bluuebunny/crossref_metadata_2025 huggingface dataset:
A Comparison of Protocols for Simulating Hemorrhage in Humans: Step vs. Ramp Lower Body Negative Pressure
Authors: Rosenberg Alexander (A.J.) , Kay Victoria L , Anderson Garen K , Sprick Justin D , Rickards Caroline A
Published: June 2021

Introduction

Lower body negative pressure (LBNP) elicits central hypovolemia, and has been used to characterize the cardiovascular and cerebrovascular responses to simulated hemorrhage in humans. LBNP protocols traditionally employ a progressive stepwise reduction in pressure that is maintained for specific time periods. More recently, however, continuous ramp LBNP protocols have been utilized to simulate the continuous nature of most bleeding injuries. The aim of this study was to compare tolerance and hemodynamic responses between a step LBNP protocol and a continuous ramp LBNP protocol until the onset of presyncope.

Methods

Healthy human subjects (N=20; 8F, 12M) participated in two LBNP protocols to presyncope: 1) Step Protocol, where chamber pressure decreased every 5‐min to −15, −30, −45, −60, −70, −80, −90 and −100 mmHg, and, 2) Ramp Protocol, where chamber pressure decreased 3 mmHg/min. Heart rate (HR), mean arterial pressure (MAP), stroke volume (SV), middle and posterior cerebral artery velocity (MCAv and PCAv), muscle and cerebral oxygen saturation (SmO2 and ScO2), and endtidal CO2 (etCO2) were measured continuously. Time to presyncope, the cumulative stress index (CSI; summation of chamber pressure*time at each pressure), and hemodynamic responses were compared between the two protocols.

Results

Time to presyncope (Step: 1611.8 ± 80.5 s vs. Ramp: 1675.4 ± 68.3 s; P=0.17), and the ensuing magnitude of central hypovolemia (%Δ SV, Step: −54.3 ± 2.5 % vs. Ramp: −51.9 ± 2.7 %; P=0.31) were similar between protocols, despite a higher CSI for the step protocol (Step: 946.5 ± 98.4 mmHg*min vs. Ramp: 836.7 ± 81.6 mmHg*min; P=0.06). While there were no differences at presyncope between protocols for the maximum change in HR, MCAv, or SmO2 (P≥0.21), the reduction in MAP was slightly less (Step: −17.1 ± 1.8 % vs. Ramp: −20.0 ± 1.4 %; P=0.02) and the reductions in PCAv, ScO2, and etCO2 (P≤0.08) were slightly greater for the step protocol compared to the ramp protocol.

Conclusion

These results suggest that step and continuous ramp LBNP protocols elicit relatively similar tolerance times, reductions in central blood volume, and subsequent reflex hemodynamic responses, despite a greater cumulative stress in young healthy adults.

Support or Funding Information

Funding for this study was provided by the U.S. Army MRMC Combat Casualty Care Research Program Grant # W81XWH‐11‐2‐0137. The content is solely the responsibility of the authors and does not necessarily represent the official views the US Department of Defense.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Ampakine pretreatment enables a single brief hypoxic episode to evoke phrenic motor facilitation
Authors: Wollman Lila Buls , Streeter Kristi , Fuller David
Published: June 2021

Glutamatergic synaptic transmission acting through the amino‐3‐hydro‐5‐methyl‐4‐isoxazolepropionate receptors (AMPARs) is critical for respiratory rhythmogenesis and activation of respiratory motor neurons. Ampakines are a class of synthetic pharmacological agents that act as positive allosteric modulators of AMPARs to enhance glutamatergic currents. Ampakines can be respiratory stimulants, especially during conditions of low respiratory motor output. We previously reported that pretreament with ampakine CX717 enhances long‐term facilitation of inspiratory bursting in the hypoglossal nerve following repeated hypoxic exposures in mice. Here, we used an adult rat model to test the hypothesis that ampakine pretreatment would enable a single bout of hypoxia to produce a sustained facilitation of phrenic nerve inspiratory burst amplitude. Phrenic nerve output was recorded in urethane anesthetized, mechanically ventilated and vagotomized male Sprague‐Dawley rats. Inspiratory burst amplitude and frequency, as well as heart rate and mean arterial pressure were evaluated in three experimental groups: 1) intravenous (IV) ampakine (Cx717, 15 mg/kg) (baseline PaCO2= 46.3±2.5 mmHg, n=8), 2) IV Cx717 followed 2 minutes later by a single, 5 min hypoxia challenge (baseline PaO2= 47.2±1.8 mmHg, n=8), and 3) IV vehicle (10% (2‐Hydroxypropyl)‐bcyclodextrin) + hypoxia (baseline PaO2= 46.7±5.2 mmHg, n=8). IV delivery of Cx717 produced a small, transient increase in phrenic burst amplitude and frequency which returned to baseline values by ~ 20 min. When IV ampakine was followed by hypoxia there was a sustained facilitation of phrenic inspiratory bursting that persisted to 90‐min following hypoxia (132±33 %baseline amplitude). There were no sustained changes in heart rate or blood pressure in any of the experimental groups. The sham+hypoxia treatment did not evoke phrenic facilitation, and to our knowledge, there is no evidence in the literature to suggest that a single brief bout of hypoxia can induce sustained facilitation of phrenic nerve bursting – rather the established “long term facilitation” model requires multiple bouts of hypoxia. We conclude that a low dose of ampakine Cx717 has a small and transient impact on cardiorespiratory function, but when paired with a single bout of hypoxia can induce sustained respiratory plasticity. This approach may have value in the context of hypoxia‐based neurorehabilitation strategies.

Support or Funding Information

1T32HL134621‐01A1 (LBW), K99 HL143207‐01 (KS), F32NS095620‐01(KS), 1R01HL139708‐01A1 (DDF)

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Deficiency of the Arginase 2 Gene Protects Neonatal Mice from Hyperoxia‐Induced Lung Injury
Authors: Nelin Leif D , Jin Yi , Meng Xiaomei D , Chen Bernadette , Liu Yusen
Published: June 2021

Bronchopulmonary dysplasia (BPD) is the most common co‐morbidity following preterm birth and is due to neonatal exposure to mechanical ventilation and/or supplemental oxygen. Neonatal mice placed in hyperoxia (85% oxygen) on post‐natal day 1 (P1) develop histological evidence of lung injury and abnormal lung development manifest by fewer and larger alveoli, otherwise termed alveolar simplification, that is reminiscent of BPD in humans. We have found in adult models of lung injury that arginase 2 induction is involved in the pathogenesis of lung injury. We tested the hypothesis that neonatal mice deficient in the Arginase 2 gene (Arg2) would have attenuated alveolar simplification. Time‐dated pregnant Arg2 deficient mice (Arg2−/−) on a C57BL/6 background and C57BL/6 (wild‐type) mice were used in these studies. Neonatal mice were placed in either room air (normoxia) or 85% oxygen (hyperoxia) by placing their cages in a Plexiglas chamber and flowing either room air or 85% oxygen through the chamber at 10 lpm. Dams were rotated daily between a normoxic litter and a hyperoxic litter to prevent oxygen toxicity in the dam. There were 4 experimental groups, wild‐type normoxia, wild‐type hyperoxia, Arg2−/− normoxia, and Arg2−/− hyperoxia. After 1, 7 or 14 days the neonatal animals were euthanized and the lungs were inflation‐fixed for H&E staining and determination of alveolar area, perimeter and number using automated software. After 1 day in hyperoxia, there were no differences between any of the four groups in terms of alveolar area (size) or alveolar number. After 7 days in hyperoxia, wild‐type mice had a substantially greater mean alveolar area than did normoxic wild‐type mice (1135±41 μm vs 465±17 μm, p<0.001). While there was no difference in normoxic wild‐type or normoxic Arg2−/− mice, hyperoxic Arg2−/− mice had significantly lower mean alveolar area (856±45 μm) than did hyperoxic wild‐type mice (p<0.001). Similar differences in alveolar perimeter were seen between the groups, i.e. hyperoxic wild‐type mice had the greatest perimeter, hyperoxic Arg2−/− were significantly lower than hyperoxic wild‐type but not as low as normoxic‐exposed animals. Furthermore, hyperoxic wild‐type mice had substantially lower mean alveolar numbers than did normoxic wild‐type mice (35±2 vs 89±3, p<0.001). There was no difference between normoxic wild‐type mice and normoxic Arg2−/− mice in mean alveolar number. Hyperoxic Arg2−/− mice had significantly more alveoli (50±2) than did hyperoxic wild‐type mice (p<0.001). These differences in alveolar area, perimeter, and number seen at 7 days remained at 14 days. Taken together these data demonstrate that deficiency of Arg2 attenuates the alveolar simplification caused by exposing neonatal mice to hyperoxia.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Transcriptional Characterization of Human Lung Cells Identifies Novel Markers for Mesenchymal Lineages
Authors: Danopoulos Soula , Bhattacharya Soumyaroop , Mariani Thomas J , Al Alam Denise
Published: June 2021

Rationale

The lung mesenchyme gives rise to multiple distinct lineages of cells in the mature respiratory system, including smooth muscle cells (SMCs) of the airway and vasculature, vascular endothelial cells and parenchymal fibroblasts. However, a thorough understanding of the specification and inter‐relationship among mesenchymally derived cell diversity in the human lung is lacking.

Methods

We completed single cell RNA sequencing analysis of human pseudoglandular and early canalicular stage human lung cells. Canonical correlation analysis, clustering, cluster marker gene identification and tSNE representation was performed in Seurat. Cell populations were annotated using Toppfun. Immunohistochemistry (IHC) and in situ hybridization (ISH) of human lung tissues from multiple developmental stages validated spatial gene expression patterns for key marker genes.

Main Results

We identified molecularly distinct populations representing “committed” fetal human lung endothelial cells, pericytes, and smooth muscle cells (SMCs). Early endothelial lineages expressed “classic” endothelial cell markers (PECAM, CDH5, VWF) while pericytes expressed PDGFRB, THY1 and basement membrane molecules (COL4, laminin, proteoglycans). We also observed a large population of “nonspecific” human lung mesenchymal progenitor (HLMP) cells characterized by expression of COL1, PDGFRA and multiple elastin fiber genes (ELN, MFAP4, FBLN1). We further characterized diversity of mesenchymal lineages defined by ACTA2 expression. Two mesenchymal cell populations, with the highest levels of ACTA2, expressed unique sets of markers associated with airway‐ or vascular‐associated SMCs. ISH for unique SMC cluster marker genes confirmed early and persistent spatial specification of airway (HHIP, MYLK, IGF1) and vascular (NTRK3, MEF2C) SMCs in the developing human lung.

Conclusion

These data add significantly to our understanding of transcriptional phenotypes of human lung cells and provide the first ever description of molecular heterogeneity at the single cell level in human fetal lung tissue. From these data, we can identify a rich and heterogeneous set of mesenchymal cell populations, and define distinct molecular profiles for fetal human lung endothelial cells, fibroblasts, pericytes and smooth muscle cells. These data also facilitated the identification of markers for molecularly distinct populations of airway‐ and vascular smooth muscle cells, which were spatially validated across fetal development.

Support or Funding Information

This work is funded by NIH/NHLBI R01HL (to DA), and the Hastings center for pulmonary research (to SD).

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Recombinant Human MG53 Mediated Protection against Injury to Neuronal Cells
Authors: Paleo Brian J , Madalena Kathryn , Mital Rohan , Bodnar Tom , Rose Aubrey , McElhanon Kevin , Lerch Jessica , Weisleder Noah
Published: June 2021

Injuries to the nervous system can cause irreversible damage to mental and motor function. This has led to extensive interest in studying mechanisms of neuronal regeneration and recovery from injury. Since many neurons are terminally differentiated, increasing the emphasis on maintaining cell survival could provide more translational potential for treatment of neuronal diseases and injuries. While several cell types are known to survive injury through plasma membrane repair mechanisms, there has been little investigation of membrane repair in neurons and even fewer efforts to target membrane repair in diseases affecting neurons. Much of the current understanding of the plasma membrane repair response comes from studies of these repair processes in striated muscle cells. Studies from our laboratory group and others demonstrated that mitsugumin 53 (MG53), a muscle‐enriched tripartite motif (TRIM) family protein also known as TRIM72, is an essential component of the cell membrane repair machinery in skeletal muscle, cardiac muscle, liver, and alveolar epithelial cells. Interestingly, recombinant human MG53 (rhMG53) when applied exogenously can increase membrane repair and integrity both in vitro and in vivo. Increasing the membrane repair capacity of neurons could potentially minimize the death of these cells and affect the progression of various neuronal diseases. While there have been previous studies of membrane repair mechanisms in skeletal muscles that are relevant to neurotrauma, there has been little investigation of membrane repair specifically in neurons. While the action of rhMG53 has been proven to be robust, lack of knowledge of membrane repair in the nervous system leads to questions of the potential efficacy of membrane repair in neurons. In this study we assess the therapeutic potential of rhMG53 in the treatment of various traumatic neuropathies. We find a robust repair response exists in various neuronal cells and that rhMG53 can increase neuronal membrane repair in vitro and in vivo. These findings provide novel, direct evidence of conserved membrane repair responses in neurons and that these repair mechanisms can be targeted as a potential therapeutic approach for the treatment of neuronal injury.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Peroxynitrite induces depolarization and impairments of respiration in isolated murine brain mitochondria
Authors: Albuck Aaron L. , Sakamuri Siva S. V. P. , Sperling Jared A. , Sure Venkata N. , Zheng Sufen , Katakam Prasad V. G.
Published: June 2021

Objective

Peroxynitrite, generated from nitric oxide and superoxide, plays a pathological role in both ischemic and neurodegenerative brain injury. Mitochondrial dysfunction is a key initiator of cerebrovascular and neurological pathologies. Thus, understanding the acute effects of peroxynitrite on brain mitochondrial physiology will help identify potential therapeutic targets. The objective of the study was to investigate the impact of peroxynitrite on respiratory function and membrane potential of isolated murine brain mitochondria.

Methods

Mitochondria were isolated from the brains of 12‐week‐old C57bl6 (n= 5–6) mice and treated with various concentrations of peroxynitrite (0.1μM, 1μM and 5μM) for one hour in assay buffer containing 10mM pyruvate and 2mM malate. Mitochondria (10μg/well) were plated into cell plates and oxygen consumption rate (OCR) was measured using Seahorse XFe24 Analyzer after serial injection of ADP (5mM), oligomycin (5μM), FCCP (5μM) and antimycin (10μM)/rotenone (2μM). Respiratory parameters were reported as OCR (pmoles/minute). Mitochondrial membrane potential was measured using fluorescence probe rhodamine 123 dye. Western Blots were then performed to detect the nitrotyrosine residues in the mitochondrial proteins.

Results

Acute peroxynitrite treatment decreased basal, or state II respiration, by 55.5 % (46.7±4.7 vs 102.5±2.5, n=6, p=0.0004) in brain mitochondria. Similarly, 5μM peroxynitrite treatment of isolated mitochondria diminished the uncoupler‐induced, or state III respiration, by 58% (94.5±12.9 vs 225.9±45.3, n=6, p=0.03). In addition, 5μM peroxynitrite reduced the mitochondrial membrane potential by 65.6% compared to untreated mitochondria (34.4 ±13.2 vs. 100, n=6, p‐value < 0.05). Notably, isolated mitochondria treated with peroxynitrite displayed increased nitrotyrosine content confirming the nitrotyrosination of the mitochondrial proteins.

Conclusions

Acute treatment with peroxynitrite depolarizes isolated mitochondria leading to impairment of mitochondrial respiration which in turn impairs the ability of mitochondria to provide energy at the basal level and in response to increasing demand. Thus, peroxynitrite exhibits direct effects on the mitochondrial respiration which has deleterious consequences for neuronal survival in ischemic and neurodegenerative brain injury.

Support or Funding Information

Undergraduate Summer Research Fellowship by American Physiological Society (Albuck); and National Institute of Health: National Institute of General Medical Sciences and National Institute of Neurological Disorders and Stroke (Katakam: R01NS094834).

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Primary Cilia of Supraoptic Nucleus Regulate Drinking and Blood pressure
Authors: Jiang Jingwei , Morgan Donald , Rahmouni Kamal
Published: June 2021

Fluid intake is tightly related to blood pressure control through balance between electrolytes and water in the body. Virtually, every mammalian cell is equipped with a primary cilium, a cell surface protrusion that is thought to act as a sensory organelle. However, the contribution of primary neuronal cilia to cardiovascular regulation and fluid homeostasis has not been addressed. We hypothesized that primary neuronal cilia contribute to the control of fluid intake and blood pressure. For this, we first examined the length of primary neuronal cilia in the brain nuclei that contribute to osmolarity and cardiovascular regulation in 3‐week DOCA‐salt treated mice vs sham controls (n=5/group). Cilia length was examined by immunostaining a ciliary protein, adenylate cyclase 3 (AC3), followed by confocal 3D reconstruction, and quantification by IMARIS imaging analysis software. We found changes in cilia length in a number of brain nuclei of DOCA‐salt mice. The supraoptic nucleus (SON), a key brain nucleus in the regulation of osmolarity and fluid homeostasis, displayed the most significant changes in cilia length (average primary cilia: 7.46±□□□□□m in DOCA‐salt mice vs 6.76±□□□□□m in sham, p=0.0509). Moreover, the number of primary cilia that are over 10□□m was significantly increased in the SON of DOCA‐salt mice by 8% (p=0.0114). On the other hand, the number of cilia that are 4–5□□m in length was significantly decreased in the SON of DOCA‐salt mice (11.73±1.70%) compared to sham controls (18.73±2.02%, p=0.0385). Next, we assessed the relevance of SON cilia by targeting the ift88 gene which encodes a protein necessary for the formation of cilia. For this, we performed targeted stereotaxic injection of AAV‐Cre into the SON of IFT88flox/flox mice. We found that disruption of SON ift88 (AAV‐Cre group had a 38% ift88 expression level relative to AAV‐GFP control group) caused impairment in water intake in response to 24h water deprivation (30min water intake: 0.78±0.06 ml in AAV‐Cre vs 1.07±0.08 ml in AAV‐GFP, p=0.0129) without affecting total daily water and food consumption. Moreover, ift88 deletion in SON increased systolic blood pressure (133.7±1.71 mmHg vs 115.8±2.33 in AAV‐GFP, p=0.0008) without altering heart rate. Our data demonstrate the importance of primary cilia in the SON in the regulation of water intake and blood pressure. We speculate that primary neuronal cilia may function as part of environmental surveillance system in the brain that ensures cardiovascular homeostasis.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Pre‐autonomic corticotropin‐releasing hormone neurons in the paraventricular nucleus of the hypothalamus may contribute to chronic unpredictable stress‐induced blood pressure hyperreactivity
Authors: Wang Lei A , Cross Sissy , Mifflin Steve
Published: June 2021

Exposure to chronic unpredictable stress (CUS) has been shown to augment the pressor response to novel stress. The neural circuits mediating the CUS induced blood pressure hyperreactivity are not fully understood. Exposure to CUS is known to increase the production of corticotropin‐releasing hormone (CRH) in the paraventricular nucleus of the hypothalamus (PVN). A subpopulation of PVN CRH neurons project to the rostroventral medulla (RVLM), a key nucleus regulating sympathetic outflow and blood pressure. To study how CUS exposure affect RVLM‐projecting CRH neurons in the PVN, we injected retrograde AAV expressing Cre‐inducible mCherry into the RVLM of CRH‐Cre mice. We then exposed mice to the a CUS paradigm. The CUS paradigm included the following stressors: hot air stream (10 min), cold temperature (4°C, 1h), fox urine exposure (10 min), hypoxia (10% oxygen, 20 min), shaker (100 oscillations/min, 1h), forced swimming (10 min). These stressors were randomly assigned to the mice twice daily for 14 days. We found that mice exposed to the CUS had significantly increased number of RVLM‐projecting CRH neurons in the PVN compared to mice that were not exposed to the CUS (CON, 32 ± 4 vs CUS 81 ± 9; p = 0.003). To study whether RVLM‐projecting CRH neurons in the PVN regulates blood pressure, we expressed Channelrhodopsin 2 (ChR2) specifically into the PVN CRH neurons by injecting AAV expressing Cre‐inducible ChR2 into the PVN of CRH‐Cre mice. Optogenetic activation of CRH fibers in the RVLM significantly increased blood pressure (7 ± 1 mmHg). These results indicate that CUS can induce CRH production in a large population of PVN preautonomic neurons that do not produce CRH in basal condition. This population of plastic RVLM‐projecting CRH neurons in the PVN may contribute to CUS induced blood pressure hyperreactivity.

Support or Funding Information

This work was supported by American Heart Association Postdoctoral Fellowship 18POST34050013 and National Heart, Lung, and Blood Institute PO1 Grant HL‐088052.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Low doses of Intrarenal Bradykinin induce a Monophasic Sympathoinhibitory Response
Authors: Hindermann Martin , Ditting Tilmann , Rodionova Kristina , Loosen Sonja , Ott Christian , Schmieder Roland , Amann Kerstin , Veelken Roland
Published: June 2021

Introduction

As recently reported intrarenally administered bradykinin induced a biphasic renal sympathetic nerve response (RSNA) with increases and decreases of RSNA but not uniform sympathoexcitation as previously assumed. The high doses necessary to evoke this biphasic response to bradykinin did not suggest a major physiological role of this observation in renal salt and water handling. Hence, we wanted to test the hypothesis that significantly lower doses of intrarenal bradykinin as previously used will induce a merely monophasic renal sympathoinhibition.

Material and Methods

Groups of anesthetized SD rats (n=6–12) were equipped with femoral catheters (blood pressure (BP) & heart rate (HR) recording, drug application), a renal arterial catheter for one time intrarenal administration (IRA) of Bradykinin (BK 5 nM, 5 μl) or Capsaicin (CAP 1 nM, 10 μl) and a bipolar electrode for RSNA recordings; eventually an intravenous (iv) bolus of the NK1‐receptor blocker RP67580 (10*10−3M, 15 μl) was given

Results

IRA Bradykinin and IRA CAP decreased RSNA from baseline 3.8±1.2 μV*sec to 1.3±0.7 μV*sec (5 μl, 5 nM BK, p<0.05) and 4.0±0.6 μV*sec to 1.6±0.4 μV*sec (10 μl, 1 nM CAP, p<0.01). After reaching the lowest point of RSNA activity at 95 min, both groups showed a slight re‐increase of RSNA within the following 55 min (from 1.3±0.7 μV*sec up to 2.4±1.3 μV*sec (BK) and from 1.6±0.4 μV*sec up to 1.9±0.4 μV*sec (CAP)). Suppressed RSNA in both groups could be unmasked by systemic (i.v.) administration of the NK1‐blocker (1.4±0.4 μV*sec to 5.0±1.9 μV*sec; p<0.05 (BK); 3.5±0.6 μV*sec to 9.7±1.7 μV*sec; p<0.01 (CAP)).

Conclusion

Bradykinin is able to reduce renal sympathetic nerve activity in doses that are significantly lower than doses inducing a biphasic sympathoexcitatory/‐inhibitory response suggesting a physiological role of bradykinin in renal sympathetic nerve control and hence in neurogenic salt and water handling.

Support or Funding Information

DFG, German Research Foundation Interdisciplinary Center for Clinical Research ‐ University of Erlangen, Erlangen, Germany

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
A Novel Sympathoexcitatory Pathway between The Deep Superior Colliculus and The Rostral Ventromedial Medulla
Authors: Lynch Erin , Turner Anita , Dempsey Bowen , Dampney Roger , Goodchild Ann , McMullan Simon
Published: June 2021

Introduction

The superior colliculus plays key roles in the immediate processing of threatening sensory stimuli and generates rapid behavioural responses that are critical for survival. We have previously found that disinhibition of the deep superior colliculus (dSC) unmasks coordinated respiratory, sympathetic and somatomotor outputs that are independent of processing in higher centres. We postulate that these effects are a result of direct innervation of autonomic and respiratory medullary neurons based on the results of neuroanatomical tracing studies that identified dSC synaptic contacts on spinally projecting neurons within the rostral ventromedial medulla (RVMM). In the present study we investigate the physiological significance of this relay and compare effects of dSC stimulation to activation of dSC‐RVMM terminals.

Methods

We used an AAV vector to express Channelrhodopsin2 (ChR2) in the dSC and chronically implanted a fibre optic cannula into the dSC. After transgene expression electrophysiology experiments were conducted under urethane anesthesia with rats instrumented to record blood pressure, diaphragmatic EMG and splanchnic sympathetic nerve activity (SNA). One subset of experiments then positioned the optrode in the RVMM to deliver light to the terminal projections from the dSC. For the other subset of experiments, extracellular recordings were made of spinally projecting RVMM neurons.

Results

dSC photoactivation evoked an increase in respiratory frequency (35±8.7%, P=<0.01, N=7) and heart rate (11±2.6 bpm, P=<0.01, N=7), and a modest change in blood pressure when compared to controls. Stimulus‐triggered averaging of SNA revealed short‐latency excitatory potentials in most cases. Photoactivation of ChR2‐expressing dSC‐RVMM terminals evoked similar effects on cardiorespiratory outputs as dSC stimulation, with an increase in respiratory frequency (14 ±9.2%, P<0.001, N=6) and blood pressure (13 ±2 mmHg P<0.01, N=7), as well as qualitatively similar sympathoexcitation. In preliminary experiments that recorded extracellular action potentials in bulbospinal RVMM neurons we recorded excitatory effects of dSC stimulation in 6/8 spontaneously active neurons.

Conclusion

We conclude that optogenetic activation of dSC neurons that project to the RVMM can drive cardiorespiratory effects that are consistent with our previous findings and that dSC stimulation evokes excitatory effects in some RVMM bulbospinal neurons. This confirms our hypothesis of the existence of a previously uncharacterized excitatory pathway between the dSC and RVMM. Ongoing experiments will functionally characterize RVMM neurons that receive dSC input.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Breathing Exercises Modulate Heart Rate Variability in Individuals with Chronic Stroke
Authors: Larson Mia , Chantigian Dan , Van de Winckel Ann , Keller‐Ross Manda
Published: June 2021

Background

Autonomic dysfunction is common after stroke and increases risk of mortality. Heart rate variability (HRV) is an index of autonomic function and is lower in individuals after a stroke compared with healthy adults. Slow, paced breathing at six breaths per minute has been shown to increase HRV and may be an important therapeutic strategy to improve autonomic function in people with stroke.

Objective

To determine the influence of a slow, paced breathing intervention on HRV after stroke.

Methods

Nine participants (3 women, 52 ± 13.2 years old) with chronic stroke attended two counterbalanced visits, where they completed a 15 min breathing exercise. The slow, paced, device‐guided breathing exercise (SB) consisted of 6 breaths/min and the device‐guided control breathing exercise (CB) was set at 12 breaths/min. A 3‐lead electrocardiogram (ADInstruments) was used to measure heart rate (HR) and calculate measures of HRV during a ten‐minute, spontaneous breathing rest period before and after the breathing exercise. HRV measures were calculated in the frequency domain and included total power (TP), high frequency percent of total power (HF%) and low frequency percent of total power (LF%).

Results

TP did not change pre to post SB (146±198.6%, p = 0.1), but did increase during CB (101.8±107.6%, p = 0.01). LF% increased more for SB compared with CB (15.1±13.98%, 6.8±7.03%, respectively, p = 0.04). HF% did not change after SB (−7.04±11.73%, p = 0.13) nor after CB (−3.23±7.61%, p = 0.26). Heart rate decreased after CB (4.03±4.31 beats/min, p = 0.03), but did not change after SB (−0.34±6.37 beats/min, p = 0.92).

Conclusion

Unexpectedly, the control breathing (12 breaths/min) resulted in a consistent, moderate increase in HRV and a decrease in heart rate. Although slow breathing (6 breaths/min) had a greater effect on HRV for some individuals, the response was much more variable, suggesting that some individuals with chronic stroke may have a difficult time maintaining the slow pace. Consequently, we found 12 breaths/min to be sufficient and more manageable for some individuals after stroke. Further, controlled device‐based breathing can increase HRV and may serve as a complimentary adjunct therapy to improve autonomic function in people after stroke.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Cerebral Blood Flow Response to a Sympathoexcitatory Stimulus in Postmenopausal Women with a History of Preeclampsia
Authors: Miller Kathleen B , Miller Virginia M , Harvey Ronee E , Ranadive Sushant M , Joyner Michael J , Barnes Jill N
Published: June 2021

A history of preeclampsia (PE) places women at higher risk for future cardiovascular disease and cognitive decline. We have previously shown that postmenopausal women with a history of PE demonstrated a reduced cerebral blood flow response to hypercapnia compared with postmenopausal women with a history of a normotensive pregnancy (NP). These findings suggest cerebrovascular effects of PE 30+ years postpartum. In addition, we have reported that women with a history of PE did not have an exaggerated pressor response to a sympathoexcitatory stimulus. However, it is possible that women with a history of PE may have an altered cerebral blood flow response to a sympathoexcitatory stimulus. In this study, middle cerebral artery blood velocity (MCAv) and beat‐to‐beat mean arterial blood pressure (MAP) were measured during isometric handgrip to fatigue (IHG) at 30% of maximal voluntary contraction followed by post‐exercise ischemia (PEI) for 90 seconds and a two minute recovery period (REC). Participants were postmenopausal women with a history of PE (n = 21; age at study = 58±5 y; years since last pregnancy = 35±3 y) and NP (n = 27; age at study = 58±5 y; years since last pregnancy = 35±3 y). Baseline MAP and MAP responses to IHG and PEI were similar between groups (p>0.05). PE women demonstrated lower MCAv at baseline (NP: 72±3 cm/s vs. PE: 57±4 cm/s), IHG (NP: 78±4 cm/s vs. PE: 63±4 cm/s), PEI (NP: 74±4 cm/s vs. PE: 60±4 cm/s), and REC (NP: 70±3 cm/s vs. PE: 58±4 cm/s) compared with NP women (p<0.05 for all). MCAv, when calculated as a percent change from baseline, was elevated during IHG in both PE and NP women as expected (p<0.05); however, PE women had an augmented MCAv response during the last third of IHG compared with NP women (NP: 9±2 % vs. PE: 13±2 %; p<0.05). PE women also had a higher MCAv response during REC compared with NP women (NP:−2±2% vs. PE: 3±2%; p<0.05). Importantly, these elevated MCAv responses occurred despite the fact that MAP was not different between groups. In summary, PE women demonstrated a lower basal cerebral blood velocity but an elevated cerebrovascular response to a sympathoexcitatory stimulus compared with NP women. This suggests that in postmenopausal women, there is an effect of pregnancy history on the sensitivity of the cerebral blood flow response to a sympathoexcitatory stimulus.

Support or Funding Information

NIH: AG 044170. This investigation was supported by the National Institutes of Health, under Ruth L. Kirschstein National Research Service Award T32 HL 007936 from the National Heart Lung and Blood Institute to the University of Wisconsin‐Madison Cardiovascular Research Center.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Muscle pump‐induced inhibition of sympathetic vasomotor outflow during leg cycling is blunted by high‐intensity muscle metaboreflex activation
Authors: Katayama Keisho , Barbosa Thales C. , Kaur Jasdeep , Young Benjamin E. , Shigehiko Ogoh , Fadel Paul J.
Published: June 2021

Muscle sympathetic nerve activity (MSNA) decreases during leg cycling at light or mild intensity (below 40%VO2max), and this is attributed to loading of the cardiopulmonary baroreceptors via muscle pump‐induced increases in venous return. In contrast, MSNA increases during leg cycling above moderate intensity, suggesting that the sympathoinhibitory effect of the cardiopulmonary baroreflex could be overridden by a powerful sympathoexcitatory drive. Skeletal muscle metaboreflex is known to elicit robust sympathoexcitation. Therefore, we tested the hypothesis that high‐intensity muscle metaboreflex activation attenuates muscle pumpinduced cardiopulmonary baroreflex‐mediated inhibition of MSNA during leg cycling. In five healthy young men (25 ± 2 yrs, mean ± SE), MSNA (radial nerve) and thoracic impedance were recorded. Subjects performed cycling at 30 watts with and without muscle metaboreflex activation. Graded isolation of the muscle metaboreflex was achieved by post‐exercise ischemia (PEI) following low and high intensity isometric handgrip at 20% (PEI‐L) and 40% (PEI‐H) maximum voluntary contraction, before the onset of leg cycling. Resting MSNA burst frequency was 23.4 ± 0.4 burst·min−1 and MSNA total activity was 1503 ± 20 AU·min−1. As expected, leg cycling alone significantly decreased MSNA burst frequency and total activity (Δtotal activity: −43.0 ± 4.9%, P < 0.05). When leg cycling was performed during muscle metaboreflex activation with PEI, MSNA burst frequency and total activity decreased significantly during PEI‐L (Δtotal activity: −32.8 ± 7.1%, P < 0.05), but not during PEI‐H (Δtotal activity: +1.6 ± 7.1%, P = 0.89). Thoracic admittance, which was calculated from thoracic impedance as an index of central blood volume, was not changed during handgrip or PEI, whereas leg cycling evoked similar increases in thoracic admittance during all conditions (ΔLeg cycling alone: +3.5 ± 1.0, ΔPEI‐L: +3.6 ± 1.0, ΔPEI‐H: +3.6 ± 0.8 S·10−4, one‐way repeated measures ANOVA: F = 0.02, P = 0.98). These preliminary findings suggest that high intensity muscle metaboreflex activation attenuates muscle pump‐induced cardiopulmonary baroreflex inhibition of sympathetic vasomotor outflow during dynamic leg cycling.

Support or Funding Information

JSPS KAKENHI Grant Number JP16KK0201, UTA College of Nursing and Health Innovation

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
The intercalated cell MR directly regulates pendrin abundance, subcellular distribution and function
Authors: Pham Truyen Derek , Verlander Jill W , Wang Yanhua , Lazo‐Fernandez Yoskaly , Chen Chao , Wall Susan M
Published: June 2021

Aldosterone increases pendrin‐dependent Cl− absorption and HCO3− secretion and activates the intercalated cell (IC) mineralocorticoid receptor (MR) in type B (ICs). Whether the IC MR modulates CCD function and whether aldosterone acts directly on the IC MR in vivo to regulate pendrin in vivo remains to be determined. The purpose of this study was to determine the effect of the IC MR on pendrin abundance, subcellular distribution and function in vivo and to determine if this was from a direct effect of the IC MR. To answer these questions, we developed mice in which MR gene ablation occurred specifically within ICs and measured Cl− absorption in cortical collecting ducts perfused in vitro, pendrin total protein abundance and pendrin subcellular distribution in these IC MR null mice and their wild type littermates. Cl− absorption was ~50% lower in cortical collecting ducts from aldosterone‐treated IC MR null relative to wild type mice. Moreover, following dietary NaCl restriction or an aldosterone infusion, pendrin total protein abundance and pendrin's relative abundance in the apical plasma membrane region were lower in the IC MR null than in wild type littermates. We asked if IC MR gene ablation modulates pendrin abundance and function through a direct effect of the MR within ICs. Since IC MR null mice express both MR positive and negative ICs, we compared pendrin label intensity and pendrin subcellular distribution in MR positive and negative ICs from the same IC MR KO mouse. We observed that pendrin label intensity and pendrin's relative abundance in the apical plasma membrane region were much lower in MR negative than MR positive ICs that were taken from aldosterone‐treated or NaCl‐restricted IC MR null mice. In conclusion, IC MR gene ablation reduces Cl− absorption in the CCD. Furthermore, IC MR gene ablation reduces pendrin total protein abundance and pendrin abundance in the region of the apical plasma membrane through a direct effect of the MR within ICs.

Support or Funding Information

Susan M. Wall, M.D.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Palm Domain Hydrophobic Residues and ENaC Gating
Authors: Wang Xueqi , Chen Jiingxin , Kleyman Thomas R. , Sheng Shaohu
Published: June 2021

Epithelial Na+ channel (ENaC) gating is allosterically modulated by both its extracellular and intracellular domains. Among several regions that have been implicated in regulating ENaC gating are the extracellular palm domains where three subunits interface. We previously identified a palm domain hydrophobic residue (γL511) as an important determinant of ENaC gating. Interestingly, its homologous residue in chicken ASIC1a (L414), located at the β11 and β12 linker, reportedly adapts flipped conformations in the rest (or open) and desensitized states. In this study, we investigated the roles of γL511 homologous resides in the α and β subunits (αL531 and βF502) in regulating ENaC gating, by mutating these residues to a hydrophilic glutamine and examining channel properties in Xenopus oocytes. ENaCs both transport Na+ and are inhibited by extracellular Na+, a response referred to as Na+ self‐inhibition that reflects a reduction in channel open probability and is often used as a surrogate for ENaC gating. Similar to γL511Q, both αL531Q and βF502Q significantly reduced the magnitude and speed of the Na+ self‐inhibition response (p<0.01 vs WT, n=6). Oocytes expressing αL531Qbg ENaCs also showed significantly greater amiloride‐sensitive currents (relative‐to‐WT currents: 1.9 ± 0.2, Mean ± SE, p<0.001, n=32). However, βF502Q expressing oocytes showed the channel currents that were similar to wild type (relative‐to‐WT currents: 0.9 ± 0.1, p>0.05, n=22). The changes caused by both αL531Q and βF502Q on the channel activity and Na+ self‐inhibition response appear to be less dramatic than the previously reported γL511Q. Our results suggest that the highly conserved palm domain hydrophobic residues in three subunits have similar roles in regulating ENaC gating.

Support or Funding Information

DK079307 DK051391

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Paracrine Communication Links Sodium Chloride Cotransporter Activity in the Distal Convoluted Tubule to Remodeling of the Aldosterone‐Sensitive Distal Nephron
Authors: Zapf Ava , Grimm PR , Delpire Eric , Welling Paul A
Published: June 2021

We recently discovered that cell‐specific expression of constitutively active SPAK (CA‐SPAK) in the early distal tubule is sufficient to drive a remodeling process of the entire distal nephron that includes structural dystrophy of the aldosterone‐sensitive distal nephron and inhibition of ROMK and ENaC (Grimm et al, JASN ’17). Here, we tested the hypothesis that SPAK‐activation of NCC stimulates the release of paracrine or autocrine factors that cause the dystrophic response in the CNT.

We found increased levels of PGE2 in kidney cortical homogenates of CA‐SPAK mice compared to control mice. Comparable elevation of PGE2 was also observed when NCC was physiologically activated by feeding wild‐type mice (WT) a low potassium diet (LKD). Administration of the NCC‐blocking diuretic, hydrochlorothiazide, to either CA‐SPAK or WT mice on LKD, restored PGE2 to control levels, but had no effect on PGE2 of WT or SPAK KO mice. Thus, PGE2 is elevated in response to super activation of NCC.

PGE2 synthesis is mediated by prostaglandin E synthase isoforms. As revealed in western blot analysis, the microsomal prostaglandin E synthase‐1(mPGES1) was elevated in the kidney cortex of CA‐SPAK mice and WT mice on LKD compared to control mice. Quantitative microscopy of mPGES1 surprisingly revealed the augmentation of mPGES1 was confined to the CNT and CCD of CA‐SPAK and LKD treated mice. No changes in mPGES1 were observed in the DCT1.

In conclusion, these studies identify PGE2 as a potential remodeling autocrine that is released from the CNT and CCD in response to activation of NCC. We speculate that decreased sodium delivery to the CNT and CCD when NCC is activated drives the response, similar to the mechanism by which PGE2 synthesis is stimulated in the macula densa with changes in sodium reabsorption in upstream tubule segments. The autocrine pathway provides a novel means to communicate potassium sensing in the DCT to potassium secretion in the ASDN.

Support or Funding Information

Supported by the NIDDK and Fondation Leducq

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Inhibition of basolateral ClC‐K2 chloride channels in collecting duct intercalated cells by adenosine
Authors: Zaika Oleg , Tomilin Viktor , Pochynyuk Oleh
Published: June 2021

Adenosine plays an important role in various aspects of kidney physiology, including vasoconstriction of afferent arterioles, modulation of renin secretion and tubule‐glomerular feedback. In addition, adenosine regulates water‐electrolyte transport in the renal tubule, while specific targets are less defined. The collecting duct has the highest expression of adenosine receptors, particularly A1R. Interstitial adenosine levels are greatly increased up to a micromolar range in response to dietary salt loading, favoring activation of its receptors and respective end‐effectors on the basolateral side. The collecting duct is composed of electrically uncoupled principal and intercalated cells having different morphology and physiological roles. We previously showed that the basolateral membrane of principal cells has primarily potassium conductance mediated by Kir4.1/5.1 (KCNJ10/16) channels to mediate K+ recycling and to set up a favorable driving force for Na+/K+ exchange. Intercalated cells lack electrogenic K+ flux but express chloride ClC‐K2 channel mediating transcellular Cl− reabsorption in the collecting duct. Using patch clamp electrophysiology in freshly isolated mouse collecting ducts, we found that acute application of adenosine reversely inhibits ClC‐K2 open probability from 0.31±0.04 to 0.17±0.06 and to 0.10±0.05 for 1 μM and 10 μM, respectively. Inhibitory effect of adenosine was abolished in the presence of A1R blocker, DPCPX (10 μM) suggesting the central role of these receptors in mediating adenosine actions. Consistently, application of A1R agonist N6‐cyclohexyladenosine (CHA, 1 μM) recapitulated the inhibitory action of adenosine on ClC‐K2 open probability. In contrast, adenosine (10 μM) had no measureable effect on Kir4.1/5.1 channel activity in principal cells of the collecting duct. Overall, we demonstrate that adenosine selectively inhibits ClC‐K2 activity in intercalated cells by targeting A1R receptors. We propose that inhibition of transcellular Cl− reabsorption in the collecting duct by adenosine would aid to augment NaCl excretion during high salt intake.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.