bluuebunny/crossref_metadata_2025 huggingface dataset:
Cardioprotective and vasomotor effects of HO activity during acute and chronic hypoxia
Authors: Hartsfield Cynthia L. , McMurtry Ivan F. , Ivy D. Dunbar , Morris Kenneth G. , Vidmar Shanda , Rodman David M. , Fagan Karen A.
Published: June 2004

Prolonged hypoxia leads to the development of pulmonary hypertension. Recent reports have suggested enhancement of heme oxygenase (HO), the major source of intracellular carbon monoxide (CO), prevents hypoxia-induced pulmonary hypertension and vascular remodeling in rats. Therefore, we hypothesized that inhibition of HO activity by tin protoporphyrin (SnPP) would exacerbate the development of pulmonary hypertension. Rats were injected weekly with either saline or SnPP (50 μmol/kg) and exposed to hypobaric hypoxia or room air for 5 wk. Pulmonary and carotid arteries were catheterized, and animals were allowed to recover for 48 h. Pulmonary and systemic pressures, along with cardiac output, were recorded during room air and acute 10% O2breathing in conscious rats. No difference was detected in pulmonary artery pressure between saline- and SnPP-treated animals in either normoxic or hypoxic groups. However, blockade of HO activity altered both systemic and pulmonary vasoreactivity to acute hypoxic challenge. Despite no change in baseline pulmonary artery pressure, all rats treated with SnPP had decreased ratio of right ventricular (RV) weight to left ventricular (LV) plus septal (S) weight (RV/LV + S) compared with saline-treated animals. Echocardiograms suggested dilatation of the RV and decreased RV function in hypoxic SnPP-treated rats. Together these data suggest that inhibition of HO activity and CO production does not exacerbate pulmonary hypertension, but rather that HO and CO may be involved in mediating pulmonary and systemic vasoreactivity to acute hypoxia and hypoxia-induced RV function.
Opening of Ca2+-activated K+ channels triggers early and delayed preconditioning against I/R injury independent of NOS in mice
Authors: Wang Xiaoyin , Yin Chang , Xi Lei , Kukreja Rakesh C.
Published: June 2004

Opening of Ca2+-activated K+ (KCa) channels has been shown to confer early cardioprotection. It is unknown whether the opening of these channels also induces delayed cardioprotection. In addition, we determined the involvement of nitric oxide synthases (NOSs), which have been implicated in cardioprotection induced by opening of mitochondrial ATP-sensitive K+ (KATP) channels. Adult male ICR mice were pretreated with the KCa-channel opener NS-1619 either 10 min or 24 h before 30 min of global ischemia and 60 min of reperfusion (I/R) in Langendorff mode. Infusion of NS-1619 (10 μM) for 10 min before I/R led to smaller infarct sizes as compared with the vehicle (DMSO)-treated group ( P < 0.05). This infarct-limiting effect of NS-1619 was associated with improvement in ventricular functional recovery after I/R. The NS-1619-induced protection was abolished by coadministration with the KCa-channel blocker paxilline (1 μM). Similarly, pretreatment with NS-1619 (1 mg/kg ip) induced delayed protection 24 h later ( P < 0.05). Interestingly, the NS-1619-induced late protection was not blocked by the NOS inhibitor Nω-nitro-l-arginine methyl ester (15 mg/kg ip). Unlike diazoxide (the opener of mitochondrial KATP channels), NS-1619 did not increase the expression of inducible or endothelial NOS. Western blot analysis demonstrated the existence of α- and β-subunits of KCa channels in mouse heart tissue. We conclude that opening of KCa channels leads to both early and delayed preconditioning effects through a mechanism that is independent of nitric oxide.
Mechanism of the delay phenomenon: tissue protection is mediated by heme oxygenase-1
Authors: Kubulus Darius , Roesken Frank , Amon Michaela , Rücker Martin , Bauer Michael , Bauer Inge , Menger Michael D.
Published: June 2004

Induction of the “delay phenomenon” by chronic ischemia is an established clinical procedure, but the mechanisms conferring tissue protection are still incompletely understood. To elucidate the role of heme oxygenase-1 [HO-1 or heat shock protein-32 (HSP-32)] in delay, we examined in the skin-flap model of the ear of the hairless mouse, 1) whether chronic ischemia (delay) is capable to induce expression of HO-1, and 2) whether delay-induced HO-1 affects skin-flap microcirculation and survival by either its carbon monoxide-associated vasodilatory action or its biliverdin-associated anti-oxidative mechanism. Chronic ischemia was induced by transsection of the central feeding vessel of the ear 7 days before flap creation. The flap was finally raised by an incision through four-fifths of the base of the ear. Microcirculatory dysfunction and tissue necrosis were studied with the use of laser Doppler fluxmetry and intravital fluorescence microscopy. HO-1 protein expression was determined with Western blot analysis. Seven days of chronic ischemia (delay) induced a marked expression of HO-1. This was paralleled by a significant improvement ( P < 0.05) of microvascular perfusion and a reduction ( P < 0.05) of flap necrosis when compared with nondelayed controls. Importantly, blockade of HO-1 activity by tin protoporhyrin-IX completely blunted the protection of microcirculation and the improvement of tissue survival. Additional administration of the vitamin E analog trolox after blockade of HO-1 to mimic exclusively the anti-oxidative action of the heat shock protein did not restore the HO-1-associated microcirculatory improvement and only transiently attenuated the manifestation of flap necrosis. Thus our data indicate that the delay-induced protection from tissue necrosis is mediated by HO-1, predominantly through its carbon monoxide-associated action of adequately maintaining nutritive capillary perfusion.
Acute local subcutaneous VEGF165 injection for augmentation of skin flap viability: efficacy and mechanism
Authors: Khan Asim , Ashrafpour Homa , Huang Ning , Neligan Peter C. , Kontos Christopher , Zhong Anguo , Forrest Christopher R. , Pang Cho Y.
Published: June 2004

Distal skin ischemic necrosis is a common complication in skin flap surgery. The pathogenesis of skin flap ischemic necrosis is unclear, and there is no clinical treatment available. Here, we used the 4 × 10 cm rat dorsal skin flap model to test our hypothesis that subcutaneous injection of vascular endothelial growth factor 165 (VEGF165) in skin flaps at the time of surgery is effective in augmentation of skin flap viability, which is associated with an increase in nitric oxide (NO) production, and the mechanism involves 1) an increase in skin flap blood flow in the early stage after surgery and 2) enhanced angiogenesis subsequently to sustain increased skin flap blood flow and viability. We observed that subcutaneous injection of VEGF165 in skin flaps at the time of surgery increased skin flap viability in a dose-dependant manner. Subcutaneous injection of VEGF165 at the dose of 2 μg/flap increased skin flap viability by 28% ( P < 0.05; n = 8). Over 80% of this effect was blocked by intramuscular injection of the NO synthase (NOS) inhibitor Nω-nitro-l-arginine (13 mg/kg) 45 min before surgery ( P < 0.05; n = 8). The VEGF165 treatment also increased skin flap blood flow (2.68 ± 0.63 ml·min−1·100 g−1) compared with the control (1.26 ± 0.10 ml·min−1·100 g−1; P < 0.05, n = 6) assessed 6 h postoperatively. There was no change in skin flap capillary density at this time point. VEGF165-induced increase in capillary density (32.2 ± 1.1 capillaries/mm2; P < 0.05, n = 7) compared with control (24.6 ± 1.4 capillaries/mm2) was seen 7 days postoperatively. There was also evidence to indicate that VEGF165-induced NO production in skin flaps was stimulated by activation of NOS activity followed by upregulation of NOS protein expression. These observations support our hypothesis and for the first time provide an important insight into the mechanism of acute local VEGF165 protein therapy in mitigation of skin flap ischemic necrosis.
Early gestation dexamethasone programs enhanced postnatal ovine coronary artery vascular reactivity
Authors: Roghair Robert D. , Lamb Fred S. , Miller Francis J. , Scholz Thomas D. , Segar Jeffrey L.
Published: June 2004

Excessive exposure of the fetus to maternally derived corticosteroids has been linked to the development of adult-onset diseases. To determine if early gestation corticosteroid exposure alters subsequent coronary artery reactivity, we administered dexamethasone (0.28 mg·kg−1·day−1) to pregnant ewes at 27–28 days gestation (term being 145 days). Vascular responsiveness was assessed in endothelium-intact coronary and mesenteric arteries isolated from steroid-exposed and age-matched control fetal sheep at 123–126 days gestation and lambs at 4 mo of age. Lambs exposed to maternal dexamethasone had higher mean arterial blood pressures than the age-matched controls (93 ± 3 vs. 83 ± 5 mmHg, P < 0.05). Mesenteric arteries from the steroid-exposed fetuses displayed diminished responses to ANG II, relative to controls. In 4-mo-old lambs, prenatal dexamethasone exposure significantly increased coronary artery vasoconstriction to ANG II, ACh, and U-46619, but not KCl. In contrast, postnatal mesenteric artery reactivity was unaltered by steroid exposure. Compared with fetal mesenteric reactivity, postnatal mesenteric reactivity to ANG II, phenylephrine, and U-46619 was diminished, whereas the response to 120 mmol/l KCl was heightened. Coronary artery ANG II receptor protein expression was not significantly altered by steroid exposure in either age group. These findings demonstrate that early-gestation glucocorticoid exposure programs postnatal elevations in blood pressure and selectively enhances coronary artery responsiveness to second messenger-dependent vasoconstrictors. Glucocorticoid-induced alterations in coronary vascular smooth muscle structure or function may provide a mechanistic link between an adverse intrauterine environment and later cardiovascular disease.
Effects of extracellular nucleotides and their hydrolysis products on regulatory volume decrease of trout hepatocytes
Authors: Pafundo D. E. , Mut P. , Pérez Recalde M. , González-Lebrero R. M. , Fachino V. , Krumschnabel G. , Schwarzbaum P. J.
Published: June 2004

In trout hepatocytes, hypotonic swelling is followed by a compensatory shrinkage called regulatory volume decrease (RVD). It has been postulated that extracellular ATP and other nucleotides may interact with type 2 receptors (P2) to modulate this response. In addition, specific ectoenzymes hydrolyze ATP sequentially down to adenosine, which may bind to type 1 receptors (P1) and also influence RVD. Accordingly, in this study, we assessed the role of extracellular nucleoside 5′-tri- and diphosphates and of adenosine on RVD of trout hepatocytes. The extent of RVD after 40 min of maximum swelling was denoted as RVD40, whereas the initial rate of RVD was called vRVD. In the presence of hypotonic medium (60% of isotonic), hepatocytes swelled 1.6 times followed by vRVDof 1.7 min−1and RVD40of 60.2%. ATP, UTP, UDP, or ATPγS (P2agonists; 5 μM) increased vRVD1.5–2 times, whereas no changes were observed in the values of RVD40. Addition of 100 μM suramin or cibacron blue (P2antagonists) to the hypotonic medium produced no effect on vRVDbut a 53–58% inhibition of RVD40. Incubation of hepatocytes in the presence of either 5 μM [γ-32P]ATP or [α-32P]ATP induced the extracellular release of [γ-32P]Pi(0.21 nmol·10−6cells−1·min−1) and [α-32P]Pi(∼8 × 10−3nmol·10−6cells−1·min−1), suggesting the presence of ectoenzymes capable of fully dephosphorylating ATP. Concerning the effect of P1activation on RVD, 5 μM adenosine, both in the presence and absence of 100 μM S-(4-nitrobenzil)-6-tioinosine (a blocker of adenosine uptake), decreased RVD40by 37–44%, whereas 8-phenyl theophylline, a P1antagonist, increased RVD40by 15%. Overall, results indicate that ATP, UTP, and UDP, acting via P2, are important factors promoting RVD of trout hepatocytes, whereas adenosine binding to P1inhibits this process.
Medullary substrate and differential cardiovascular responses during stimulation of specific acupoints
Authors: Tjen-A-Looi Stephanie C. , Li Peng , Longhurst John C.
Published: June 2004

Electroacupuncture (EA) at P5–P6 acupoints overlying the median nerve reduces premotor sympathetic cardiovascular neuronal activity in the rostral ventral lateral medulla (rVLM) and visceral reflex pressor responses. In previous studies, we have noted different durations of influence of EA comparing P5–P6 and S36–S37 acupoints, suggesting that point specificity may exist. The purpose of this study was to evaluate the influence of stimulating P5–P6 (overlying the median nerve), LI4–L7 (overlying branches of the median nerve and the superficial radial nerve), LI6–LI7 (overlying the superficial radial nerve), LI10–LI11 (overlying the deep radial nerves), S36–S37 (overlying the deep peroneal nerves), or K1–B67 (overlying terminal branches of the tibial nerves) specific acupoints, overlying deep and superficial somatic nerves, on the excitatory cardiovascular reflex and rVLM responses evoked by stimulation of chemosensitive receptors in the cat's gallbladder with bradykinin (BK) or direct splanchnic nerve (SN) stimulation. We observed point-specific differences in magnitude and duration of EA inhibition between P5–P6 or LI10–LI11 and LI4–L7 or S36–S37 in responses to 30-min stimulation with low-frequency, low-current EA. EA at LI6–LI7 and K1–B67 acupoints as well as direct stimulation of the superficial radial nerve did not cause any cardiovascular or rVLM neuronal effects. Cardiovascular neurons in the rVLM, a subset of which were classified as premotor sympathetic cells, responded to brief (30 s) stimulation of the SN as well as acupoints P5–P6, LI10–LI11, LI4–L7, S36–S37, LI6–LI7, or K1–B67, or underlying somatic pathways in a fashion similar to the reflex responses. In fact, we observed a significant linear relationship ( r2 = 0.71) between the evoked rVLM response and reflex change in mean arterial blood pressure. In addition, EA stimulation at P5–P6 and LI4–L7 decreased rVLM neuronal activity by 41 and 12%, respectively, for >1 h, demonstrating that prolonged input into the medulla during stimulation of somatic nerves, depending on the degree of convergence, leads to more or less inhibition of activity of these cardiovascular neurons. Thus EA at acupoints overlying deep and superficial somatic nerves leads to point-specific effects on cardiovascular reflex responses. In a similar manner, sympathetic cardiovascular rVLM neurons that respond to both visceral (reflex) and somatic (EA) nerve stimulation manifest graded responses during stimulation of specific acupoints, suggesting that this medullary region plays a role in site-specific inhibition of cardiovascular reflex responses by acupuncture.
A putative H+-K+-ATPase in the Atlantic stingray,Dasyatis sabina: primary sequence and expression in gills
Authors: Choe Keith P. , Verlander Jill W. , Wingo Charles S. , Evans David H.
Published: June 2004

In mammals, the gastric H+-K+-ATPase (HKα1) mediates acid secretion in the stomach and kidneys. Like mammals, elasmobranchs also secrete acid from their stomachs, but unlike mammals they primarily use their gills for systemic acid excretion instead of their kidneys. The purpose of this study was to determine if an HKα1 orthologue exists in an elasmobranch (Atlantic stingray, Dasyatis sabina), to determine if it is expressed in gills and, if so, to localize its expression and determine if its expression is regulated during hypercapnia or freshwater acclimation. A polyclonal antibody made against an HKα1 peptide detected HKα1 immunoreactivity in protein isolates and tissue sections of stingray stomachs and gills. Immunohistochemistry demonstrated that HKα1 immunoreactivity was present in a subpopulation of epithelial cells in both organs. Double-labeling experiments in the gills showed that HKα1 immunoreactivity occurred in Na+-K+-ATPase-rich cells and not in V-type H+-ATPase-rich cells. RT-PCRs were used to deduce the primary sequence of a putative H+-K+-ATPase from the stomach of Atlantic stingrays. The 3,421-base pair cDNA includes a coding region for a 1,025-amino acid protein that is over 80% identical to HKα1 of mammals. RT-PCRs were then used to demonstrate that this transcript is also expressed in the gills. To our knowledge, this is the first H+-K+-ATPase sequence reported for any elasmobranch and the first full-length sequence for any fish. We also provide the first evidence for its expression in the gills of any fish and demonstrate that its expression increased during freshwater acclimation but not exposure to hypercapnia.
The 1- to 2-Hz oscillations in muscle force are exacerbated by stress, especially in older adults
Authors: Christou Evangelos A. , Jakobi Jennifer M. , Critchlow Ashley , Fleshner Monika , Enoka Roger M.
Published: June 2004

Although force fluctuations during a steady contraction are often heightened in old adults compared with young adults and are enhanced in young adults during the stress response, the mechanisms underlying the augmentation are uncertain. The purpose of the study was to compare the effect of a stressor on the plasma concentrations of selected stress hormones and on the force fluctuations experienced by young and old adults during the performance of a precision grip. Thirty-six men and women (19–86 yr) participated in a protocol that comprised anticipatory (30 min), stressor (15 min), and recovery periods (25 min). The stressor was a series of noxious electrical stimuli applied to the dorsal surface of the left hand. Subjects sustained a pinch-grip force with the right hand at 2% of the maximal voluntary contraction force. The fluctuations in pinch-grip force, the interference electromyogram (EMG) of six muscles, and the spectra for the force and EMG were quantified across the 70-min protocol. The stressor increased the force fluctuations, largely due to an enhancement of the power at 1–2 Hz in the force spectrum ( r2 = 0.46). The effect was greatest for the old adults compared with young and middle-aged adults. The plasma concentrations of the stress hormones (adrenocorticotropin, epinephrine, and norepinephrine) were elevated to similar levels for all three age groups, and the changes were not associated with modulation of the force fluctuations. Furthermore, the heightened EMG activity exhibited by the old adults during all periods was not related to the changes in the force fluctuations or the 1- to 2-Hz force oscillations. The absence of a change in the mean pinch-grip force during the protocol and the lack of an association between elevation of the plasma concentrations for the stress hormones and modulation of the force fluctuations suggest that the enhanced force fluctuations caused by the stressor was due to an increase in the low-frequency output of the spinal motor neurons.
Differential displacement of the human soleus and medial gastrocnemius aponeuroses during isometric plantar flexor contractions in vivo
Authors: Bojsen-Møller Jens , Hansen Philip , Aagaard Per , Svantesson Ulla , Kjaer Michael , Magnusson S. Peter
Published: June 2004

The human triceps surae muscle-tendon complex is a unique structure with three separate muscle compartments that merge via their aponeuroses into the Achilles tendon. The mechanical function and properties of these structures during muscular contraction are not well understood. The purpose of the study was to investigate the extent to which differential displacement occurs between the aponeuroses of the medial gastrocnemius (MG) and soleus (Sol) muscles during plantar flexion. Eight subjects (mean ± SD; age 30 ± 7 yr, body mass 76.8 ± 5.5 kg, height 1.83 ± 0.06 m) performed maximal isometric ramp contractions with the plantar flexor muscles. The experiment was performed in two positions: position 1, in which the knee joint was maximally extended, and position 2, in which the knee joint was maximally flexed (125°). Plantarflexion moment was assessed with a strain gauge load cell, and the corresponding displacement of the MG and Sol aponeuroses was measured by ultrasonography. Differential shear displacement of the aponeurosis was quantified by subtracting displacement of Sol from that of MG. Maximal plantar flexion moment was 36% greater in position 1 than in position 2 (132 ± 20 vs. 97 ± 11 N·m). In position 1, the displacement of the MG aponeurosis at maximal force exceeded that of the Sol (12.6 ± 1.7 vs. 8.9 ± 1.5 mm), whereas in position 2 displacement of the Sol was greater than displacement of the MG (9.6 ± 1.0 vs. 7.9 ± 1.2 mm). The amount and “direction” of shear between the aponeuroses differed significantly between the two positions across the entire range of contraction, indicating that the Achilles tendon may be exposed to intratendinous shear and stress gradients during human locomotion.
Vasodilatory mechanisms in contracting skeletal muscle
Authors: Clifford Philip S. , Hellsten Ylva
Published: June 2004

Skeletal muscle blood flow is closely coupled to metabolic demand, and its regulation is believed to be mainly the result of the interplay of neural vasoconstrictor activity and locally derived vasoactive substances. Muscle blood flow is increased within the first second after a single contraction and stabilizes within ∼30 s during dynamic exercise under normal conditions. Vasodilator substances may be released from contracting skeletal muscle, vascular endothelium, or red blood cells. The importance of specific vasodilators is likely to vary over the time course of flow, from the initial rapid rise to the sustained elevation during steady-state exercise. Exercise hyperemia is therefore thought to be the result of an integrated response of more than one vasodilator mechanism. To date, the identity of vasoactive substances involved in the regulation of exercise hyperemia remains uncertain. Numerous vasodilators such as adenosine, ATP, potassium, hypoxia, hydrogen ion, nitric oxide, prostanoids, and endothelium-derived hyperpolarizing factor have been proposed to be of importance; however, there is little support for any single vasodilator being essential for exercise hyperemia. Because elevated blood flow cannot be explained by the failure of any single vasodilator, a consensus is beginning to emerge for redundancy among vasodilators, where one vasoactive compound may take over when the formation of another is compromised. Conducted vasodilation or flow-mediated vasodilation may explain dilation in vessels (i.e., feed arteries) not directly exposed to vasodilator substances in the interstitium. Future investigations should focus on identifying novel vasodilators and the interaction between vasodilators by simultaneous inhibition of multiple vasodilator pathways.
Muscle pressor reflex: potential role of vanilloid type 1 receptor and acid-sensing ion channel
Authors: Li Jianhua , Maile Michael D. , Sinoway Adam N. , Sinoway Lawrence I.
Published: June 2004

Reflex cardiovascular responses to muscle contraction are mediated by mechanical and metabolic stimulation of thin muscle afferent fibers. Metabolic stimulants and receptors involved in responses are uncertain. Capsaicin depolarizes thin sensory afferent nerves that have vanilloid type 1 receptors (VR1). Among potential endogenous ligands of thin fibers, H+ has been suggested as a metabolite mediating the reflex muscle response as well as a potential stimulant of VR1. It has also been suggested that acid-sensing ion channels (ASIC) mediate H+, evoking afferent nerve excitation. We have examined the roles of VR1 and ASIC in mediating cardiovascular reflex responses to acid stimulation of muscle afferents in a rat model. In anesthetized rats, injections of capsaicin into the arterial blood supply of triceps surae muscles evoked a biphasic response ( n = 6). An initial fall in mean arterial pressure (from baseline of 95.8 ± 9.5 to 70.4 ± 4.5 mmHg, P < 0.05 vs. baseline) was followed by an increase (to 131.6 ± 11.3 mmHg, P < 0.05 vs. baseline). Anandamide (an endogenous substance that activates VR1) induced the same change in blood pressure as did capsaicin. The pressor (but not depressor) component of the response was blocked by capsazepine (a VR1 antagonist) and section of afferent nerves. In decerebrate rats ( n = 8), H+ evoked a pressor response that was not blocked by capsazepine but was attenuated by amiloride (an ASIC blocker). In rats ( n = 12) pretreated with resiniferatoxin to destroy muscle afferents containing VR1, capsaicin and H+ responses were blunted. We conclude that H+ stimulates ASIC, evoking the reflex response, and that ASIC are likely to be frequently found on afferents containing VR1. The data also suggest that VR1 and ASIC may play a role in processing of muscle afferent signals, evoking the muscle pressor reflex.
Malleability of human skeletal muscle Na+-K+-ATPase pump with short-term training
Authors: Green H. J. , Barr D. J. , Fowles J. R. , Sandiford S. D. , Ouyang J.
Published: June 2004

To investigate the hypothesis that short-term submaximal training would result in changes in Na+-K+-ATPase content, activity, and isoform distribution in skeletal muscle, seven healthy, untrained men [peak aerobic power (peak oxygen consumption; V̇o2 peak) = 45.6 ml·kg−1·min−1 (SE 5.4)] cycled for 2 h/day at 60–65% V̇o2 peak for 6 days. Muscle tissue, sampled from the vastus lateralis before training (0 days) and after 3 and 6 days of training and analyzed for Na+-K+-ATPase content, as assessed by the vanadate facilitated [3H]ouabain-binding technique, was increased ( P < 0.05) at 3 days (294 ± 8.6 pmol/g wet wt) and 6 days (308 ± 15 pmol/g wet wt) of training compared with 0 days (272 ± 9.7 pmol/g wet wt). Maximal Na+-K+-ATPase activity as evaluated by the 3- O-methylfluorescein phosphatase assay was increased ( P < 0.05) by 6 days (53.4 ± 5.9 nmol·h−1·mg protein−1) but not by 3 days (35.9 ± 4.5 nmol·h−1·mg protein−1) compared with 0 days (37.8 ± 3.7 nmol·h−1·mg protein−1) of training. Relative isoform distribution, measured by Western blot techniques, indicated increases ( P < 0.05) in α2-content by 3 days and β1-content by 6 days of training. These results indicate that prolonged aerobic exercise represents a potent stimulus for the rapid adaptation of Na+-K+-ATPase content, isoform, and activity characteristics.
Ideas about control of skeletal and cardiac muscle blood flow (1876–2003): cycles of revision and new vision
Authors: Rowell Loring B.
Published: June 2004

This perspective examines origins of some key ideas central to major issues to be addressed in five subsequent mini-reviews related to Skeletal and Cardiac Muscle Blood Flow. The questions discussed are as follows. 1) What causes vasodilation in skeletal and cardiac muscle and 2) might the mechanisms be the same in both? 3) How important is muscle's mechanical contribution (via muscle pumping) to muscle blood flow, including its effect on cardiac output? 4) Is neural (vasoconstrictor) control of muscle vascular conductance and muscle blood flow significantly blunted in exercise by muscle metabolites and what might be a dominant site of action? 5) What reflexes initiate neural control of muscle vascular conductance so as to maintain arterial pressure at its baroreflex operating point during dynamic exercise, or is muscle blood flow regulated so as to prevent accumulation of metabolites and an ensuing muscle chemoreflex or both?
Influenza virus-induced sleep responses in mice with targeted disruptions in neuronal or inducible nitric oxide synthases
Authors: Chen Lichao , Duricka Deborah , Nelson Scott , Mukherjee Sanjib , Bohnet Stewart G. , Taishi Ping , Majde Jeannine A. , Krueger James M.
Published: June 2004

Influenza viral infection induces increases in non-rapid eye movement sleep and decreases in rapid eye movement sleep in normal mice. An array of cytokines is produced during the infection, and some of them, such as IL-1β and TNF-α, are well-defined somnogenic substances. It is suggested that nitric oxide (NO) may mediate the sleep-promoting effects of these cytokines. In this study, we use mice with targeted disruptions of either the neuronal NO synthase (nNOS) or the inducible NO synthase (iNOS) gene, commonly referred to as nNOS or iNOS knockouts (KOs), to investigate sleep changes after influenza viral challenge. We report that the magnitude of viral-induced non-rapid eye movement sleep responses in both nNOS KOs and iNOS KOs was less than that of their respective controls. In addition, the duration of rapid eye movement sleep in nNOS KO mice did not decrease compared with baseline values. All strains of mice had similar viral titers and cytokine gene expression profiles in the lungs. Virus was not isolated from the brains of any strain. However, gene expression in the brain stem differed between nNOS KOs and their controls: mRNA for the interferon-induced gene 2′,5′-oligoadenylate synthase 1a was elevated in nNOS KOs relative to their controls at 15 h, and IL-1β mRNA was elevated in nNOS KOs relative to their controls at 48 h. Our results suggest that NO synthesized by both nNOS and iNOS plays a role in virus-induced sleep changes and that nNOS may modulate cytokine expression in the brain.
Variable Hypocoagulant Effect of Fish Oil Intake in Humans
Authors: Vanschoonbeek Kristof , Feijge Marion A.H. , Paquay Martine , Rosing Jan , Saris Wim , Kluft Cornelis , Giesen Peter L.A. , de Maat Moniek P.M. , Heemskerk Johan W.M.
Published: June 2004

Objective—

The beneficial effect of dietary fish oil, rich in omega-3 polyunsaturated fatty acids (PUFAs), on cardiovascular disease is multifactorial and may partly rely on their anticoagulant action. We studied how fish oil intake influenced thrombin generation in plasma and which factors were involved herein.

Methods and Results—

Twenty-five healthy males with borderline overweight received 3.0 g omega-3 PUFAs daily for 4 weeks. Fish oil intake reduced plasma triglycerides and lowered platelet integrin activation, as well as plasma levels of fibrinogen and factor V, but had no effect on vitamin K-dependent coagulation factors. Before fish oil intake, thrombin generation (reflecting the coagulant potential) considerably varied between plasmas from individual subjects, which were partly explained by variation in prothrombin, antithrombin, fibrinogen, and factor V levels. Fish oil intake reduced thrombin generation in the presence and absence of platelets. This reduction correlated with the fish oil effect on fibrinogen and factor V levels. Interestingly, the lowering effect of fish oil on thrombin generation and fibrinogen clustered around subjects with high fibrinogen carrying a structural fibrinogen α-chain polymorphism.

Conclusions—

Dietary omega-3 PUFAs provoke a hypocoagulant, vitamin K-independent effect in humans, the degree of which may depend on fibrinogen level.
Ceramide Triggers Weibel–Palade Body Exocytosis
Authors: Bhatia Rinky , Matsushita Kenji , Yamakuchi Munekazu , Morrell Craig N. , Cao Wangsen , Lowenstein Charles J.
Published: June 2004

The sphingolipid ceramide mediates a variety of stress responses, including vascular inflammation and thrombosis. Activated endothelial cells release Weibel-Palade bodies, granules containing von Willebrand factor (vWF) and P-selectin, which induce leukocyte rolling and platelet adhesion and aggregation. We hypothesized that ceramide induces vascular inflammation and thrombosis in part by triggering Weibel-Palade body exocytosis. We added ceramide to human aortic endothelial cells and assayed Weibel-Palade body exocytosis by measuring the concentration of vWF released into the media. Exogenous ceramide induces vWF release from endothelial cells in a dose-dependent manner. Activators of endogenous ceramide production, neutral sphingomyelinase, or tumor necrosis factor-α also induce Weibel-Palade body exocytosis. We next studied NO effects on ceramide-induced Weibel-Palade body exocytosis because NO can inhibit vascular inflammation. The NO donor
S
-nitroso-
N
-acetylpenicillamine decreases ceramide-induced vWF release in a dose-dependent manner, whereas the NO synthase inhibitor
N
G
-nitro-
l
-arginine methyl ester increases ceramide-induced vWF release. In summary, our findings show that endogenous ceramide triggers Weibel-Palade body exocytosis, and that endogenous NO inhibits ceramide-induced exocytosis. These data suggest a novel mechanism by which ceramide induces vascular inflammation and thrombosis.
IQGAP1, a Novel Vascular Endothelial Growth Factor Receptor Binding Protein, Is Involved in Reactive Oxygen Species—Dependent Endothelial Migration and Proliferation
Authors: Yamaoka-Tojo Minako , Ushio-Fukai Masuko , Hilenski Lula , Dikalov Sergey I. , Chen Yuqing E. , Tojo Taiki , Fukai Tohru , Fujimoto Mitsuaki , Patrushev Nikolay A. , Wang Ningning , Kontos Christopher D. , Bloom George S. , Alexander R. Wayne
Published: June 2004

Endothelial cell (EC) proliferation and migration are important for reendothelialization and angiogenesis. We have demonstrated that reactive oxygen species (ROS) derived from the small GTPase Rac1-dependent NAD(P)H oxidase are involved in vascular endothelial growth factor (VEGF)–mediated endothelial responses mainly through the VEGF type2 receptor (VEGFR2). Little is known about the underlying molecular mechanisms. IQGAP1 is a scaffolding protein that controls cellular motility and morphogenesis by interacting directly with cytoskeletal, cell adhesion, and small G proteins, including Rac1. In this study, we show that IQGAP1 is robustly expressed in ECs and binds to the VEGFR2. A pulldown assay using purified proteins demonstrates that IQGAP1 directly interacts with active VEGFR2. In cultured ECs, VEGF stimulation rapidly promotes recruitment of Rac1 to IQGAP1, which inducibly binds to VEGFR2 and which, in turn, is associated with tyrosine phosphorylation of IQGAP1. Endogenous IQGAP1 knockdown by siRNA shows that IQGAP1 is involved in VEGF-stimulated ROS production, Akt phosphorylation, endothelial migration, and proliferation. Wound assays reveal that IQGAP1 and phosphorylated VEGFR2 accumulate and colocalize at the leading edge in actively migrating ECs. Moreover, we found that IQGAP1 expression is dramatically increased in the VEGFR2-positive regenerating EC layer in balloon-injured rat carotid artery. These results suggest that IQGAP1 functions as a VEGFR2-associated scaffold protein to organize ROS-dependent VEGF signaling, thereby promoting EC migration and proliferation, which may contribute to repair and maintenance of the functional integrity of established blood vessels.
Right Ventricular Myocardium Derives From the Anterior Heart Field
Authors: Zaffran Stéphane , Kelly Robert G. , Meilhac Sigolène M. , Buckingham Margaret E. , Brown Nigel A.
Published: June 2004

The mammalian heart develops from a primary heart tube, which is formed by fusion of bilateral cardiac territories in which myocardial and endothelial cells have already begun to differentiate from splanchnic mesoderm. A population of myocardial precursors has been identified in pharyngeal mesoderm, anterior to the early heart tube. Cell labeling studies have indicated that this novel territory, called the anterior heart field (AHF), gives rise to the myocardial wall of the outflow tract. We now report that not only the myocardium of the outflow tract but also myocardial cells of the embryonic right ventricle are derived from this source. Explants of pharyngeal mesoderm or of the early heart tube were cultured from transgenic mice in which transgene expression marks different regions of the heart. Pharyngeal mesoderm from 5 to 7 somite embryos gives rise to cardiomyocytes with right ventricular and outflow tract identities, whereas the heart tube as this stage has an essentially left ventricular identity. DiI labeling confirms that the early heart tube is destined to contribute to the embryonic left ventricle and indicates that right ventricular myocardium is added from extracardiac mesoderm. Retrospective clonal analysis of the heart at embryonic day (E) 10.5 reveals the existence of a clonal boundary in the interventricular region, which appears during ventricular septation, underlining different origins of the two ventricular compartments. This study demonstrates the differences in the embryological origin of right and left ventricular myocardium, which has important implications for congenital heart disease.
Cardiac Voltage-Gated Sodium Channel Na
v
1.5 Is Regulated by Nedd4-2 Mediated Ubiquitination
Authors: van Bemmelen Miguel X. , Rougier Jean-Sébastien , Gavillet Bruno , Apothéloz Florine , Daidié Dorothée , Tateyama Michihiro , Rivolta Ilaria , Thomas Marc A. , Kass Robert S. , Staub Olivier , Abriel Hugues
Published: June 2004

Na
v
1.5, the cardiac isoform of the voltage-gated Na
+
channel, is critical to heart excitability and conduction. However, the mechanisms regulating its expression at the cell membrane are poorly understood. The Na
v
1.5 C-terminus contains a PY-motif (xPPxY) that is known to act as binding site for Nedd4/Nedd4-like ubiquitin-protein ligases. Because Nedd4-2 is well expressed in the heart, we investigated its role in the ubiquitination and regulation of Na
v
1.5. Yeast two-hybrid and GST-pulldown experiments revealed an interaction between Na
v
1.5 C-terminus and Nedd4-2, which was abrogated by mutating the essential tyrosine of the PY-motif. Ubiquitination of Na
v
1.5 was detected in both transfected HEK cells and heart extracts. Furthermore, Nedd4-2–dependent ubiquitination of Na
v
1.5 was observed. To test for a functional role of Nedd4-2, patch-clamp experiments were performed on HEK cells expressing wild-type and mutant forms of both Na
v
1.5 and Nedd4-2. Na
v
1.5 current density was decreased by 65% upon Nedd4-2 cotransfection, whereas the PY-motif mutant channels were not affected. In contrast, a catalytically inactive Nedd4-2 had no effect, indicating that ubiquitination mediates this downregulation. However, Nedd4-2 did not alter the whole-cell or the single channel biophysical properties of Na
v
1.5. Consistent with the functional findings, localization at the cell periphery of Na
v
1.5-YFP fusion proteins was reduced upon Nedd4-2 coexpression. The Nedd4-1 isoform did not regulate Na
v
1.5, suggesting that Nedd4-2 is a specific regulator of Na
v
1.5. These results demonstrate that Na
v
1.5 can be ubiquitinated in heart tissues and that the ubiquitin-protein ligase Nedd4-2 acts on Na
v
1.5 by decreasing the channel density at the cell surface.
Preferential Expression and Function of Voltage-Gated, O
2
-Sensitive K
+
Channels in Resistance Pulmonary Arteries Explains Regional Heterogeneity in Hypoxic Pulmonary Vasoconstriction
Authors: Archer Stephen L. , Wu Xi-Chen , Thébaud Bernard , Nsair Ali , Bonnet Sebastien , Tyrrell Ben , McMurtry M. Sean , Hashimoto Kyoko , Harry Gwyneth , Michelakis Evangelos D.
Published: June 2004

Hypoxic pulmonary vasoconstriction (HPV) is initiated by inhibition of O
2
-sensitive, voltage-gated (Kv) channels in pulmonary arterial smooth muscle cells (PASMCs). Kv inhibition depolarizes membrane potential (E
M
), thereby activating Ca
2+
influx via voltage-gated Ca
2+
channels. HPV is weak in extrapulmonary, conduit pulmonary arteries (PA) and strong in precapillary resistance arteries. We hypothesized that regional heterogeneity in HPV reflects a longitudinal gradient in the function/expression of PASMC O
2
-sensitive Kv channels. In adult male Sprague Dawley rats, constrictions to hypoxia, the Kv blocker 4-aminopyridine (4-AP), and correolide, a Kv1.x channel inhibitor, were endothelium-independent and greater in resistance versus conduit PAs. Moreover, HPV was dependent on Kv-inhibition, being completely inhibited by pretreatment with 4-AP. Kv1.2, 1.5, Kv2.1, Kv3.1b, Kv4.3, and Kv9.3. mRNA increased as arterial caliber decreased; however, only Kv1.5 protein expression was greater in resistance PAs. Resistance PASMCs had greater K
+
current (I
K
) and a more hyperpolarized E
M
and were uniquely O
2
− and correolide-sensitive. The O
2
-sensitive current (active at −65 mV) was resistant to iberiotoxin, with minimal tityustoxin sensitivity. In resistance PASMCs, 4-AP and hypoxia inhibited I
K
57% and 49%, respectively, versus 34% for correolide. Intracellular administration of anti-Kv1.5 antibodies inhibited correolide’s effects. The hypoxia-sensitive, correolide-insensitive I
K
(15%) was conducted by Kv2.1. Anti-Kv1.5 and anti-Kv2.1 caused additive depolarization in resistance PASMCs (Kv1.5>Kv2.1) and inhibited hypoxic depolarization. Heterologously expressed human PASMC Kv1.5 generated an O
2
− and correolide-sensitive I
K
like that in resistance PASMCs. In conclusion, Kv1.5 and Kv2.1 account for virtually all the O
2
-sensitive current. HPV occurs in a Kv-enriched resistance zone because resistance PASMCs preferentially express O
2
-sensitive Kv-channels.
Overexpression of CuZn-SOD Prevents Lipopolysaccharide-Induced Endothelial Dysfunction
Authors: Didion Sean P. , Kinzenbaw Dale A. , Fegan Pamela E. , Didion Lisa A. , Faraci Frank M.
Published: June 2004

Background and Purpose—

Inflammation is thought to be a major contributor to carotid artery disease. Lipopolysaccharide (LPS) activates inflammatory mechanisms thought to contribute to endothelial dysfunction by mechanisms that are not well defined. The goal of this study was to determine whether overexpression of CuZn-SOD protects against LPS-induced increases in superoxide and endothelial dysfunction.

Methods—

Carotid arteries from CuZn-SOD transgenic (SOD-Tg) and nontransgenic (non-Tg) littermates were examined in vitro. Superoxide levels were measured using lucigenin-enhanced chemiluminescence.

Results—

In non-Tg mice, LPS (0.5 μg/mL for 22 hours) produced marked impairment of vasorelaxation in response to the endothelium-dependent dilator acetylcholine (ACh). For example, 100 μmol/L ACh relaxed carotid arteries from non-Tg mice by 86±6% and 38±8% after treatment with vehicle and LPS, respectively. In contrast, LPS did not significantly impair responses of carotid artery to ACh in SOD-Tg mice, and LPS had no effect on relaxation responses to the endothelium-independent dilator nitroprusside in carotid artery from non-Tg or SOD-Tg mice. LPS-induced increases in superoxide, as measured using lucigenin-enhanced chemiluminescence, were higher in vessels from non-Tg mice than from SOD-Tg mice.

Conclusions—

These results indicate that LPS increases superoxide and impairs endothelium-dependent relaxation. Overexpression of the CuZn isoform of SOD effectively prevents LPS-induced oxidative stress and endothelial dysfunction in the carotid artery.
Predictors of Cerebral Infarction in Aneurysmal Subarachnoid Hemorrhage
Authors: Rabinstein Alejandro A. , Friedman Jonathan A. , Weigand Stephen D. , McClelland Robyn L. , Fulgham Jimmy R. , Manno Edward M. , Atkinson John L.D. , Wijdicks Eelco F.M.
Published: June 2004

Background—

Clinical and radiologic predictors of cerebral infarction occurrence and location after aneurysmal subarachnoid hemorrhage have been seldom studied.

Methods—

We evaluated all patients admitted to our hospital with aneurysmal subarachnoid hemorrhage between 1998 and 2000. Cerebral infarction was defined as a new hypodensity located in a vascular distribution on computed tomography (CT) scan.

Results—

Fifty-seven of 143 patients (40%) developed a cerebral infarction. On univariate analysis, occurrence of cerebral infarction was associated with a worse World Federation of Neurological Surgeons grade (
P
=0.01), use of ventriculostomy catheter (
P
=0.01), preoperative vasospasm (
P
=0.03), surgical clipping (
P
=0.02), symptomatic vasospasm (
P
<0.01), and vasospasm on transcranial Doppler ultrasonography (TCD) or repeat angiogram (
P
<0.01). On multivariable analysis, only presence of symptoms ascribed to vasospasm (
P
<0.01) and evidence of vasospasm on TCD or angiogram predicted cerebral infarction (
P
<0.01). TCD and angiogram agreed on the diagnosis of vasospasm in 73% of cases (95% CI, 63% to 81%), but the diagnostic accuracy of this combination of tests was suboptimal for the prediction of cerebral infarction occurrence (sensitivity, 0.72; specificity, 0.68; positive predictive value, 0.67; negative predictive value, 0.72). Location of the cerebral infarction on delayed CT was predicted by neurological symptoms in 74%, by aneurysm location in 77%, and by angiographic vasospasm in 67%.

Conclusions—

Evidence of vasospasm on TCD and angiogram is predictive of cerebral infarction on CT scan but sensitivity and specificity are suboptimal. Cerebral infarction location cannot be predicted in one quarter to one third of patients by any of the studied clinical or radiological variables.
Probability of Cortical Infarction Predicted by Flumazenil Binding and Diffusion-Weighted Imaging Signal Intensity
Authors: Heiss Wolf-Dieter , Sobesky Jan , Smekal Ulrich v. , Kracht Lutz W. , Lehnhardt Fritz-Georg , Thiel Alexander , Jacobs Andreas H. , Lackner Karl
Published: June 2004

Background and Purpose—

The differentiation of reversible from irreversible ischemic damage is essential for identifying patients with acute ischemic deficits who may benefit from therapeutic interventions. Diffusion-weighted imaging (DWI) has become the method of choice to detect ischemic lesions. Positron emission tomography (PET) of the central benzodiazepine receptor ligand 11C flumazenil (FMZ) has been shown to be a reliable marker of neuronal integrity. These 2 imaging parameters were compared with respect to the probability to predict cortical infarction in early ischemic stroke.

Methods—

In 12 patients with acute stroke, results from DWI (median, 6.5 hours after symptom onset) and FMZ–PET (interval, 85 minutes between DWI and PET) were compared with infarct extension 24 to 48 hours after onset of stroke on T2-weighted magnetic resonance imaging (T2-MRI). Probability curves predictive of eventual infarction were computed using respective DWI, FMZ, and apparent diffusion coefficient (ADC) values for voxels of interest (VOI) later classified as representing infarcted or noninfarcted tissue.

Results—

Ninety-five percent limits predictive of cortical infarction were determined for relative FMZ binding (≤3.2), DWI signal intensity (≥1.18), and ADC values (≤0.83). Cortical regions with values beyond these 95% limits did not necessarily overlap with nor were fully congruous with final cortical infarct volumes. The respective median volumes for these regions were FMZ median 10.9, range 0 to 99.7 cm
3
; DWI median 15.2, range 0 to 116.0 cm
3
; ADC median 12.4, range 0 to 112.7 cm
3
; and final infarct median 14.9, range 0 to 114.7 cm
3
. Overall, 83.5% of the final infarct, on average, was predicted by decreased FMZ binding, 84.7% by increased DWI signal intensity, and 70.9% by a decreased ADC value. The portions of the final infarct not predicted in the early investigation (false-negatives) were 4.8 cm
3
(median) for FMZ, 3.7 cm
3
for DWI, and 6.0 cm
3
for ADC. The false-positive volumes not included in the final infarct were 0 cm
3
(median) for FMZ, 5.1 cm
3
for DWI, and 3.6 cm
3
for ADC.

Conclusions—

These results indicate that FMZ–PET and DWI are comparable in the prediction of probability of ischemic cortical infarction, but FMZ–PET carries a lower probability of false-positive prediction. The final infarcts include tissue not identified by these imaging modalities; at the time of the study, these tissue compartments are viable and could benefit from treatment. The discrepancy in predictive probability could be related to the fundamental difference of the measured variables: benzodiazepine receptor activity is a reliable marker of neuronal integrity in the cortex, and movement of water molecules in the extracellular space might be a more variable indicator of tissue damage.