bluuebunny/crossref_metadata_2025 huggingface dataset:
GPER activation ameliorates vascular remodeling in salt‐sensitive mRen2.Lewis rats (867.7)
Authors: Lindsey Sarah , Liu Liu , Chappell Mark
Published: June 2021

The mRen2.Lewis female rat is an estrogen‐ and salt‐sensitive model of hypertension which may reflect the higher pressure and salt sensitivity associated with menopause. In salt‐loaded mRen2.Lewis, activation of the novel estrogen receptor GPER reduces renal hypertrophy, proteinuria, and oxidative stress but not blood pressure (Lindsey et al. 2011). The current study hypothesized that vascular remodeling is also improved by GPER activation in the absence of changes in blood pressure. Intact mRen2.Lewis females were fed a normal (0.5% Na+) or high salt diet (4% Na+) for 10 weeks and treated with vehicle or the selective GPER agonist G‐1 for the last two weeks of the study. Systolic blood pressure was measured by tail cuff plethysmography, and aortic sections were mounted on a wire myograph or formalin‐fixed for histological analysis. Systolic blood pressure increased with the high salt diet (137 ± 2 mmHg, n=7 to 224 ± 8, n=9; P<0.001) and remained unchanged with G‐1 (216 ± 4, n=9; P>0.05). While aortic reactivity to phenylephrine and acetylcholine were not different between groups (P>0.05), chronic G‐1 treatment reduced vasoconstriction to angiotensin II (P<0.05) and enhanced G‐1 vasorelaxation ex vivo (P<0.01) in both endothelium‐intact and denuded aortic rings. Analysis of aortic sections revealed that while the high salt diet significantly increased the media to lumen ratio (0.43 ± 0.02, n=6 vs. 0.83 ± 0.05, n=7; P<0.001), two‐week G‐1 treatment partially reversed the salt effect (0.62 ± 0.02, n=6, P<0.005). We conclude that GPER exerts vascular protective actions in the aorta of salt‐loaded mRen2.Lewis females independently of blood pressure. The anti‐remodeling actions of this novel estrogen receptor may be effective in postmenopausal cardiovascular disease by targeting the vascular renin‐angiotensin system.

Grant Funding Source: Supported by NIH (HL56973, HL51952, HL103974) and the American Heart Association (25515E, 3080005)
Overexpression of miRNA‐195 induced by hyperthyroidism is attenuated by the AT1 receptor blockage (868.6)
Authors: Diniz Gabriela , Barreto‐Chaves Maria Luiza
Published: June 2021

Introduction: miRNA‐195 is increased during cardiac hypertrophy, and it´s overexpression is sufficient to drive cardiac growth. We demonstrated that the AT1 receptor (AT1R) participates of the thyroid hormone (TH)‐induced cardiac hypertrophy. However, the potential modulation of the miRNA‐195 in TH‐mediated cardiac growth, as well as the possible contribution of the AT1R in this event is unknown.

Objective: The aim of this study was to investigate the cardiac miRNA‐195 levels in TH‐induced cardiac hypertrophy, as well as the potential role of the AT1R in this response.

Methods: The hyperthyroidism was induced by injections of T4 for 14 days. The T4 group was subdivided into two groups: T4 and T4 plus Losartan (an AT1R blocker). Cardiac hypertrophy was assessed by heart weight (HW) to tibia length (TL) ratio and by the ANF and skeletal‐alpha actin gene expression. The analysis of the miRNA levels was assessed by miRNA Stem Loop RT‐PCR. Data are presented as mean±SD (n=5).

Results: The hyperthyroid group presented cardiac hypertrophy, as evaluated by the higher HW to TL length ratio, as well as by the increased ANF and skeletal‐alpha actin mRNA levels. However, the AT1R blocker prevented the cardiac hypertrophy induced by T4. In addition, the cardiac growth in response to TH was accompanied by higher cardiac levels of miRNA‐195 (4.9±0.7, P<0.01 vs control), which were attenuated by the use of the AT1R blocker (2.1±0.5, P<0.01 vs T4).

Conclusions: These results evidence for the first time that T4‐induced cardiac hypertrophy is accompanied by a cardiac overexpression of miRNA‐195, and that the AT1R participates at least in part of this response.

Grant Funding Source: Supported by FAPESP (Foundation for the Support of Research in the State of Sao Paulo) and by CNPq.
Cardiac specific TIMP‐4 dictates cardiomyocyte contractility through Serca‐2a in cardiac dysfunction: a novel mechanism (868.7)
Authors: Chaturvedi Pankaj , Metreveli Naira , Tyagi Suresh
Published: June 2021

Although previous studies show MMP9 (Matrix Metalloprotease 9) dysregulation in cardiac disorders, the mechanisms how an MMP9 modulator (TIMP4‐Tissue Inhibitor of Matrix Metalloprotease 4) regulates contractility and regeneration remain elusive. Providing an insight into this mechanism, we hypothesize that TIMP4 regulates cardiomyocyte contractility by regulating the expression of Serca‐2a (Sarcoplasmic Reticulum Calcium ATPase) through microRNAs. To check this hypothesis, we overexpressed TIMP4 in cardiomyocytes along with its inhibition through siRNA, to elucidate the contractility profiles and establish regulatory effects of TIMP4. We checked the expression of serca‐2a and microRNA 122 with Western, RT‐PCR, qRT‐PCR. DesRed2 plasmid carrying the TIMP4 gene was used to overexpress TIMP4 in mouse cardiomyocytes while GFP plasmid was used as control to standardize transfections. Contractility measurements were executed by ION OPTIX instrument and transfection studies were done by confocal microscopy. The expression studies of TIMP4 were performed by Western, RT‐PCR, qRT‐PCR and additionally confirmed in HL‐1 cardiomyocyte cell line. We found increased contractility with increase in the amplitude and peak height with TIMP4 overexpression while with siRNA, the results were reversed. Intrestingly, we found increased expression of serca‐2a along with downregulation of microRNA 122. The data suggested a novel regulatory effect of TIMP4 on contractile function by regulating serca‐2a through microRNA 122.

Grant Funding Source: NIH grant HL‐74185 and HL‐108621
Stimulation of mitochondrial superoxide in lung epithelial cells using a novel stretching device (869.5)
Authors: Rapalo Gabriel , Roan Esra , Schwingshakl Andreas , Eckstein Eugene , Waters Christopher
Published: June 2021

Mechanical ventilation of patients with acute lung injury can cause further damage to the lung epithelium, leading to ventilator‐associated lung injury. We and others have previously demonstrated that cyclic stretch of bronchial and alveolar epithelial cells stimulates increased production of reactive oxygen species (ROS) and other proinflammatory cytokines, but the mechanism is not completely understood. To investigate, we designed a motor‐driven device to apply linear strains up to 20% on cells grown on an elastic substrate while imaging under a confocal microscope. We hypothesized that distention of the lung epithelium triggers an oxidative stress response of mitochondrial superoxide through deformation of the mitochondria and actin filaments. Cultures of an immortalized human airway epithelial cell line (16HBE) and human alveolar type II cell line (A549) were tonically stretched while the production of mitochondrial superoxide was measured using MitoSox. Stretch in 16HBE cells caused a significant increase in production of superoxide, while treatment with Tiron or Cytochalasin D both prevented the stretch‐induced increase in superoxide. A549 cells did not show a significant increase in production of superoxide compared to treatment with the inhibitors. A single tonic deformation of the mitochondria via actin filaments directly stimulated the release of superoxide.

Grant Funding Source: Supported by NIH grant HL094366 (CMW)
Expression of BMP‐2 in the pulmonary neuroepithelial body microenvironment (869.7)
Authors: Verckist Line , Lembrechts Robrecht , Schnorbusch Kathy , Pintelon Isabel , Timmermans Jean‐Pierre , Brouns Inge , Adriaensen Dirk
Published: June 2021

Recently, members of the bone morphogenetic protein (BMP) family, in particular BMP‐2, have been reported to promote adult lung tissue repair after injury. Following severe experimental intrapulmonary airway epithelial damage, the neuroepithelial body (NEB) microenvironment (ME) has been shown to be involved in epithelial regeneration, and today may be regarded as an airway stem cell niche. The NEB ME consists of highly innervated groups of pulmonary neuroepithelial endocrine cells, which are specifically covered by Clara‐like cells with supposed stem cell characteristics.

Here we studied the expression of BMP‐2 in the NEB ME of healthy postnatal mouse lungs. After collection of ‘NEB sample’ by selective isolation of GFP‐fluorescent NEBs via laser microdissection from frozen sections of GAD67‐GFP mouse lungs, (q)RT‐PCR revealed a significantly higher expression of BMP‐2 in the NEB ME, compared to control airway epithelium. Localization of the BMP‐2 protein in postnatal mouse lungs by multiple immunofluorescence, revealed that BMP‐2 was expressed by vagal sensory nerve fiber endings that selectively contact NEB cells.

The presented data reveal a strong expression of BMP‐2 in the NEB ME and strengthen the suggestion of an important role in postnatal airway epithelial repair after injury.

Grants: FWO (G.0589.11), Hercules Foundation (AUAH‐09‐001); UA (GOA BOF 2007)
IgE and H. pylori titers in severe asthmatic subjects versus controls: antigen hyperreactivity or etiology? (870.1)
Authors: Varanasi Laalasa , Lankala Shreya , Quinn Tim , Herndon Betty
Published: June 2021

Nearly 25 million Americans are asthmatic, and several triggers have been implicated for producing symptoms. Gastroesophageal reflux (GER), often caused by H. pylori, is common and may trigger asthma exacerbations. The hyperreactivity to antigens in atopic asthma may have effects on H. pylori‐associated GER. The rationale for this study was to determine, in a population of asthmatics and controls, if the H. pylori serum titer is related to elevated IgE (atopy). Sera from 58 severe asthma subjects and 24 control sera were used for IgE and H. pylori specific ELISAs to test for titer based on a 450 nm read of 2.000 or greater (high) or a read below 0.25 (low). Asthmatic IgE samples were 44.83% high, 12.06% low while controls were 8.3% high and 16.7% low. Asthmatic sera H. pylori titers, read on the same scoring system, showed 16.07% high and 26.7% low. Control sera had no H. pylori high values and 61.9% low. 77.8% of the asthmatic subjects with high serum H. pylori titers had high serum IgE titers also. Only 11.1% of the asthmatic subjects with low H. pylori titers also had low IgE. No steroid use data were available for either group. We conclude that, in this patient population diagnosed as severe asthmatics by our hospital pulmonology dept, H. pylori is significantly more prevalent than in controls, although total IgE in asthmatics appears higher than H. pylori‐specific IgE.

Grant Funding Source: Supported by Student Sarah Morrison grant to LV
Downregulation of NGF attenuates AHR persistence induced by respiratory syncytial virus infection in BALB/c mice (870.3)
Authors: Zang Na , Zhou Na , Deng Yu , Xie Xiaohong , Xu Fadi , Liu Enmei
Published: June 2021

RSV infection causes acute morbidity and recurrent wheezing that could persist up to several years, but the relevant mechanisms remain unclear. Because NGF is involved in developing AHR during acute phase of RSV infection, we asked whether RSV infection also induced a chronic AHR in mice, and if so, what role pulmonary NGF played. Mice at day 7, 14, 21, 30 and 60 after intranasal inoculation of RSV were either placed in a whole body plethysmograph to measure their Penh responses to increasing doses of aerosol methacholine or euthanized for collecting BALF to detect pulmonary NGF. We found that AHR and NGF/BDNF were gradually increased and persisted 60 days after RSV infection. To define whether NGF was responsible for the AHR persistence via TLRs‐TRIF signaling pathway, the AHR was tested in other mice after administration of anti‐NGF neutralized antibody (ip) once on day 14 postinfection and resveratrol (inhibitor of TRIF, ip) daily from day 0 to day 21 postinfection, respectively. The RSV‐induced AHR and NGF were markedly attenuated by both treatments. Additionally, RSV infection also produced chronic airway inflammation 60 days postinfection that was diminished by resveratrol but not by anti‐NGF. We conclude that NGF is involved in generating the RSV‐induced AHR probably via activation of TLRs‐TRIF signaling pathway, while the latter but not NGF participates the RSV‐induced chronic airway inflammation.
Visualization of dynamic airway bronchoconstriction in live rats (870.6)
Authors: Phung Thien‐Khoi , Sinclair Scott , Makena Patrudu , Molthen Robert , Waters Christopher
Published: June 2021

Airway hyperresponsiveness severely limits gas exchange in patients with asthma. Imaging studies have shown that bronchoconstriction causes patchy patterns of ventilation defects throughout the lungs, and computational models have predicted that these regions are due to constriction of small airways. Since these imaging approaches are limited in their ability to capture dynamic changes in small airways, we utilized tantalum dust and microfocal X‐ray imaging of rat lungs to obtain dynamic images of airways in an intact animal model. The flexiVent system was used to simultaneously measure airway resistance. Custom software was used to track changes in airway diameters up to 13 generations (~0.3 mm to 3 mm in diameter). The objective of the study was to identify the size and location of airways with changes in diameter during bronchoconstriction in response to methacholine. In contrast with the model predictions, we observed significantly greater percent constriction in larger airways in response to methacholine challenge. Although there was a dose‐dependent increase in airway resistance, the percent change in airway diameters was similar for increasing doses. Paradoxically, a deep inspiration following methacholine caused greater percent constriction in smaller airways. These measurements allow us to directly quantify dynamic changes in airways during bronchoconstriction.

Grant Funding Source: Supported by NIH HL094366 (CMW) and APS UGSRF (TNP)
Loss of M2 and M3 muscarinic receptor function in double mutant mice eliminates ovalbumin‐induced airway hyperresponsiveness (870.7)
Authors: Fisher John , Vincent Sandra , Wannas Paul , Wess Jurgen
Published: June 2021

Mutant murine models lacking specific muscarinic receptor (MR) subtypes are a primary tool to explore MR biology in health and disease, due to the lack of selective antagonists. Since both M2R and M3R subtypes are involved in the vagal control of airway responsiveness (AR; FASEB J 18:711,2004), we examined the impact of combined loss of M2R and M3R function on the development of AHR in M2R‐/‐/M3R‐/‐ double mutant mice. AR was assessed in 12 mutant mice divided into an ovalbumin (OVA) sensitization group (n=6; OVA‐Alum adjuvant i.p. day 0 and 7 plus aerosol exposure days14‐16) and an Alum adjuvant control group (n=6; Alum). Since M2R‐/‐/M3R‐/‐ double mutants show no cholinergic contractile responses, we performed a 5‐hydroxytryptamine (5‐HT) i.v. dose‐response challenge (5‐100 ug/kg) for respiratory system resistance (Rrs) to assess AR and bronchioalveolar lavage (BAL) to assess inflammatory cells. The Rrs response was not different between groups (max increase OVA 339%+32 sem vs Alum 301%+28; p>0.05), although the BAL eosinophil count was significantly greater in the OVA group (25% + 5) compared to the Alum (0.1% + 0.08). Our results suggest that M2R/M3R functions are critically involved in the development of OVA‐induced AHR.

Grant Funding Source: Supported by Canadian Institutes for Health Research (MOP81211), Intramural Research Pgm NIDDKD, NIH
Age‐related loss of neurotrophic signaling at the diaphragm muscle (871.4)
Authors: Greising Sarah , Ermilov Leonid , Sieck Gary , Mantilla Carlos
Published: June 2021

The age‐related mechanisms of diaphragm muscle (DIAm) sarcopenia are largely unknown. We hypothesize that sarcopenia may be related to neurotrophic signaling, specifically that of brain derived neurotrophic factor (BDNF) acting through the high‐affinity tropomyosin‐related kinase receptor subtype B (TrkB). In total 47, adult male TrkBF616A mice were examined at 6, 18, and 24 months of age (mo), representing survival rates of 100, 90, and 75%. At each age, ex vivo contractility of the DIAm was assessed to determine maximal force as well DIAm‐phrenic nerve neuromuscular transmission failure under the following preincubation conditions; control (vehicle) solution, BDNF, 1NMPP1 (inhibiting TrkB kinase activity), or co‐treatment of BDNF and 1NMPP1. Specific DIAm maximal twitch and isometric tetanic force, in addition to fatigue index were all negatively impacted at 24 mo (P蠄0.02). A significant interaction between age and neurotrophic treatment was found for DIAm‐phrenic nerve neuromuscular transmission failure (P蠄0.01). Specifically, at 6 and 18 mo BDNF decreased failure, while this was lost at 24 mo. Conversely, TrkB kinase inhibition increased failure at 6 mo, but this relationship was lost at both 18 and 24 mo. Taken together, neurotrophic signaling is key to the physiological function of the DIAm, however there is likely an age‐related loss of BDNF and TrkB signaling that detrimentally affects the DIAm.

Grant Funding Source: Supported by R01‐AG044615, T32‐HL105355, and the Mayo Clinic.
The Ca2+‐activated K+ channels play a braking role in locus coeruleus neurons during CO2 exposure (872.1)
Authors: Patrone Luis Gustavo , Bicego Kenia , Putnam Robert , Gargaglioni Luciane
Published: June 2021

The LC has an important role in central chemoreception and K+ channels have been suggested to be sensitive to changes of pH. Studies have shown that some ion channels present in the cell membrane of LC neurons, among them the Ca2+‐activated K+ channels (KCa), are activated by high levels of CO2. These channels appear to hyperpolarize LC neurons and limit increased firing rate in response to hypercapnic acidosis. We evaluated the participation of KCa in the CO2‐drive to breath in unanesthetized adult rats by bilaterally microinjecting Paxiline (KCa blocker, 100µM or 500µM / 0.1µL) or vehicle (aCSF) intra‐LC. Bilateral KCa blockade in LC neurons did not affect resting ventilation. However, the increase in ventilation produced by hypercapnia (7% CO2) was amplified by 39 and 33% after Paxilline (100µM or 500µM, respectively) injection (1305.2 ± 164.2 mL.kg‐1.min‐1 in vehicle group vs 1822.0 ± 201.7 in Paxilline 100µM group and 1735.3 ± 116.5 in Paxilline 500µM group) due largely to a increase in tidal volume. The results suggest that KCa++ modulate the hypercapnic ventilatory response as braking pathway, but that they are not involved in respiratory control during normocapnia, in the LC of adult rats.

Grant Funding Source: FAPESP and CNPq
State‐dependent regulation of breathing by the retrotrapezoid nucleus (872.10)
Authors: Burke Peter , Kanbar Roy , Hodges Walter , Viar Kenneth , Coates Melissa , Stornetta Ruth , Guyenet Patrice
Published: June 2021

In this study we stimulated the retrotrapezoidal neurons (RTN) by optogenetics in behaving animals to determine ventilatory responses across sleep and wake states. We targeted the rostral RTN region (n=12) with a PRSX8 promoter lentivirus and limited ChannelRhodopsin expression to RTN neurons and noradrenergic A5 neurons. A subset of rats (n=6) also received intraspinal anti‐DBH‐saporin, a catecholaminergic‐specific neurotoxin, to eliminate A5 neurons. We observed no significant differences for the resting or evoked ventilatory behavior after effective lesion. In quiet awake and non‐rapid eye movement (nREM; quiet asleep) states, RTN stimulation increased respiratory frequency and tidal volume, and could entrain respiratory rate between 1.2Hz ‐ 2Hz. We also observed active expiration in the wake state only. The increase in breathing by RTN stimulation was occluded by hypercapnia (6% CO2), but not hypoxia (12% O2). Surprisingly, RTN stimulation in REM (active sleep) failed to increase or pace respiratory rate and only tidal volume was significantly increased. This REM phenomenon draws an intriguing parallel to voluntary breathing (active wake), where homeostatic mechanisms are limited to adjusting the depth of each cortically‐triggered breath.

Grant Funding Source: Supported by NIH (HL28785, HL74011)
Antagonism of orexin receptors significantly lowers blood pressure and exaggerated CO2 chemoreflex in spontaneously hypertensive rats (872.2)
Authors: Li Aihua , Hindmarch Charles , Paton Julian , Nattie Eugene
Published: June 2021

The orexin system is involved in cardiorespiratory function as well as wake‐sleep regulation. We recently showed that in spontaneously hypertensive rats (SHRs): 1) blocking both orexin receptors by oral administration of an antagonist, almorexant (almxt), significantly lowers blood pressure in wakefulness and NREM sleep in both dark and light periods of the diurnal cycle, 2) the anti‐hypertension effect is accompanied by decreased sympathetic vasomotor tone and noradrenaline levels in cerebrospinal fluid and plasma, 3) there is a strong trend towards increased orexin‐A mRNA expression in the rostral ventrolateral medulla, a projection site for orexinergic lateral hypothalamic (LHA) neurons, 4) there is an exaggerated CO2 ventilatory chemoreflex, which can be normalized by blocking orexin receptors with almxt. Antagonism of orexin receptors had no effect on resting blood pressure in normotensive WKY rats. We interpret these results to suggest that an overactive orexin system may play a significant role in the development and maintenance of high blood pressure and enhanced CO2 chemoreflex in SHRs. Modulation of orexin and the hypothesized overactive central chemoreflex could be a potential target in treating hypertension.

Grant Funding Source: NIH HL 28066
TASK‐2 channels contribute to pH sensitivity of retrotrapezoid nucleus chemoreceptor neurons (872.4)
Authors: Kumar Natasha , Wang Sheng , Benamer Najate , Zanella Sebastian , Shi Yingtang , Bevengut Michelle , Penton David , Guyenet Patrice , Lesage Florian , Gestreau Christian , Barhanin Jacques , Bayliss Douglas
Published: June 2021

Phox2b‐expressing neurons of the retrotrapezoid nucleus (RTN) function as central respiratory chemoreceptors; they are directly activated by CO2/H+, via an unidentified pH‐sensitive background K+ channel, to drive breathing. Here, we test the effect of TASK2 (K2P5, an alkaline‐activated background K+ channel) channel deletion on the pH sensitivity of RTN neurons and on CO2 modulation of central respiratory output. For patch clamp recordings in brainstem slices, individual RTN neurons were identified by GFP expression (driven by the Phox2b promoter) or β‐galactosidase (from the gene trap used for TASK2 deletion). Whereas 95% of RTN cells from control mice were pH sensitive, only 56% from TASK2‐/‐ mice were classified as pH sensitive; the remaining cells were pH insensitive (44%). The alkaline‐activated background K+ currents were reduced in amplitude in pH‐sensitive RTN neurons from TASK2‐/‐ mice but were absent from pH‐insensitive cells. TASK2 was strongly expressed in 63% of Phox2b‐expressing RTN neurons (by X‐gal staining), and in 85% (by a more sensitive scPCR approach). Using an in situ working heart‐brainstem preparation, we found diminished inhibition of phrenic burst amplitude by alkalization in TASK2‐/‐ mice, with apneic threshold shifted to higher pH levels. We conclude that TASK2 channels represent a molecular substrate for pH sensing in RTN respiratory chemoreceptor neurons.

Grant Funding Source: Supported by: NIH HL074011 and NIH HL108609; and ANR‐RESPITASK and ANR‐11‐LABX‐0015‐01
Coping with acute hypertensive stressors after selective carotid body ablation in conscious spontaneously hypertensive rats (873.10)
Authors: Pijacka Wioletta , McBryde Fiona , Paton Julian
Published: June 2021

We have previously shown that bilateral carotid sinus denervation in SHR reduces arterial pressure (AP). This technique, however, also removes carotid sinus baroreceptors. We hypothesized that the fall in basal AP with selective bilateral CBA would not impair the normal cardiovascular responses to hypoxia, exercise and restraint stress. SHR were implanted with telemetry devices to record AP, and responses to hypoxia (10% O2), exercise (10m/min) and restraint stress were tested. The chemoreflex was tested with i.v. sodium cyanide (120μg.kg‐1); this response was abolished after CBA (n=4) but preserved in Shams (n=3). Basal systolic AP fell significantly after 14 days in CBA (‐18.2±2mmHg) compare to Sham (2.5±0.6mmHg), (P<0.05). SHR with CBA showed a depressor effect to 10% oxygen (‐15.3±7mmHg) whereas a pressor response was seen in Sham (15.9±6mmHg; P<0.029); heart and respiratory rates were not different. The change in AP was similar between groups during both exercise (CBA 12±7 vs. Sham 23±8mmHg, respectively; NS) and restraint stress (CBA 28±8 vs. Sham 17±9mmHg; NS). We conclude that chronic CBA prevents the pressor response to hypoxia but does not affect cardiovascular adjustment to exercise and restraint stress.

Grant Funding Source: Supported by the British Heart Foundation and Cibiem
Peripheral chemoreceptor contribution to ventilatory control during steady‐state exercise (873.2)
Authors: Peinado Ana , Johnson Blair , Taylor Jennifer , Joyner Michael
Published: June 2021

We tested the hypothesis that the carotid body chemoreceptors (CB) contribute to ventilatory control during steady‐state exercise. Six men completed two randomized bouts of cycling exercise at 65% VO2max for up to 2 hours during an IV infusion of dopamine (D; 2 µg/kg/min) to acutely inactivate the CB, or a saline infusion (S). Expired gases were analyzed at baseline, resting‐infusion, and during exercise (25%, 50%, 75% and 100% of total exercise time). A hypoxic ventilatory response (HVR) test was performed after exercise during the infusion condition. Subjects were classified as responders (RS; n=3) if their HVR was lower during D vs. S (1.03 ± 0.6 vs. 2.14 ± 0.4 L/min/%SaO2). In RS, partial pressure of end‐tidal CO2 had a greater delta from baseline at 75% (2.9 ± 2.1 vs. 1.1 ± 1.9 mmHg) and 100% (0.9 ± 3.4 vs. ‐0.3 ±3.3 mmHg) during D than S, and the increase in oxygen uptake at 100% was greater during D vs. S (34.7 ± 2.6 vs. 33.0 ± 2.6 mL/min/kg). In RS, there were no differences in the changes from baseline for ventilation (VE), VE/VCO2, respiratory rate, tidal volume, and respiratory exchange ratio between conditions. In the non‐responders group, there were no differences between D and S in any variables. When the data was pooled, HVR accounted for 37% of the variance in VE at 100%. Despite evidence that CB chemosensitivity was blunted, the CB appear to play a minor role in controlling ventilation during steady state exercise.

Grant Funding Source: NIH R01DK090541
Electrophysiological properties of RVLM pre‐sympathetic neurons modulated by the respiratory network in rats (875.11)
Authors: Moraes Davi , da Silva Melina , Bonagamba Leni , Mecawi André , Zoccal Daniel , Antunes‐Rodrigues José , Varanda Wamberto , Machado Benedito
Published: June 2021

The respiratory pattern generator modulates the sympathetic outflow which strength is enhanced by hypoxia. This coupling is due to the respiratory‐modulated pre‐sympathetic neurons in the RVLM, but the underlining electrophysiological mechanisms remain unclear. For a better understanding of the central generation of respiratory‐sympathetic coupling, we combined immunofluorescence, single cell qRT‐PCR and whole cell patch clamp recordings of the pre‐sympathetic neurons in in situ preparations from normal and rats submitted to chronic intermittent hypoxia (CIH). Our results show that the spinally projected C1‐ and non‐C1 respiratory‐modulated pre‐sympathetic neurons (inspiratory‐modulated: 22; post‐I‐modulated: 23; inspiratory‐inhibited: 11; non‐modulated: 20) present distinct electrophysiological properties, synaptic inputs and expression of ionic currents, albeit all neurons presented iNaP‐dependent intrinsic pacemaker after synaptic blockade. Only the non‐C1 post‐I‐modulated neurons presented enhanced excitatory synaptic inputs from the respiratory network after CIH (n=14), which may contribute to the increased sympathetic activity observed in CIH rats. We conclude that the different respiratory‐modulated pre‐sympathetic neurons contribute to the central generation of respiratory‐sympathetic coupling and in response to the challenges produced by CIH may contribute to respiratory‐related increase in the sympathetic activity and hypertension observed in this experimental model.

Grant Funding Source: Supported by CAPES and FAPESP
Cardiovascular responses to disinhibition of rostral ventrolateral medulla (RVLM) following unilateral RVLM blockade in sedentary or physically active, sinoaortic denervated rats (875.13)
Authors: Dombrowski Maryetta , Azar Toni , Mueller Patrick
Published: June 2021

The RVLM is a bilateral region in the brainstem important for sympathetic control of arterial pressure (AP). Although tonically active, both RVLMs receive strong γ‐amino‐butyric acid (GABA)‐mediated inhibition, due in part to tonic arterial baroreceptor input. Previous studies report that unilateral inhibition of the RVLM in sinoaortic denervated (SAD) rabbits leads to dramatic decreases in AP and sympathetic nerve activity (SNA), demonstrating the importance of the baroreflex to resting AP. Following SAD, a strong GABAergic input to RVLM persists and likely suppresses the activity of the RVLM in the absence of baroreceptor input. However, whether this inhibition is altered in sedentary (SED) and active conditions is unknown. The current study was aimed at testing the hypothesis that inhibition of one RVLM in SAD rats would result in enhanced non‐barosensitive GABAergic input in the remaining intact RVLM, and this input would be greater in the SEDs compared to the active rats (i.e., 12 weeks of chronic wheel running on average =WR ). In Inactin‐anesthetized SEDs (n=4) and WRs (n=4), AP and splanchnic SNA were recorded following surgical SAD. Following inhibition of one RVLM (muscimol 2mM, 90nl), bicuculline (5mM, 90nl) was microinjected into the contralateral RVLM. Preliminary data indicate that changes in mean AP appeared to be higher in the SEDs (∆60±12mmHg) compared to the WRs (∆44±15mmHg). Due to small sample size, however, we were unable to support our finding statistically. Splanchnic SNA responses between SEDs (∆52%±16) and WRs (∆49%±23) did not appear to differ. These preliminary data suggest that in the absence of baroreceptor input, MAP and SNA are significantly reduced due to strong non‐barosensitive GABAergic input to the intact RVLM. However, non‐barosensitive GABAergic input may be altered in response to sedentary and active conditions.

Grant Funding Source: Supported by R01‐HL096787; R01‐ HL096787‐S1
The relationship between muscle sympathetic nerve activity and hemodynamics in women taking oral contraceptive pills (875.2)
Authors: Harvey Ronee , Hart Emma , Charkoudian Nisha , Curry Timothy , Carter Jason , Fu Qi , Minson Christopher , Joyner Michael , Barnes Jill
Published: June 2021

The relationship between mean arterial pressure (MAP), muscle sympathetic nerve activity (MSNA), and total peripheral resistance (TPR) is altered in young women versus men and postmenopausal women due to endogenous sex hormone differences. Exogenous sex hormones in the form of oral contraceptive pills (OCPs) may further affect this relationship. We performed a retrospective review of 136 premenopausal women in whom MSNA and arterial pressure were measured, including 78 women with natural menstrual cycles in the early follicular phase (26±1 yr) and 58 women in the placebo phase of OCPs (24±1 yr). TPR and cardiac output (CO) were measured in a subset of women in whom continuous intra‐arterial blood pressure was recorded. Compared to those with natural menstrual cycles, women on OCPs had greater MAP (89±1 vs. 84±1 mmHg, p<0.01) and tended to have lower MSNA (15±1 vs. 18±1 bursts/100 heart beats, p=0.06). MAP and MSNA were positively correlated (r=0.22, p<0.05), TPR and MSNA were positively associated (r=0.41, p<0.05) and CO and MSNA (r=‐0.44, p<0.05) were inversely associated in naturally cycling women (n=26) but not in women on OCPs (r=0.15, r=0.11, r=‐0.01, respectively; p>0.05; n=38). In contrast to naturally cycling women, there appears to be no relationship between MSNA and hemodynamics in women on OCPs, suggesting that these women may regulate their blood pressure differently than women.

Grant Funding Source: Supported by NIH
Chronic neuroinflammation and cognitive impairment following transient global cerebral ischemia: role of fractalkine/CX3CR1 signaling (877.1)
Authors: Briones Tess , Woods Julie , Wadowska Magdalena
Published: June 2021

Neuroinflammation has been studied extensively in cerebral ischemia. A major participant in neuroinflammation is microglia its activation is usually regulated by the chemokine fractalkine (CX3CL1) and its receptor, CX3CR1. Here, we examined the involvement of CX3CR1 on ischemia‐induced chronic neuroinflammation and cognitive function using small interfering RNA (siRNA). Forty adult male Wistar rats were included in the study and received either ischemia or sham surgery then randomized to receive either CX3CR1 siRNA or scrambled RNA as control starting at 7 days after reperfusion. Behavioral testing commenced 28 days after and all rats were euthanized after behavioral testing. Our data showed that: (i) transient global cerebral ischemia significantly decreased fractalkine/CX3CR1 signaling in the hippocampus; (ii) inhibition of CX3CR1 function exacerbated the ischemia‐induced chronic increased in microglial activation and pro‐inflammatory cytokine levels; (iii) inhibition of CX3CR1 function worsened ischemia‐induced chronic cognitive impairment; (iv) inhibition of CX3CR1 function in sham rats resulted in increase IL‐1β expression and impaired behavioral performance. No significant effect of CX3CR1 on ischemia‐induced neurodegeneration was seen. This study provides important insight to understanding the role of CX3CR1 in chronic neuroinflammation and cognitive impairment.

Grant Funding Source: Supported in part by the National Institutes of Health, NINR grant # RO1 NR005260 and the Wayne Stat
Delayed neuronal death in swine following cardiac arrest and resuscitation (877.12)
Authors: Nguyen Anh , Cherry Brandon , Ryou Myoung‐Gwi , Williams Arthur , Hollrah Roger , Baker Charla , Choudhury Gourav , Olivencia‐Yurvati Albert , Mallet Robert
Published: June 2021

Cardiac arrest, a leading cause of death in the U.S., kills >90% of its victims, and survivors often are disabled by permanent brain injury inflicted by ischemia‐reperfusion. Purkinje cells of the cerebellum and CA1 neurons of the hippocampus are especially vulnerable to post‐ischemic neuronal death. We tested the hypothesis that cardiac arrest in a swine model caused delayed neuronal death. Yorkshire swine (25‐35 kg) were subjected to cardiac arrest‐resuscitation (n = 9) or non‐arrest sham (n = 5) protocols. Ventricular fibrillation was induced by electrical pacing. Precordial compressions (100/min) were given at 6‐10 min arrest, and then sinus rhythm was restored with transthoracic countershocks. NaCl was infused iv at 0.1 mmol/kg/min during CPR and the first 60 min after return of spontaneous circulation (ROSC). At 7 d ROSC, brain regions were fixed in 4% paraformaldehyde and H&E stained. More than 70% of the Purkinje cells were shrunken, lacked dendrites and displayed condensed cytoplasm at 7 d ROSC; in contrast, in shams the majority of Purkinje cells retained the characteristic thick dendrites and well‐defined nuclei. Thus, cardiac arrest‐resuscitation produced marked changes in cerebellar neurons evident 7d after acute insult.

Grant Funding Source: NINDS 076975
The effects of melatonin and therapeutic exercise on brain to spinal cord network organization after ischemic brain injury in rats (877.15)
Authors: Lee Seunghoon , Won Jinyoung , Hong Yonggeun
Published: June 2021

The corticospinal tract (CST), long axons of the cortical pyramidal neurons extending to the spinal cord, connecting with the spinal motor neurons directly or indirectly, is the primary transmission tract from the sensorimotor cortex and, thus, forms the neuroanatomical basis for brain‐controlled voluntary movements of the peripheral muscles, In the view point of this research, we investigated whether CST altered, and affected following melatonin and/or therapeutic exercise intervention from acute to recovery phase after focal cerebral ischemia by MCAo surgery. Following the MCAo, the 8‐week‐old Sprague‐Dawley (SD) rats were treated twice daily with MT (10 mg/kg) at 7:00 and 19:00 hr and control group animals were treated with physiological saline (Veh) instead of MT. Also, the rats were trained twice daily with Ex (20 m/min) at 17:00 and 22:00 hr. Melatonin and/or therapeutic exercise not only showed the increment of neuronal cell in penumbra and contralateral region (p<.01), but also the overexpression of Dcx and NG2‐positive cells (p<.01). Moreover, dually applications of melatonin with therapeutic exercise have a prophylactic effect on neural cell death through Golgi‐cox staining as well as promote to neurite outgrwoth. Moreover, MCAo+MT and MCAo+MT+Ex groups revealed a strengthening of projection fiber from cerebral cortex to spinal motor neuron on both corticospinal and rubrospinal tracts. Finally, melatonin and/or therapeutic exercise might induce the blockage of muscle atrophy through MuRF1/MAFbx signaling pathway as well as the activation of hypertrophy signal pathway. In conclusion, melatonin and therapeutic exercise contributed to faster motor recovery and led to reduced stroke‐induced muscle atrophy. Especially, combined therapy has the most efficiency for both neural network and functional recovery.

Kew words: Focal cerebral ischemia, Melatonin, Therapeutic exercise, Brain‐spinal cord network.
Inhibition of DAPK‐1 induces neuronal cell rescue which is linked to ER stress in stroke animal model (877.16)
Authors: Won Jinyoung , Lee Seunghoon , Hong Yonggeun
Published: June 2021

Background: Death associated protein kinase (DAPK) was identified as key player of multiple cell death. DAPK‐1 related apoptosis is the prototypic member of DAPK family that is activated by signaling pathway such as endoplasmic reticulum (ER) stress. Activation of ER stress response has been found to be associated with neurodegenerative disease. ER stress by the unfolded protein response interfere ER homeostasis and promotes neuronal cell death.

Purpose: Here, we are going to determine that cerebral ischemia induces ER stress by reperfusion brain injury following middle cerebral artery occlusion (MCAo) in rats. Additionally, we tried to identify that DAPK‐1 is an essential component in the programmed cell death pathway.

Results: Focal cerebral ischemic rats by MCAo are exhibited ER stress response and neuronal cell death. However, brain injury volume was reduced in MCAo rats treated with DAPK inhibitor compared to MCAo vehicles suggesting inhibition of DAPK has not only protective effect on neuronal cells death but also resistant to cerebral ischemic injury.

Suggestion: Accordingly, DAPK linked to ER stress may regulate the neuronal cell death. Furthermore, down regulation of DAPK represents a potential therapeutic target for transient focal ischemia and neurodegenerative diseases.
Melatonin plus treadmill exercise synergistically promotes neurogenesis and reduce apoptosis in focal cerebral ischemic rats (877.17)
Authors: Lee Minkyung , Lee Seunghoon , Hong Yonggeun
Published: June 2021

Ischemic stroke results from sudden decrease or loss of blood flow to the brain, resulting in damage to neurological function and neuronal networks, cognition. The purpose of this study, we determined whether melatonin and/or treadmill exercise could induce neurogenesis, thereby promoting functional recovery in ischemic brain. Following the MCAo, Sprague‐Dawley (SD) rats were randomly divided into control (Con), MCAo, MCAo+Exercise (MCAo+Ex), MCAo+Melatonin (MCAo+MT), Melatonin combined Exercise in MCAo (MCAo+MT+Ex). The 8 week old SD rats were treated twice daily with melatonin (10 mg/kg) at 17:00 and 19:00, and trained twice daily with treadmill exercise (20 m/min, 6 days/week) at 17:00 and 22:00. At the Golgi‐cox staining, dendrite density in the MCAo+MT+Ex group was higher than in MCAo group. We confirmed that MCAo+MT+ME group was significantly recovered the neurological symptom in behavioral assessment (p<.01). Moreover, MCAo+MT and MCAo+MT+Ex were significantly increased both DCx and GFAP mRNA expression. In conclusion, we suggest that melatonin with and without treadmill exercise intervention enhanced neurogenesis and neuronal cell reconstruction in damaged brain lesions, thereby induced improvement of neurological dysfunction in animal model.

Key words: Melatonin, Treadmill exercise, Neurogenesis, Apoptosis, Focal cerebral ischemia
Neuron‐targeted Cav‐1 as a novel therapy for motor cortex and hippocampal traumatic brain injury (877.3)
Authors: Posadas Edmund , Cui Weihua , Schilling Jan , Kassan Adam , Niesman Ingrid , Patel Piyush , Head Brian
Published: June 2021

Traumatic Brain Injury (TBI) is the leading cause of death in young adults in the Western world. Membrane/lipid rafts (MLR) are plasmalemmal microdomains enriched in sphingolipids, cholesterol, and the protein caveolin (Cav). We have previously shown that primary neurons in vitro transfected with neuron‐targeted Cav‐1 (SynCav1) significantly enhanced expression of pro‐survival and pro‐growth signaling components and increased dendritic growth and arborization. We therefore extended our studies to an in vivo mouse traumatic brain injury (TBI) model utilizing a controlled cortical impact (CCI; 5 m/sec; +1.5 to ‐2.5 bregma, right hemisphere). CCI was performed on a transgenic mouse that overexpresses Cav‐1 only in neurons (SynCav1TG). We performed motor and cognitive tests on sham (craniotomy only) and CCI TG positive (TGpos) and negatives (TGneg). Motor tests were performed on a weekly basis post CCI. Fear conditioning was performed 35 days post CCI. Preliminary data shows a deficit in the inverted wire grid and hippocampal dependent fear conditioning in CCI TGneg mice. We are currently testing the TGpos mice in our CCI model as well as AAV‐SynCav1 intravenous injections in WT mice 24 hr post CCI. Such data could indicate that neuron‐targeted Cav‐1 may be a novel therapy to limit brain injury after TBI and may be an effective therapeutic intervention for other neurodegenerative diseases.

Grant Funding Source: Supported by VA Merit (B.P. Head) and NINDS (B.P. Head)
Detraining minimizes the protective effects of exercise on stroke recovery in mice (877.9)
Authors: Poinsatte Katherine , Ortega Sterling , Partin Alex , Meeuwissen Anouk , Kong Xiangmei , Zhang Rong , Stowe Ann
Published: June 2021

We previously found that 3‐weeks of voluntary exercise downregulated peripheral cytotoxic CD8 T cells and phenotypically altered resident B cells compared to sedentary controls, but only in animals that maintained exercise (duration, frequency and intensity). Mice that decreased their level of exercise with each passing week were phenotypically closer to sedentary mice, indicating that the neuroprotective effects of exercise may depend on sustained physical conditioning. We hypothesized that the protective changes to immune mechanisms seen immediately following exercise would be diminished after detraining. We developed a detraining paradigm with 3 weeks of voluntary exercise followed by a 2‐week sedentary period, prior to a 60‐min transient middle cerebral artery occlusion in adult male Swiss Webster mice. While exercise significantly reduced infarct volumes (p<0.05), 2 weeks of detraining intensified stroke severity when compared to both exercise animals (p<0.0001) and sedentary controls (p<0.05). Furthermore, detraining resulted in a larger resident CD8 T cell population in the spleen compared to sedentary controls. In conclusion, voluntary exercise does not appear to produce either long‐term neuroprotection against stroke or sustained alterations to immune mechanisms. All the changes seen in the immune system immediately following exercise were lost, and potentially exacerbated, after 2 weeks of detraining. Therefore, termination of exercise prior to injury may worsen the outcome of stroke.
Gender differences in vascular responses to carotid baroreflex stimulation (879.14)
Authors: Lloyd Matthew , Der Andrew , Claydon Victoria
Published: June 2021

The relationships between sympathetic nerve activity, arterial blood pressure and cardiac output are strikingly different in young men and women, and further modified by aging. We hypothesised that some of these differences may be mediated by altered baroreflex control of arterial pressure. Full baroreflex response curves are seldom constructed, particularly for vascular resistance responses, and possible gender differences in these responses are unclear.

We, therefore, used a variable‐pressure neck collar system to deliver a range of positive and negative pressures to the carotid sinus in young men (n=5) and women (n=5). Beat‐to‐beat mean arterial pressure (MAP), R‐R interval, and brachial blood flow velocity (BBFV) were recorded noninvasively. From these data we constructed full cardiac and forearm vascular resistance (FVR=MAP/BBFV) baroreflex response curves. Gain, set point, threshold and saturation were calculated.

Resting heart rates and blood pressures were similar in men and women. Cardiac and FVR baroreflex gains, threshold and saturation were also similar in men and women. The set point for the cardiac responses in women (105±10 mmHg) was greater than in men (77±7 mmHg, p<0.05); the set point for FVR responses was similar. In both groups the latency of the maximal MAP response to negative pressure (13 beats) was larger than positive pressure (7 beats, p<0.05). Overall there was a significant correlation between the set point of the blood pressure response and neck circumference (r= ‐0.64, p<0.05).

These data suggest there may be subtle but physiologically relevant differences in carotid baroreflex control in young men and women. Given the impact of gender and age on susceptibility to hypertension, further study is required to evaluate these responses in older men and women.

Grant Funding Source: None
Effects of life history on oxygen delivery in the grasshopper jumping leg (879.22)
Authors: Zembrzuski Deanna , Kirkton Scott
Published: June 2021

Although the insect tracheal system is extremely efficient, oxygen delivery is reduced during the intermolt period. Intermolt growth results in compressed tracheae, reduced tracheal volume, and reduced femoral air sac ventilation rates. Gravidity reduces tracheal volume in adult female grasshoppers; however, it is unknown whether gravid grasshoppers also have lower ventilation rates. Using micro‐dissections and video analysis, we examined the tracheal system of the abdomen and femur of ten (5 females; 5 males) American locust grasshoppers (Schistocerca americana). We found that abdominal compression rates correlated with the inflation of proximal and distal femoral air sacs. Females (1.99 to 2.95 g) had higher compression rates in both their abdomens (36.2 ± 3.62 min‐1; mean ± s.e.m.) and femoral air sacs (36.8 ± 3.73 min‐1) than male grasshoppers (1.07 to 1.27 g; abdomen 28.6 ± 2.04 min‐1 and femoral air sacs 26.8 ± 1.85 min‐1). The most gravid females showed the highest abdominal and femoral air sac compression rates. These findings suggest that gravid females will compensate for reduced tracheal volumes by increasing tracheal ventilation rates. Current research is examining how gravidity affects tibial and tarsal air sac compression rates.

Grant Funding Source: Supported by Union College Undergraduate Research Grants