bluuebunny/crossref_metadata_2025 huggingface dataset:
Pufferfish Slc4a11 functions as a borate channel for borate secretion
Authors: Kato Akira , Kimura Yuuri , Kurita Yukihiro , Chang Min‐Hwang , Kasai Koji , Fujiwara Toru , Hirata Taku , Doi Hiroyuki , Hirose Shigehisa , Romero Michael F.
Published: June 2021

Boron is a vital micronutrient and is toxic at high concentrations, however, little is known about whole‐body boric acid homeostasis in animals. Slc4a11 was reported to function as a Na+‐coupled borate transporter in mammals, presumably for borate absorption. However, seawater (SW) contains 0.4 mM boric acid, and the bladder urine of a euryhaline pufferfish mefugu (Takifugu obscures) in SW contains ~20 mM boric acid. Therefore, mefugu kidney is a good model to study a borate efflux system. In the mefugu kidney, a paralog of Slc4a11 (Slc4a11A) was markedly induced after transfer to SW and localized to the apical membrane of renal tubules. When Xenopus oocytes expressing Slc4a11A were voltage‐clamped at a holding potential of −60 mV and exposed medium containing borate, intracellular pH was increased and an outward current (anion influx) was observed. The borate current was not altered when Na+ was replaced with other cations such as choline and Li+, but was eliminated when Na+ was replaced with borate chelating agent NMDG. The borate (boron) influx was confirmed by elemental analysis of the oocytes. These results indicate that Slc4a11A is a Na+‐independent B(OH) 4− channel which is suitable for borate secretion, and clarify at first the mechanism of the borate efflux system in animal that prevent marine fishes from the toxic effects of borate.
Real‐time electrochemical detection of endogenous substance release in freshly isolated organs
Authors: Palygin Oleg , Levchenko Vladislav , Ilatovskaya Daria V , Pavlov Tengis S , Ryan Robert P , Cowley Allen W , Staruschenko Alexander
Published: June 2021

We established a novel approach to measure acute changes in endogenous substances in the freshly isolated kidney. This method is based on enzymatic microelectrode biosensors and utilizes amperometry technique. In addition to quantitation of basal levels this technique allows real‐time detection of variety of substances, reflecting changes and fluctuations in their concentrations in freshly isolated organs during vehicle or drug perfusion. Basal levels of H2O2 in the kidney cortex and medulla of Sprague Dawley (SD) rats were measured by biosensors and conventional interstitial fluid collection method. Real‐time changes in ATP and H2O2 concentrations in response to angiotensin II (Ang II) were determined. Freshly isolated kidneys of SD and Dahl salt‐sensitive (SS) rats were perfused with 1 μM Ang II under constant laminar flow. This produced a rapid simultaneous release of both ATP and H2O2 in the kidney cortex area with a strong enhancement in SS rats, especially when animals were fed a high‐salt diet. Biosensors specificity revealed by kidney perfusion with AT1 receptor antagonist losartan and decomposition of H2O2 with catalase enzyme. Established method represents a unique powerful approach for the real‐time monitoring of the endogenous substance level in organs or tissue and could be used for ex vivo and in vivo measurements during normal or pathological conditions. This research was supported by NHLBI.
Anatomically unbiased analysis of renal BOLD magnetic resonance images reveals disruption of corticomedullary gradient during chronic angiotensin II infusion
Authors: Menzies Robert Ian , Zammit‐Mangion Andrew , Hollis Lyam M , Lennen Ross J , Jansen Maurits A , Webb David J , Mullins John J , Dear James W , Sanguinetti Guido , Bailey Matthew A
Published: June 2021

Oxygenation defects may contribute to renal disease progression but are difficult to define in vivo without invasive methods. BOLD MRI provides a non‐invasive alternative but the signal is physiologically complex and data analysis often rely on region of interest selection, excluding biological information.

We present an analysis pipeline that compartmentalizes voxels into distinct clusters. In control rats, low R2* clustering was located predominantly within the cortex and higher R2* clustering within the medulla, consistent with a cortico‐medullary oxygen gradient. An intravenous bolus of acetylcholine caused a transient reduction of the R2* signal in both clustered segments. This was nitric oxide dependent and temporally distinct from hemodynamic effects. Rats were then chronically infused with angiotensin II and rescanned three days later. Clustering demonstrated a disruption of the cortico‐medullary gradient, producing less distinctly segmented mean R2* clusters. The acetylcholine‐induced attenuation of the R2* signal was abolished by chronic angiotensin II infusion, indicative of reduced nitric oxide bioavailability

Our more inclusive analysis pipeline may be more robust in defining deficits in renal oxygenation than the absolute magnitude of R2* in small, manually selected and averaged regions of interest.
Angiotensin II stimulates basolateral 10 pS Cl channels, a main type of Cl channels in the thick ascending limb (TAL)
Authors: Wang Lijun , Gu Ruimin , Wu Peng , Wang Mingxiao , Luan Haiyan , Fan Lili , Wang Wen‐Hui
Published: June 2021

Cl channel in the basolateral membrane plays a key role for Cl absorption in the TAL. The aim of the present study is to test whether AngII increases Cl absorption in the TAL by stimulated the basolateral 10 pS Cl channel in the TAL. The patch clamp experiments were performed in the TAL to record Cl channels in cell‐attached mode. AngII (10–100 nM) stimulated the 10 pS Cl channel, an effect was blocked by losartan (AT1R antagonist) but not by PD123319 (AT2R antagonist). Inhibition of phopsholipase C (PLC) or PKC also abolished the stimulatory effect of AngII on Cl channels. Moreover, stimulation of PKC with phorbol 12‐ myristate 13‐acetate (PMA) mimicked the effect of AngII and increased Cl channel activity. However, the stimulatory effect of AngII on Cl channels was absent in the TAL pretreated with DPI, an inhibitor of NADPH‐dependent oxidase (NOX) while DPI had no significant effect on Cl channels. Moreover, treatment of the TAL with DPI also blocked the effect of PMA on the Cl channels. Western blot demonstrated that incubation of isolated TAL with AngII increased phosphorylation of P47phox at Ser304, suggesting that AngII stimulates the basolateral Cl channels by increasing NOX‐dependent superoxide generation. This notion was also supported by the observation that H2O2 significantly increased 10 pS Cl channel activity in the TAL. We conclude that stimulation of AT1R increased the basolateral Cl channels by activating PKC‐dependent NOX pathway. The stimulatory effect of AngII on the basolateral Cl channel may contribute to AngII‐induced increase in NaCl reabsorption in the TAL.
Angiotensin II rapidly stimulates the short‐circuit current in opossum kidney cells expressing rat Na,K‐ATPase
Authors: Yingst Douglas R. , Hou Xia , Li Quanwen , Mattingly Raymond R. , Sun Fei
Published: June 2021

The binding of angiotensin II (Ang II) to the angiotensin receptor (AT1) stimulates Na,K‐ATPase activity that drives sodium reabsorption in the proximal tubule, which reabsorbs 2/3 of the filtered sodium. To determine how quickly Ang II can stimulate Na,K‐ATPase in the proximal tubule, the α‐1 subunit of rat kidney Na,K‐ATPase and the AT1a receptor were stably co‐expressed in opossum kidney cells. Cells were grown on permeable supports and mounted in an Ussing Chamber containing Ringer's solution. The addition of 10 μM Ang II to the basolateral side increased the short‐circuit current (Isc), which is proportional to net sodium transport, in ≤ ~10 sec. Isc peaked at ~20 sec and declined to a plateau above baseline for over 3 min. The subsequent addition of 200 μM nystatin to increase the sodium permeability of the apical membrane, and thereby the concentration of intracellular sodium, increased Isc to the level of the Ang II‐dependent peak. Subsequently adding 600 μM ouabain, which inhibits Na,K‐ATPase, reduced Isc to its original baseline value. Thus, Ang II significantly stimulated Na,K‐ATPase activity and transcellular sodium transport in 10 seconds or less. We hypothesize that such rapid Ang II‐dependent stimulation of sodium reabsorption in the proximal tubule is part of a mechanism to help the body to recover from a sudden drop in blood pressure. Supported in part by HL096800 to F.S.
Acute and chronic adaptation of renal phosphate transporters to dietary phosphate intake in mice demonstrates importance of NaPi‐IIa and upregulation despite high FGF23 levels
Authors: Bourgeois Soline , Weston‐Capuano Paola , Stange Gerti , Murer Heini , Biber Juerg , Wagner Carsten A.
Published: June 2021

Renal inorganic phosphate (Pi) reabsorption is mediated by the sodium‐dependent phosphate transporters, NaPi‐IIa, NaPi‐IIc, and Pit2 in the brush border membrane of proximal tubule (BBM). Dietary Pi intake regulates these transporters, partly via hormones such as PTH and FGF23. We investigated the time course of adaption in WT mice and the relative contribution of NaPi‐IIa in the adaptive response in NaPiIIa KO mice. Mice received 5 days a high Pi (HPD) or low Pi (LPD) diet. On day 5, some mice were switched for 4 hrs to LPD or HPD. Serum Pi was similar under chronic diets in WT and KO, but KO mice reduced their serum Pi when acutely switched to LPD. Urinary Pi excretion was similar in WT and KO mice under HPD, but under LPD KO mice exhibited a constant urinary Pi loss compensated by a higher intestinal Pi absorption. During the acute HPD‐to‐LPD switch, KO mice exhibited a delayed decrease in urinary Pi excretion. In WT mice PTH and NaPi‐IIa plasma membrane expression were acutely regulated by Pi diet, whereas c‐term FGF23 levels were still high after 8 hrs HPD‐to‐LPD switch. In BBM vesicles, Pi transport activity and plasma membrane NaPi‐IIa but not NaPi‐IIc or Pit2 abundance acutely adapted to diets in WT mice. In KO, BBM vesicles transport activity was lower under all conditions and did not adapt. Thus, only NaPi‐IIa mediates the fast adaptation to Pi intake and is upregulated during the adaptation to low Pi diet despite high FGF23 levels.
Impaired airway epithelial barrier function in cystic fibrosis related diabetes
Authors: Koval Michael , Molina Samuel A , Hunt W Randy , Hanson Jason M , McCarty Nael A
Published: June 2021

Cystic Fibrosis (CF)‐Related Diabetes (CFRD) impacts ~50% of adult CF patients. CFRD is associated with abnormally high airway glucose and an accelerated decline in pulmonary function, indicating a failure in the airway epithelial barrier to regulate glucose permeability. Thus, we examined tight junctions of airway cells expressing wild type CF Transmembrane Conductance Regulator (CFTR) to cells expressing mutant ΔF508‐CFTR. Increasing basolateral glucose from 100 to 450 mg/dL compromised tight junctions of ΔF508‐CFTR cells by ~25% vs. control cells which were significantly less affected. Impaired barrier function was associated with decreased expression of claudin‐4 and displacement of junctional ZO‐1. Using a dual‐luciferase‐ based ER stress reporter assay, cells expressing ΔF508‐ CFTR exhibited higher levels of ER stress than control cells. ER stress was further induced upon exposure to high glucose medium. Thus, ER stress amplified by expression of ΔF508‐CFTR impairs the airway glucose barrier and may predispose the lungs of CF patients to the detrimental consequences of systemic diabetes. Support: Emory+Children's Center for CF Research, Emory URC, CFF, NIH‐NIAAA, NIH‐NHLBI.
Hydrogen peroxide modulates TRPV4‐mediated Ca2+ entry in human coronary artery endothelial cells
Authors: Zheng Xiaodong , Nishijima Yoshinori , Zhang David X.
Published: June 2021

Endothelial TRPV4 has been implicated as an important Ca2+ entry channel contributing to shear‐ and/or agonist‐induced vasodilation in various species and vascular beds, including human coronary arterioles. However, the mechanism responsible for extracellular stimulus‐induced TRPV4 activation is not well understood. Here we investigated the role of hydrogen peroxide (H2O2), an endogenous vasoactive factor produced in human coronary artery endothelial cells (HCAECs), in regulating TRPV4‐mediated Ca2+ entry. In both native and TRPV4‐overexpressing HCAECs loaded with fura‐2, low concentrations of H2O2 (10–100 μM) did not affect intracellular Ca2+ concentration ([Ca2+]i), whereas a high concentration of H2O2 (1 mM) induced a transient increase in [Ca2+]i (372 ± 98 nM of transfected cells). At the subthreshold concentration (10 μM), H2O2 enhanced TRPV4 agonist 4α‐PDD‐evoked [Ca2+]i increase (3896 ± 520 nM vs. 2279 ± 339 nM of control) in transfected HCAECs. In addition, H2O2 (10 μM) augmented TRPV4‐mediated Ca2+ response to the lipid mediator arachidonic acid (1696 ± 201 nM vs. 597 ± 195 nM of control). In conclusion, H2O2 at physiological concentrations does not directly activate endothelial TRPV4, but rather sensitizes TRPV4 to channel agonists. The molecular mechanism involved in this H2O2‐ mediated TRPV4 sensitization remains to be determined.
Deletion of myostatin improves vascular function in obese mice
Authors: Qiu Shuiqing , Mintz James , Nernpermpisooth Nitirut , Salet Christina , Fulton David J , Stepp David W
Published: June 2021

Exercise improves endothelial function in obese patients. Salutatory effects of exercise include increases in muscle size and quality, reduction in fat mass and alterations of the plasma milieu. The relationships between these changes and improvements in vascular function are poorly defined.

Hypothesis

Increasing muscle mass by deletion of muscle growth negative regulator myostatin, improves vascular function in mesenteric arteries from obese db/db mice. Myostatin deletion increases muscle mass in both lean (gastrocnemius 57.93%, gluteus maximus 60.95%, Triceps 57.64%) and obese mice (gastrocnemius 79.64%, gluteus maximus 112.32%, Triceps 103.61%). Myostatin deletion increases muscle fiber size in lean and obese mice (p< 0.05) but has no significant effects on adipocyte size in obese mice. Fasting glucose, HbA1c and glucose tolerance are improved in obese myostatin null mice. Obese mice demonstrate superoxide‐mediated impairment of ACh‐induced vasodilation compared to lean mice. Deletion of myostatin in obese mice improved ACh‐induced vasodilation in mesenteric arteries without effects in lean mice. This improvement was blunted by L‐NAME. PGI2 and EDHF mediated vasodilation were preserved in obese mice, and unaffected by myostatin deletion. Taken together, these data suggested that increasing muscle mass by deletion of myostatin improves NO not PGI2 or EDHF mediated vasodilation in obese mice.
Adaptive cytoprotective responses of motor neuron cells to reactive oxygen species generation by muscle cells, in co‐culture
Authors: Scullion Siobhán Mary Josephine , Vasilaki Aphrodite , McArdle Anne , Jackson Malcolm J
Published: June 2021

Hydrogen peroxide (H2O2) is produced by contracting skeletal muscle although the effect of this on neuronal cells is unknown. The aims of this study were to determine the effect of increased extracellular H2O2 on adaptive responses in neuronal cells and to determine the effect of electrical stimulation of H‐2kb muscle cells on catalase and superoxide dismutases (SODs) in NSC‐34 myotubes using a neuron‐myotube co‐culture system. NSC‐34 motor neuron cells alone were treated with 5, 20 or 100 μM H2O2 for 15 mins or H‐2kb myotubes co‐cultured with NSC‐34 cells, electrically stimulated using platinum electrodes, for 15 mins (0.5 sec every 5 sec). Expression of catalase and MnSOD was determined by western blotting. Both catalase and MnSOD expression were significantly increased in these cells at 24 h following 15 min exposure to 5, 20 or 100 μM H2O2. Following contractions in co‐culture dishes, catalase and MnSOD content of H‐2kb cells was increased 1.5 fold (p<0.01) and 1.4 fold (p<0.05) respectively and in NSC‐34 cells, catalase and MnSOD content was increased 1.6 fold and 2.6 fold (p<0.05). These findings demonstrated that increased extracellular H2O2 results in increased content of catalase and MnSOD in NSC‐34 cells and that muscle cell release of H2O2 during contractions may result in a similar increase in defence protein content of motor neuron cells. Financial support from the BBSRC is acknowledged.
Protein S‐nitrosylation regulates Ca2+ handling and myofilament Ca2+ sensitivity in beta‐adrenergic signaling
Authors: Irie Tomoya , Sips Patrick , Tokuda Kentaro , Ichinose Fumito
Published: June 2021

Objective

To assess the impact of increased rotein denitrosylation on beta‐adrenergic receptor (b‐AR)‐dependent signaling and to evaluate effects of augmented denitrosylation during nitrosative stress in cardiomyocytes.

Methods and Results

We studied cardiomyocytes isolated from wild‐type (WT) and cardiomyocytespecific S‐nitrosoglutathione reductase (GSNOR) overexpressing transgenic (GSNOR‐CMTg) mice. Stimulation of the b‐AR with isoproterenol (ISO)increased sarcomere shortening in both genotypes, while systolic Ca2+ release and sarcoplasmic Ca2+ load only increased in WT. This effect was correlated with increased myofilament Ca2+ sensitivity in ISO‐treated GSNOR‐CMTg cells. Both genotypes showed similar increases in phosphorylation of phospholamban (PLN) and troponin I after b‐AR stimulation while S‐nitrosylation of PLN and troponin C was only increased in WT. Exposure of ISO‐stimulated WT cells to S‐nitrosocysteine severely depressed contractility and systolic Ca2+ release, associated with sarcoplasmic Ca2+ leak and increased ryanodine receptor (RyR2) S‐nitrosylation. GSNOR overexpression prevented RyR2 S‐nitrosylation and preserved contractile function during nitrosative stress.

Conclusion

These results suggest that b‐AR‐induced S‐nitrosylation of excitation‐contraction coupling proteins is required for b‐AR‐induced augmentation of cardiomyocyte function.
The dual‐specificity phosphatase 8 (Dusp8) regulates cardiac hypertrophic response in vitro and in vivo
Authors: Liu Ruijie
Published: June 2021

Background

Phosphorylation and dephosphorylation of mitogen‐activated protein kinases (MAPKs) play important roles in regulating cardiac hypertrophy, remodeling, and heart failure. Dual‐specificity phosphatases (Dusps) directly dephosphorylate and inactivate each of the MAPK terminal kinases (ERK, p38, JNK) within 30 to 60 minutes activation. In this study, we generated mice and mouse embryonic fibroblasts (MEFs) lacking DUSP8 gene and determined the role of Dusp8 in regulating c‐Jun N‐terminal kinases (JNK1/2) signaling in neonatal myocyte culture and mouse heart. We hypothesized that knockout of Dusp8 will lead to increased JNK phosphorylation and decreased cardiac hypertrophy.

Methods

DUSP8 null mice were generated using traditional gene‐targeting approach to delete exon 5 and 6. Phenylephrine was injected at final concentration of 10 mg/kg using 2‐month‐old mice.

Result

s: Adenovirus mediated Dusp8 overexpression in cultured cardiomyocytes resulted in decreased JNK1/2 phosphorylation but not that of p38 and ERK1/2, indicating Dusp8 is specific for JNK dephosphorylation. In contrast, injection of Dusp8 null mice with phenylephrine led to sustained JNK phosphorylation compared to that of WT mice, which was also true for the JNK signaling in Dusp8 null MEF cells. Dusp8 null mice had reduced left ventricular weight/body weight ratio at both baseline and angiotensin II/phenylephrine stimulation condition. At cellular level, we found that overexpression of Dusp8 led to increased nuclear accumulation of NFAT and expression of hypertrophy gene ANF.

Conclusions

The results of the present study indicate that knockout of Dusp8 is cardioprotective through increasing JNK phosphorylation to prevent NAFT signaling induced cardiomyopathy.
Functional implications of Kv7 channels expression in guinea pig urinary bladder smooth muscle contractility
Authors: Afeli Serge A. Y. , Petkov Georgi V.
Published: June 2021

The physiological roles of Kv7 channel family have been well studied in various tissues. However, knowledge about the functional role of Kv7 channels in the urinary bladder smooth muscle (UBSM) is limited. We investigated Kv7 channels molecular expression in guinea pig UBSM and further assessed their functional role using isometric DSM tension recordings and Kv7 channels modulators. RT‐PCR experiments revealed mRNA expression for Kv7.1, Kv7.2, Kv7.3, Kv7.4, and Kv7.5 channel subunits in guinea pig UBSM tissue. Quantitative PCR analysis showed that in UBSM tissue, Kv7 channel subunits relative mRNA expression level was as follows: Kv7.1 ≈ Kv7.2 >; Kv7.3 ≈ Kv7.5 >; Kv7.4. Retigabine, a Kv7 channel activator, and L‐364,373, a selective Kv7.1 channel activator, induced relaxation of spontaneous phasic contraction amplitude, force integral, duration, and frequency as well as the 10 Hz electrical field stimulation (EFS)‐induced contractions amplitude and force of UBSM isolated strips (IC50 = 3 – 6 μM). Linopiridine and XE991, two well‐established Kv7 channel inhibitors, increased the spontaneous phasic contraction amplitude and force as well as the 10 Hz EFS‐induced contractions amplitude and force of UBSM isolated strips (IC50 = 3 – 4 μM). In summary, we revealed the expression and the regulatory role of Kv7 channels in the spontaneous and nerve‐evoked contractions of guinea pig UBSM. Supported by NIH DK084284 to GVP.
ChemR23 Receptor signals through pro‐contractile signaling pathways
Authors: Torres Rosa Yazmin , Watts Stephanie W.
Published: June 2021

(Pro)Chemerin is a peptide secreted from fat and the liver. Chemerin activates ChemR23 to elicit arterial contraction when endothelium is dysfunctional. The signaling of ChemR23 receptor in smooth muscle is unknown. Therefore, we tested the hypothesis that the ChemR23 receptor signals through pro‐contractile pathways using the thoracic aorta of the female rat. Immunohistochemistry supported the presence of ChemR23 receptor in aortic media. Aortic rings lacking endothelium were mounted in tissue baths for measurement of isometric contraction and cumulative responses to Chemerin‐9, an agonist of ChemR23, developed. Tissues were incubated with a signaling inhibitor or vehicle for one hour. Nifedipine reduced the maximum contraction to chemerin‐9 [Veh = 47% ± 12% maximum phenylephrine contraction vs Nif (1 uM) =5% ± 0.7%], as did the Rho Kinase inhibitor Y27632 [Veh = 32% ± 16%, Y27632 (10 uM) =0.8% ± .3%], protein kinase C inhibitor Chelerythrine [Veh = 57%± 12%, Chel (10 uM) =0.1% ± 2%] and p38 MAPK inhibitor SB203580 [Veh= 56% ± 12%, SB203580 (10 uM) =19% ± 5%]. The phosphoinoside‐3 kinase inhibitor LY294002 also reduced contraction [Veh = 57± 16%, LY294002 (10 uM) =26% ± 8]. By contrast, Erk MAPK inhibitor PD09859 did not reduce chemerin‐9 induced contraction. These findings suggest the ChemR23 receptor signals through pro‐contractile signaling pathways. Supported by REPID.
Ion channel mechanisms underlying ethanol‐mediated relaxation in detrusor smooth muscle
Authors: Malysz John , Afeli Serge A.Y. , Petkov Georgi V.
Published: June 2021

Various ion channels are affected by ethanol (EtOH). EtOH evokes relaxation of detrusor smooth muscle (DSM) and increases urinary bladder volume, but the underlying mechanisms remain unknown. In this study, we investigated whether the large conductance Ca2+‐activated K+ (BK) and L‐type voltage‐gated Ca2+ channels, key regulators of DSM function, are the molecular targets for EtOH in guinea pig DSM. Experiments were conducted using patch‐clamp electrophysiology (whole‐cell and single‐channel) and DSM contractions. EtOH at 0.3% (~50 mM) enhanced outward K+ currents at ≥ +20 mV that were attenuated by the selective BK channel blocker paxilline. In excised patches, EtOH (0.1–0.3%) increased single BK channel open probability, number of events and open dwell‐time constants without altering amplitude and mean single channel conductance. In DSM isolated strips, EtOH (0.03–0.1%) inhibited 20 mM KCl‐evoked phasic contraction frequency and at 0.3% also the amplitude and force; these effects were attenuated by paxilline. EtOH at 1% evoked similar relaxation in the presence and absence of paxilline and reduced L‐type voltage‐gated Ca2+ channel currents. In summary, our study reveals involvement of BK and L‐type Ca2+ channels in mediating the effects of EtOH in DSM causing relaxation and increase in bladder volume accommodating alcohol‐induced diuresis. Supported by NIH DK084284 to GVP and USC ASPIRE‐I to JM and GVP.
Defining the physiology of cerebral autoregulation
Authors: Hamner Jason W. , Taylor J. Andrew , Tan Can Ozan
Published: June 2021

Cerebrovascular autoregulation is critical for neurophysiologic health; however, our understanding of the mechanisms is limited. Recent studies show the importance of noradrenergic (NA) and cholinergic (ACh) pathways, though less so for myogenic (MYO). There have been no attempts to consolidate these effectors of cerebral flow into a comprehensive model. Moreover, most work relies on linear approaches that are limited when applied to this inherently nonlinear function. Therefore, we used our projection pursuit method to explore alterations in the relation between pressure (Finapres) and cerebral flow (Doppler) in unblocked (n=48) and blocked (NA, n=11; ACh, n=9; MYO, n=16) states. Differences in autoregulation caused by blockade were used to derive a predictive model for autoregulation in the unblocked state. Projection pursuit provided gain and range of autoregulation as well as the pressure‐flow relations above and below this range. NA blockade markedly linearized the autoregulatory response whereas ACh blockade primarily increased the autoregulatory range by 50%. In contrast to what has been found with linear assessments, MYO blockade reduced the pressure‐flow relations above and below the autoregulatory region. Thus, the predictive model used the contribution of each individual mechanism to achieve an integrated understanding of cerebral autoregulatory control. Support: NIH HL093113.
Impairment of Neurovascular Coupling by Chronic Stress
Authors: Longden Thomas , Dabertrand Fabrice , Hammack Sayamwong E , Nelson Mark T
Published: June 2021

Chronic stress (CS) is a contributory factor in a wide range of diseases. To date, no studies have focused on the effects of chronic stress on neurovascular coupling (NVC). NVC matches neuronal activity with an increase in local blood flow, ensuring that the metabolic demands of the active tissue are satisfied.

We studied NVC in a rat model of CS. Rats were exposed to one of 5 stressors each day for 7 days; we have previously shown that this produces an anxious behavioral phenotype. NVC was impaired by CS: vasodilation of parenchymal arterioles evoked by electrical field stimulation in brain slices was greatly reduced, whereas evoked astrocyte endfoot [Ca2+] was enhanced. In isolated amygdalar arterioles of CS rats, dilation evoked by increasing [K+]o was diminished, suggesting an impairment of inward‐rectifier K+ (Kir) channels. In myocytes from CS rats we observed a decrease in Kir current density. Corticosterone delivery produced a similar NVC phenotype to CS, suggesting that this molecule might mediate NVC impairment. These data suggest that CS causes a decrease in Kir channel number in myocytes of amygdalar parenchymal arterioles, possibly through the actions of corticosterone, rendering the vessel less able to respond to small increases in [K+]o released from astrocyte endfeet and resulting in impaired vasodilation after neuronal activity. This impairment may contribute to CNS disorders with a stress component.
Cardiorespiratory control as a function of wake‐sleep behavior and diet in mice lacking CB1 cannabinoid receptors
Authors: Silvani Alessandro , Bastianini Stefano , Berteotti Chiara , Cohen Gary , Martire Viviana Lo , Mazza Roberta , Quarta Carmelo , Pagotto Uberto , Zoccoli Giovanna
Published: June 2021

Cannabinoid type 1 receptors (CB1R) are widely expressed in the brain. We investigated the effects of chronic CB1R deficiency on cardiorespiratory control as a function of wake‐sleep behavior and diet. Adult male CB1R knock‐out (KO) and wild‐type (WT) control mice (n=9–10 per group) were fed standard diet or HFD for 3 months and instrumented with electrodes and a telemetric blood pressure (BP) transducer. Sleep was recorded with either BP and heart rate (HR) or ventilation (whole‐body plethysmography). We found that KO slept less than WT during the dark (active) period and retained clear‐cut effects of sleep on cardiovascular and respiratory control. HFD increased HR in KO and WT. KO fed HFD had higher values of BP and HR during the dark period and more frequent apneas during non‐rapid‐eye‐movement sleep than WT. These results suggest that CB1R are not necessary for the occurrence of sleep‐dependent cardiovascular and respiratory changes. However, CB1R are necessary for physiological control of wake‐sleep behavior and of respiratory variability during sleep. Finally, chronic lack of endocannabinoid signaling through CB1R may enhance cardiovascular consequences of HFD. This effect is of interest because lack of CB1R signaling has been shown to protect from metabolic derangements induced by HFD by increasing sympathetic activity to brown adipose tissue.
Differential effects of selective versus nonselective alpha adrenergic receptor blockade on cardiovascular (CV) parameters and heart rate variability (HRV) in rats
Authors: Bhatt Siddhartha , Northcott Carrie Annalice , Ringer Laura , Wisialowski Todd , Flynn Declan , Steidl‐Nichols Jill
Published: June 2021

HRV analysis is an important noninvasive tool for investigating autonomic nervous system influence on the heart. The objective of the study was to compare CV profiles of doxazosin (selective á1 blocker) at 10 mg/kg, p.o. and phentolamine (non selective á blocker) at 20 and 100 mg/kg. p.o. in conscious rats using telemetry and subsequent HRV analysis, in addition plasma norepinephrine (NE) was also measured. Doxazosin caused a sustained decrease in blood pressure (BP): maximum change −12 mmHg occurred at 8–16 hours post dose (hpd), induced reflex tachycardia (65 bpm) and increased plasma NE levels by 3 fold. HRV analysis revealed no change in high (HF) and low (LF) frequency power compared to vehicle. Phentolamine decreased BP: maximum changes of −9.2 mmHg (20 mg/kg) and −29.1 mmHg (100 mg/kg) occurred at 0–2 hpd with reflex tachycardia (71 and 111 bpm respectively), reduced body temperature and increased NE 10 fold compared to vehicle. HRV analysis revealed a significantly reduced LF/HF ratio with a significant increase in HF but no change in LF at 0–2 hpd. 22 hpd there were no changes in any parameters measured, suggesting the changes observed were treatment‐mediated. It is hypothesized that the increased parasympathetic tone (HF) with phentolamine could be compensatory in response to reflex tachycardia and immediate NE release. In conclusion, doxazosin and phentolamine show differential effects on HRV and CV parameters
Central Neuropeptide W administration elicits a pressor response due, in part, to mesenteric vasoconstriction in conscious rats
Authors: Gould Amy , Langasek Julie , Willingham Laura A. , Yosten Gina , Samson Willis K , Knuepfer Mark M
Published: June 2021

Neuropeptide W (NPW) is a centrally acting peptide that activates the central melanocortin system and increases arterial pressure in conscious rats. We investigated the mechanism by which NPW evokes hemodynamic responses. Rats were instrumented for recording arterial pressure and for measuring blood flow changes in the abdominal aorta (hindquarters) and the superior mesenteric artery using pulsed Doppler flowmetry. Intracerebroventricular NPW (0.3 nmol and 1 nmol) elicited a pressor response of 9.6±3 and 10.6±4.9 mmHg, respectively, occurring between 90 and 180 seconds after administration. There was a concomitant increase in heart rate (27±10 and 30±12 b/min) and increase in mesenteric vascular resistance (26±8 and 28±10%). Therefore, the increase in arterial pressure due to central administration of NPW is at least partially dependent on increased mesenteric vascular resistance. Hindquarters vascular responses were minimal and highly variable. We conclude that central NPW evokes mesenteric vasoconstriction and tachycardia contributing to a pressor response similar to a classical behavioral stress response. Supported by USPHS DA017371.
Corticotropin Releasing Factor Receptors in the Rostral Ventrolateral Medulla are Required to Initiate but not Sustain Glucose‐Induced Sympathoexcitation
Authors: Bardgett Megan E. , Toney Glenn M.
Published: June 2021

Elevated glucose and insulin increase sympathetic nerve activity (SNA) through a hypothalamic paraventricular nucleus (PVN) melanocortin receptor (MCR) dependent mechanism. Corticotropin releasing factor (CRF) co‐localizes with PVN MC4R and increases arterial blood pressure when delivered into RVLM. Here, the role of RVLM CRF receptors in glucose‐induced increases of SNA was determined. First, RVLM injection of the CRF receptor antagonist astressin (10 pmol, 60 nL) was determined to prevent the pressor effect of local CRF (25 pmol, 11±2 mmHg vs. 2±0 mmHg, P<0.05). Next, effects of CRF receptor blockade on responses to systemic glucose were tested by injecting astressin bilaterally into RVLM 15 min before or 30 min after the start of glucose infusion (150 mg/kg, 10 min, IV). In control rats (n=6) that received bilateral RVLM injections of vehicle (aCSF, 60nL), lumbar (134±9%) and splanchnic (147±6%) SNA were increased (P<0.05) 45 min post glucose infusion. In rats pre‐injected with RVLM astressin (n=4), responses were significantly smaller (lumbar SNA: 109±1%, splanchnic SNA: 120±5%, P<0.05). By contrast, RVLM astressin failed to reverse SNA responses when delivered 30 min post‐infusion (n=5). We conclude that CRF receptor activation in RVLM is required to initiate the increase of SNA by elevated glucose, but ongoing CRF signaling is not required to sustain sympathetic network activity. HL102310 & 088052 (GMT)
Loss of muscle sympathetic nerve activity and blood pressure phase synchronization in postural vasovagal syncope
Authors: Schwartz Christopher E , Lambert Elisabeth , Medow Marvin S , Stewart Julian M
Published: June 2021

Withdrawal of muscle sympathetic nerve activity (MSNA) may not be necessary for the precipitous fall of peripheral arterial resistance and blood pressure (BP) during vasovagal syncope (VVS). We tested the hypothesis that the MSNA‐BP baroreflex entrainment is disrupted prior to VVS regardless if MSNA is withdrawn. We measured phase synchronization during 70° head‐up tilt (HUT) in 14 individuals including 8 VVS and 6 healthy control (C). Heart rate (HR), BP and MSNA were measured during supine baseline and at early, mid, and late stages of HUT. The phase synchronization index (PhSI), the rate of change of MSNA phase with BP phase (neural arc) and that of BP with MSNA (peripheral arc) were computed. During HUT, HR was greater in VVS compared to C (time × group; p=.04). BP did not change during the early stage, but significantly decreased during mid and late stages in VVS only (time × group; p=.001). MSNA was not different between groups during HUT (time × group; p=.700). However, PhSI significantly decreased during mid and late stages in VVS but not C (time × group; p<.001) indicating breakdown of MSNA‐BP phase locking, and therefore interaction, in VVS. In addition, the neural arc was significantly affected more than the peripheral arc prior to syncope. This provides a mechanism for the loss of vasoconstriction and drop in BP independent of late sympathetic nerve activity at the time of vasovagal faint. Supported by NIH (HL‐074873).
Ultrasonic actuation (UA) reduces the brain inflammatory response to neural microelectrode insertion
Authors: Clark Catharine Grace , Chen Po‐Cheng , Gilmour Robert F. , Lal Amit , Schaffer Chris B.
Published: June 2021

Neural electrodes are increasingly being used for the treatment of disease conditions (e.g. control of prosthetic limbs, brain disorders, hypertension). Over time neural electrodes fail, likely due to inflammatory responses, limiting the utility of this treatment approach. We tested the hypothesis that ultrasonic actuation (UA) would reduce the insertion force and brain inflammatory response of neurally implanted electrodes. A custom electrode was fabricated which allows for measurement of force, recording of neural signals and UA (V) of the electrode. Acute insertion in the mouse cortex demonstrated that UA reduced the implantation force in a voltage dependent manner (9.6 ± 0.7 vs 3.8 ± 0.5 mN, 0 vs 30V, p<0.05). UA also resulted in a significant decrease in the area of damaged brain tissue (56246 ± 6889 vs 34141 ± 5613 pixels, 0V vs 30V, p<0.05). Using transgenic mice to selectively examine microglia revealed a reduction in the inflammatory response surrounding the electrode (e.g. 56 ± 6 vs 34 ± 7 # microglia, 0 vs. 30V, p<0.05), such that UA was not different from control mice (56 ± 6 # microglia). These findings demonstrate that UA decreases the insertion force and brain inflammatory response of implanted neural electrodes and represents a novel means to increase the functionality of brain electrodes.

DARPA MTO under the auspices of Dr. J. Judy through the Space and Naval Warfare Systems Center Grant N66001–11‐1–4011.
Central Sympathoinhibition Ameliorates the Reduction of Splenic Regulatory T Cells with Decreased IL‐17 Production in Hypertensive Rats
Authors: Katsuki Masato , Hirooka Yoshitaka , Kishi Takuya , Sunagawa Kenji
Published: June 2021

Background

Regulatory T cells (Tregs) have been reported to prevent organ damages in hypertension. This effect might be caused by anti‐inflammatory process. We examined Treg expression and number focusing on spleen where T cells are abundant. The aim of the present study was to determine whether Tregs is altered in the spleen, and blockade of brain Ang II type 1 receptors (AT1R) for central sympathoinhibition affects splenic Tregs in hypertensive rats.

Methods and Results

Stroke‐prone spontaneously hypertensive rats (SHRSP) with sympathetic hyperactivity were treated with intracerebroventricular infusion of an AT1R blocker (losartan, 1mg/kg/day for 14 days, ARB) or vehicle (VEH). Wister Kyoto rats (WKY) served as control. Splenic Foxp3 (an essential transcription factor of Tregs) mRNA and CD4+CD25+Foxp3+ cells were lower in SHRSP‐VHE than in WKY (Foxp3 mRNA: 0.47±0.03 vs. 1.02±0.03, P<0.05). Splenic Foxp3 mRNA expression levels were upregulated in ARB‐SHRSP (ARB: 0.44±0.04 vs. VEH: 0.31±0.02, P<0.05), while systolic blood pressure and 24‐hour urinary norepinephrine excretion were decreased. IL‐17 mRNA in the spleen was significantly lower in SHRSP‐ARB than in SHRSP‐VEH (ARB: 0.27±0.04 vs. VEH: 0.47±0.03, P<0.01).

Conclusion

Central sympathoinhibition induced by blockade of brain AT1R ameliorates the reduction of splenic Tregs possibly via decreased IL‐17 production in SHRSP.
Effects of Social Isolation on Connexin 43 Expression in the Left Ventricle of Female Prairie Voles
Authors: Firkins Rachel Marie , Dotson Ashley , McNeal Neal , Schultz Rachel , Scotti Melissa L. , Wardwell Joshua , Henry Matthew K. , Moffitt Julia A. , Grippo Angela J.
Published: June 2021

Social stress plays a key role in the relationship between cardiovascular disease (CVD) and depression. Previous studies in animals have shown that social isolation leads to depressive like behaviors and increased incidence of arrhythmias. Alterations in the expression of left ventricular (LV) connexin43 (Cx43) has been identified in the pathogenesis of cardiac arrhythmias in some forms of CVD. Prairie voles, which are socially monogamous rodents, were used to investigate the hypothesis that social isolation leads to increased depressive like behaviors and decreased LV Cx43 expression. Adult prairie voles were paired with or isolated from a sibling for 4 weeks. Then, half of the paired and isolated animals were exposed to 7 days of chronic mild stressors (CMS) while the other half remained undisturbed. The animals were subjected to the forced swim test (FST) as an index of depressive behavior. Following the FST, hearts were collected and LV Cx43 expression was determined by Western blotting. Data indicate a significant increase in depressive behavior during the FST exhibited by the paired+CMS, isolated, and isolated+CMS groups compared to the paired group. The expression of LV Cx43 was significantly lower in all groups as compared to the paired group. We conclude that a reduction in the expression of LV Cx43 may be a mechanism for the increased arrhythmogenesis in animals that exhibit depressive like behaviors. MH77581, HL112350, IOER 3726
Attenuated Heart Rate Recovery is Associated with Sympathetic Hyperactivity in Patients with Metabolic Syndrome and Obstructive Sleep Apnea
Authors: Cepeda Felipe Xerez , Toschi‐Dias Edgar , Godoy Daniel Martinez , Maki‐Nunes Cristiane , Rondon Maria Urbana P.B. , Alves Maria Janieire N.N. , Braga Ana Maria F.W. , Drager Luciano F. , Lorenzi‐Filho Geraldo , Negrao Carlos E. , Trombetta Ivani C.
Published: June 2021

Obstructive sleep apnea (OSA) and metabolic syndrome (MetS) are independently associated with impaired heart rate recovery (HRR) after maximal exercise. We tested the hypotheses that: 1) MetS+OSA would have an additive effect on attenuation of HRR; 2) The sympathetic hyperactivation would be involved on slower HRR. Newly diagnosed MetS (ATP III) patients, sedentary were divided into: MetS+OSA (n=18, 49±2y) and MetS‐OSA (n=21, 44±2y). Normal individuals were also studied (C, n=13, 47±2y). OSA was characterized by apnea‐hypopnea index (AHI) >;15events/h (polysomnography). Muscle sympathetic nerve activity (MSNA, microneurography), heart rate (EKG), low (LF) and high (HF) frequency power bands and sympatho‐vagal balance (LF/HF‐Spectral analysis) were measured. The HRR was calculated (HRpeak minus HHR at 1st, 2nd, 4th min). MetS+OSA had higher MSNA, LF and LF/HF compared with MetS‐OSA and C. MSNA was higher in MetS‐OSA than in C. HF power was lower in MetS+OSA compared with MetS‐OSA and C. HRR at 1st, 2nd min was lower in MetS+OSA compared with C (P<0.01) and lower at 4th min compared with Met‐OSA and C (P<0.01). Further analyses showed an association between MSNA levels and HRR (P<0.01). In conclusion, OSA exacerbates the attenuation of HRR after maximal exercise in patients with MetS. In addition, the attenuation of HRR is associated with sympathetic hyperactivation.(FAPESP#2011/17533–6)
Electrical stimulation of the aortic depressor nerve in conscious state elicits similar hemodynamic responses in heart failure and control rats
Authors: Teixeira‐Pinto Tomás , de Toledo Durand Marina , Lataro Renata Maria , Castania Jaci Airton , Silva Carlos Alberto Aguiar , Fazan Rubens , Salgado Helio Cesar
Published: June 2021

Heart Failure (HF) is characterized by attenuation of the baroreflex. The present study investigated the hypotensive and bradycardic responses produced by electrical stimulation of the aortic depressor nerve (ADN) in HF rats. HF was induced by left coronary artery ligation. Six weeks after ligation (n=11), or sham surgery (control rats; n=12), the animals were anesthetized for femoral artery and vein cannulation and implantation of bipolar electrodes for ADN stimulation. Twenty‐four hours later, electrical stimulation of the ADN was applied during 20s (5, 15, 30, 60 and 90 Hz; 1mA; 2ms duration). Baseline mean arterial pressure (105 ± 3 vs 100 ± 4 mmHg in HF) and heart rate (341±10 vs 324±13 bpm in HF) were similar between HF and control rats. Electrical stimulation of the ADN (5 to 90 Hz) elicited similar hypotensive (14±2 to 49±6 vs 10±3 to 41±3 mmHg in HF rats) and bradycardic (17±6 to 75±15 vs 21±6 to 67±9 bpm in HF rats) responses in both groups. Taken into account that HF is characterized by baroreflex attenuation, these findings indicate a derangement of the baroreceptor endings and/or the electrical stimulation overriding the baroreflex attenuation in HF rats.

Financial Support: FAPESP, CNPq.
Apnea Alters the Pattern of Intermittent Hypoxia‐Induced Long Term Facilitation in Anesthetized Mice
Authors: ElMallah Mai Kamal , Sandhu Milap S , Lee Kun‐Ze , Baekey David M , Fuller David D
Published: June 2021

Intermittent hypoxia (IH) can trigger persistent increases in efferent hypoglossal (XII) motor output (i.e. long‐term facilitation, LTF) in several species, but this response has not been investigated in the mouse. As a prelude to studies in genetic “knockout” mice, our first objective was to determine if the pattern of XII LTF evoked by IH in anesthetized 129SVE mice was similar to that reported in other species. Our second purpose was to test the hypothesis that the expression of XII LTF would be altered if IH was preceded by a period of apnea‐induced XII inactivity. Mice were anesthetized with urethane, tracheotomized and vagotomized, and XII nerve activity was recorded bilaterally. A “time control” group (N=4) showed no changes in efferent XII output over a 90 min recording period with PETCO2 maintained constant at or near 35 mmHg. In another group (N=6), baseline was followed by IH (3 × 1 min hypoxia, 10%O2, separated by 3 min of hyperoxia, 50% O2). These mice showed robust facilitation, but surprisingly this was evident immediately following the final bout of hypoxia. In contrast, mice that experienced a brief (~ 3 min) period of apnea (N=4) before IH showed a gradual increase in XII bursting over the 90 min post‐IH period. These preliminary results indicate that IH triggers robust XII LTF in urethane anesthetized mice, and a brief period of apnea can alter the pattern of LTF expression.

Funding: Parker B. Francis Fellowship (MKE), 2R01HD052682–06A1 (DDF), Craig H. Neilsen Foundation #220521 (MSS)
Retrograde transynaptic tracing of the inspiratory intercostal circuitry following cervical spinal cord injury in rats
Authors: Gonzalez‐Rothi Elisa Janine , Dougherty Brendan J , Baekey David M , Lane Michael A , Reier Paul J , Fuller David D
Published: June 2021

Unilateral cervical (C2) spinal hemisection (C2Hx) causes persistent deficits in ipsilateral (IL) phrenic output but only transiently diminishes IL inspiratory intercostal (IC) EMG activity. Neurophysiological studies indicate an extensive population of propriospinal thoracic interneurons that receive respiratory‐related synaptic inputs. Our working hypothesis is that thoracic interneurons are part of the anatomical substrate for a “crossed‐IC” pathway which can relay inspiratory drive to ipsilateral IC motoneurons following C2Hx. Ongoing experiments are utilizing retrograde transynaptic tracing methods to define the distribution of spinal interneurons which innervate IC motoneurons. Pseudorabies virus (PRV) was injected into the rostral external IC muscles of uninjured and C2Hx male Sprague‐Dawley rats and spinal cord histology was examined 72–96 hrs later. In the uninjured spinal cord, pre‐motor interneurons were located throughout the cervical and thoracic spinal cord in the dorsal, intermediate and ventral gray matter. Preliminary assessment of tissue from C2Hx rats revealed robust interneuronal labeling in the same regions. These data confirm that IC motoneurons are robustly innervated by propriospinal interneurons, and are consistent with our hypothesis that these cells play a role in IC motor recovery after C2Hx.