitor captopril had no effect on the AITC-evoked pulmonary-cardiac reflexes in the SH rat. Here, we investigated the impact of chronic ACE inhibitor treatment (captopril, 20mg/kg/ ad libitum) or chronic AT1R treatment (losartan, (15mg/kg/ ad libitum) from 6-15 weeks of age in SH rats on AITC-evoked pulmonary-cardiac reflexes. Both chronic captopril and losartan normalized BP and prevented AITC-evoked tachy-arrhythmia (% of tachycardic beats and PVCs) in SH rats at 15 weeks of age. As such, following antiRAS treatment, AITC only evoked reflex bradycardia in SH rats (similar to our previous studies of normotensive rats). These findings suggest that the remodeling of the irritant-evoked pulmonary-cardiac reflexes can be prevented by targeting the renin-angiotensin system. Reducing tachyarrhythmia evoked by the inhalation of irritants/pollution in susceptible individuals may reduce the risk of CVD exacerbations associated with air pollution.

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.
Characterizing the role of pulmonary veins in pulmonary hypertension
Authors: Bazaz Abhishek , Chen Yen-Lin , Daneva Zdravka , Sonkusare Swapnil
Published: May 2023

Pulmonary veins (PVs) transport oxygenated blood from lungs back to the heart. Small PVs can play a crucial role in the pathogenesis of pulmonary hypertension. Studies have also shown PV remodeling in pulmonary hypertension and heart failure. However, unlike pulmonary arteries (PAs), which have been a major focus of pulmonary vascular studies, the signaling mechanisms in PVs have not been investigated under normal conditions and in pulmonary hypertension. We hypothesized that the vascular wall composition and cellular signaling mechanisms in PVs are distinct from PAs. We imaged 4th order PVs (~ 50 mm) from C57BL6 mice using spinning disk confocal imaging along the z-axis. In direct contrast to PAs, where a single endothelial cell (EC) layer is surrounded by alpha actin-positive smooth muscle cells (SMCs), PV walls did not show SMCs. Cardiac troponin C staining of PVs identified cardiac myocyte-like (CML) cells with striations surrounding the EC layer. The CML layer was not observed in small PAs, vena cava, and mesenteric veins. Confocal calcium imaging showed spontaneous calcium signals in CML cells from PVs that were inhibited by ryanodine (1 mM), a ryanodine receptor inhibitor. Cannulated PVs in pressure myography experiments showed spontaneous pulsatile constrictions, which were also inhibited by ryanodine. Collectively, our results suggest that ryanodine receptor calcium signals in CML cells underlie spontaneous contractions in small PVs. Our findings also indicate that signaling mechanisms in CML cells could be potential targets for therapy in pulmonary hypertension.

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.
Neonatal Alveolar Type 1 Cells Produce a Pro-inflammatory Response to LPS and TNF-alpha
Authors: Mohammadi Atefeh , Higazy Randa , Ivanovska Julijana , Tran Harvard , Ivanovski Nikola , Gauda Estelle
Published: May 2023

Introduction: Bronchopulmonary dysplasia (BPD) is the most common lung disease in premature infants. Inflammation and oxidative stress during the early stages of lung development damage alveolar Type 1 (AT1) and Type 2 (AT2) epithelial cells that line the alveoli, leading to decreased alveolarization and surfactant production. Unlike AT2 cells, the role of AT1 cells in inflammation is not understood. Since inflammation initiates BPD pathogenesis, we sought to characterize the profile of inflammatory markers produced by neonatal AT1 cells in response to lipopolysaccharide (LPS) and tumour necrosis factor α (TNF-α). Methods: AT1 cells were isolated from neonatal rats on postnatal day 4 using fluorescence activated cell sorting. Cells were serum starved for 24hrs and then exposed to LPS (5, 10, 25μg/mL), TNF-α (5, 10, 25ng/mL), or a combination of LPS and TNF-α (10μg/mL and 5ng/mL) for 18h in vitro. Conditioned media was collected for cytokine and chemokine analysis by multiplex ELISA. Results: AT1 cells exposed to both LPS and TNF-α produced a larger inflammatory response than those exposed to LPS or TNF-α alone. After 18h, AT1 cells from the combined LPS and TNF-α group increased the production of IL-6 by 243-fold, CCL2 by 34-fold, and CCL3 by 9-fold in comparison to the control (all p<0.05, n=4). IL-1β and IFN-γ, and the anti-inflammatory cytokines, IL-4 and IL-10, were not detected in any experimental groups at this time point. Conclusion: This is the first study that characterizes the response of neonatal AT1 cells to inflammatory stimuli. Our preliminary findings show that LPS and TNF-α stimulate the production of the pro-inflammatory cytokines and chemokines IL-6, CCL2, and CCL3 in primary AT1 cells. These inflammatory markers are strongly associated with the development of BPD and are predictors of adverse pulmonary outcomes in neonates. Developing therapies that regulate the inflammatory response produced by AT1 cells may be beneficial in mitigating BPD pathogenesis.

This work was funded by the Women's Auxiliary of the Hospital for Sick Children.

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.
Role of Piezo2 in mechanosensation by vagal esophageal afferent fibers
Authors: Nair Sanjay , Hadley Stephen , Patil Mayur , Taylor-Clark Thomas
Published: May 2023

The esophagus is densely innervated by extrinsic sensory afferents projected from the dorsal root and vagal ganglia capable of sensing mechanical and chemical stimuli. In particular a major subset of vagal afferents innervating the esophagus are low-threshold mechanoreceptors whose activation by esophageal stretch regulates homeostatic processes like peristalsis and upper and lower sphincter muscle movements in addition to swallowing and vomiting reflexes. Importantly, the identity of specific proteins that mediate mechanotransduction within the esophageal afferents is not known. Previous studies have identified Piezo2 as a mechanosensitive ion channel expressed in various sensory afferent subsets that is required for sensing lung and bladder inflation. Hence, we hypothesize that esophageal afferent fibers from the vagal ganglia sense mechanical distention through mechanically-gated Piezo2. Here, we used two-photon imaging of neuronal GCaMP6s fluorescence in an ex vivo vagal-esophageal preparation to measure the activation of esophageal afferents in response to pressure-evoked distension of the esophagus. For control studies, GCaMP6s was expressed from the knockin ROSA26 gene under the control of cre recombinase (B6.129S6-Gt(ROSA)26Sor-tm96(CAG-CGaMP6s)Hze), following cross with the Pirt-cre (which expresses Cre in all sensory neurons). To assess the role of Piezo2, we injected retrograde AAV (rgAAV2) vector encoding Cre recombinase with a tdTomato reporter into the esophagus of Piezo2fl/fl mice, combined with an intranodose injection of an AAV9 vector encoding GCaMP6s Ca2+-indicator. Only tdTomato-expressing neurons (i.e. cre expressing) were included in the GCaMP6s analysis of the Piezo2fl/fl mice. Our preliminary data from PIRT-GCaMP6s mice studies show that distention of the esophagus with 5, 10 and 30 mmHg caused profound Ca2+ influx in a select population of neurons in the vagal (nodose) ganglia (118 neurons analyzed), consistent with our previous electrophysiological studies of tension mechanoreceptors. However, we saw a reduction of >85% in the Ca2+ influx in response to esophageal distention (5, 10 and 30 mmHg) in the tdTomato+ esophageal afferent population following cre-mediated knockout of the Piezo2 channel (99 neurons analyzed). Our findings support our hypothesis that Piezo2 is required for sensing mechanical distention by esophageal afferents arising from the nodose ganglia.

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.
Nlrp3 inflammasome in the heart of hypothyroid mice
Authors: Parletta Aline , Vieira Junior Denival , Senger Nathalia , Barreto-Chaves Maria Luiza
Published: May 2023

Cardiovascular diseases are the most common cause of death worldwide and thyroid diseases are frequently associated with cardiac function impairment. Hypothyroidism is related to dilation of the ventricles, fibrosis and consequent reduction in cardiac contractility, as well as an increased risk of heart failure. Recent studies have demonstrated the participation of the NLRP3 inflammasome, an important mechanism of the innate immune response, in cardiovascular diseases, such as myocardial infarction and atherosclerosis. There is evidence in the literature that hypothyroidism is accompanied by increased levels of cytokines and oxidative stress, processes that can activate the inflammasome. The aim of the present study is to evaluate the role of the NLRP3 inflammasome in the heart of hypothyroid mice. For this, Wild Type (WT) mice treated with methimazole (0.05% added to drinking water for 12 weeks) were used (N=4-6). All protocols used in this study were approved by the Animal Ethics Committee of the Institute of Biomedical Sciences (9768300119). Thyroid hormones were quantified by bead-based immunoassay Luminex to confirm the efficacy of experimental model. Fibrosis was analyzed by histology, using picrosirius red staining. Cardiac function was evaluated in mice by plethysmography and in isolated hearts after ischemia/reperfusion by the Langendorff method. Protein expression of NLRP3 inflammasome components was evaluated by Western Blotting. Data were analyzed by one-way ANOVA or Student's t test. Results are presented as mean ± standard error of the mean, and p<0.05 was considered statistically significant. Increased TSH levels (control=92±33, hypo=369±62 (pg/ml)) and reduced T4 (control=1.16±0.38, hypo=0.03±0.01 (ng/ml)) confirmed the hypothyroid status of mice. The heart weight/tibial length ratio and fibrosis were not altered by hypothyroidism. Heart rate was reduced in hypothyroid mice (control=663±27, hypo=541±28 (bpm)). No differences were detected in the cardiac function parameters after Ischemia-Reperfusion injury: left ventricular developed pressure (LVDP), maximum (dP/dtmax) or minimum first derivative (dP/dtmin). Concerning to inflammasome, NLRP3 expression was reduced (control=100±16, hypo=29±14 (%)) and pro-caspase 1 increased (control=100±17, hypo=259±43 (%)) in hypothyroid hearts. These preliminary results showed characterization data for experimental hypothyroidism and that this condition can modulate inflammasome expression. Additional experiments are underway to confirm the role of the NLRP3 inflammasome on cardiac function in hypothyroid mice.

FAPESP (grants nº 2021/06151-7 and 2019/17031-2)

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.
Development of an inquiry-based physiology lab activity analyzing salivary cortisol
Authors: Walton Kristen
Published: May 2023

Many of the classic lab activities in undergraduate animal or human physiology courses use specialized physiology recording equipment and software, and provide valuable illustrations of complex physiological processes in nerves, muscles, and the heart. However, some of the basic skills and equipment typical of many bench research labs are not as frequently represented. With a goal of increasing the diversity of common laboratory skills gained by students in our upper-division animal physiology course, as well as expanding lab activity options in endocrine physiology, a lab activity centered on measuring salivary cortisol was designed, using a commercially available, 96-well plate-based, relatively inexpensive enzyme immunoassay (EIA) kit. This activity was designed to give students the opportunity to learn the principles of EIA, which is commonly used for measuring hormones or other soluble molecules, as well as gain experience in pipetting, using a plate reader, and graphing data. This project was approved by the Missouri Western State University IRB. Students who volunteered to provide saliva samples were given saliva collection vials and instructions on sample collection. Samples were stored and processed according to the manufacturer’s instructions. The next week, student groups performed the cortisol immunoassay according to the kit instructions and calculated cortisol concentrations in the saliva samples based on the standard curve. In one semester, students collected saliva samples in the morning and evening, and determined whether their measured cortisol levels aligned with the expected diurnal variation in cortisol levels. In the next two course offerings, in a more inquiry-based design, students were given unlabeled samples and were asked to predict, based on the measured cortisol concentration, whether the samples may have been collected in the morning or evening. Other possible directions for a more inquiry-based lab activity could include testing samples collected before and after an intervention such as heavy exercise, studying for an exam, or relaxation or meditation techniques. When surveyed about their opinions on this lab activity, 63% of students agreed or strongly agreed that they liked the activity, and 69% agreed or strongly agreed that they learned new skills and/or understood concepts better after this lab activity. Students who disliked the activity mainly commented about the long wait time (a 45 minute incubation). In summary, this salivary cortisol enzyme immunoassay is a relatively inexpensive lab activity that is relatively well-received by students, and is feasible in the context of an undergraduate physiology lab. This activity adds to the body of options for lab activities in endocrine physiology.

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.
The effect of low-density lipoprotein cholesterol on T-cell mitochondrial respiration
Authors: DeConne Theodore , Ward Houston , Edwards David , Fancher Ibra , Trott Daniel , Martens Christopher
Published: May 2023

Aging is associated with an increase in chronic, sterile, low-grade inflammation, known as “inflammaging” that is implicated in the development of multiple diseases, including cardiovascular disease. Immune cells are the primary source of inflammaging, of which T-cells play a major contributing role. Mitochondria are essential for maintaining proper T-cell function and reduced mitochondrial respiration (MitoRESP) induces T-cell dysfunction and shifts T-cells towards a pro-inflammatory phenotype. Elevated low-density lipoprotein cholesterol (LDL-C) is cross-sectionally associated with lower MitoRESP in circulating immune cells and may contribute to the inflammaging phenotype, but whether LDL-C is causally related to impaired MitoRESP in T-cells is not known. We hypothesized that treating T-cells in vitro with a high concentration of exogenous LDL-C would mimic accelerated aging by impairing T-cell MitoRESP from young adults. Seven young adults were recruited for this study (6 female / 1 male, age: 23 ± 2 years, BMI: 23.29 ± 6.4 kg/m2, total cholesterol: 154 ± 16 mg/dl, high-density lipoprotein cholesterol: 59 ± 17 mg/dL, & LDL-C: 78 ± 14 mg/dL). CD4+ and CD8+ T-cells were separated from peripheral blood mononuclear cells isolated using magnetic bead separation (Miltenyi). CD4+ and CD8+ T-cells were treated exogenously with a physiologically normal (1.8 mMol/L) and high (4.9 mMol/L) concentration of LDL-C for 24 hours. MitoRESP was assessed using extracellular flux analysis (Agilent). Compared with the low LDL condition, the high LDL-C condition lowered basal (0.47 ± 0.06 vs. 0.35 ± 0.10 pMol/min/105 cells, p=0.031) and ATP-linked oxygen consumption rate (OCR) (0.51 ± 0.13 vs. 0.35 ± 0.12 pMol/min/105 cells, p=0.016) in CD8+ T-cells. High LDL-C also lowered non-mitochondrial OCR (0.71 ± 0.22 vs. 0.57 ± 0.20 pMol/min/105 cells, p=0.016) and augmented basal glycolysis (0.097 ± 0.049 vs. 0.14 ± 0.036 pMol/min/105 cells, p=0.031) in CD8+ T-cells. High LDL-C lowered ATP-linked OCR (0.69 ± 0.23 vs. 0.54 ± 0.22 pMol/min/105 cells, p=0.031) and non-mitochondrial OCR (0.80 ± 0.17 vs. 0.68 ± 0.21 pMol/min/105 cells, p=0.016), but did not statistically alter basal OCR (0.47 ± 0.056 vs. 0.35 ± 0.10 pMol/min/105 cells, p=0.063) or basal glycolysis (0.14 ± 0.031 vs. 0.18 ± 0.028 pMol/min/105 cells, p=0.063) in CD4+ T-cells. Treatment with a high LDL-C accelerated aging in T-cells from young adults by impairing MitoRESP and shifting T-cells towards glycolytic metabolism. Future studies should explore the relation between age-related impairments in T-cell MitoRESP and cardiovascular function in humans.

Funding Sources: P20GM113125

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.