bluuebunny/crossref_metadata_2025 huggingface dataset:
Protection Against Pulmonary Injury after Exposure to Blast by N‐Acetylcysteine Amide
Authors: Chavko Mikulas , Adeeb Saleena , Ahlers Stephen T. , McCarron Richard M.
Published: June 2021

Lung contusion is a common problem from blunt chest trauma caused by mechanical forces and by exposure to blast overpressure, often with fatal consequences. Lung contusion is also a risk factor for the development of pneumonia and severe clinical acute lung injury (ALI) and acute respiratory distress syndrome (ARDS). Infiltrating neutrophils are considered to be central mediators of lung injuries after blunt trauma. The purpose of this study was to examine the effect of the antioxidant N‐acetylcysteine amide (NACA, ThiolTech, NY) on progression of lung inflammation after exposure to a moderate level of blast overpressure 140 kPa.

Rats were administered with NACA (i.p.100 mg/kg) or placebo (PBS) 30 and 60 min after exposure and then 24 h later. Non‐blasted sham‐injected animals served as controls.

Myeloperoxidase (MPO) activity in the lung was significantly increased 2 days after blast and returned to controls at 8 days. This increase corresponded with activation of CD11b mRNA and lung inflammatory chemokine mRNA expression (MIP‐1, MCP‐1, and CINC‐1). At 8 days, all inflammatory mediators returned to control levels. In addition, expression of heme oxygenase‐1 (HO‐1) mRNA increased at 2 days after exposure.

NACA significantly reduced infiltration of neutrophil and CD11b mRNA activation in lungs, and completely blocked activation of MIP‐1, MCP‐1 and CINC‐1 mRNA. The relatively higher inhibition of chemokine mRNAs compared with reduction in MPO activity and CD11b is in accordance with an antioxidant effect of NACA on reactive oxygen species (ROS) accumulation, rather than by an effect on neutrophil sequestration. The inhibition of HO‐1 mRNA activation after blast was likely also related to the drug antioxidant effect.

This work was supported by Office of Naval Research Work Unit 601153N.04508.518.A0406.
Prevention of In Utero Nicotine‐Induced Bone Marrow Mesenchymal Stem Cell (BMMSC) Myofibroblast Differentation by Augmenting Lipofibroblast Phenotype
Authors: Rehan Virender K. , Guo Pinzheng , Cruz Emmanuel , Chap Kimberley , Lee Wai‐Nang Paul , Torday John S.
Published: June 2021

Alveolar lipofibroblasts (LIFs) are critical for lung homeostasis and injury/repair. We have shown that in utero nicotine exposure drives the differentiation of LIFs to myofibroblasts (MYFs). Whether BMMSCs can be preferentially driven to LIF phenotype is not known. We hypothesized that PPARγ agonists can prevent in utero nicotine‐induced MYF differentiation of BMMSCs and drive these cells to a LIF phenotype. Sprague Dawley dams were given 1 mg/kg nicotine from day 3 of pregnancy until term. After spontaneous delivery, pups were given either 1 or 3 mg/kg RGZ or saline (control) from postnatal day 1 to 3 weeks, when pups were sacrificed and BMMSCs were isolated and characterized for morphologic, molecular, and functional characteristics by Oil Red O staining, Western blotting, RT‐PCR, triolein uptake, and 1,2‐13 C‐glucose labeling. In utero nicotine exposure resulted in decreased Oil Red O staining, triolein uptake, oxidative/non‐oxidative ribose synthesis, PPARγ expression by BMMSCs, but markedly increased α‐SMA expression; all features, suggestive of in utero nicotine‐induced LIF‐to‐MYF transdifferentiation. Concomitant treatment with RGZ virtually blocked all of these nicotine‐induced morphologic, molecular, and functional changes, suggesting a possible targeted molecular preventive and/or therapeutic approach to prevent in utero nicotine‐induced lung injury (TRDRP 17R0170).
A structure‐guided design strategy to develop ligands with subtype selectivity for human nicotinic acetylcholine receptors
Authors: Yamauchi John G , Nemecz Ákos , Ho Kwok‐Yiu , Fotsing Joseph R. , Weide Timo , Grimster Neil , Stump Bernhard , Park John F. , Nayyar Deepika , Talley Todd T. , Fokin Valery V. , Sharpless K. Barry , Taylor Palmer
Published: June 2021

Nicotinic acetylcholine receptors (nAChRs) are pentameric ligand gated cationic channels expressed in both neurons and muscles, where they mediate action potentials and neurotransmitter release. Obtaining structural information is limited due to the nature of the receptors being membrane bound. However, discovery of soluble acetylcholine binding proteins (AChBPs) identified in mollusks (Aplysia Californica and Lymnaea Stagnalis), have been amenable as structural surrogates for the receptor extracellular domain. In collaboration with K. Barry Sharpless, we have generated a small library of compounds (~500) utilizing chemical synthesis or in situ "Freeze‐Frame Click Chemistry" on the AChBP templates. The building blocks emphasized starting with bicyclic amines (tropanes), isoquinoline, and cumarine chemical entities. Here we report the binding constants of these compounds on the AChBP and functional responses with different neuronal receptors. The binding affinities (kd) for the AChBP ranged from 3547‐0.04 nM. Our studies continued to with x‐ray crystallography for compounds showing tight affinities to the AChBPs.

Support from (R37‐GM18360, UO1‐NS 06856, GM07752‐29)
Women Respond Differently to Theophylline than do Men
Authors: Henkin Robert I. , Velicu Irina , Schmidt Loren
Published: June 2021

Gender differences in drug response are well known and reflect important aspects of drug treatment. We treated 178 women and 134 men with smell loss in a clinical trial in which all patients received oral theophylline for 2‐8 months at daily doses of 200, 400, 600 and 800mg. At the end of each period smell function was measured by subjective responses and standard psychophysical techniques and plasma theophylline was measured by a fluorescence polarization method. Plasma theophylline was consistently higher among women than men at each theophylline dose and differences increased as theophylline dose increased: 200mg, men ‐ 3.6±0.2, women ‐ 4.4±0.3 mg/L (Mean±SEM); 400mg, men ‐ 4.0±0.4, women ‐ 10.2±0.7; 600mg, men ‐ 8.2±0.6, women ‐ 10.8±0.9; 800mg, men ‐ 9.6±0.8, women ‐ 12.5±0.7. Subjective smell improvement at 600 and 800mg in women was consistently higher than in men. Responses to psychophysical testing in women were consistently better than in men at 400, 600 and 800mg and were statistically significant after 600mg. These differences may relate to a relative decreased theophylline metabolic clearance rate in women than in men caused by gonadal hormone differences. The decreased clearance rate could cause slower drug absorption with increased drug blood and tissue levels resulting in a better response of their smell loss. This apparently is the first report of gender differences in theophylline blood levels.
Lack of Drug‐Drug Interaction Between ATI‐5923, a Novel Vitamin K Epoxide Reductase Inhibitor, and Amiodarone, in Beagle Dog
Authors: Choppin Agnes , Irwin Ian , Lach Lach , Davies Adrian , Shao Lei , Becker Cyrus , Palme Monica P , Paliard Xavier , Bowersox Scott , Dennis Donn M , Druzgala Pascal
Published: June 2021

ATI‐5923 is a novel oral anticoagulant and like warfarin, is a vitamin K epoxide reductase inhibitor. Unlike warfarin, which is metabolized by CYP2C9 and CYP3A4, ATI‐5923 is metabolized by carboxylesterases. These studies compared the efficacy of ATI‐5923 to that of warfarin, when administered either intravenously (IV) or once a day orally, to produce stable anticoagulation in beagle dogs. The drug‐drug interaction potential was assessed using co‐administration with amiodarone, an inhibitor of CYP3A4 and CYP2C9, with either ATI‐5923 or warfarin. Effects on coagulation were assessed by measuring Factor VII and X levels and prothrombin times (PT). Continuous IV infusions and oral administration of ATI‐5923 or warfarin caused a dose‐dependent decrease in plasma concentrations of Factor VII and Factor X, and an associated increase in PT. On a dose basis, ATI‐5923 was a more potent anticoagulant than warfarin. Intravenous fresh frozen canine plasma or subcutaneous vitamin K1 treatment reversed the anticoagulant effects of orally‐administered ATI‐5923 in beagle dogs. Co‐administration of amiodarone (40mg/kg for 2 days, 20mg/kg for 6 days) significantly increased the plasma levels and anticoagulation effect of warfarin (0.25mg/kg) in beagle dogs, as measured by an increase in PT and decrease in factor VII and X. In contrast, amiodarone had no effect on the anticoagulation parameters and plasma levels of ATI‐5923 (0.3mg/kg). We conclude that ATI‐5923, via a vitamin K‐dependent mechanism, causes changes in key parameters of hemostasis in beagle dogs that are consistent with effective anticoagulation without the important drug‐drug interaction commonly observed with amiodarone in warfarin‐treated patients.
Effects of Peritoneal Dialysis Solutions on the Murine Peritoneal Microcirculation
Authors: Quinn Kathleen , Fox‐Robichaud Alison
Published: June 2021

Peritoneal dialysis is often used as a treatment for patients with end‐stage renal disease, however it is limited by the development of progressive fibrosis. This study uses intravital microscopy to examine the acute impact of clinically available dialysis solutions on the peritoneal inflammatory response.

To test whether dialysis fluids elicit, or inhibit an inflammatory response, mice (C57Bl/6) were given either saline or TNF intraperitoneally (IP). Three hours later, the test solution was administered IP. After a 60‐minute dwell, the peritoneal microvasculature was examined. This study compares dextrose‐based, lactate buffered Dianeal 1.5%, 2.5%, and 4.25%, as well as the glucose‐based, bicarbonate‐buffered Physioneal 3.86% to Ringer's Lactate.

In non‐TNF stimulated mice, Dianeal 4.25% was associated with a 5‐fold increase in leukocyte rolling, compared to Ringer's Lactate (9.1± 3.7 vs 49.1± 11.2, p<0.0001). This effect was not seen with Physioneal 3.86%. In the TNF‐stimulated mice, reduced leukocyte rolling was observed in all groups, except for Physioneal 3.86%. The cytokine‐induced increase in leukocyte adhesion within the peritoneal venules was not altered by the presence of TNF, except when treated with the Physioneal 3.86%.

The acute presence of lactate‐buffered high glucose peritoneal dialysis fluid is associated with a marked increase in leukocyte rolling, but not adhesion, in the peritoneal microcirculation. This effect is not exacerbated in the presence of the cytokine TNF. The use of a bicarbonate buffer alleviates the pro‐inflammatory properties of high glucose and TNF stimulation.
Selective CB2 receptor activation attenuates cerebral microcirculatory dysfunction during cerebral ischemic/reperfusion injury
Authors: Zhang Ming , Adler Martin , Razdan Raj , Ganea Doina , Tuma Ronald
Published: June 2021

Cannabinoid CB2 Receptor (CB2) activation has been shown to have immunomodulatory properties and our previous studies indicated that selective CB2 activation could attenuate cerebral ischemia/reperfusion (I/R) injury. The goal of current study is to investigate the possible mechanisms of this target during I/R injury, primarily focusing on the cerebral microcirculatory function. Middle cerebral artery occlusion/reperfusion (MCAO/R) was performed in male mice. A CB2 agonist was administered at 3 different doses and times during MCAO/R. Cerebral infarction and neurological function were determined 24 hours after MCAO. Cerebral microcirculatory function was evaluated by measuring of leukocyte/endothelial interactions (in vivo), cell adhesion molecules (ICAM‐1 and Mac‐1) expression (histology) and blood‐brain barrier disruption (Evans Blue extravasation). The results showed a bell shaped dose dependent response after CB2 agonist treatment; CB2 agonist decreased cerebral infraction when even given 3 hours after MCAO. Transient cerebral ischemia increased leukocyte/endothelial interactions, adhesion molecules expression and blood‐brain barrier disruption which were all significantly attenuated by treatment with CB2 agonist (p<0.05). In conclusion, CB2 activation is neuroprotective in cerebral I/R injury partially through the protection of cerebral microcirculatory function.
Intravital Microscopy of the Murine Urinary Bladder Microcirculation
Authors: Kowalewska Paulina , Burrows Lori , Fox‐Robichaud Alison
Published: June 2021

Catheter associated urinary tract infection (UTI) is one of the most common nosocomial infections. The objective of this study was to establish an in vivo mouse model of the urinary bladder microcirculation to study the inflammatory response to UTI.

Female C57Bl/6 mice were challenged intravesically with 0.1, 0.5, 1, 5 or 7 mg/kg of Escherichia coli lipopolysaccharide (LPS) or saline. 3.5 hrs later, the right jugular vein was cannulated, a catheter was inserted into the bladder, the urine drained and 100 μl of warm saline injected. The bladder was gently exteriorized and the bladder microcirculation was recorded for the next hour. In some mice, leukocyte endothelial cell interactions were observed after exposure to anti‐P‐selectin or anti‐α4‐integrin Ab.

LPS at a dose of 5 and 7 mg/kg resulted in a significant increase in leukocyte adhesion and rolling. Leukocyte adhesion at 4 and 4.5 hrs post saline stimulation was 0.9±1.0 and 2.7±1.0 cells/field of view, respectively, and the flux of rolling leukocytes was 21±6.4 and 47±22 cells/min. Leukocyte adhesion at the two time points after 5 mg/kg of LPS stimulation was 17±7.8 and 21±8.0 cells/field of view, respectively, and the flux of rolling leukocytes was 74±17 and 116±56 cells/min. Blockage of α4‐integrin had no effect on leukocyte rolling, whereas the use of a P‐selectin Ab significantly inhibited leukocyte rolling (54.0± 26.0 to 1.25±0.97 cells/min, p<0.05).

These results demonstrate that baseline conditions are maintained within the first half‐hour of microcirculatory observations. Leukocyte rolling within the bladder microcirculation is P‐selectin dependent but α4‐integrin independent. This model can be used to decipher the molecular mechanisms of leukocyte recruitment into the bladder as well as to examine in vivo responses to catheter materials and bacterial infection.
Intestinal ischemia‐reperfusion injury: reversible and irreversible damage imaged in vivo
Authors: Guan Yanfang , Worrell Roger T , Pritts Timothy A. , Montrose Marshall H.
Published: June 2021

Early events in an intestinal ischemic episode have been difficult to evaluate. In vivo microscopy was used to continually report the effects of short (15 min) or long (50 min) ischemia with subsequent reperfusion (IR) on mouse jejunal epithelium. IR was imposed by an inflatable vascular occluder. Blood flow was monitored with confocal imaging. During short ischemia, villus NAD(P)H contents rapidly increased (23%; confocal autofluorescence) and the intracellular pH (BCECF) showed a biphasic response, quickly alkalinizing from 6.8+0.1 to 7.1+0.1 but then strongly acidifying to 6.3+0.1. Upon reperfusion, the pH normalized. In contrast, during long ischemia one‐third of the villus cells remained viable with reversible changes upon reperfusion, but remaining cells lost membrane integrity (Lucifer Yellow uptake) and were shed from the villus. This damage became more pronounced during reperfusion. Irreversible effects of IR mainly occurred at the anti‐mesenteric jejunal side, most likely due to incomplete restoration of blood flow upon reperfusion (stereo microscopy).

In conclusion, adverse effects of short ischemia on the jejunum mucosa are fully reversible by reperfusion but after long ischemia, reperfusion cannot reverse such effects in the majority of cells, which deteriorate further. Our results may help defining the mechanism causing cells to transit from a reversible to an irreversible state during IR.
Computational Analysis of Cardiac Energetics during Ischemia and Reperfusion in Buffer‐Perfused Rabbit Hearts
Authors: Wu Fan , Vinnakota Kalyan C , Beard Daniel A
Published: June 2021

To determine the mechanisms by which nucleotide concentrations change in the heart during ischemia and reperfusion, data from Kroll et al. [Am. J. Physiol. 272:H2567‐2576, 1997] were analyzed using a computational model of cardiac energy metabolism and metabolite transport. In the experiments of Kroll et al., rabbit hearts were subjected to 10 minutes of perfusion at basal conditions; then 45 minutes of ischemia with 95% flow reduction; and then reperfusion. Phosphate metabolite concentrations were measured by 31P‐Magnetic Resonance Spectroscopy (31P‐MRS) and oxygen flux by arterial‐venous differences in oxygen tension during the ischemia reperfusion protocol. Data were analyzed by integrating open adenosine kinetics into a previously developed computational model [Wu et al., J. Physiol. 586:4193‐4208, 2008]. Our analyses find that during ischemia/anoxia the heart actively down‐regulates ATP hydrolysis rate in the cytoplasm, while inhibiting mitochondrial ATP consumption. Down‐regulation of mitochondria ATP consumption during anoxia may be associated with inhibition of F1Fo‐ATPase reversal or reduction of mitochondrial proton leak. During the reperfusion period, the mitochondrial ATP synthesis capacity is largely restored, while mitochondrial proton leak is significantly magnified resulting in a slightly impaired efficiency of mitochondrial oxidative phosphorylation. The loss of adenosine nucleotides during ischemia does not limit the mitochondrial oxidative capacity during reperfusion.
Epicatechin‐induced cardiac protection is opioid receptor dependent
Authors: Kawaraguchi Yoshitaka , Horikawa Yousuke T. , Tsutsumi Yasuo M. , Yokoyama Takaakira , Panneerselvam Mathivadhani , Villarreal Francisco J. , Patel Piyush M. , Roth David M. , Patel Hemal H.
Published: June 2021

Introduction

Epicatechin, a polyphenolic plant metabolite and primary constituent of dark chocolate, has antioxidant properties at high doses and can reduce injury caused by stroke and cardiac ischemia; however, a receptor mediated effect has not been proposed. Epicatechin and other flavonoids are structurally similar to opioids and have opioid receptor activity. Therefore, we tested the hypothesis that low‐dose (sub‐antioxidant) epicatechin mediates cardiac protection via opioid receptors.

Methods

Mice were randomly assigned to the following four groups: control, epicatechin, naloxone (non‐selective opioid antagonist), and epicatechin+naloxone. Epicatechin was administered by oral gavage daily (1mg/kg) for 10 days. Naloxone was administered by intraperitioneal injection daily (5mg/kg) for 10 days. On day 11, mice were subjected to 30 min coronary artery occlusion followed by 2 h of reperfusion. Infarct size was determined via planimetry.

Results

Infarct size was significantly reduced in epicatechin treated mice compared to control (17.8+2.5 vs. 35.5+1.9%, n=8, P<0.001, respectively). Epicatechin‐induced protection was blocked by naloxone (39.6+1.7%).

Conclusion

We show that sub‐antioxidant doses of epicatechin produce cardiac protection via opioid receptors, thus defining a receptor mediated mechanism of epicatechin action.
Roles of Ang‐2 KO in Experimental IBD
Authors: Alexander Jonathan Steven , Jennings Merilyn H , Daley Sarah , Bernas Michael , Witte Marlys , Mathis J Michael , Jordan Paul A , Wang Yuping
Published: June 2021

Angiopoietin‐2 (Ang‐2) is known to influence blood and lymphatic vessel remodeling, but how Ang‐2 gene knockout influences experimental inflammatory bowel disease (IBD) is not known. We used Ang‐2 gene knockout and control mice in the 3% dextran sulfate model of IBD to investigate how the lack of Ang‐2 affects vascular remodeling and indices of disease activity (over 7d). Ang‐2 +/+ and ‐/‐ mice showed equivalent levels of disease activity (stool blood, form and wasting) and weight loss was similar in both groups. There was a signficant increase in both blood (MECA‐32+) and lymphatic (VEGFR‐3+) vessel densities in colitic (DSS‐treated) groups (p<0.05). These increases were significantly reduced in Ang‐2 KO mice, and were also accompanied by decreased concentrations of gut neutrophils (myeloperoxidase activity). Colitic inflamed Ang‐2 ‐/‐ gut tissue show evidence of dysplastic MECA32+/VEGFR3+ networks which were not observed in non‐colitic Ang‐2 KO's. Histopathological scores in DSS colitis were partially, but not completely, resolved by Ang‐2 gene deficiency, and suggest that Ang‐2 plays complex roles in the initiation of IBD possibly by controlling tissue remodeling and leukocyte infiltration of tissues.
Reduction of Erythrocyte (RBC) Deformability Attenuates Low Oxygen Tension (pO2) Induced ATP Release from the RBC
Authors: Sridharan Meera , Adderley Shaquria , Hanson Madelyn , Bowles Elizabeth , Stephenson Alan , Sprague Randy
Published: June 2021

The mechanism by which decreased pO2 stimulates ATP release from RBCs has not been established. Here we evaluated the hypothesis that RBC deformability is required for ATP release from rabbit RBCs in response to exposure to low pO2. Deformability of RBCs, measured with a St. George's Blood Filtrometer, was decreased by incubation of RBCs with diamide, an agent that promotes cytoskeletal cross‐linking. Incubation of RBCs with diamide (100 uM) resulted in a 7.67 ± 1.19 % reduction in RBC deformability by 30 min that was stable for 1 hr (n=4). In untreated RBCs, changes in pO2 from 121.9 ± 2.7 to 13.7 ± 1.2 mmHg resulted in a significant 8.5 ± 2.9 fold increase in ATP release (p<0.01, n=5). In contrast, in diamide treated RBCs, changes of pO2 from 125.8 ± 6.1 to 13.5 ± 0.6 mmHg resulted in only a 2.9 ± 1.0 fold increase in ATP release, which was significantly lower than the increase in untreated RBCs (p<0.05, n=5). To ensure that effects of diamide on low pO2‐induced ATP release were selective for that signaling pathway and was not due to a general inhibition of ATP release, RBCs were incubated with iloprost (ILO), to stimulate ATP release in the absence or presence of diamide. Diamide did not inhibit ILO‐induced ATP release (n=2). These results are consistent with the hypothesis that membrane deformability is required for low pO2‐induced ATP release from rabbit RBCs. (ADA grant RA‐133 and NIH grants HL‐64180 & HL‐89094)
Gender‐related differences in synaptic ecto‐ATPase in skeletal muscle arteries
Authors: Stone Audrey Jon , Evanson Kirk W. , Kluess Heidi A.
Published: June 2021

The purpose of this study was to determine the activity of ecto‐ATPase in the vasculature of males and females. The hypothesis was that females would have greater ecto‐ATPase activity. The left and right femoral arteries, saphenous arteries, and gastrocnemius‐ 1A arterioles (G1A) were removed from F344 rats (male n=8; female n=8). Vessel segments (2mm) were incubated at room temperature in a PiBlue Reagent for 30 minutes. Free phosphate was measured at an absorbance of 620nm. No differences were found between genders for phosphate concentration (males: 1111±517 pM; females: 1046±445 pM). There was no difference in phosphate concentration for left and right vessels (left: 1067±483 pM; right: 1091±483 pM). However, there was a significant difference in vessel type (Femoral: 1292±476 pM; Saphenous: 1160±411 pM; G1A: 784±412 pM; p<0.05). Within males, phosphate concentration differed between femoral arteries and G1A arterioles (1405± 557; 774±414 pM; p<0.05). Within females, phosphate concentration differed between femoral arteries and G1A arterioles (1179±362; 793±423 pM; p<0.05) and differed between saphenous and G1A arterioles (1165±457; 793±423 pM; p<0.05). Although gender does not appear to influence the amount of synaptic ecto‐ATPase in skeletal muscle arteries, vessel type may be influential. Research supported by Arkansas Biosciences Institute and the National Institute on Aging.
Endothelin‐1 constricts cerebral arteries by stimulating physical coupling between type 1 IP3 receptors and TRPC3 channels in myocytes
Authors: Adebiyi Adebowale , Zhao Guiling , Narayanan Damodaran , Thomas Candice M , Jaggar Jonathan H
Published: June 2021

Vasoconstrictors elevate inositol 1,4,5‐trisphophate (IP3), but signaling mechanisms activated by this second messenger in arterial myocytes are poorly understood. Recently, we described that IP3‐induced vasoconstriction can occur independently of intracellular Ca2+ release and via IP3 receptor (IP3R) and TRPC3 channel activation (Xi et al., Circ Res. 2008; 102:1118‐26). Here, we tested the hypothesis that IP3Rs stimulate TRPC channels due to physical coupling between these proteins in cerebral artery myocytes. An IP3R1 antibody co‐immunoprecipitated both IP3R1 and TRPC3 channels from cerebral artery lysate. IP3R1 and TRPC3‐bound fluorescent (Cy2, Cy3) secondary antibodies generated significant immunofluorescence resonance energy transfer (N‐FRET) in myocytes. In contrast, antibodies bound to IP3R1 and TRPM4 generated weak N‐FRET. Endothelin‐1 (ET‐1), a PLC‐coupled receptor agonist, increased N‐FRET between IP3R1 and TRPC3‐bound fluorescent antibodies. A peptide corresponding to the TRPC channel C‐terminus calmodulin and IP3R binding domain (CIRB) reduced ET‐1 and IP3‐induced ICat activation in myocytes. Similarly, a membrane‐permeant (HIV‐1 TAT) CIRB peptide reduced ET‐1‐induced constriction in pressurized arteries. Data suggest that ET‐1 enhances physical coupling between the CIRB domain of IP3R1 and TRPC3, leading to cation current activation and vasoconstriction. R01 HL67061.
The role of the submentalis muscle in neuromechanical control of tongue movements in Bufo marinus
Authors: Mitton Teri , Puljan Dan , Mandal Rakesh , Anderson Curt
Published: June 2021

Our lab is studying the feeding behavior in the marine toad, Bufo marinus, as a model system for the neuronal regulation of coordinated movement. Previous studies have suggested that the brainstem coordinates precise movements through a muscular 'trigger' that signals and initiates pre‐loaded muscle activation. We investigated a small distal muscle in the lower jaw, the submentalis, as the trigger for the feeding behavior in toads. The innervation of this muscle was identified, as well as the distribution of trigeminal nuclei innervating the submentalis. Using high speed videography, we analyzed jaw‐tongue movements before and after denervation of the submentalis. Following denervation, the toads were unable to completely protract their tongues. However, tongue protraction gradually increased and full tongue protraction and successful feeding behavior was observed after 4‐6 attempts, suggesting a potential model for motor learning. Additionally, we have anatomically identified the presence of muscle spindles in the submentalis, suggesting that proprioceptive afferents are regulating the motor output. These data support the idea of the submentalis muscle acting as a trigger for timed feeding movements and the importance of the submentalis in coordinating jaw‐tongue movement.
Characterization of fast oscillatory rhythms in multiple respiratory‐related motor outputs during eupnea in decerebrate adult rat in vivo
Authors: Ono Kenichi , Chun Hyun Hye , Solomon Irene C , Reid‐Martin Inefta
Published: June 2021

Respiratory motor discharges exhibit both slow and fast oscillatory rhythms. The slow rhythms correspond to inspiratory (I), expiratory (E), or phase‐spanning behaviors while the fast rhythms appear to be exclusive to motor outputs exhibiting inspiratory‐related discharges. To further evaluate these respiratory‐related rhythms, we examined basal phrenic (PHR), hypoglossal (XII), and recurrent laryngeal (RLN) nerve bursts recorded simultaneously from decerebrate (unanesthetized) paralyzed mechanically‐ventilated adult rats in vivo. PHR discharge served as an index of I activity, and burst duration (TI) and the duration between bursts (TE) were determined from PHR traces. XII activity exhibited pre‐I and I phase discharge and RLN activity exhibited pre‐I, I, and E phase discharge. In each respiratory‐related motor output, high frequency oscillations (HFO) were identified but they were restricted to the I phase of each burst (i.e., no significant HFO activity was seen during pre‐I or E phases). PHR and RLN HFO peaks were located between 100‐150 Hz, and XII activity exhibited several distinct peaks with a broadband power distribution. HFO onset occurred at ~50% TI for PHR activity and somewhat earlier for XII and RLN activities; RLN HFO activity spanned most of the TI period. These data suggest that fast oscillatory rhythms in respiratory‐related motor discharges underlie only the I phase. Supported by NS049310.
Mapping of serotoninergic and nonserotoninergic connections from the medullary midline to diaphragm motoneurons in the cat
Authors: Rice Cory D , Lois James H , Yates Bill J
Published: June 2021

Prior work in the cat showed that brainstem medial reticular formation (MRF) neurons are infected following injection of the transneuronal tracer rabies virus (RV) into the diaphragm. In the present study, we ascertained whether this population of neurons is serotoninergic. Felines were euthanized at a variety of times following injections of RV into the diaphragm, and brainstem sections were processed using antibodies to detect RV and tryptophan hydroxylase‐2 (TPH2). TPH2+ neurons were concentrated in the raphe nuclei, MRF region dorsal to the inferior olive, lateral reticular formation especially the area near the lateral reticular nucleus (LRN), and dorsal pontine reticular core. At short survival times following RV injections, only a small fraction of the TPH2+ neurons were dual‐labeled, although RV‐infected cells were present in MRF and LRN. At longer survival times, a large fraction of TPH2+ neurons in the medullary raphe nuclei, MRF and LRN were infected by RV, as were TPH2‐ cells in the same areas. However, a much smaller proportion of pontine TPH+ neurons was infected by RV. These data show that both serotoninergic and non‐serotoninergic medullary midline neurons participate in regulating diaphragm activity, mainly through multisynaptic connections.
Exposure to intermittent hypoxia (IH) for 7 days reduces the number of tyrosine hydroxylase‐immunoreactive (TH‐ir) neurons in the caudal NTS
Authors: Aditham Anupama , Andrade Maryann , Cunningham Rebecca , Herrera‐Rosales Myrna , Vitela Melissa , Mifflin Steve
Published: June 2021

In neonates, hypoxia‐ischemia results in cell loss within the NTS A2 cell group. To determine if exposure to IH in adult rats resulted in a loss within the A2 cell group, we counted the number of TH‐ir neurons in the NTS in normoxic rats (n=11) and rats exposed to IH (between 8am and 4pm alternate 21% and 10% O2 every 3 min for 7 days, n=7). TH‐ir was revealed by treatment of sections with a monoclonal TH antibody (Chemicon) followed by a secondary labeled with Alexa Fluor 594. Photomicrographs of NTS sections were counted by at least 3 investigators blind as to the origin of the sections. In normoxic controls, sections of caudal NTS (Bregma ‐14.3) had 21±2 TH‐ir neurons/section compared to 11±3 TH‐ir neurons/section in IH exposed rats (p=.008). Sections of intermediate NTS (Bregma ‐13.6) had 27±3 TH‐ir neurons/section in normoxic rats compared to 14±5 TH‐ir neurons/section in IH exposed rats (p=.04). Sections of rostral NTS (Bregma ‐12.9) had 16±6 TH‐ir neurons/section in normoxic rats compared to 13+6 TH‐ir neurons/section in IH exposed rats (p=.17). These results indicate that exposure to IH reduces the number of TH‐ir neurons in caudal regions of the NTS. A reduction in the number of catecholaminergic neurons might contribute to the pathophysiology associated with exposure to IH.
Progressive augmentation and long term facilitation of ventilation is enhanced in participants with sleep apnea compared to control and is mitigated by reductions in oxidative stress
Authors: Lee D S , Badr M S , Miller R E , Hadden T J , Mateika J H
Published: June 2021

Purpose

To determine whether progressive augmentation (PA) and long‐term facilitation of minute ventilation (vLTF) is enhanced in participants with obstructive sleep apnea (OSA) compared to control. Moreover, to determine whether treatment with an antioxidant cocktail (AOX) mitigates PA and vLTF in OSA participants.

Methods

13 OSA and 13 controls completed two trials. During both trials participants were exposed to intermittent hypoxia (IH) which included 12‐4 minute episodes of hypoxia (PETO2 ‐ 50 mmHg, PETCO2 ‐ 4 mmHg above baseline) followed by 30 minutes of recovery. During the trials participants were administered, in a randomized fashion, either an AOX or a PLB before IH.

Results

During the PLB trial, baseline minute ventilation was similar between OSA and control (18.8 ± 2.1 vs. 14.8 ± 0.5 L/min). PA was evident in both groups; however it was enhanced in the OSA group compared to control (last hypoxic episode ‐ 36.9 ± 2.8 vs. 27.7 ± 2.2 L/min; p = 0.002). During end‐recovery vLTF was greater in the OSA group compared to control (27.7 ± 2.2 vs. 20.2 ± 1.2 L/min; p = 0.01). PA and vLTF were reduced in the OSA group following AOX compared to PLB (PA ‐ 30.6 ± 2.0 vs. 36.9 ± 2.8 L/min, p = 0.02; vLTF 22.3 ± 1.3 vs. 27.7 ± 2.2 L/min, p = 0.002).

Conclusions

PA and vLTF is enhanced in participants with OSA compared to control. PA and vLTF is mitigated after treatment with an antioxidant cocktail.

Funding: NIH (HL‐085537)
Arterial baroreflex control of sympathetic nerve activity in multiple sclerosis
Authors: Davis Scott L , Keller David M , Fadel Paul J , Remington Gina M , Korkmas Melissa A , Frohman Elliot M
Published: June 2021

Up to 50% of patients with multiple sclerosis (MS) experience symptoms of orthostatic intolerance. The aim of this study was to determine if this higher incidence is potentially due to altered arterial baroreflex (ABR) control of muscle sympathetic nerve activity (MSNA) or heart rate (HR). Five patients with relapsing‐remitting MS (4 females/1 male) and 3 healthy age‐matched controls (2 females/1 male) were instrumented for MSNA, arterial blood pressure (Finometer), and HR. ABR gain was calculated from linear regression slopes of both MSNA (relative burst amplitude) and HR plotted against diastolic and systolic pressure (3‐mmHg bins), respectively, during venous administration of sodium nitroprusside followed by phenylephrine (i.e., modified Oxford technique). Baseline burst frequency (number of bursts/min; BF) and burst incidence (number of bursts/100 heartbeats; BI) were significantly lower (P<0.01) in MS patients (BF=5±1, BI =8±1;n=5) compared to controls (BF=25±10, BI=41±13;n=3). ABR control of MSNA may be attenuated (P=0.09) in MS patients (gain=‐1.1±0.7; n=4) compared to controls (gain=‐2.1±0.7; n=3). ABR control of HR is similar (P=0.61) in MS patients (gain=‐0.7±0.3; n=5) and controls (gain=‐0.6±0.4; n=3). These initial findings suggest that impaired ABR control of MSNA may contribute to the higher incidence of orthostatic intolerance in MS patients.

Funded by National MS Society (PP1440)
Skeletal Muscle Overexpression of SOD Normalizes the Exaggerated Exercise Pressor Reflex in Rats with Heart Failure
Authors: Wang Hanjun , Wang Weizhong , Gao Lie , Zucker Irving H. , Wang Wei
Published: June 2021

An sensitized exercise pressor reflex (EPR) in chronic heart failure (CHF), is believed to contribute to the exaggerated sympatho‐excitation observed during exercise in the CHF state. Accumulating evidence also shows that the exaggerated EPR in CHF may be due to an enhanced skeletal muscle mechanoreflex. However, the cause of the exaggerated EPR and the enhanced mechanoreflex in CHF are not well understood. Here, we hypothesized that muscle oxidative stress in CHF contributes to the exaggerated EPR and the enhanced mechanoreflex. We used the technique of adenoviral gene transfer to overexpress SOD proteins (AdSOD1 or AdSOD2, 1×1012 particles in saline/ml, 50 μl, Adempty as a control) in the triceps surae muscle of CHF rats (2‐4 weeks). The EPR was induced by static contraction in response to electrical stimulation of the ventral roots (L4/L5) in decerebrate rats. The mechanoreflex was induced by passive stretch in decerebrate rats. Table 1 shows the effect of overexpression of SOD proteins on the EPR and mechanoreflex in CHF rats. These data indicate that muscle oxidative stress contributes to the exaggerated EPR and the enhanced mechanoreflex in CHF.

Effect of overexpression of SOD on the pressor response evoked by static contraction (EPR) or passive stretch in CHF rats.

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Values are Mean±SE, n=6 in each group,*P<0.05 vs Sham, ?P<0.05 vs CHF+Empty.
The effect of season on the mood, concentration of interleukin‐6, adrenocorticotrophic hormone, metabolites of catecholamine and cortisol level in healthy volunteers.
Authors: Kanikowska Dominika , Sato Maki , Inukai Yoko , Shimizu Yuuki , Nishimura Naoki , Iwase Satoshi , Sugenoya Junichi
Published: June 2021

We investigated seasonal changes in interleukin‐6 (IL‐6), adrenocorticotrophic hormone (ACTH), metabolites of catecholamine (VMA, HVA, and 5‐HIAA) and cortisol in humans. Eight volunteers were investigated at four times during the year at latitude 35°N (i.e., February, May, August and November). The mean atmospheric temperature in the collection periods was higher in the order of summer > autumn ≈ spring > winter. Changes in mood were also monitored by a profile of mood states (POMS) questionnaire. The concentration of IL‐6 was significantly higher in the winter and summer than in the spring and autumn. The concentrations of ACTH, HVA and VMA were significantly higher in summer. No seasonal variation was detected in cortisol. There were significant differences among the seasons in the subscale for tension and anger in the POMS questionnaire; the tension subscale showed significant differences between spring and autumn, with a higher score in spring. The results demonstrate that Il‐6, ACTH, HVA and VMA exhibit statistically significant seasonal rhythms, which might have important diagnostic and therapeutic implications.
Intracellular recording from cardiac ganglion neurons receiving functional vagal inputs
Authors: Pickering Anthony Edward , McAllen Robin M , Harper Alexander A , Paton Julian F
Published: June 2021

Cardiac vagal tone is an important indicator of cardiovascular health, and its loss is an independent risk factor for arrhythmias and cardiac mortality. Thus, it is important to elucidate the factors that modulate cardiac vagal ganglionic transmission. Using the working heart‐ brainstem preparation (Paton 1996, J.Neurosci Meth. 65, 63‐), we report the first intracellular recordings from functionally connected cardiac vagal ganglion cells in situ. The atria were dissected from the ventricles, pinned out, stabilized with a nylon mesh foot, and ganglion cells (n=33) were impaled with sharp microelectrodes (0.5M KCl; 80‐120 MOhm). Spontaneous EPSPs and action potentials (AP) occurred most commonly in post‐inspiration. Activation of bradycardic reflexes (baro‐, chemo‐, nasotrigeminal) increased dramatically EPSP frequency producing AP and atrial rhythm slowing. Spontaneous and reflexly‐evoked AP were mostly triggered by suprathreshold unitary EPSPs rather than by summation of subthreshold EPSPs. Stimulation of the right cervical vagus evoked EPSPs with a latency of 30‐40 ms. Thus, it is possible to record intracellularly from functionally intact cardiac vagal ganglion cells, which exhibit appropriate patterns of ongoing and reflex excitatory synaptic drives. Future studies will elucidate how cardiac vagal transmission may be modified in health and disease.

RMcA holds an NHMRC Principal Research Fellowship; JFRP holds a Royal Society Wolfson Research Merit Award; AEP is a Wellcome Trust Advanced Clinical Fellow
Hyperosmotic evoked sympathoexcitation is blocked by overexpression of macrophage inhibitory migration factor (MIF) in the paraventricular nucleus of hypothalamus (PVN)
Authors: Colombari Eduardo , Colombari Debora Simoes Almeida , Li Hongwei , Sumners Colin , Raizada Mohan K , Murphy David , Paton Julian F. R.
Published: June 2021

In anaesthetized rats, systemic hyperosmotic stimulation (HS) evokes increases in sympathetic nerve activity (SNA) that are mediated by activation of AT1 receptors in the PVN. Further, MIF can antagonize ANG II‐induced cardiovascular effects mediated via the PVN. This inhibitory effect of MIF is due to its intrinsic thiol‐protein oxidoredutase (TPOR) activity. In this study, we evaluated the effect of overexpressing either MIF or C60SMIF (which lacks TPOR activity) in the PVN on the sympathoexcitation induced by HS. Male Wistar rats (65‐85 g) were decorticated to make insentient and perfused intra‐arterially. HS was performed by raising the perfusate osmolality from 290 to 380 mOsmol for 40 s. Adeno associated 2 viruses for over expression of MIF (1.0x 108), C60SMIF (1.0x 108) or eGFP (8.3 x 108) were injected bilaterally (500 nl/side) into the PVN as was saline as a control. Seven to 10 days later the HS‐induced sympathoexcitation in both the saline and eGFP groups (increases of 27 ± 4 and 25 ± 4%, respectively) was not observed in the MIF group (4 ± 5%). Conversely, the HS induced SNA response was potentiated (45 ± 6%) in the C60SMIF group. We propose that MIF is a major counter regulator of HS induced sympathoexcitation, which is dependent on its TPOR activity. Further, manipulation of TPOR may have therapeutic potential in restricting salt‐induced hypertension. Funded by CNPq (Brazil), CAPES (Brazil), The Royal Society.
Activated microglia in the hypothalamic paraventricular nucleus contribute to sympathoexcitation in heart failure
Authors: Kang Yu‐Ming , Su Yu‐Kun , Ma Ying , Qin Da‐Nian , Guo Zheng , Lei Jing‐Hui , Guo Zhou‐Wei , Zheng Jin‐Ping , Francis Joseph
Published: June 2021

Recent studies have demonstrated that the elevation of proinflammatory cytokines (PIC) in the paraventricular nucleus of the hypothalamus (PVN) contributes to sympathoexcitation in heart failure (HF). However, the origin of these cytokines is unknown. We hypothesized that microglia are activated in the PVN, and that contribute significantly to release the PIC that stimulate sympathoexcitatory systems in HF. Adult male Sprage‐Dawley rats underwent left anterior descending coronary artery ligation to induce HF or sham surgery (SHAM). Subsequently, animals received bilateral PVN infusion of the nuclear factor κB (NF‐κB) inhibitor pyrrolidine dithiocarbamate (PDTC, 5µg/hr), or vehicle for 4 weeks. Compared with SHAM rats, HF rats had higher levels of TNF‐α and more activated microglia in the PVN, higher TNF‐α protein expression in myocardium cells, and augmented renal sympathetic nerve activity. HF rats also had higher left ventricular end‐diastolic pressure (LVEDP), and higher right ventricle/body weight (BW) and lung/BW ratio. PDTC ameliorated cardiac dysfunction in HF rats, decreased TNF‐α in the PVN and cardiac tissues, reduced the numbers of activated microglia, and downregulated renal sympathetic drive. These findings suggest that activated microglia in the PVN contribute to the sympathoexcitation by releasing cytokines in HF. Supported by NIH HL‐080544.
RAGE Contributes to Aldose Reductase mediated myocardial ischemia‐reperfusion Injury
Authors: Li Qing , Ananthakrishnan Radha , Shang Linshan , Quadri Noseerudin , Vedantham Srinivasan , Yan Shi Fang , Schmidt Ann Marie , Ramasamy Ravichandran
Published: June 2021

The efficacy of reperfusion therapy in limiting infarction and improving recovery of contractile function depends on the amount of irreversible damage occurring prior to initiating reperfusion. In this context, our earlier studies demonstrated that the activity of myocardial aldose reductase (AR) (a key regulatory enzyme in polyol pathway) is increased during ischemia and that inhibition of AR protected ischemic myocardium. Mechanisms by which increased AR mediates ischemia‐reperfusion (IR) injury is evolving. We have hypothesized that AR contributes to generation of advanced glycation end product (AGE) precursors, thereby facilitating the interaction of AGEs with its receptor (RAGE) and subsequent signaling for cellular injury in myocardial infarction. To determine the contribution of AR pathway towards generation of key AGE precursors, 3‐deoxyglucosone (3‐DG) and methylglyoxal (MG), we measured myocardial levels of 3‐DG and MG in Wild type (WT), and human AR expressing (hARTg) transgenic mice. To determine if AR modulates IR injury via increases in AGE‐RAGE interaction, we performed LAD occlusion (30 min) followed by 48 hours of reperfusion in WT, hARTg, RAGE knockout (RKO), RAGE knockout bred into hARTg background (AR‐RKO) mice hearts. 3‐DG and MG levels were significantly greater in hAR Tg hearts than in WT (p<0.05), with corresponding increases in AGE levels and RAGE transcripts. Infarct area was significantly greater and functional recovery was poor in the hARTg mice compared to WT mice hearts (p<0.05). Significant reduction in infarct area was observed in ARI treated hARTg, RKO, and AR‐RKO vs WT mice hearts (p<0.05). These data indicate that genetic deletion of RAGE reduces infarct area in hARTg and WT mice hearts, and that AR mediates IR injury via RAGE‐dependent and RAGE‐independent pathways.
Transient alkalosis during early reperfusion blocks helium preconditioning against myocardial infarction: restoration of protection by cyclosporin A
Authors: Weihrauch Dorothee , Krolikowski John G , Pagel Paul S
Published: June 2021

Maintenance of acidosis during early reperfusion after coronary artery occlusion produces cardioprotection. We tested the hypotheses that brief alkalosis during reperfusion blocks helium (He) preconditioning and that the mitochondrial permeability transition pore (mPTP) inhibitor cyclosporin A (CsA) restores this protection in vivo. Barbiturate‐anesthetized rabbits were instrumented for hemodynamic measurement and subjected to 30 min coronary occlusion and 3 h reperfusion. Rabbits were randomly assigned to receive 0.9% saline or 3 cycles of 70% He‐30% O2 administered for 5 min interspersed with 5 min of an air‐O2 mixture before coronary occlusion with or without transient alkalosis (pH = 7.5 to 7.6) produced by sodium bicarbonate (10 mEq) administered 2 min before reperfusion. Other He preconditioned rabbits received CsA (5 mg/kg) and transient alkalosis or CsA alone. He reduced infarct size (25±4% of left ventricular area at risk; TTC staining; P<0.05) compared with saline (44±6%). Alkalosis did not alter infarct size alone (46±2%), but blocked He‐induced cardioprotection (45±3%). CsA restored He preconditioning during alkalosis (28±6%; P<0.05), but did not affect infarct size alone (43±6%). Transient metabolic alkalosis during early reperfusion abolishes He preconditioning and CsA restores this cardioprotection. The results suggest that He preconditioning may inhibit mPTP formation by maintaining acidosis during early reperfusion in vivo.
Obesogenic Western diet changes fatty acid composition in failing rat heart
Authors: Harmancey Romain , Wright Nathan R. , Algahim Mohamed F. , Wilson Christopher R. , Taegtmeyer Heinrich
Published: June 2021

We have previously found that 'Western' diet (WD), but not high fat diet (HFD), adversely affects cardiac function in rats. We now tested the hypothesis that the deleterious effects of WD may be driven by unique alterations in cardiac lipid content. We analyzed intracardiac lipid parameters after 1 week (acute), 8 weeks (short), 24 weeks (intermediate) and 48 weeks (long term) of diet.

Results

triacylglycerol and diacylglycerol levels increased transiently with WD and HFD, while ceramide and cholesterol levels were unchanged. Lipid peroxide levels increased both with WD and HFD, and were accompanied by reduced plasma membrane fluidity. However, only WD led to an increase of the saturated and monounsaturated long chain fatty acyl‐coenzyme A species. This change was followed by a decrease in the ratio of unsaturated to saturated fat, and by greatly reduced levels of the cytoprotective, insulin‐sensitizing, palmitoleate species (p<0.05).

Conclusion

WD induces changes in cardiac fatty acid composition, with a robust enrichment in saturated species and a loss of protective monounsaturated fat. We speculate that these changes contribute to contractile dysfunction. (Supported by NIH‐R01‐HL073162)