bluuebunny/crossref_metadata_2025 huggingface dataset:
Tonic GABAergic signaling from prostaglandin EP3 receptor-expressing preoptic neurons bidirectionally controls body temperature
Authors: Nakamura Kazuhiro , Nakamura Yoshiko , Yahiro Takaki , Fukushima Akihiro , Kataoka Naoya , Hioki Hiroyuki
Published: May 2023

The principal controller neurons of the thermoregulatory center in the preoptic area (POA) have yet to be determined. In the present study using rats, we discovered that prostaglandin EP3 receptor-expressing POA neurons (POAEP3R neurons) are a pivotal bidirectional controller in the central thermoregulatory circuit mechanism. POAEP3R neurons were activated in response to elevated ambient temperature, but inhibited by prostaglandin E2, a pyrogenic mediator. In newly generated Ptger3 (EP3R)-tTA transgenic rats, chemogenetic stimulation of POAEP3R neurons at room temperature reduced body temperature by eliciting skin vasodilation (enhancing heat dissipation), whereas inhibition of them elicited hyperthermia involving brown adipose tissue thermogenesis and tachycardia, mimicking fever. Furthermore, we revealed that POAEP3R neurons innervate sympathoexcitatory neurons in the dorsomedial hypothalamus (DMH) via tonic inhibitory signaling. Although many POAEP3R neuronal cell bodies expressed a glutamatergic mRNA marker, paradoxically, most of their axons in the DMH contained GABAergic synaptic proteins. Interestingly, such GABAergic terminals of POAEP3R→DMH axons were increased by chronic heat exposure of rats, indicating a synaptic alteration to enhance heat tolerance. Consistent with the anatomical observation, slice patch-clamp recordings from DMH neurons combined with optogenetic stimulation of POAEP3R→DMH axon terminals showed that these terminals predominantly form GABAergic synapses onto DMH neurons. These findings demonstrate that tonic GABAergic inhibitory signaling from POAEP3R neurons is a fundamental determinant of body temperature for thermal homeostasis and fever.

NEXT Program from JSPS (LS070 to K.N.); MEXT KAKENHI (JP21K06767, JP17K08568, JP26860159, JP23790271 to Y.N.; JP22K06844 to A.F.; JP22K06470, JP19K06954, JP16K19006 to N.K.; JP21H02592 to H.H.; JP20H03418, JP16H06276 (AdAMS), JP16H05128, JP15H05932, JP26118508, JP26713009, JP22689007 to K.N.); PRESTO (JPMJPR13M9 to K.N.), FOREST (JPMJFR204D to H.H.), Moonshot R&D (JPMJMS2024 to H.H.; JPMJMS2023 to K.N.) of JST; AMED (JP21wm0525002 to N.K.; JP21dm0207112 to H.H.; JP21gm5010002 to K.N.); Hori Sciences and Arts Foundation (to Y.N.); Kato Memorial Bioscience Foundation, Foundation of Kinoshita Memorial Enterprise (to N.K.); Takeda Science Foundation (to T.Y. and K.N.); Nakajima Foundation, Uehara Memorial Foundation, Ono Medical Research Foundation, Brain Science Foundation, Kowa Life Science Foundation (to K.N.).

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.
Metabolic regulation of the two-pore domain potassium channel TASK-1
Authors: Yu Yang , Curry John , Hu Keli
Published: May 2023

TASK-1 is a two-pore potassium ion channel and implicated in the maintenance of cell membrane potential. Little is known about the traffic regulation of TASK-1 under metabolic stress. The present study was designed to explore whether cell surface expression of TASK-1 is modulated by hypoxia. The trafficking of recombinant TASK-1 channels was studied in HEK293T cells under hypoxia induced by sodium cyanide (NaCN). Incubation of cells with NaCN significantly reduced TASK-1 fluorescence intensity at cell periphery, and this effect was inhibited by pretreatment with bisindolylmaleimide (BIM), a specific protein kinase C (PKC) inhibitor. Similar results were confirmed by quantifying extracellular HA-tagged TASK-1 on the cell surface using chemiluminescence assay. NaCN-induced reduction in TASK-1 at cell periphery was also prevented by co-expressing the dominant-negative dynamin 1 (K44E), or cholesterol depletion with MβCD, indicating a potential role of cholesterol and dynamin-dependent endocytosis in PKC-mediated reduction of cell surface TASK-1 by NaCN. Mutagenesis studies further showed that NaCN-induced reduction of TASK-1 at cell periphery was observed in cells expressing wild-type TASK-1 but not mutant TASK-1 with mutation of the dileucine motif (263-264 LL-AA) or the PKC phosphorylation consensus site (T400A). Moreover, Western blot analysis demonstrated that NaCN reduced the amount of wild-type TASK-1 but not the TASK-1 mutants (263-264 LL-AA and T400A) in the membrane fraction. In conclusion, our results demonstrate that cell surface TASK-1 channels are downregulated under metabolic inhibition, possibly through PKC-mediated endocytosis in a cholesterol and dynamin-dependent manner.

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.
What really matters mobility of middle-aged and older adults: low muscle mass or obesity?
Authors: Chen Chiao-Nan , Hsu Kuo-Jen , Chen Shu-Chen , Chien Kuei-Yu
Published: May 2023

Study objective: This study explored which body size–adjusted skeletal muscle indices (SMI) has a better correlation with mobility and cardiometabolic health. Additionally, the roles of low muscle mass and obesity in the mobility and cardiometabolic health of individuals were identified. We hypothesized that body mass index (BMI)-or body weight (Wt)-adjusted SMI had a better correlation with mobility in middle-aged and older adults with obesity than did body height (Ht)-adjusted SMI. Methods: 427 community-dwelling middle-aged and older adults (age: 66.0±9.0 years) underwent assessments of body composition (dual-energy X-ray absorptiometry and waist circumference [WC]), grip strength, and mobility (timed up-and-go test and chair stand test). WC was used as a surrogate for obesity. Chi-square test and one-way analysis of variance were used to compare grip strength, mobility, and cardiometabolic health among individuals with normal body composition (N), only low muscle mass (S), only obesity (O), and low muscle mass plus obesity (SO). Pearson’s correlation coefficient was used to examine the correlation between muscle strength/mobility and different body size–adjusted SMI. Regression models were used to examine the factors independently associated with muscle strength, mobility, and cardiometabolic health. The significance level was set at α<0.05. Results: The O group, but not the S group, had poorer mobility than the N group, regardless of the SMI used to define low muscle mass. The O and SO groups had a higher prevalence of cardiometabolic diseases. The relationship between muscle mass and mobility existed in individuals with obesity, but not in individuals without obesity. After adjusting for age, sex, and WC, ASM/BMI was the only SMI that was correlated with grip strength. Similarly, only ASM/BMI was positively correlated with performance on the timed up-and-go test in the population with obesity (p<0.05). When age and sex were controlled, WC, but not SMI (regardless of adjusting for Ht, Wt, or BMI), was associated with mobility and cardiometabolic health. Conclusion: BMI-adjusted SMI had a better correlation with mobility in middle-aged and older adults with obesity than did Ht- and Wt-adjusted SMI. Obesity plays a more independent role in mobility and cardiometabolic health than low muscle mass in middle-aged and older adults.

This study was supported by the Ministry of Science and Technology, Taiwan

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.
Mediators of steatosis in HepG2 cells
Authors: Rida Reem , Rida Reem , Kreydiyyeh Sawsan
Published: May 2023

Hepatic lipid accumulation or steatosis has become nowadays a major public health problem. The disease can progress into serious liver damage that can lead eventually to hepatic failure and cancer. Fatty liver may result from an increase in lipid acquisition or a decrease in lipid disposal, or both. Although lipid metabolism is known to be under the control of multiple enzymes and signaling molecules, the ones dysregulated in steatosis are still ill defined. The aim of this study was to identify the key molecules altered during hepatic lipid accumulation. Steatosis was induced by incubating HepG2 cells for 24 hrs with 1 mM Oleic and Palmitic acid. Cellular lipid content was quantified using a lipid extraction kit in which lipids were stained with Oil red O, then extracted and quantified colorimetrically. The fatty acids, which are known to activate Gq-linked fatty acid receptors, increased significantly lipid accumulation. This increase was not maintained in presence of specific inhibitors of Gq, PI3K, mTOR, SREBP, and PPARϒ. Gq was found to be upstream of all the other intermediates. It was concluded that saturated fatty acids activate a Gq signaling pathway involving kinases and transcription factors implicated in lipogenesis. The location of the different intermediates with respect to each other needs still to be determined.

University Research Board- American University of Beirut

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.
Effects of home-based high interval inspiratory muscle training on inspiratory muscle strength and exercise capacity in individuals with chronic heart failure
Authors: Wu Yi-Chen , Chen Yung-Hsin , Huang Wei-Ming , Chen Chiao-Nan
Published: May 2023

Study objective: Heart failure (HF) is a global disease that affects people worldwide. Individuals with HF have symptoms such as dyspnea, fatigue, and exercise intolerance. Inspiratory muscle strength plays an important role in individuals with chronic heart failure (CHF). Recent studies have shown that a long duration (15-30 mins/session) of low-intensity inspiratory muscle training (LI-IMT) improves inspiratory muscle function, exercise capacity, and quality of life in individuals with CHF. However, the long duration of training decreases the compliance of patients. Recently, a short duration (5 mins/session) of high-intensity interval IMT(HI-IMT) was found to improve inspiratory strength and exercise endurance in healthy individuals and individuals with obstructive sleep apnea. To date, the effectiveness of HI-IMT in individuals with HF is unknown. The aim of the study was to examine the effects of HI-IMT in individuals with CHF. The hypothesis of the study was HI-IMT improves inspiratory muscle strength, exercise capacity, and quality of life in individuals with CHF. Methods: This is a randomized control trial. Thirty-six individuals with HF (68% male) were randomly allocated to the HI-IMT group and the control group. Both groups of participants received home-based IMT 6 days/week for 12 weeks. Every session consisted of 30 breaths (5 breaths/set, 6 sets). The intensity of HI-IMT and the control group was 60% and 0% of maximal inspiratory pressure (MIPmax), respectively. Exercise capacity (VO2 peak and 6-minute walking distance, 6MWD), MIPmax, and quality of life (Minnesota living with heart failure questionnaire, MLHFQ) were examined before and after training. Analysis of covariance and paired T-tests were used to examine the training effects. P<0.05 was considered different. Results: Thirty-six individuals with CHF were included in the study. The average age and the left ventricular ejection fraction were 64.7±11.9 years old and 44.5±12.3%, respectively. The improvements of MIPmax, 6MWD, VO2 peak, and MLHFQ were greater in the HI-IMT group than in the control group (p<0.05). The HI-IMT group, but not the control group, showed significant improvements in oxygen uptake efficiency slope after the intervention (p<0.05). Conclusion: 12 weeks of HI-IMT was an effective intervention to improve inspiratory muscle strength, exercise capacity, and quality of life in individuals with CHF.

This study was supported by Yen Tjing Ling Medical Foundation, Taiwan.

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.
Setmelanotide, a melanocortin-4-receptor agonist, increases the hypercapnic ventilatory response and relieves hypoventilation in obese mice
Authors: Aung O T , Ramos Amorim Mateus , Anokye-Danso Frederick , Y Polotsky Vsevolod
Published: May 2023

Introduction: Obesity leads to high cardiorespiratory morbidity and mortality. Obese individuals can develop obesity hypoventilation syndrome (OHS) characterized by daytime hypercapnia in the absence of alternative underlying causes. OHS leads to daytime sleepiness, cardiovascular complications, and cyanosis. There is no effective pharmacotherapy for OHS. Leptin, an adipose-produced hormone, improves breathing in OHS, but obese individuals and rodents are resistant to the beneficial effects of leptin. The melanocortin axis is a downstream target of leptin but effects of this axis on control of breathing are poorly studied. Setmelanotide is an FDA-approved drug for weight loss that specifically binds to the melanocortin 4 receptor (MC4R). We have previously demonstrated that C57BL/6J mice with diet induced obesity (DIO) hypoventilate during sleep and exhibit all features of OHS. We hypothesized that the activation of MC4R by setmelanotide augments the control of breathing during sleep in DIO mice. Methods: We performed a proof-of-concept single dose randomized trial of intraperitoneal setmelanotide (1.0 mg/kg) vs vehicle in DIO C57BL/6J mice using a cross-over design. The injections were followed by sleep studies, and measurements of the hypercapnic ventilatory response, oxygen consumption (VO2), and carbon dioxide production (VCO2). Results: Activation of MC4R by setmelanotide increased inspiratory flow, respiratory rate, and minute ventilation during NREM and REM sleep compared to vehicle. Activation of MC4R by setmelanotide increased the hypercapnic ventilatory response. Setmelanotide also increased VCO2, and VO2, but the ventilation was increased out of proportion to the upregulation of metabolism, which was evident from increases in the VE/VO2 and VE/VCO2 ratios. Conclusion: Activation of MC4R by setmelanotide relieved hypoventilation during sleep and increased the hypercapnic ventilatory response in obese mice. Our study identified MC4 as a promising therapeutic target in OHS.

NIH (R01 HL128970, R01 HL133100, and R01 HL138932), AHA (#915167).

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.
Activation of Transient Receptor Potential Vanilloid 4 (TRPV4) channels opens Connexin Hemichannels in Lens Epithelial Cells
Authors: Ek-Vitorin Jose , Shahidullah Mohammad , Lopez Rosales Joaquin , Delamere Nicholas
Published: May 2023

ATP release from the lens via connexin hemichannels has been hypothesized as a response to TRPV4 activation when the lens is subjected to osmotic swelling. To explore the apparent linkage between TRPV4 activation and hemichannel opening we performed patch-clamp recordings on cultured mouse lens epithelial cells exposed to the TRPV4 agonist GSK1016790A (GSK) in the presence or absence of the TRPV4 antagonist HC067047 (HC). GSK was found to cause a fast, variable and generally large non-selective increase of whole cell membrane conductance evident as a larger membrane current (Im) over a wide voltage range. This response was prevented by HC. The GSK-induced Im increase was proportionally larger at negative voltages and coincided with fast depolarization and the simultaneous disappearance of an outward current, likely a K+ current. The initial presence of this outward current in control conditions appeared to be a reliable predictor of a cell’s response to GSK treatment. In some studies, recordings from single cells combining cell-attached and whole-cell patch clamp configurations, were obtained. This novel approach revealed events with a channel conductance 180-270 pS following GSK application through the patch pipette on the cell-attached side. The findings are consistent with TRPV4-dependent opening of connexin hemichannels. Ongoing studies are aimed at confirming the hemichannels identity.

NIH: 5R01EY029171-04 and 2R01EY009532-28A1.

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.
Vitamin D Receptor Attenuates Osteo-sarcopenia by Suppressing the Ferroptosis
Authors: Li Wenxiong , Chen Feifei , Zhang Kuaiqiang , Sun Zhiping , Zhang Yan , Yang Feng
Published: May 2023

Objective: Vitamin D deficiency is associated with a range of skeleton and muscle disorders, including osteoporosis and sarcopenia, but its functional role and the underlying mechanism in skeleton and muscle physiology remain unclear. We investigated the effect of vitamin D receptor (VDR) deletion on skeleton and muscle using a mice model with dexamethasone-induced osteo-sarcopenia (OS). Methods: The wildtype and VDR(-/-) mice were treated with vehicle and dexamethasone (i.p., once daily) for 14 consecutive days. The murine myoblast cell line C2C12 and bone mesenchymal stem cells (BMSCs) were used as an in vitro model in this study. Results: After challenging to dexamethasone, VDR-null mice exhibited more severe amyotrophy and bone rarefaction compared with wildtype counterparts, manifested by marked weight loss and significant decrease in muscle wet weight coefficient of the tibialis anterior muscle and gastrocnemius, accompanying with a decrease in bone mineral density (BMD). In addition, the limb grip and weight-loading swimming time were also fall off. Similarly, dystrophin fluorescence staining and haematoxylin eosin staining showed the decrease in cross-sectional area of skeletal muscle in the VDR-null mice. Alizarin red staining and TRAP staining showed that the differentiation of osteoblasts was significantly decreased, while the activity of osteoclasts was significantly increased in the VDR-null mice. VDR knockout mice significantly accelerated protein degradation and bone destructions, but inhibited protein synthesis and bone construction in skeleton and muscle tissue. In addition, we further assessed whether increased Fe2+ level and decreased GPX4 in VDR knockout muscle accelerated the muscle injury, since the deletion of VDR activated ferroptosis, giving rise to the accumulated of Fe2+ in mice and Depleting GSH and inhibiting GPX4 expression. Treatment with the inhibitor of ferroptosis Ferrostatin-1 eliminated dexamethasone-induced atrophy and osteoporosis of myotube cells and BMSCs, reflected by alleviating myotube cell atrophy and BMSCs bone rarefaction via MHC fluorescent staining, Alizarin red staining and TRAP staining. It is suggested that ferroptosis may be one of the pathogenesis of OS. Conclusion: These results provided evidence that VDR attenuated dexamethasone-induced OS at least in part by suppressing the ferroptosis.

This work was sponsored by the Program of Natural Science Foundation of China (81973889), the Program of Shaanxi Education Department (19JC013 to FY & 22JK0346 to WL), Youth Innovation Team in Shaanxi universities, and the Program of Health Commission of Shaanxi Province (2022B002).

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.
HDAC and p300/CBP activities regulate Reactive Oxygen species dependent increase in Histone 3 acetylation in response to intermittent hypoxia
Authors: Nanduri Jayasri , Wang Ning , Prabhakar Nanduri
Published: May 2023

Histone3 (H3) lysine acetylation levels regulate transcriptional activation through a dynamic process involving equilibrium between histone acetylases (KATs) and histone deacetylases (KDACs). We have previously shown that lysine acetylation of H3 is an early epigenetic mechanism associated with Hypoxia inducible transcription factor (HIF-1) activation and sympathetic excitation in rodent models of intermittent hypoxia (IH), a hallmark feature of sleep apnea (Wang et al, Front Physiol 2021). The goal of the present study was to determine the molecular mechanisms underlying IH increased H3 acetylation. We tested the hypothesis that IH decreased KDAC activity leads to increased KAT activity, thereby increasing H3 acetylation. Pheochromocytoma (PC)-12 cells exposed to IH (1.5% O2 for 30 sec followed by 20% O2 for 5 min) exhibited reduced KDAC and increased KAT enzyme activity along with increased global acetylation of H3 in a stimulus-dependent and reversible manner. Increased H3 acetylation by IH was blocked by an antioxidant. Conversely, reactive oxygen species (ROS) mimetics, like IH, increased H3 acetylated protein expression. Treating cells with Trichostatin A (TSA), an inhibitor of KDACs mimicked IH-increased H3 acetylation under normoxic conditions. Pharmacological blockade p300/CBP (KAT) with CTK7A abolished IH-induced increased H3 acetylation. Similar to PC12 cells, decreased KDAC activity lead to augmented p300/CBP mediated H3 acetylation in both rats exposed to IH and heme-oxygnease null mice (HO-2-/-) which exhibit high incidence of apneas. In summary, interplay between KATs and KDACs regulate ROS-dependent H3 acetylation and the ensuing IH induced transcriptional activity leading to sympathetic activation.

NIH-PO1-HL90554

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 reduction in Homeodomain Interacting Protein Kinase-2 in lung fibroblasts ameliorates lung fibrosis
Authors: Hewawasam Sankha , Liang Yurong , Hang Chaoqun , Dang Quanjin , Liu Lin
Published: May 2023

Idiopathic pulmonary fibrosis is a chronic, progressive, and scarring lung disease with an unknown etiology. Homeodomain Interacting Protein Kinase 2 ( HIPK2) is a co-transcriptional regulator involved in kidney fibrosis. We hypothesize that a decrease in HIPK2 expression attenuates the activation and proliferation of lung fibroblasts through inhibition of TGF-β1 and Wnt 3a pathways. HIPK2 knockdown using a lentiviral shRNA in human pulmonary fibroblasts (HPFs) reduced TGF-β1- and WNT3a-induced expression of fibrotic markers including α-smooth actin, fibronectin, and collagen 1A1 and 3A1 as determined by Western blots and real-time PCR. Immunostaining of α-smooth actin showed that HIPK2 knockdown inhibited fiber formation in HPFs. Furthermore, HIPK2 knockdown suppressed fibroblast proliferation. HIPK2 knockdown in HPFs also resulted in a reduction in TGF-β1-induced SMAD 2/3 phosphorylation and WNT3a-induced activated β-Catenin expression, suggesting the involvement of HIPK2 regulation on above pathways. To investigate the effects of HIPK2 knockdown on lung fibrosis in mice, HIPK2 was knocked down (KD) in the lungs of mice using an adenovirus expressing 4 shRNAs targeting HIPK2 and lung fibrosis was induced by an intratracheal instillation of bleomycin (1U/kg body weight) for 14 days. HIPK2 KD mice lost less body weight compared to the control mice. Analysis of bronchoalveolar lavage (BAL) indicated a significant reduction in total infiltrated immune cells in Hipk2 KD mice. Lung fibrosis was reduced in Hipk2 KD mice as demonstrated by fibrotic score, Sirius red staining, fibronectin mRNA level, and collagen content as measured by hydroxyproline assay. Lung functions including elastance, compliance, resistance, and inspiratory capacity, as determined by Flexivent, were improved in Hipk2 KD mice. Real-time PCR and ELISA revealed a significantly reduced mRNA expression of Tgf-β1 in the lung tissues and a decreased TGF-β1 level in BAL fluids of Hipk2 KD mice. Immunofluorescence indicated reduced SMAD-3 and activated β-Catenin levels in the fibrotic regions of Hipk2 KD mice. These findings clearly suggest that reduced HIPK2 expression in the lung attenuates lung fibrosis through the suppression of TGF-β1 and WNT3a signaling pathways.

R01 HL157450, R01HL135152 and P20GM103648

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.
Effect of aging on aortic arterial stiffness and wave reflection in healthy mid-life adults
Authors: Chang Keng-Yu , Hibner Brooks , Phillips Shane , Piano Mariann , Liu Zhaoli , Hwang Chueh-Lung
Published: May 2023

Introduction: Aging is associated with arterial stiffening which increases pressure reflection to the heart and contributes to the development of cardiovascular disease. However, most vascular aging studies have focused on comparing young adults versus adults 65 or older, and comorbidities, such as hypertension and obesity, confounded the findings. The extent of vascular aging in healthy mid-life adults (age 50-64) is unclear. Therefore, the purpose of this study was to compare arterial stiffness and wave reflection in healthy young adults (18-35 years) and healthy mid-life adults. Methods: Twenty-six apparently healthy, nonsmoking, adults underwent assessments of arterial stiffness and wave reflection. Young adults (YA; n=15, 6 eumenorrheic women; age: 28±4 yr; systolic blood pressure [SBP]: 109±8 mmHg; body mass index [BMI]: 24.1±2.5 kg/m2; mean±SD) were compared to mid-life adults (MA; n=11, 8 postmenopausal women; age: 59±4 yr; SBP: 109±10 mmHg; BMI: 25.6±4.1 kg/m2). Using SphygmoCor XCEL (AtCor Medical), aortic arterial stiffness was measured as carotid to femoral pulse wave velocity (cfPWV) and wave reflection was measured as augmentation index normalized at heart rate of 75 (Alx75), augmented pressure (AP), backward wave pressure (Pb), and reflection magnitude (RM). Results: No significant differences between YA and MA were found in brachial and aortic SBP, diastolic blood pressure, and pulse pressure (P≥0.07). Compared to YA, MA had higher cfPWV (MA vs. YA: 6.8±0.9 m/s vs. 5.8±0.6 m/s, p=0.001), Alx75 (30±10% vs. 4±10%, p<0.001), AP (13±4 mmHg vs. 4±3 mmHg, p<0.001), Pb (16±2 mmHg vs. 13±2 mmHg, p=0.001), and RM (67.7±6.8% vs. 48.0±5.6%, p<0.001). Conclusions: Our findings suggest that mid-life adults exhibit increased arterial stiffness and wave reflection compared to young adults. These data suggest that vascular aging may already be present in mid-life adults, despite normal peripheral blood pressure. These findings help extend our understanding of the sequence of age-related changes in vascular function throughout the lifespan.

This work was supported by NIAAA K99/R00 AA028537 to CLH.

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.
Gender affirming hormone therapy in the male rat is associated with differential effects on cardiovascular risk factors
Authors: Crudup Breland , Mallette Jordan , Oudomrath Speyrer Adrian , Rawls Adam , Cockrell Kathy , Davenport Kacey , Willis Alex , Yanes Cardozo Licy , Shawky Noha , Alexander Barbara
Published: May 2023

Gender-Affirming Hormone Therapy (GAHT) in transgender females (male to female) involves administration of 17ß-estradiol (E2) with an anti-androgen (AA) or castration (CTX). GAHT is used to treat individuals that are born one sex but identify as the other gender. Numerous clinical studies indicate that cardiovascular (CV) risk is elevated in GAHT transgender females. Yet, the mechanisms involved remain unknown. Thus, the purpose of this study was to develop an experimental model of the transgender female and to test the hypothesis GAHT is associated with increased CV risk. Methods: Male Sprague Dawley rats were randomly assigned to the following groups: Control (n=8), E2 [5 mg/day, pellet sc, replaced every 3 weeks, N=6], E2+AA [E2 + Spironolactone, 10 ng/kg/day, diet, N=8], and E2+CTX (N=8). AA and E2 were initiated at 14 weeks of age; CTX rats were exposed to AA for 2 weeks prior to CTX at 16 weeks of age. At 28 weeks of age, EchoMRI for measurement of body weight, fat and lean mass was followed by 24-hour metabolic studies. E2 (ELISA), testosterone (RIA) and analytes (Chemistry Analyzer) were analyzed end of study at 28 weeks of age. Data were analyzed using GraphPad Prism 9 with one-way ANOVA with multi-group comparisons. Statistical significance was set at a P value of <0.05. Results: ( Table 1 ) E2 alone was sufficient to decrease testosterone in male rats. Administration of E2 in the male rat was associated with a significant decrease in body weight, lean mass and average kidney and testes weight. Creatinine was decreased in E2 alone. Yet, BUN was decreased in E2+CTX. E2 + AA was associated with a significant increase in total cholesterol compared to Control; an increase not observed in E2 or E2+CTX. HDL was elevated in E2+AA, whereas LDL-C was decreased in E2, E2+AA and E2+CTX mimicking what is observed in the transfemale population. Collectively, these results demonstrate differential effects of E2+AA versus E2+CTX on CV risk factors in male rats indicating that long-term studies are needed to determine the effect of GAHT on CV risk across the lifespan.

[Table: see text]

Funding: HL143459, P20-GM-104357, P20-GM121334, T32-HL105324, and LGBTQ Fund of MS

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.
Airway luminal area in long COVID patients compared to acute COVID-19 patients
Authors: Zaremba Solomiia , Miller Alex , Ovrom Erik , Senefeld Jonathon , Wiggins Chad , Dominelli Paolo , Joyner Michael , Welch Brian , Ramsook Andrew
Published: May 2023

Background: Patients with respiratory manifestations of acute COVID-19 infection have enlarged luminal areas of central conducting airways demonstrated on computed tomography imaging and abnormal pulmonary function. In some cases, COVID-19 symptoms may persist for more than 4 weeks, referred to as long COVID, and the effect on airway luminal area remains unknown. Herein, we report airway luminal cross-sections areas among a long COVID group, compared to individuals with an acute COVID-19. We aim to investigate if large conducting airways sizes are different between acute COVID-19 and long COVID patients. Methods: A cohort of 181 people with long COVID were screened, of which 20 people (9 men, 11 women; 55.9 ± 14.1 years) were included in analyses. Twenty age, height, and sex matched patients with an acute (< 4 weeks of persistent symptoms from diagnosis) COVID-19 infection (55.7 ± 15.2 years) served as a comparison group. We used a three-dimensional reconstruction of computed tomography imaging to measure luminal areas of the trachea, right and left main bronchi, bronchus intermediate, right and left upper lobe, and left lower lobe bronchi at proximal, middle, and distal cross-sectional points in both groups. Airway luminal area was taken as the average of the three points. We compared average lumen area of long COVID patients with acute COVID-19 individuals, using an independent samples t-test. Results: We found no significant difference comparing measurements of the following airway luminal cross-sectional areas: trachea (277.7 ± 100.4 vs. 268.0 ± 70.5 mm2 p=0.73), right main bronchus (191.0 ± 66.1 vs. 190.1 ± 49.0 mm2, p=0.96), bronchus intermediate (112.3 ± 33.4 vs. 109.3 ± 27.9 mm2, p=0.76), right upper lobe bronchus (74.4 ± 21.0 vs. 73.7 ± 25.5 mm2, p=0.92), left main bronchus (129.1 ± 50.1 vs. 136.9 ± 38.0 mm2, p=0.58), left lower lobe bronchus (57.8 ± 18.2 vs. 59.0 ± 13.4 mm2, p=0.82), and left upper lobe bronchus (80.1 ± 19.5 vs. 77.9 ± 22.9 mm2, p=0.75) in the long COVID and acute COVID-19 groups respectively. Conclusion: In patients with long COVID, compared to those with an acute COVID-19 infection, we did not see differences in central airway luminal areas. Further analysis of a larger cohort is needed to determine whether enlarged airway luminal areas, associated with acute COVID-19, persist in long COVID. This research will expand knowledge on functional consequences for patients with long COVID.

Funding: F32HL154320 to JWS; 5R35HL139854 to MJJ.

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.
Direct activation of Airway Sensory C-fibers by SARS-CoV-2 Spike protein
Authors: Kim Joyce , Ru Fei , Meeker Sonya , Undem Bradley
Published: May 2023

Activation of sensory nerves, in particular nociceptive C-fibers, is a feature of most respiratory viruses. Evidence of such activation is found in the classical consequences of C-fiber activation including sneezing, sore throat, coughing, and reflex secretions. As well as causing the troubling symptoms of viral infection, the activation of these nerves allows viruses to escape the body and be transmitted to other hosts, i.e. nociceptor activation amplifies spread in a community. In addition, activation of airway vagal C-fibers can lead to strong reflex bronchoconstriction and excessive secretions that likely contribute to the exacerbation of asthma particularly in children. In theory, viral infection leads to C-fiber activation by two general mechanisms. The first is that viral infection of epithelial cells leads to the production of a mediator(s) that stimulates the C-fiber terminals. The second is that the virus itself directly activates the nerves. We used three orthogonal approaches to address our hypothesis that the coronavirus spike protein directly activates (evokes action potential discharge) vagal C-fibers in mouse airways. We used male C57/BL6 mice to evaluate whether coronavirus spike protein interacts; directly with C-fiber terminals in an isolated vagus-trachea/lung preparation. Action potentials were recorded using extracellular electrodes positioned near the relevant cell bodies in the vagal sensory nodose/jugular ganglion complex. Spike protein S1 (4ug/ml) was perfused through the mouse trachea into the receptive field of an identified C-fiber (c.v. <1 m/s). 40% of C-fibers (12 out of 30) responded acutely to the S1 protein with action potential discharge. In the same type of ex-vivo experimental set-up, two-photon imaging technique was also used to evaluate the % of fibers activated by spike protein. Consistent with our electrophysiological studies we found that 45% of the C-fibers (241 fibers out of 538 fibers in 4 experiments) were stimulated by the S1protein. To determine whether spike protein was indirectly activating C-fibers in the lung, i.e. requiring other cells within the airway tissue, we evaluated its effect on neurons dissociated from the vagal ganglia with Fura-2 calcium imaging. Again, ~ 40% of the isolated neurons (65 out of 163 neurons) responded to spike protein. These data support our hypothesis that the spike protein can directly activate neurons independently of airway tissue. This supports the hypothesis that some of the nerve-related effects of SARS CoV-2 infection may involve the direct activation of sensory C-fibers.

R35 HL144887

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.
Exploring sex differences in inhibitory synaptic transmission in the kolliker-fuse of rett syndrome mice
Authors: Whitaker-Fornek Jessica , Levitt Erica
Published: May 2023

Rett Syndrome (RTT) is a rare neurodevelopmental disorder characterized by severe motor impairments including breathing problems. RTT is caused by mutations in the methyl CpG binding protein 2 (MeCP2) gene located on the X chromosome. Accordingly, girls and women account for the majority of RTT patients. The genetic nature of RTT lends itself to study in mice with loss of function mutations in the MeCP2 gene. Importantly, RTT mice display breathing irregularities similar to human patients. Insufficient inhibitory neurotransmission in the brainstem respiratory network is thought to contribute to the breathing problems in RTT. In the pontine Kölliker-Fuse (KF) area of RTT mice, there are fewer GABAergic neurites and injection of a GABA reuptake antagonist alleviates breathing irregularity in vivo. However, no one has examined the physiology of inhibitory synaptic transmission in the KF of RTT mice. In the present work, we tested the hypothesis that inhibitory synaptic transmission was deficient in the respiratory-controlling KF of MeCP2Bird mice. Although girls and women make up the majority of RTT patients, most studies examining synaptic physiology in the brainstem have been performed using male MeCP2-deficient mice. Hemizygous male MeCP2Bird mice lack MeCP2 protein and exhibit rapid onset of severe RTT symptoms which leads to short life spans. Due to partial X-inactivation, MeCP2 expression in heterozygous female MeCP2-deficient mice is mosaic, leading to variable symptom severity and delayed onset, which more closely resembles the human condition. In the present study, we examined inhibitory synaptic transmission in the KF of female heterozygous MeCP2Bird mice compared to male hemizygous MeCP2Bird mice and wild type littermates. Whole-cell voltage-clamp recordings from KF neurons contained in acute brain slices were used to measure inhibitory currents. Most notably, the frequency of spontaneous inhibitory postsynaptic currents is reduced in the KF of male, but not female, MeCP2Bird mice compared to wild type littermates. To better understand this sex difference in inhibitory neurotransmission, we assessed the influence of estrous cycle stage and MeCP2 expression status in the recorded neuron. The outcomes of this research will increase our understanding of synaptic function in respiratory control neurons in female Rett Syndrome mice.

This research was supported by the International Rett Syndrome Foundation Basic Research Grant 3608.

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.
Association Between Pain Perception and Blood Pressure Responsiveness During a Cold Pressor Test in Young Adults
Authors: Miller Kayla , Hanson Brady , Casey Darren
Published: May 2023

The cold pressor test (CPT) has been widely used as a stressor in laboratory settings to examine blood pressure (BP) responsiveness in both healthy and patient populations. Current literature indicates considerable inter-individual variability in BP responsiveness to a CPT. This variability may be due to the level of perceived pain associated with the cold stress. Therefore, in the present study we tested the hypothesis that BP responsiveness would demonstrate a positive association with ratings of perceived pain during the CPT in young adults. Since sex-related differences in BP responsiveness to a CPT have been reported, we also examined if the hypothesized association between BP responsiveness and pain perception during a CPT differs between young males and females. 26 young adults (13M/13F; 21±4 yr) underwent a CPT consisting of a 2-min baseline followed by immersion of the left hand in ice water (~4°C) for 2-min. Beat-by-beat BP was recorded continuously and averaged every 30s of the CPT. Subjects provided verbal pain scores (0-10 scale) every 30s. Two-way repeated measures ANOVAs were used to assess BP and pain over time and to determine if responses differed between sex. Additionally, Pearson correlations were used to examine the association between BP responsiveness (change (Δ) in diastolic (DBP), systolic (SBP) and mean arterial pressure (MAP)), and rating of perceived pain for the entire cohort as well as for each sex. All indices of BP and perceived pain increased in response to the CPT in young adults (P<0.05 for all), with no sex-related differences noted. ΔDBP, ΔSBP, ΔMAP were not associated with pain perception for the group as a whole. When examined within sex, no association was observed between ΔDBP, ΔSBP, ΔMAP and pain perception for males at any point during the CPT. However, within females, ΔDBP, ΔSBP, and ΔMAP were positively associated with pain perception at 1-min (r=0.77, 0.57, 0.66, respectively; P<0.05 for all) but not at 2-min (r=0.36, 0.24, 0.33, respectively; P>0.05 for all) of the CPT. Contrary to previous findings, we did not observe sex-related differences in BP responsiveness to a CPT in young adults. Additionally, BP responsiveness is not associated with pain perception during the CPT in our group as a whole. However, when analyzed by sex, an association between BP responsiveness and pain perception was found in females at the early stages of the CPT (i.e. 1-min).

Funding: American Physiological Society SURF Program (K.A.M.)

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.
Orphan GPRC5B activation modulates airway contractility
Authors: Varadharajan Subashini , Balraj Premanand , Kale Narendra , Campos Luana Janaina de , Conda Sheridan Martin , Mallik Sanku , Pabelick Christina M , YS Prakash , Sathish Venkatachalem
Published: May 2023

Background: Multiple asthma phenotypes are driven by pathobiological heterogeneity. Almost 25% of asthma patients do not respond well to current treatments like inhaled corticosteroids, urging researchers and clinicians to propose novel treatment targets and strategies. Orphan type C G-protein coupled receptor GPRC5B is one such novel target abundantly expressed in various tissues, but there are no known ligands for GPRC5B. Previously, we demonstrated that orphan type C G-protein coupled receptor GPRC5B is increasingly expressed in human asthmatic airways and plays a crucial role in regulating intracellular calcium ([Ca2+]i) in human airway smooth muscle (ASM) cells. However, the role of GPRC5B activation on airway contraction is not identified yet. Our objective for this study is to design and synthesize a novel ligand for GPRC5B and determine the effect of GPRC5B activation on [Ca2+]i responses in ASM cells and airway contractility in ex vivo murine airways. Methods: ZincPharmer was used for pharmacophore search. After energy minimization, the top hit compound was synthesized using a standard amide coupling reaction, purified by flash chromatography, and characterized by NMR spectroscopy. Human ASM cells loaded with Fura-2 AM were imaged for [Ca2+]i responses. Silencer select siRNA was used for transient silencing of GPRC5B in ASM cells. Myosin light chain (MLC) phosphorylation was measured using immunoblot analysis. Precision-cut lung slices (PCLS) (~200 μm thickness) of wild-type C57BL6 mice were used for contractile studies. Results: Based on molecular docking data, the top compound N-(5-chloro-2-hydroxybenzyl)-2-((tetrahydro-2H-pyran-4-yl)oxy) acetamide (L1) was synthesized. Concentration-dependent [Ca2+]i responses were observed on human ASM cells stimulated with L1 (1 nM - 1 μM) that were comparable to the known contractile agonist histamine (10 μM). L1-induced [Ca2+]i response was significantly reduced in GPRC5B knockdown ASM cells. GPRC5B knockdown in ASM cells was verified by Western analysis. Further, L1 exposure increased the MLC phosphorylation (Thr18/Ser19) in human ASM cells compared to vehicle. Acute exposure of L1 (1 μM) increased airway contractility of PCLS (~17%-25%) compared to vehicle exposure. Conclusion: These findings suggest that GPRC5B activation modulates intracellular calcium and airway contractility in human ASM cells and murine PCLS. Thus, GPRC5B could potentially be considered for a novel therapeutic regimen to tackle asthma.

This work was partly supported by NIH grants R01-HL146705 (Venkatachalem), R01-HL088029 (Prakash), and R01-HL142061 (Pabelick, Prakash).

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.