bluuebunny/crossref_metadata_2025 huggingface dataset:
Ionic mechanism of electrical alternans
Authors: Fox Jeffrey J. , McHarg Jennifer L. , Gilmour Robert F.
Published: March 2015

Although alternans of action potential duration (APD) is a robust feature of the rapidly paced canine ventricle, currently available ionic models of cardiac myocytes do not recreate this phenomenon. To address this problem, we developed a new ionic model using formulations of currents based on previous models and recent experimental data. Compared with existing models, the inward rectifier K+ current ( I K1) was decreased at depolarized potentials, the maximum conductance and rectification of the rapid component of the delayed rectifier K+ current ( I Kr) were increased, and I Kr activation kinetics were slowed. The slow component of the delayed rectifier K+current ( I Ks) was increased in magnitude and activation shifted to less positive voltages, and the L-type Ca2+ current ( I Ca) was modified to produce a smaller, more rapidly inactivating current. Finally, a simplified form of intracellular calcium dynamics was adopted. In this model, APD alternans occurred at cycle lengths = 150–210 ms, with a maximum alternans amplitude of 39 ms. APD alternans was suppressed by decreasing I Ca magnitude or calcium-induced inactivation and by increasing the magnitude of I K1, I Kr, or I Ks. These results establish an ionic basis for APD alternans, which should facilitate the development of pharmacological approaches to eliminating alternans.
Rat carotid artery dilation by PTCA balloon catheter induces neointima formation in presence of IEL rupture
Authors: Indolfi Ciro , Torella Daniele , Coppola Carmela , Stabile Eugenio , Esposito Giovanni , Curcio Antonio , Pisani Alfonso , Cavuto Luigi , Arcucci Oreste , Cireddu Manuela , Troncone Giancarlo , Chiariello Massimo
Published: March 2015

The best animal angioplasty model is the porcine model, which is expensive and not available in all laboratories. The aim of this study was to describe a new rat model of angioplasty. An injury was induced with the use of a standard percutaneous transluminal coronary angioplasty (PTCA) 1.5-mm balloon catheter. The neointimal tissue, arterial dimensions, and the injury index were assessed following angioplasty. Ki-67 expression was detected to evaluate cell turnover after balloon angioplasty. In contrast with the standard Clowes model, a significant neointimal formation was detected only in the presence of ruptured internal elastic lamina (IEL). A positive correlation between the percentage of ruptured IEL and the amount of neointimal tissue was also demonstrated. The percentage of IEL fracture correlates with the proliferation index by anti-Ki-67 immunolabeling 7 and 14 days after the angioplasty. Significant arterial negative remodeling was observed following PTCA balloon dilation. In conclusion, our inexpensive animal model of restenosis after angioplasty may have great relevance toward a better understanding of the mechanisms and toward assessment of new therapeutical strategies for this phenomenon.
Expression of VEGF and angiopoietins-1 and -2 during ischemia-induced coronary angiogenesis
Authors: Matsunaga Toshiro , Warltier David C. , Tessmer John , Weihrauch Dorothee , Simons Michael , Chilian William M.
Published: March 2015

The mechanisms underlying coronary capillary growth in response to ischemia are undefined. We hypothesized that the expression of vascular endothelial growth factor (VEGF) and angiopoietin (Ang)/Tie-2 were involved in capillary growth as an adaptation to ischemia. To test this hypothesis we measured capillary density, and the expressions of VEGF, Ang-1, Ang-2, and the Tie-2 receptor and its phosphorylation state during repetitive episodes of myocardial ischemia in chronically instrumented canines. Repetitive episodes of ischemia were induced by multiple (once/hour; 8/day), brief (2 min) occlusions of the left anterior descending coronary artery for 1, 7, 14, or 21 days. A sham group received the same instrumentation as the experimental groups but not the occlusion protocol. Collateral blood flow (microspheres) progressively increased from 9 ± 3 to 83 ± 10 ml · min-1· 100 g-1on day 21. Capillary density increased at day 7 from 2,378 ± 53 (sham) to 2,962 ± 60/mm2, but it decreased to 2,594 ± 39/mm2at day 21. Both VEGF and Ang-2 expression in myocardial interstitial fluid (Western analyses) peaked at day 3 of the repetitive occlusions but waned thereafter. In contrast the expression of Ang-1 remained relatively constant at all times in the occlusion groups. In shams, the expression of VEGF and Ang-2 was low and constant at all times. Tie-2 phosphorylation myocardial decreased decreased at day 7 but increased at 21 days of occlusions ( P < 0.05). Our results indicate that capillary density was augmented by myocardial ischemia, but after development of collaterals and restoration of flow to the ischemic zone, capillary density returned to control levels. The change in capillary density paralleled with VEGF and Ang-2 expression but was inversely related to Tie-2 phosphorylation. We speculate the coronary angiogenesis is a coordinated event involving the expression of both VEGF and Ang-2 and that therapeutic angiogenic strategies may ultimately require treatment with more than a single factor.
Expression and coassociation of ERG1, KCNQ1, and KCNE1 potassium channel proteins in horse heart
Authors: Finley Melissa R. , Li Yan , Hua Fei , Lillich James , Mitchell Kathy E. , Ganta Suhasini , Gilmour Robert F. , Freeman Lisa C.
Published: March 2015

In dogs and in humans, potassium channels formed by ether-a-go-go-related gene 1 protein ERG1 (KCNH2) and KCNQ1 α-subunits, in association with KCNE β-subunits, play a role in normal repolarization and may contribute to abnormal repolarization associated with long QT syndrome (LQTS). The molecular basis of repolarization in horse heart is unknown, although horses exhibit common cardiac arrhythmias and may receive drugs that induce LQTS. In horse heart, we have used immunoblotting and immunostaining to demonstrate the expression of ERG1, KCNQ1, KCNE1, and KCNE3 proteins and RT-PCR to detect KCNE2 message. Peptide N-glycosidase F-sensitive forms of horse ERG1 (145 kDa) and KCNQ1 (75 kDa) were detected. Both ERG1 and KCNQ1 coimmunoprecipitated with KCNE1. Cardiac action potential duration was prolonged by antagonists of either ERG1 (MK-499, cisapride) or KCNQ1/KCNE1 (chromanol 293B). Patch-clamp analysis confirmed the presence of a slow delayed rectifier current. These data suggest that repolarizing currents in horses are similar to those of other species, and that horses are therefore at risk for acquired LQTS. The data also provide unique evidence for coassociation between ERG1 and KCNE1 in cardiac tissue.
Phasic contractions of the rat portal vein depend on intracellular Ca2+release stimulated by depolarization
Authors: Burt Richard P.
Published: March 2015

The phasic contraction to phenylephrine of the rat isolated portal vein was investigated using functional studies. Phasic contractions to phenylephrine and caffeine could be produced after several minutes in Ca2+-free Krebs solution, which were inhibited by cyclopiazonic acid or ryanodine. The phenylephrine and caffeine contractions were abolished, however, within 10 min in Ca2+-free Krebs solution and by nifedipine. This indicated the Ca2+stores were depleted in the absence of Ca2+influx through voltage-gated channels. The phasic contraction to phenylephrine was also abolished by niflumic acid even in Ca2+-free Krebs solution. This showed that the response depended on intracellular Ca2+release stimulated directly by depolarization, resulting from opening of Ca2+-activated Cl−channels, but did not require Ca2+influx. In support of this, K+-induced phasic contractions were also produced in Ca2+-free Krebs solution. The phenylephrine but not K+-induced phasic contractions in Ca2+-free Krebs solution were inhibited by ryanodine or cyclopiazonic acid. This would be consistent with Ca2+release from more superficial intracellular stores (affected most by these agents), probably by inositol 1,4,5-trisphospate, being required to stimulate the phenylephrine depolarization.
Endotoxemia stimulates skeletal muscle Na+-K+-ATPase and raises blood lactate under aerobic conditions in humans
Authors: Bundgaard Henning , Kjeldsen Keld , Suarez Krabbe Karen , van Hall Gerrit , Simonsen Lene , Qvist Jesper , Hansen Christian Muff , Møller Kirsten , Fonsmark Lise , Madsen Per Lav , Pedersen Bente Klarlund
Published: March 2015

We assessed the hypothesis that the epinephrine surge present during sepsis accelerates aerobic glycolysis and lactate production by increasing activity of skeletal muscle Na+-K+-ATPase. Healthy volunteers received an intravenous bolus of endotoxin or placebo in a randomized order on two different days. Endotoxemia induced a response resembling sepsis. Endotoxemia increased plasma epinephrine to a maximum at t = 2 h of 0.7 ± 0.1 vs. 0.3 ± 0.1 nmol/l ( P < 0.05, n = 6–7). Endotoxemia reduced plasma K+reaching a nadir at t = 5 h of 3.3 ± 0.1 vs. 3.8 ± 0.1 mmol/l ( P < 0.01, n = 6–7), followed by an increase to placebo level at t = 7–8 h. During the declining plasma K+, a relative accumulation of K+was seen reaching a maximum at t = 6 h of 8.7 ± 3.8 mmol/leg ( P < 0.05). Plasma lactate increased to a maximum at t = 1 h of 2.5 ± 0.5 vs. 0.9 ± 0.1 mmol/l ( P < 0.05, n = 8) in association with increased release of lactate from the legs. These changes were not associated with hypoperfusion or hypoxia. During the first 24 h after endotoxin infusion, renal K+excretion was 27 ± 7 mmol, i.e., 58% higher than after placebo. Combination of the well-known stimulatory effect of catecholamines on skeletal muscle Na+-K+-ATPase activity, with the present confirmation of an expected Na+-K+- ATPase-induced decline in plasma K+, suggests that the increased lactate release was due to increased Na+-K+-ATPase activity, supporting our hypothesis. Thus increased lactate levels in acutely and severely ill patients should not be managed only from the point of view that it reflects hypoxia.
Participation of intracellular Ca2+stores in arteriolar conducted responses
Authors: Yashiro Yasuaki , Duling Brian R.
Published: March 2015

We examined the role played by intracellular Ca2+stores in conducted vasomotor responses induced by phenylephrine (PE) in isolated hamster cremasteric arterioles. When applied briefly (∼1 s) to isolated, cannulated arterioles by using pressure-pulse ejection from a micropipette, PE produced a strong local vasoconstriction and a very small biphasic conducted response (a small constriction followed by a dilation) that propagated several hundred micrometers along the vessel length. The conducted vasomotion was associated with a monophasic elevation of the endothelial cell intracellular Ca2+concentration ([Ca2+]i) at the site of stimulation, as measured with the Ca2+indicator fura 2. The Ca2+pump inhibitor thapsigargin was used to limit filling of Ca2+stores in smooth muscle and endothelial cells. Thapsigargin reduced baseline diameter and elicited a strong dilator component at the local site while enhancing both the constrictor and dilator components of the PE-induced conducted response. The enhanced conducted constrictor component induced by thapsigargin was mimicked by extraluminal application of tetraethylammonium or charybdotoxin but not by iberiotoxin, apamin, glibenclamide, barium, or 4-aminopirydine. Thapsigargin increased the estimated basal endothelial cell [Ca2+]iby ∼60 nM and converted the PE-induced change in [Ca2+]ifrom monotonic to biphasic with a late elevation of [Ca2+]iabove baseline that coincided with the increased dilatory component of the conducted response. Luminal application of charybdotoxin plus apamin significantly reduced the dilatory component of the conducted response. These results indicate that intracellular Ca2+stores play a dynamic role in regulating conducted vasomotor responses apparently through modulation of KCachannels in both cell types.
Characterization of myocardium, isolated cardiomyocytes, and blood pressure in WKHA and WKY rats
Authors: Deschepper Christian F. , Picard Sylvie , Thibault Gaétan , Touyz Rhian , Rouleau Jean-Lucien
Published: March 2015

We previously reported that the left ventricular (LV) mass of Wistar-Kyoto (WKY)-derived hyperactive (WKHA) rats was higher than that of WKY rats in the absence of a difference in systolic blood pressure. To extend these earlier observations, we conducted a series of functional and morphological investigations on both strains. Analysis of tissue sections revealed that the surface of ventricular tissue from WKHA rats was higher than that of WKY rats, without any enlargement of the cavity area. Analysis of isolated adult cells showed that cell width (as well as cell volume) of ventricular cardiomyocytes was significantly higher in WKHA than WKY rats. However, LV of WKHA rats contained ∼33% less cardiomyocytes than those from WKY rats. Mean intracellular free calcium concentration of cardiomyocytes was also higher in WKHA than WKY rats. Hemodynamic measurements revealed that the values of the maximum rates of pressure change (dP/d t) were higher in LV from WKHA rats. However, these differences were reduced (−dP/d t) or abolished (+dP/d t) when the values were normalized for both the number and mean cross-sectional area of ventricular cardiomyocytes. Mean levels of systolic and diastolic blood pressure (corresponding to the 24-h average of measurements obtained continuously in conscious unrestrained animals using radiotelemetric implants) were not different between strains. However, circadian rhythm was more evident in WKY rats, because the difference between morning and night values of systolic and diastolic blood pressure was greater (by 3 mmHg) in WKY rats. Altogether, our data validate the use of WKHA rats as models of predominantly concentric LV hypertrophy developing in the absence of increased mean levels of hemodynamic cardiac load and show that the hypertrophy phenotype is more pronounced in isolated cardiomyocytes than at the level of the whole ventricle.
Phosphorylation of eNOS initiates excessive NO production in early phases of portal hypertension
Authors: Iwakiri Yasuko , Tsai Ming-Hung , McCabe Timothy J. , Gratton Jean-Philippe , Fulton David , Groszmann Roberto J. , Sessa William C.
Published: March 2015

Akt, also known as protein kinase B, is a serine/threonine kinase. Akt becomes active when phosphorylated by the activation of receptor tyrosine kinases, G protein-coupled receptors, and mechanical forces such as shear stress. Studies in vitro have shown that Akt can directly phosphorylate endothelial nitric oxide (NO) synthase (eNOS) and activate the enzyme, leading to NO production. The aim of this study was to test the hypothesis that the phosphorylation of eNOS plays a role in the enhanced NO production observed in early portal hypertension. Male Sprague-Dawley rats were subjected to either sham or portal vein ligation (PVL), and mesenteric arterial beds were used for ex vivo perfusion studies. Mesenteric arterial beds from PVL rats had an approximately 60–70% decrease in response to methoxamine (an α1-agonist and vasoconstrictor) compared with the sham group ( P < 0.01). When N G-monomethyl-l-arginine (a NOS inhibitor) was added to the perfusion, the difference in perfusion pressure between the two groups was abolished, suggesting that enhanced NO production in the PVL group blunted the response to the vasoconstrictor. The reduced responsiveness in PVL was not due to changes in eNOS expression but was due to an increase in enzyme-specific activity, suggesting posttranslational modification of eNOS. The phosphorylation of eNOS at Ser1176 was significantly increased by twofold ( P < 0.05) in the PVL group. Furthermore, PVL significantly increased Akt phosphorylation (an active form of Akt) by threefold ( P< 0.05). When vessels were treated with wortmannin (10 nM) to block the phosphatidylinositol-3-OH-kinase/Akt pathway, NO-induced vasodilatation was significantly reduced. These results suggest that the phosphorylation of eNOS by Akt activates the enzyme and may be the first step leading to an initial increase in NO production in portal hypertension.
Increased myofibrillar protein phosphatase-1 activity impairs rat aortic smooth muscle activation after hypoxia
Authors: Teoh Hwee , Zacour Mary , Wener Avraham D. , Gunaratnam Lakshman , Ward Michael E.
Published: March 2015

We hypothesized that increased myofibrillar type 1 protein phosphatase (PP1) catalytic activity contributes to impaired aortic smooth muscle contraction after hypoxia. Our results show that inhibition of PP1 activity with microcystin-LR (50 nmol/l) or okadaic acid (100 nmol/l) increased phenylephrine- and KCl-induced contraction to a greater extent in aortic rings from rats exposed to hypoxia (10% O2) for 48 h than in rings from normoxic animals. PP1 inhibition also restored the level of phosphorylation of the 20-kDa myosin light chain (LC20) during maximal phenylephrine-induced contraction to that observed in the normoxic control group. Myofibrillar PP1 activity was greater in aortas from rats exposed to hypoxia than in normoxic rats ( P < 0.05). Levels of the protein myosin phosphatase-targeting subunit 1 (MYPT1) that mediates myofibrillar localization of PP1 activity were increased in aortas from hypoxic rats (193 ± 28% of the normoxic control value, P < 0.05) and in human aortic smooth muscle cells after hypoxic (1% O2) incubation (182 ± 18% of the normoxic control value, P < 0.05). Aortic levels of myosin light chain kinase were similar in normoxic and hypoxic groups. In conclusion, after hypoxia, increased MYPT1 protein and myofibrillar PP1 activity impair aortic vasoreactivity through enhanced dephosphorylation of LC20.
Hypoxia in the thymus: role of oxygen tension in thymocyte survival
Authors: Hale Laura P. , Braun Rod D. , Gwinn William M. , Greer Paula K. , Dewhirst Mark W.
Published: March 2015

Our previous studies using oxygen microelectrodes showed that the thymus is grossly hypoxic under normal physiological conditions. We now have investigated how oxygen tension affects the thymus at the cellular and molecular level. Adducts of the hypoxia marker drug pimonidazole accumulated in foci within the cortex and medulla and at the corticomedullary junction, consistent with the presence of widespread cellular hypoxia in the normal thymus. Hypoxia-associated pimonidazole accumulation was decreased but not abrogated by oxygen administration. Genes previously reported to be induced by hypoxia were expressed at baseline levels in the normal thymus, indicating that physiological adaptation to hypoxia occurred. Despite changes in thymus size and cellularity, thymic Po 2 did not change with age. Combined assays for hypoxia and cell death showed that hypoxia achieved using either hypoxic gas mixtures or high-density culture in normoxia decreased spontaneous thymocyte apoptosis in vitro. Taken together, these data suggest that regulatory mechanisms exist to maintain thymic cellular hypoxia in vivo and that oxygen tension may regulate thymocyte survival both in vitro and in vivo.
Is there an A-type K+ current in guinea pig ventricular myocytes?
Authors: Findlay Ian
Published: March 2015

A unique transient outward K+ current ( I to) has been described to result from the removal of extracellular Ca2+ from ventricular myocytes of the guinea pig (15). This study addressed the question of whether this current represented K+-selective I to or the efflux of K+ via L-type Ca2+ channels. This outward current was inhibited by Cd2+, Ni2+, Co2+, and La3+ as well as by nifedipine. All of these compounds were equally effective inhibitors of the L-type Ca2+ current. The current was not inhibited by 4-aminopyridine. Apparent inhibition of the outward current by extracellular Ca2+ was shown to result from the displacement of the reversal potential of cation flux through L-type Ca2+ channels. The current was found not to be K+ selective but also permeant to Cs+. The voltage dependence of inactivation of the outward current was identical to that of the L-type Ca2+ current. It is concluded that extracellular Ca2+ does not mask an A-type K+current in guinea pig ventricular myocytes.
Serotonin 5-HT3 receptors on mechanosensitive neurons with cardiac afferents
Authors: Linz Peter , Veelken Roland
Published: March 2015

In rats, the mechanosensitive cardiorenal baroreflex influencing renal excretory function might be impaired by serotonin occurring in coronary arteries, e.g., in hypertension. Because the afferent limb of this reflex could be affected, we investigated the responses of nodose ganglion cells (one neuron of reflex) to osmotic, mechanical stress in presence or absence of the serotonin 5-HT3 receptor agonist phenylbiguanide (PBG). Current-voltage relationships (from −100 to +50 mV) were obtained using cell patch recordings while the cells were exposed to control or hypoosmotic solutions to induce mechanical stress. This protocol was repeated after low doses of PBG (10 μM), angiotensin II (10 nM), or the stretch-activated channel blocker gadolinium (20 μM) were added to the extracellular medium (EM). Hypoosmotic EM induced significant changes in cellular conductance. The full-range current-voltage relationship allowed for the calculation of a mean reversal potential of −13 ± 1.2 mV with respect to this change in cellular conductance ( n= 44). This increase in conductance was impaired after addition of either PBG or gadolinium to the EM,which was statistically evaluated at a voltage of −80 mV, where influences of voltage-gated channels are not likely to interfere with the responses recorded. The serotonin 5-HT3 receptor antagonist tropisetron (10 nM) prevented the PBG effect on conductance responses. Angiotensin II had no influence. Hence, serotonin might decrease the mechanical sensitivity of afferent cardiac nerves controlling renal sympathetic nerve activity.
NONOates regulate KCl cotransporter-1 and -3 mRNA expression in vascular smooth muscle cells
Authors: Di Fulvio Mauricio , Lauf Peter K. , Shah Shalin , Adragna Norma C.
Published: March 2015

Nitric oxide (NO) donors regulate KCl cotransport (KCC) activity and cotransporter-1 and -3 (KCC1 and KCC3) mRNA expression in sheep erythrocytes and in primary cultures of rat vascular smooth muscle cells (VSMCs), respectively. In this study, we used NONOates as rapid and slow NO releasers to provide direct evidence implicating NO as a regulator of KCC3 gene expression at the mRNA level. In addition, we used the expression of KCC3 mRNA to further investigate the mechanism of action of these NO donors at the cellular level. Treatment of VSMCs with rapid NO releasers, like NOC-5 and NOC-9, as well as with the direct NO-independent soluble guanylyl cyclase (sGC) stimulator YC-1, acutely increased KCC3 mRNA expression in a concentration- and time-dependent manner. The slow NO releaser NOC-18 had no effect on KCC3 gene expression. A specific NO scavenger completely prevented the NONOate-induced KCC3 mRNA expression. Inhibition of sGC with LY-83583 blocked the NONOate- and YC-1-induced KCC3 mRNA expression. This study shows that in primary cultures of rat VSMCs, the fast NO releasers NOC-9 and NOC-5, but not the slow NO releaser NOC-18, acutely upregulate KCC3 mRNA expression in a NO/sGC-dependent manner.
Rate-dependent [K+]o accumulation in canine right atria in vivo: electrophysiological consequences
Authors: Miyata Akira , Dowell Joshua D. , Zipes Douglas P. , Rubart Michael
Published: March 2015

Sudden increases in heart rate cause accumulation of K+ in the extracellular space. However, the exact relationship between rate and extracellular K+ concentration ([K+]o) in vivo is unknown. We measured [K+]o in right atria of anesthetized dogs by using K+-sensitive electrodes. Peak increase in [K+]o ranged from 0.18 ± 0.04 mM [means ± SE; cycle length (CL) = 350 ms] to 0.80 ± 0.09 mM (CL = 250 ms) above baseline (3.50 ± 0.08 mM at CL = 380 ms; n = 5). During rapid pacing-induced atrial fibrillation, peak increase in [K+]oaveraged 0.80 ± 0.07 mM ( n = 5). Whole cell current-clamp measurements in single right atrial myocytes ( n = 5) showed that raising [K+]o from 3 to 5 mM in 1-mM steps progressively depolarized resting membrane potential and reduced both phase 0 action potential amplitude and maximal upstroke velocity. Multisite epicardial mapping ( n = 4) demonstrated that sudden rate increases changed longitudinal conduction velocity (CVL) by −3.6 ± 1.8% to −5.9 ± 1.2% over a CL range of 330 to 250 ms. Our observations suggest that rate-related [K+]o accumulation in vivo is of sufficient magnitude to modulate those cellular electrophysiological properties that determine atrial CVL.
Tissue window chamber system for validation of implanted oxygen sensors
Authors: Makale Milan T. , Lin Joe T. , Calou Richard E. , Tsai Amy G. , Chen Peter C. , Gough David A.
Published: March 2015

An experimental system is described for validating electrochemical oxygen sensors implanted in tissues. The system is a modified hamster window chamber in which a thin layer of vascularized tissue is held between two plates, one plate having an observation window and the other plate having an array of oxygen sensors. This arrangement permits simultaneous recording of oxygen sensor signals and nondestructive visualization of the tissue adjacent to the sensors over periods of 1 mo or more, without the inhibitory effects of anesthesia. The system provides a means for study of the effects of spatial and temporal oxygen distributions on the sensor signals and adaptation of the tissue structure over time. Examples are given of sensor recordings and images of tissues with implanted oxygen sensor arrays.
Role of the paraventricular nucleus in renal excretory responses to acute volume expansion: role of nitric oxide
Authors: Li Yi-Fan , Mayhan William G. , Patel Kaushik P.
Published: March 2015

Acute volume expansion (VE) produces a suppression of renal sympathetic nerve discharge (RSND) resulting in diuresis and natriuresis. Recently, we have demonstrated that the endogenous nitric oxide (NO) system within the paraventricular nucleus (PVN) produces a decrease in RSND. We hypothesized that endogenous NO in the PVN is involved in the suppression of RSND leading to diuretic and natriuretic responses to acute VE. To test this hypothesis, we first measured the VE-induced increase in renal sodium excretion and urine flow with and without blockade of NO, with microinjection of NG-monomethyl-l-arginine (l-NMMA; 200 pmol in 200 nl), within the PVN of Inactin-anesthetized male Sprague-Dawley rats. Acute VE produced significant increases in urine flow and sodium excretion, which were diminished in rats treated with l-NMMA within the PVN. This effect of NO blockade within the PVN on VE-induced diuresis and natriuresis was abolished by renal denervation. Consistent with these data, acute VE induced a decrease in RSND (52% of the baseline level), which was significantly blunted by prior administration of l-NMMA into the PVN (28% of the baseline level) induced by a comparable level of acute VE. Using the push-pull perfusion technique, we found that acute VE induced a significant increase in NOx concentration in the perfusate from the PVN region. Taken together, these results suggest that acute VE induces an increase in NO production within the PVN that leads to renal sympathoinhibition, resulting in diuresis and natriuresis. We conclude that NO within the PVN plays an important role in regulation of sodium and water excretions in the volume reflex via modulating renal sympathetic outflow.
Endothelin receptor blockade has an oxygen-saving effect in Dahl salt-sensitive rats with heart failure
Authors: Noguchi Teruo , Chen Zengyi , Bell Stephen P. , Nyland Lori , LeWinter Martin M.
Published: March 2015

The effects of endothelin (ET) receptor blockade on energy utilization in heart failure (HF) are unknown. We administered ET type A (ETA), ET type B (ETB), and ETA/ETB antagonists to isolated hearts from Dahl salt-sensitive (DS) rats with HF and controls. Contractile efficiency was assessed as slope–1 of myocardial O consumption (V̇o2)-pressure-volume area relation. In HF, ETA and ETA/ETB but not ETB blockade decreased the contractility index ( Emax)(–15 ± 3% and –17 ± 2%, P < 0.05), excitation-contraction (E-C) coupling V̇o2 (–39 ± 4% and –37 ± 5%, P < 0.01), and efficiency (–15 ± 4% and –17 ± 2%, P < 0.05). Despite decreased efficiency, ETA and ETA/ETB blockade decreased total V̇o2 (–24 ± 3% and –22 ± 2%, P < 0.05). Na+/H+ exchanger inhibition decreased Emax and E-C coupling V̇o2 similar to ETA and ETA/ETB blockade, but did not alter efficiency. In HF, endogenous ET-1 maintains contractility at expense of increased V̇o2 through ETA receptor activation, likely mediated by Na+/H+ exchange.
Acute exercise reduces the response to colon distension in T5 spinal rats
Authors: Collins Heidi L. , Dicarlo Stephen E.
Published: March 2015

Individuals with spinal cord injuries above thoracic level 6 (T6) experience life-threatening bouts of hypertension, termed autonomic dysreflexia (AD). AD is mediated by peripheral α-adrenergic receptor supersensitivity as well as a reorganization of spinal pathways controlling sympathetic preganglionic neurons. A single bout of dynamic exercise may be a safe therapeutic approach to reduce the severity of AD because mild-to-moderate dynamic exercise reduces postexercise α-adrenergic receptor responsiveness, lowers postexercise sympathetic nerve activity, and reduces the postexercise response to stress. Therefore, this study was designed to test the hypothesis that mild-to-moderate dynamic exercise attenuates the postexercise response to colon distension (mechanism to elicit AD). To test this hypothesis, six male Wistar rats (406 ± 23 g), 5 wk post-T5 spinal cord transection, were instrumented with an arterial catheter. After recovery, the response to graded colon distension (10, 30, 50, and 80 mmHg, in random order) was determined before and after a single bout of mild-to-moderate dynamic exercise (9–12 m/min, 0% grade for 40 min). After exercise, the pressor response to graded colon distension was significantly attenuated (preexercise change: 2 ± 1, 9 ± 1, 14 ± 1, and 24 ± 2 vs. postexercise change: 2 ± 1, 2 ± 1, 9 ± 1, and 12 ± 3 mmHg). Thus acute exercise is a safe, therapeutic approach to reduce the severity of AD in paraplegic subjects.
Human SM22α BAC encompasses regulatory sequences for expression in vascular and visceral smooth muscles at fetal and adult stages
Authors: Xu Rui , Ho Ye-Shih , Ritchie Raquel P. , Li Li
Published: March 2015

The SM22α gene has widely been used to study the regulatory mechanisms of smooth muscle cell (SMC) gene expression during cardiovascular development. To determine the regulatory mechanisms for the evolutionarily conserved human SM22α (h SM22α) gene, we demonstrated that 445 bp upstream DNA sequences of h SM22α gene exhibited a high transcriptional activity in arterial SMC, not in venous nor in visceral SMCs during embryogensis. However, this promoter was gradually turned off in adulthood. Inclusion of the first intron in this promoter suppressed the promoter activity in pulmonary trunk arterial SMCs, whereas the expression in other systemic vasculature remained similar to that of the h SM22-445 promoter during the fetal and adult stages. To determine whether additional sequences are required for SM22α expression in all subtypes of SMCs, we examined the expression of a bacterial artificial chromosome containing the h SM22α locus in transgenic mice. The h SM22α transgene showed similar developmental expression patterns as the endogenous mouse SM22α gene, suggesting that this bacterial artificial chromosome contains essential regulatory sequences for its expression in arterial, venous, and visceral tissues during development.
Optimal ventricular rate slowing during atrial fibrillation by feedback AV nodal-selective vagal stimulation
Authors: Zhang Youhua , Mowrey Kent A. , Zhuang Shaowei , Wallick Don W. , Popović Zoran B. , Mazgalev Todor N.
Published: March 2015

Although the beneficial effects of ventricular rate (VR) slowing during atrial fibrillation (AF) are axiomatic, the precise relationship between VR and hemodynamics has not been determined. We hypothesized that selective atrioventricular node (AVN) vagal stimulation (AVN-VS) by varying the nerve stimulation intensity could achieve precise graded slowing and permit evaluation of an optimal VR during AF. The aims of the present study were the following: 1) to develop a method for computerized vagally controlled VR slowing during AF, 2) to determine the hemodynamic changes at each level of VR slowing, and 3) to establish the optimal anterograde VR during AF. AVN-VS was delivered to the epicardial fat pad that projects parasympathetic nerve fibers to the AVN in 14 dogs. Four target average VR levels, corresponding to 75%, 100%, 125%, and 150% of the sinus cycle length (SCL), were achieved by computer feedback algorithm. VR slowing resulted in improved hemodynamics and polynomial fit analysis found an optimum for the cardiac output at VR slowing of 87% SCL. We conclude that this novel method can be used to maintain slow anterograde conduction with best hemodynamics during AF.
On the mechanism of the probabilistic nature of ventricular defibrillation threshold
Authors: Yashima Masaaki , Kim Young-Hoon , Armin Sean , Wu Tsu-Juey , Miyauchi Yasushi , Mandel William J. , Chen Peng-Sheng , Karagueuzian Hrayr S.
Published: March 2015

The probabilistic nature of the ventricular defibrillation threshold (DFT) remains poorly understood. We hypothesized that shock outcome is a function of the amount of myocardium in its vulnerable period (VP). The endocardial surface of five isolated, perfused swine right ventricles was mapped with 477 bipolar electrodes during ventricular fibrillation (VF). Shock parameters and VF cycle length were not significantly different in the successful (S; n = 26) and failed (F; n= 26) trials. At the instant of the shock, the number of sites with 45- to 55-ms recovery was significantly smaller in the S trials than the F trials ( P < 0.04). No significant difference in the number of sites with recovery intervals outside the 45- to 55-ms range was seen in S and F shocks. Endocardial action potential showed that a recovery time of 45–55 ms corresponded to the VP spanning −15 to −60 mV in 92% of the regenerative action potentials. We conclude that the probabilistic nature of the DFT is related to the amount of myocardium in its VP.
Angiogenic growth factor expression in rat skeletal muscle in response to exercise training
Authors: Lloyd Pamela G. , Prior Barry M. , Yang Hsiao T. , Terjung Ronald L.
Published: March 2015

Angiogenesis occurs in skeletal muscle in response to exercise training. To gain insight into the regulation of this process, we evaluated the mRNA expression of factors implicated in angiogenesis over the course of a training program. We studied sedentary control ( n = 17) rats and both sedentary ( n = 18) and exercise-trained ( n = 48) rats with bilateral femoral artery ligation. Training consisted of treadmill exercise (4 times/day, 1–24 days). Basal mRNA expression in sedentary control muscle was inversely related to muscle vascularity. Angiogenesis was histologically evident in trained white gastrocnemius muscle by day 12. Training produced initial three- to sixfold increases in VEGF, VEGF receptors (KDR and Flt), the angiopoietin receptor (Tie-2), and endothelial nitric oxide synthase mRNA, which dissipated before the increase in capillarity, and a substantial (30- to 50-fold) but transient upregulation of monocyte chemoattractant protein 1 mRNA. These results emphasize the importance of early events in regulating angiogenesis. However, we observed a sustained elevation of the angiopoietin 2-to-angiopoietin 1 ratio, suggesting continued vascular destabilization. The response to exercise was (in general) tempered in high-oxidative muscles. These findings place importance on cellular events coupled to the onset of angiogenesis.
Effects of nicotine and dietary salt on a learned blood pressure response in Dahl-S rats
Authors: Kuo Justin H. , Speakman Richard O. , Sprinkle Aletia G. , Li Sheng-Gang , Brown David R. , Randall David C.
Published: March 2015

We examined the effects of chronic nicotine exposure and dietary salt on the arterial blood pressure (BP) changes learned in response to an acute behavioral stress in the Dahl salt-sensitive rat. Four groups were tested: low salt + vehicle; low salt + nicotine; high salt + vehicle; and high salt + nicotine. Rats were fed a low-salt (0.08% NaCl) or a high-salt (8% NaCl) diet for 4 wk; 2.4 mg · kg−1 · day−1nicotine or vehicle was given via an implanted osmotic minipump for the last 2 wk. All rats were conditioned by following one tone (CS+) with a 0.5-s tail shock; another tone (CS−) was never followed by shock. CS+ in low salt + vehicle and high salt + vehicle-treated rats evoked an initial arterial BP increase (C1), a component of the startle response, and an ensuing, smaller, but more sustained, pressor response (C2), which is acquired with training. In these rats, both C1 and C2 evoked by CS− were significantly smaller than those to CS+, demonstrating that these groups discriminated between the two tests. Conversely, although the low salt + nicotine-treated rats had both the C1 and C2 components of the conditional arterial pressure response, they did not discriminate between CS+ and CS−. Finally, the high salt + nicotine group failed to both discriminate between tones and acquire (i.e., learn) the C2 response. The unconditional response to shock did not differ between groups. We conclude that combined exposure to high salt and to nicotine inhibits the salt-sensitive animal's acquisition of a learned conditional BP response, perhaps because nicotine acts preferentially on those central processes required for associative learning versus those involved in orientating to external stimuli.
Estrogen modulation of eNOS activity and its association with caveolin-3 and calmodulin in rat hearts
Authors: Wang Xu , Abdel-Rahman Abdel A.
Published: March 2015

Previous studies have shown that estrogen modulation of endothelial nitric oxide (NO) synthase (eNOS) may confer protection against heart disease. Here, we demonstrate an association between reductions in baroreflex-mediated bradycardia and in cardiac NOS activity in ovariectomized (Ovx) rats compared with controls. The latter resulted, at least in part, from a reduction in cardiac eNOS protein. eNOS-derived NO and its biological effects are determined by the levels of eNOS protein and by eNOS catalytic activity; the latter is regulated partly through the dynamic interaction with an inhibitory protein (caveolin) and a stimulatory protein (calmodulin). The association of eNOS immunoprecipitated with caveolin-3 and calmodulin was examined. Caveolin-3 and calmodulin binding with eNOS was increased and decreased, respectively, in Ovx rats. 17β-Estradiol replacement restored, to within normal levels, the baroreflex-mediated bradycardic responses along with eNOS activity, eNOS expression, and the association of eNOS with caveolin-3 and calmodulin. Our findings may help to elucidate the molecular mechanism underlying the favorable effects of estrogen on cardiac responses to baroreflex activation.
Oxytocin does not directly affect vascular tone in vessels from nonpregnant and pregnant rats
Authors: Miller M. E. , Davidge S. T. , Mitchell B. F.
Published: March 2015

Recent evidence suggests oxytocin (OT) may regulate vascular tone. OT and its receptor (OTR) have been identified in the rat heart and great vessels. Expression of OT and OTR is increased in some tissues during pregnancy. We hypothesized that the OT/OTR system may be a physiological regulator of vascular tone and mediate the decreased vascular resistance noted during pregnancy. Using a wire myograph system, we measured changes in vascular tone in response to OT in small mesenteric arteries, uterine arcuate arteries, and thoracic aorta from nonpregnant and pregnant rats. Additionally, we used reverse transcriptase-polymerase chain reaction (RT-PCR) to measure mRNA for OTR in these vascular tissues. Although OTR mRNA was identified by RT-PCR, OT did not elicit a vasodilatory effect in any of the vessels studied. High concentrations of OT (>10−8 M) caused vasoconstriction that was eliminated by a specific vasopressin V1a receptor antagonist. Although it may have an indirect effect in regulation of peripheral resistance, we conclude that OT is unlikely to play a direct role in the physiological regulation of vascular tone.
Conducted dilations initiated by purines in arterioles are endothelium dependent and require endothelial Ca2+
Authors: Duza Tasmia , Sarelius Ingrid H.
Published: March 2015

The signaling pathways underlying the regulation of vascular resistance by purines in intact microvessels and particularly in communication of remote vasomotor responses are unclear. One process by which remote regions of arterioles communicate is via transmission of signals axially along the vessel wall. In this study, we identified a pathway for local and conducted dilations initiated by purines. Adenosine (Ado) or ATP (bind P1 and P2 purinergic receptors, respectively) was micropipette applied to arterioles (maximum diameter ∼40 μm) in the cheek pouch of anesthetized hamsters. Observations were made at the site of stimulation (local) or ∼1,200 μm upstream along the same vessel. P2 antagonists (pyridoxalphosphate-6-azophenyl-2′,4′-disulfonic acid tetrasodium and suramin) inhibited local constriction to ATP, whereas local and upstream dilations were unaffected. In contrast, during inhibition of P1 receptors (with xanthine amine congener) the local constriction was unchanged, whereas both local and upstream dilations to ATP were inhibited. Hydrolysis of ATP to Ado is implicated in the dilator response as blocking 5′-ectonucleotidase (with α,β-methyleneadenosine 5′-diphosphate) attenuated ATP-induced dilations. After endothelium denudation, constriction to ATP was unchanged, but dilations to both ATP and Ado were inhibited, identifying endothelial cells (ECs) as the primary target for P1-mediated dilation. Purines increased EC Ca2+ locally and upstream. Chelation of EC Ca2+ (with BAPTA) abolished the local and upstream dilations to P1 receptor stimulation. Collectively, these data demonstrate that stimulation of P1 receptors on ECs produces a vasodilation that spreads to remote regions. There is an associated increase in EC Ca2+, which is a required signaling intermediate in the manifestation of both the local and axially communicated arteriolar dilations.
Calcium sparks in mouse ventricular myocytes at physiological temperature
Authors: Ferrier Gregory R. , Smith Robin H. , Howlett Susan E.
Published: March 2015

In cardiac muscle, Ca2+is released from the sarcoplasmic reticulum (SR) in units called Ca2+sparks. Ca2+spark characteristics have been studied almost entirely at room temperature. This study compares characteristics of spontaneous sparks detected with fluo 3 in resting mouse ventricular myocytes at 22 and 37°C. The incidence and frequency of Ca2+sparks decreased dramatically at 37°C compared with 22°C. Also, spark amplitudes and times to peak were significantly reduced at 37°C. In contrast, spatial width and decay times were unchanged. During field stimulation, peak spatially averaged transients were similar at 22 and 37°C, and experiments with fura 2 demonstrated that diastolic and systolic Ca2+concentrations were unchanged. However, SR Ca2+content decreased significantly at 37°C. Restoration of SR Ca2+by superfusion with 5 mM Ca2+increased spark frequency but did not reverse the effects of temperature on spark parameters. Thus effects of temperature on spark frequency may reflect changes in SR stores, whereas changes in spark amplitude and rise time may reflect known effects of temperature on ryanodine receptor function.
Hu protein R-mediated posttranscriptional regulation of VEGF expression in rat gastrocnemius muscle
Authors: Tang Kechun , Breen Ellen C. , Wagner Peter D.
Published: March 2015

Hypoxic exercise increases VEGF expression and the formation of new capillaries. In addition to hypoxia-inducible factor regulation at the transcriptional level, VEGF message stabilization is also a key regulatory step for VEGF expression. In vitro experiments have identified Hu protein R (HuR) as a potential posttranscriptional regulator of VEGF gene expression. Here, we report that in rat skeletal muscle (gastrocnemius muscle), 1) HuR binds to a known regulatory sequence located in the VEGF mRNA 3′-untranslated region (1,631–1,678 bp); 2) HuR specifically binds to the A/U-rich element AUUUUA (1,665–1,670 bp) and an additional A/U-rich region containing the consensus sequence UUUUUUA (1,658–1,664 bp); 3) binding of HuR to VEGF mRNA is seen already 5 min after acute ischemia, remaining elevated throughout a 60-min ischemic period; 4) a second inducible HuR-VEGF mRNA binding factor is evident 30 and 60 min postischemia; and 5) VEGF mRNA and protein levels are increased 20 and 30 min, respectively, after acute ischemia. These findings suggest that acute ischemia induces a rapid binding of HuR to the VEGF mRNA 3′-untranslated region. In skeletal muscle, this specific protein-RNA interaction may be an important posttranscriptional regulatory mechanism for increasing VEGF expression in response to hypoxia or acute ischemia.
Attenuation of heat shock-induced cardioprotection by treatment with the opiate receptor antagonist naloxone
Authors: Patel Hemal H. , Hsu Anna , Gross Garrett J.
Published: March 2015

Whole body hyperthermia induces heat shock proteins (HSPs), which confer cardioprotection. Several opioid receptor subtypes are expressed in the heart and are linked to cardioprotection; however, no one has attempted to link the protection elicited by heat stress (HS) to opioids. Therefore, we investigated the effect of an opiate receptor antagonist, naloxone, on HS-induced cardioprotection. Anesthetized Sprague-Dawley rats were subjected to HS (42°C for 20 min) with and without naloxone pretreatment and were allowed to recover for 48 h. They then underwent 30 min of ischemia followed by 2 h of reperfusion. An acute HS group was given an intravenous bolus of naloxone (3 mg/kg) 10 min before index ischemia. Infarct size (IS), expressed as a percentage of the area at risk (IS/AAR), was determined. The right heart was excised for analysis of HSP content by Western blot. Heat-shocked rats showed significant reductions in IS/AAR versus control (16 ± 3 vs. 58 ± 4%, P < 0.001). Pretreatment with naloxone before HS attenuated the protective effects in a dose-dependent fashion, with significant attenuation of protection occurring at 15 mg/kg naloxone versus heat shock (42 ± 6 vs. 16 ± 3%, P < 0.001). Acute treatment with naloxone (3 mg/kg) 48 h after recovery from HS also significantly attenuated the delayed protective effect (47 ± 4 vs. 16 ± 3%, P < 0.001). No difference was seen in the level of HSP70 induced in the different groups. We conclude that heat shock-induced cardioprotection can be attenuated by naloxone, an opiate receptor antagonist, without reducing the levels of certain HSPs. These results suggest there may be a link between the endogenous release of opioids and HS that mediates cardioprotection.