bluuebunny/crossref_metadata_2025 huggingface dataset:
Inflammatory cell-derived NO modulates cardiac allograft contractile and electrophysiological function
Authors: Worrall N. K. , Pyo R. T. , Botney M. D. , Misko T. P. , Sullivan P. M. , Alexander D. G. , Lazenby W. D. , Ferguson T. B.
Published: December 2017

We previously demonstrated that inhibition of inducible nitric oxide (NO) synthase (iNOS) ameliorated acute cardiac allograft rejection. This study used a rat cardiac transplant model to characterize contractile and electrophysiological dysfunction during early acute rejection, further examine the role of NO and iNOS in this process, and determine which cells expressed iNOS during early rejection. During early acute rejection, before significant myocyte necrosis, allograft papillary muscles had reduced tension development and rates of tension development and decline during beta-adrenergic, adenylate cyclase, and calcium stimulation compared with isograft and normals [e.g., tension of 36 (allograft) vs. 73 (isograft) mN/mm2 during calcium stimulation, P< 0.001]. Allografts had resting membrane potential depolarization and reduced action potential amplitude and upstroke velocity. iNOS mRNA was expressed in infiltrating inflammatory cells but not in allograft myocytes, endothelial cells, or isografts. Corticosteroids attenuated allograft contractile and electrophysiological dysfunction and inhibited iNOS enzyme activity. Direct iNOS inhibition with aminoguanidine inhibited NO production and prevented allograft contractile and electrophysiological dysfunction (e.g., tension of 64 mN/mm2 during calcium stimulation, P < 0.001). We conclude that 1) early allograft rejection caused contractile and electrophysiological dysfunction that was largely mediated by iNOS expression in infiltrating inflammatory cells, 2) corticosteroid-mediated amelioration of allograft contractile and electrophysiological dysfunction may reflect inhibition of iNOS, and 3) iNOS inhibition may offer an alternative in management of immune-mediated myocardial dysfunction.
Newborn piglet cerebral microvascular responses to epoxyeicosatrienoic acids
Authors: Leffler C. W. , Fedinec A. L.
Published: December 2017

The present study on the newborn pig cerebral microcirculation determined the vasoactive properties of epoxyeicosatrienoic acids (EETs) and the contributions of prostaglandin cyclooxygenase to these properties. Pial arterioles of anesthetized piglets were observed through closed cranial windows, EETs were applied topically, and artificial cerebrospinal fluid from beneath the cranial windows was collected for the determination of adenosine 3',5'-cyclic monophosphate and 6-ketoprostaglandin F1 alpha. EETs caused dilation of pial arterioles and increased adenosine 3',5'-cyclic monophosphate. 5,6-EET produced a dose-dependent dilation at 10(-8) M and above, whereas 10(-6) M was required for 8,9-EET, 11,12-EET, and 14,15-EET. Indomethacin abolished pial arteriolar dilation to the EETs. However, EETs did not increase cortical 6-ketoprostaglandin F1 alpha concentration. Treatment of indomethacin-treated piglets with iloprost (10(-12) M topically) restored dilation to 5,6-EET. Neither isoproterenol nor sodium nitroprusside allowed vasodilation to 5,6-EET in indomethacin-treated piglets. Therefore, in the newborn pig cerebral microvasculature. EETs are potent vasodilators and prostacyclin-receptor agonists are necessary to allow this dilation to occur.
Meandering and unstable reentrant wave fronts induced by acetylcholine in isolated canine right atrium
Authors: Ikeda T. , Wu T. J. , Uchida T. , Hough D. , Fishbein M. C. , Mandel W. J. , Chen P. S. , Karagueuzian H. S.
Published: December 2017

The mechanism(s) by which acetylcholine (ACh) increases atrial vulnerability to reentry and maintains its activity for longer durations remains poorly defined. In the present study we used high-resolution activation maps to test the hypothesis that ACh promotes meandering of atrial reentrant wave fronts, resulting in breakup and the generation of new wave fronts that sustain the activity. Reentry was induced in 11 isolated canine right atrial tissues (3.8 x 3.2 cm) by a premature point stimulus (S2) before and after superfusion with ACh (15 x 10(-6) M). Endocardial isochronal activation maps were constructed with the use of 509 bipolar electrodes (1.6-mm spatial resolution), and the dynamics of the activation wave fronts were visualized with animation. A vulnerable period was found during which an S2 current strength > 4.4 +/- 2.5 mA [lower limit of vulnerability (LLV)] and < 26 +/- 13 mA [upper limit of vulnerability (ULV)] induced a single stationary reentrant wave front that lasted 3 +/- 2.5 s with a period of 159 +/- 17 ms (16 episodes). AC shortened the refractory period from 100 +/- 12 to 59 +/- 9 ms (P < 0.001) and increased vulnerability to reentry induction by simultaneous decrease in the LLV (0.7 +/- 0.2 mA, P < 0.001) and an increase in the ULV (82 +/- 24 mA, P < 0.01). ACh accelerated the rate (period of 110 +/- 16 ms, P < 0.001) and converted the stationary reentrant wave front to a nonstationary (meandering) reentrant wave front showing polymorphic electrograms, i.e., “fibrillation-like” activity (22 episodes). Rapid meandering of the reentry tip led to wave front breakup (18 episodes) and the generation of new wave fronts (19 episodes). These wave front dynamics also led to sustained (76 +/- 224 s, P < 0.001) fibrillation-like electrograms. We conclude that ACh increases the ULV and promotes meandering of a single reentrant wave front, leading to breakup and the generation of new wave fronts. Single meandering and complex wave front dynamics cause fibrillation-like activity and sustain the activity for longer duration.
Diabetic human platelets release a substance that inhibits platelet-mediated vasodilation
Authors: Oskarsson H. J. , Hofmeyer T. G.
Published: December 2017

This study was performed to investigate the mechanism for impaired vasodilation in response to activated diabetic human platelets. As observed previously, diabetic platelets failed to cause vasorelaxation, whereas normal platelets produced normal vasodilation. However, when activated and perfused through quiescent, NG-nitro-L-arginine-pretreated arteries, diabetic and normal platelets caused similar degrees of vasoconstriction. Inhibition of serotonergic and thromboxane A2 receptors in preconstricted normal arteries also failed to improve vasodilatory responses to diabetic platelets. The amount of ADP released into the supernatant from activated diabetic and normal platelets was similar. Concomitant perfusion of activated diabetic platelets impaired vasodilation produced by abluminally applied acetylcholine but perfusion of normal platelets did not. Whereas activated diabetic platelets failed to produce vasodilation, supernatant from the same platelets caused normal vasorelaxation. Dimethylthiourea and Tiron, intracellular free radical scavengers, normalized the vasodilatory response to diabetic platelets, whereas superoxide dismutase, catalase, and mannitol did not. We conclude that the impaired vasorelaxation in response to activated diabetic platelets is caused by an unidentified, short-acting, platelet-derived substance(s) that interferes with the normal dilatory response.
Organ-specific endothelial cell uptake of cationic liposome-DNA complexes in mice
Authors: McLean J. W. , Fox E. A. , Baluk P. , Bolton P. B. , Haskell A. , Pearlman R. , Thurston G. , Umemoto E. Y. , McDonald D. M.
Published: December 2017

This study identified the organ and cellular distribution of cationic liposome-DNA complexes injected intravenously into CD-1 mice for gene delivery. DOTIM-cholesterol liposomes were labeled with the fluorescent dye CM-Dil and complexed with plasmid DNA encoding the chloramphenicol acetyltransferase reporter gene. The distribution of the complexes was examined in 29 organs and tissues by fluorescence, confocal, and electron microscopy from 5 min to 24 h after injection. The complexes formed clusters in blood, which were cleared within 20 min. Complexes visible by fluorescence microscopy were taken up by endothelial cells, leukocytes, and macrophages and did not leave the vasculature except in the spleen. At 5 min, the complexes formed a patchy coating on the endothelial surface, but by 4 h, they were internalized into endosomes and lysosomes in organ- and vessel-specific patterns. Uptake by capillary endothelial cells was greatest in the lung, ovary, and anterior pituitary, less in muscle and the heart, and nearly absent in the brain and pancreatic islets. In lymph nodes and intestinal Peyer's patches, the uptake was sparse in capillaries but abundant in high endothelial venules. In the liver and spleen, most of the uptake was in Kupffer cells and macrophages. Measurements of chloramphenicol acetyltransferase reporter gene expression were generally consistent with the pattern of uptake by endothelial cells. The uptake and gene expression were accompanied by a decrease in circulating leukocytes and platelets. Overall, our results showed that the complexes were internalized by endothelial cells in organ- and vessel-specific patterns that did not match any previously identified properties of the microvasculature. The unusual distribution of endothelial cell uptake may be explained by a heterogeneously distributed membrane receptor for which the complexes are ligands.
Motor responses of cultured rat cerebral vascular smooth muscle cells to intra- and extracellular pH changes
Authors: Apkon M. , Weed R. A. , Boron W. F.
Published: December 2017

Alkalizing perivascular fluid constricts, whereas acidification dilates, cerebral arterioles. It is not known whether vascular smooth muscle cells (VSMCs), endothelium, or neuronal elements sense pH changes. We hypothesized that VSMCs themselves transduce extracellular pH (pHo) changes. We examined the motor responses of cultured adult rat middle cerebral arterial VSMCs during pHo and intracellular pH (pHi) changes. Motor responses were inferred from the deformation pattern of a silicone substratum, dimethylpolysiloxane, which wrinkles as cells contract. pHi was measured with 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein. Cultured VSMCs retained motor responses to vasoconstrictors (5-hydroxytryptamine and K+), and to sodium nitroprusside, which were typical of intact arterioles. VSMCs contracted with increasing and relaxed with decreasing pHo. Hypocapnia contracted VSMCs when the pHo increased, and hypercapnia relaxed VSMCs when the pHo decreased. However, at a constant pHo, changes in PCO2 caused opposite responses despite equivalent changes in pHi. Thus VSMCs contract with increased pHo and relax with decreased pHo just as intact arterioles do. These responses do not reflect changes in pHi or PCO2. pHi changes paradoxically alter VSMC tone in the direction opposite that caused by pHo changes.
Numerical simulation of mass transfer in porous media of blood vessel walls
Authors: Huang Z. J. , Tarbell J. M.
Published: December 2017

The tunica media of a blood vessel wall is modeled as a heterogeneous medium composed of a periodic array of cylindrical smooth muscle cells and a continuous interstitial fluid phase of proteoglycan and collagen fibers. By applying Brinkman's model to describe the behavior of the interstitial flow, we obtain an analytical solution for the transmural flow field through the periodic array of smooth muscle cells in the form of a power series, making it possible to compute the convection of solutes in the interstitial phase. With reaction of solutes at the surface of smooth muscle cell membranes being treated as boundary conditions and the diffusion of species being limited to the interstitial fluid phase only, mass transfer in the media of blood vessel walls is simulated numerically using Cray supercomputers. It is found that the Sherwood number (the dimensionless mass-transfer coefficient) is not only constant for all interior smooth muscle cells but also minimally sensitive to changes of parameters controlling the relative rates of diffusion and convection in the interstitial fluid phase and the reaction on the smooth muscle cell surface. In addition, the Sherwood number is not very sensitive to changes in the volume fraction of smooth muscle cells. A homogeneous, one-dimensional model (effective-medium model) is also developed to predict the bulk concentration profile in the media, based on the equivalent properties of the effective medium derived from the heterogeneous medium. A comparison of results from the one-dimensional model and two-dimensional simulation is quite satisfactory for all practical ranges of parameters. It is also determined that, for a small molecule such as ATP, the mass transfer to the surface of smooth muscle cells is “reaction limited” as assumed previously in the literature, whereas, for a large molecule such as low-density lipoprotein, the mass transfer might not be reaction limited.
Substantial cardiac parasympathetic activity exists during heavy dynamic exercise in dogs
Authors: O’Leary Donal S. , Rossi Noreen F. , Churchill Paul C.
Published: December 2017

We investigated the extent of functional parasympathetic and sympathetic activity to the heart at rest and during mild to heavy dynamic exercise in conscious dogs. The animals were chronically instrumented to monitor mean arterial pressure (MAP), heart rate (HR), and terminal aortic blood flow (TAQ) and trained to run on a motor-driven treadmill. MAP, HR, and TAQ were monitored at rest and during steady-state dynamic exercise ranging from mild [3.2 kilometers per hour (kph), 0% grade] to heavy exercise (8 kph, 15% grade). Experiments were performed before and after blocking the effects of either the parasympathetic nerves (atropine 0.2 mg/kg iv) or sympathetic nerves (atenolol 2.0 mg/kg iv) to the heart. In addition, blood samples were taken at rest and at steady state during exercise, and plasma levels of vasopressin and renin activity were assessed. At rest and during all levels of exercise, muscarinic cholinergic receptor blockade caused a marked increase in HR over control (saline treated) levels with little effect on MAP or TAQ. β-Adrenergic receptor blockade had no significant effect on HR at rest and during mild exercise. At moderate to heavy workloads, β-receptor blockade significantly reduced MAP, HR, and TAQ and increased plasma vasopressin levels. We conclude that, even during heavy dynamic exercise, significant functional parasympathetic tone to the heart exists. Thus, over a wide range of exercise workloads, HR is under the tonic control of both sympathetic and parasympathetic nerves.
Cardiorenal epinephrine kinetics: evidence for neuronal release in the human heart
Authors: Johansson Mats , Rundqvist Bengt , Eisenhofer Graeme , Friberg Peter
Published: December 2017

There are experimental data suggesting that epinephrine (Epi) may act as a cotransmitter in sympathetic nerves, stimulating presynaptic β2-receptors and enhancing norepinephrine (NE) release. To examine neuronal Epi release, patients with congestive heart failure and hypertension and healthy subjects were examined with the isotope-dilution method. At baseline, small cardiac and renal Epi spillovers were found in patients. During intense supine exercise, cardiac NE and Epi spillovers increased concomitantly with similar magnitude, whereas no renal Epi spillover could be detected. Blockade of neuronal uptake 1 caused a consistent decrease in both cardiac and renal fractional extractions of NE and Epi. The present study demonstrates baseline cardiorenal Epi release in patients with congestive heart failure and renal Epi release in hypertensive patients. Furthermore, Epi is removed by neuronal uptake in both the heart and kidney, and cardiac Epi spillover increases during exercise. This study, in contrast to other results, provides evidence for cardiac neuronal Epi release.
Enhancement of spontaneous baroreflex by antisense c-fosoligonucleotide treatment in the NTS of the rat
Authors: Chan Julie Y. H. , Shih Cheng-Dean , Chan Samuel H. H.
Published: December 2017

We evaluated the hypothesis that basal Fos protein at the nucleus tractus solitarii (NTS), the primary terminal site for baroreceptor afferents, exerts a tonic inhibitory modulation on the spontaneous baroreceptor reflex (BRR) control machinery, which is responsible for beat-to-beat regulation of resting systemic arterial pressure (SAP). In adult male Sprague-Dawley rats anesthetized and maintained with pentobarbital sodium, microinjection bilaterally into the caudal NTS of a 15-mer antisense oligonucleotide that targets against the initiation codon of c- fosmRNA (5′-129 to 143-3′) significantly enhanced the spontaneous BRR response, as determined by transfer function analysis of SAP and heart rate signals. The same treatment also diminished baseline Fos-like immunoreactivity in the absence of acute cardiovascular perturbation. Control treatments with artificial cerebrospinal fluid, sense cDNA, or antisense oligonucleotides that either target against a different site of the c- fos mRNA (5′-135 to 149-3′) or with three mismatched nucleotides in the antisense sequence, were ineffective. These observations support the notion that, under minimal cardiovascular perturbation, basal expression of Fos protein in the NTS may represent an early step in the cascade of intracellular events that leads to long-term inhibitory modulation of beat-to-beat baroreflex control of blood pressure.
Flow- and agonist-mediated nitric oxide- and prostaglandin-dependent dilation in spinal arteries
Authors: Yashiro Yasuaki , Ohhashi Toshio
Published: December 2017

Isolated rabbit spinal resistance-sized arteries (∼100 μm in diameter and ∼3 mm long) were cannulated at both ends with glass micropipettes and perfused at constant pressure (60 mmHg). An increase of flow rate corresponding to a change of pressure gradient (ΔP) ranging from 0 to 20 mmHg produced a flow-dependent vasodilation. Treatment with 50 μM aspirin or 10 μM indomethacin produced a significant reduction of the flow-dependent vasodilation only at ΔP of 5 mmHg. In contrast, treatment with N ω-nitro-l-arginine methyl ester (l-NAME, 30 μM) produced no significant change. In the presence of 10 μM indomethacin, however, 30 μMl-NAME caused a marked decrease in the arterial diameter at ΔP of 5 mmHg, which was completely reversed with additional administration of 1 mMl-arginine. Acetylcholine (ACh) produced a dose-dependent increase in the arterial diameter. The ACh-induced vasodilation was significantly reduced by 10 μM indomethacin or 50 μM aspirin and partially suppressed by 30 μMl-NAME. Pretreatment with both indomethacin and l-NAME completely reduced the ACh-induced vasodilation. In the presence of 10 μM indomethacin, additional treatment with 1 mMl-arginine significantly reversed the l-NAME-induced inhibition of the ACh-mediated vasodilation. Endothelial removal with Triton X-100 significantly reduced the ACh-induced vasodilation. Isocarbacyclin (a stable prostaglandin I2 analogue), prostaglandin E2, and arachidonic acid caused a dose-dependent dilation in the small arteries. These findings suggest that prostanoids play a major role in the flow- or ACh-induced vasodilation in the rabbit spinal resistance-sized small arteries.
Estrogen regulates myogenic tone in pressurized cerebral arteries by enhanced basal release of nitric oxide
Authors: Skarsgard Peter , Van Breemen Cornelis , Laher Ismail
Published: December 2017

Second-order middle cerebral arteries (135.0 ± 4.6 μm ID) from male, female, ovariectomized female (no endogenous estrogen), and estrogen-treated ovariectomized female Sprague-Dawley rats were harvested and mounted in a pressure myograph. Myogenic response was recorded over a pressure range of 10–100 mmHg and was repeated in the presence of N ω-nitro-l-arginine methyl ester (l-NAME; 2 × 10−4 M), an inhibitor of nitric oxide (NO) synthase, and after endothelium removal, to examine the contribution of NO to net myogenic tone. With intact endothelium, there were no differences in myogenic tone between the groups, but in the presence of l-NAME and after endothelium removal, estrogen-exposed vessels developed significantly greater tone at high transmural pressure. There were no differences in sensitivity to sodium nitroprusside, an NO donor, or A-23187, a calcium ionophore. These results suggest an increase in basal release of NO in cerebral arteries exposed to estrogen, without change in NO sensitivity or maximally stimulated NO release.
Clearance receptors and endopeptidase: equal role in natriuretic peptide metabolism in heart failure
Authors: Rademaker Miriam Tessa , Charles Christopher John , Kosoglou Teddy , Protter Andrew A. , Espiner Eric Arnold , Nicholls Michael Gary , Richards Arthur Mark
Published: December 2017

The effects of separate and combined endopeptidase inhibition (by SCH-32615) and natriuretic peptide receptor C blockade [by C-ANP-(4—23)] on the clearance and bioactivity of atrial (ANP) and brain (BNP) natriuretic peptides was investigated in eight sheep with heart failure. SCH-32615 and C-ANP-(4—23) administered separately induced significant and proportionate dose-dependent rises in plasma ANP, BNP, and guanosine 3′,5′-cyclic monophosphate (cGMP) levels. Associated with these changes were reductions in arterial pressure, left atrial pressure, and peripheral resistance and increases in cardiac output, urine volume, sodium excretion, and creatinine clearance. SCH-32615 induced greater diuresis and natriuresis than C-ANP-(4—23). Combined administration of SCH-32615 and C-ANP-(4—23) induced greater than additive rises in plasma ANP, BNP, and cGMP concentrations, with enhanced hemodynamic effects, diuresis, and natriuresis and reduced plasma aldosterone levels. In conclusion, we find that the enzymatic and receptor clearance pathways contribute equally to the metabolism of endogenous ANP and BNP in sheep with heart failure. Combined inhibition of both degradative pathways was associated with enhanced hormonal, hemodynamic, and renal effects and may have greater potential therapeutic value than either agent separately.
Protein kinase A does not alter unloaded velocity of sarcomere shortening in skinned rat cardiac trabeculae
Authors: Janssen Paul M. L. , De Tombe Pieter P.
Published: December 2017

Whether β-adrenergic stimulation affects the cross-bridge cycling rate independently of its effect on Ca2+ handling by the cardiac myocyte is still unknown. An increase in cross-bridge cycling rate may result in increased unloaded velocity of sarcomere shortening ( V o). To test this hypothesis directly, skinned rat cardiac trabeculae were attached between a silicon strain gauge (∼3.5 kHz resonant frequency) and a fast displacement motor. V o was measured by a modified “Edman slack test” during a single maximal activation using seven to eight sarcomere-length step releases (measured by laser diffraction) ranging between 0.12 and 0.20 μm (15.0 ± 0.1°C). β-Adrenergic stimulation was mimicked by exposing the trabeculae to the catalytic subunit of protein kinase A (PKA). Treatment with PKA (3 μg/ml; 45 min) caused a significant ( P < 0.01) increase (41 ± 13%) in the Ca2+concentration required for half-maximal steady-state tension development. Neither maximum tension nor V o was affected by treatment with PKA, suggesting that β-adrenergic stimulation does not affect the rate-limiting step of cross-bridge cycling during unloaded shortening in myocardium.
Electrical properties of iridial arterioles of the rat
Authors: Hirst G. David S. , Edwards Frank R. , Gould Dianna J. , Sandow Shaun L. , Hill Caryl E.
Published: December 2017

When intracellular recordings were made from iridial arterioles, the cells had membrane potentials of about −65 mV and perivascular nerve stimulation evoked a membrane depolarization. When these cells were labeled with lucifer yellow, all cells that responded to perivascular nerve stimulation had the morphological characteristics of smooth muscle cells. Cells with the morphological characteristics of endothelial cells were never stained. When impaled with two separate recording electrodes, the smooth muscle layer was shown to form an electrical syncytium with a membrane time constant of ∼80 ms and an electrical length constant of ∼900 μm. At the ultrastructural level, areas of close apposition were frequently observed between adjacent smooth muscle cells and between adjacent endothelial cells. On the other hand, at contacts between smooth muscle and endothelial cells, the membranes characteristically had much larger separations. The observations show that individual smooth muscle cells are electrically coupled to their neighbors, but the morphological studies raise the possibility that in these arterioles the endothelial and muscle layers are electrically separate.
Ionic basis of ryanodine’s negative chronotropic effect on pacemaker cells isolated from the sinoatrial node
Authors: Li Jin , Qu Jihong , Nathan Richard D.
Published: December 2017

Spontaneous electrical activity and indo 1 fluorescence ratios were recorded simultaneously in cultured pacemaker cells isolated from the rabbit sinoatrial node. Ryanodine (10 μM) reduced the amplitude of action potential-induced intracellular Ca2+([Formula: see text]) transients by 19 ± 3%, increased the time constant for their decay by 51 ± 5%, and slowed spontaneous firing by 32 ± 3%. 1,2-Bis(2-aminophenoxy)ethane- N, N, N′, N′-tetraacetic acid (BAPTA)-acetoxymethyl ester (AM; 25 μM) inhibited the [Formula: see text] transients and slowed spontaneous firing by 28 ± 4%. Ryanodine did not alter hyperpolarization-activated or time-independent inward current, but it reduced the sum of L- and T-type Ca2+currents ( ICa,Land ICa,T) in both the presence and absence of BAPTA-AM. In contrast, ICa,Lwas unchanged by ryanodine. Slow inward current tails, presumed to be Na/Ca exchange current ( INa/Ca), were abolished by BAPTA or ryanodine. The results suggest that a decrement of ICa,T, due to reduction of the intracellular Ca2+concentration or a direct effect of ryanodine on T-type Ca2+channels, contributes to the negative chronotropic effect. Another possibility, based primarily on theory and results in other preparations, is that a reduction of INa/Ca, as a consequence of the smaller action potential-induced[Formula: see text] transients, contributes to the effect of ryanodine.
Measurement of cardiac output in small laboratory animals using recordings of blood conductivity
Authors: Vogel Johannes
Published: December 2017

No method exists which enables easy, frequent, and, at the same time, reliable cardiac output (CO) measurements in mice. To validate a simple indicator-dilution method suitable for frequent measurements of CO in small laboratory animals, a 5% glucose solution was injected as a bolus into femoral veins of mice and rats. The corresponding blood conductivity was measured continuously between an intra-aortic and a rectal electrode. The resulting conductivity-dilution curves were used to calculate CO in mice during hypervolemia and hypovolemia and in conscious as well as halothane-anesthetized mice and rats. In rats, conductivity-dilution curves and time courses of plasma glucose concentration were recorded simultaneously. Compared with CO in awake animals, CO in both species was slightly, but not significantly, reduced during halothane anesthesia. CO was significantly and gradually reduced in hypovolemic mice (up to 58 ml blood/kg body wt), whereas hypervolemia (23 ml saline/kg body wt) had no significant effect. Simultaneous recordings of conductivity-dilution curves and time courses of plasma glucose concentration yielded corresponding values of CO ( P < 0.001). Measurement of blood conductivity appears to be a reliable, simple, and convenient method for quantification of CO in small animals.
TNF-α causes reversible in vivo systemic vascular barrier dysfunction via NO-dependent and -independent mechanisms
Authors: Worrall Neil K. , Chang Kathy , Lejeune Wanda S. , Misko Thomas P. , Sullivan Patrick M. , Ferguson T. Bruce , Williamson Joseph R.
Published: December 2017

Tumor necrosis factor (TNF-α) and nitric oxide (NO) are important vasoactive mediators of septic shock. This study used a well-characterized quantitative permeation method to examine the effect of TNF-α and NO on systemic vascular barrier function in vivo, without confounding endotoxemia, hypotension, or organ damage. Our results showed 1) TNF-α reversibly increased albumin permeation in the systemic vasculature (e.g., lung, liver, brain, etc.); 2) TNF-α did not affect hemodynamics or blood flow or cause significant tissue injury; 3) pulmonary vascular barrier dysfunction was associated with increased lung water content and impaired oxygenation; 4) TNF-α caused inducible nitric oxide synthase (iNOS) mRNA expression in the lung and increased in vivo NO production; 5) selective inhibition of iNOS with aminoguanidine prevented TNF-α-induced lung and liver vascular barrier dysfunction; 6) aminoguanidine prevented increased tissue water content in TNF-α-treated lungs and improved oxygenation; and 7) nonselective inhibition of NOS with NG-monomethly-l-arginine increased vascular permeation in control lungs and caused severe lung injury in TNF-α-treated animals. We conclude that 1) TNF-α reversibly impairs vascular barrier integrity through NO-dependent and -independent mechanisms; 2) nonselective NOS inhibition increased vascular barrier dysfunction and caused severe lung injury, whereas selective inhibition of iNOS prevented impaired endothelial barrier integrity and pulmonary dysfunction; and 3) selective inhibition of iNOS may be beneficial in treating increased vascular permeability that complicates endotoxemia and cytokine immunotherapy.
Daily exercise attenuates the sympathetic component of the arterial baroreflex control of heart rate
Authors: Collins Heidi L. , Dicarlo Stephen E.
Published: December 2017

The influence of daily spontaneous running (DSR) on the sympathetic (SC) and parasympathetic components of the arterial baroreflex control of heart rate (HR) was examined in 16 female Long Evans rats [8 sedentary (SED) and 8 DSR]. After 8–9 wk of SED control or DSR, animals were chronically instrumented with arterial and venous catheters. DSR resulted in an increased heart weight-to-body weight ratio (2.71 ± 0.11 vs. 3.09 ± 0.09 g/kg) and a resting bradycardia (378 ± 6 vs. 330 ± 5 beats/min). Arterial baroreflex function was examined during ramp infusions of phenylephrine and sodium nitroprusside under the following three experimental conditions: 1) control, 2) after β1-adrenergic receptor blockade (β1-X), and 3) after muscarinic-cholinergic receptor blockade (M-X). Arterial baroreflex function parameters were compared between SED and DSR rats. In the control condition, DSR attenuated the range (182 ± 15 vs. 124 ± 18 beats/min), maximum HR (464 ± 9 vs. 394 ± 15 beats/min), and maximal gain (Gmax; 5.57 ± 0.42 vs. 3.2 ± 0.45 beats ⋅ min−1 ⋅ mmHg−1). Similarly, after M-X, DSR attenuated the range (84 ± 5 vs. 62 ± 8 beats/min), maximum HR (449 ± 11 vs. 412 ± 11 beats/min), and Gmax (2.73 ± 0.37 vs. 1.57 ± 0.32 beats ⋅ min−1 ⋅ mmHg−1). In contrast, after β1-X, DSR did not alter the range (61 ± 13 vs. 70 ± 7 beats/min), maximum HR (326 ± 9 vs. 313 ± 7 beats/min), or Gmax (3.04 ± 0.54 vs. 3.75 ± 0.52 beats ⋅ min−1 ⋅ mmHg−1). Results demonstrate that DSR attenuated the arterial baroreflex control of HR by reducing the SC.
Central gain of the cardiac sympathetic afferent reflex in dogs with heart failure
Authors: Ma Rong , Zucker Irving H. , Wang Wei
Published: December 2017

Previous studies from our laboratory have shown that the cardiac sympathetic afferent reflex is enhanced in dogs with experimental heart failure. The aim of the present study was to determine if the central gain of the cardiac sympathetic afferent reflex was also enhanced in dogs with heart failure. Fifteen dogs with pacing-induced heart failure were used in this study. Seventeen sham-operated dogs served as control. At the time of the acute experiment the dogs were anesthetized with α-chloralose. Arterial blood pressure, heart rate, and renal sympathetic nerve activity were recorded. After sinoaortic denervation and cervical vagotomy, a thoracotomy was performed in the second intercostal space. The left stellate ganglion was identified, and the left cardiac sympathetic nerves were cut. The central end of the left cardiac sympathetic nerves was placed on bipolar stimulating electrodes. The renal sympathetic nerve activity responses to electrical stimulation (30 Hz, 1 ms with varying voltages from 1 to 10 V; or 10 V, 1 ms with varying frequencies from 1 to 30 Hz) of the afferent cardiac sympathetic nerves were compared between sham and heart failure groups. Reflex renal sympathetic nerve activity responses to stimulation of the cardiac sympathetic nerves were significantly greater in the heart failure group compared with that in the sham group (21.4 ± 3.2 vs. 9.8 ± 2.9% at 10 V, 30 Hz and 27.7 ± 4.5 vs. 9.9 ± 3.4% at 30 Hz, 10 V, heart failure vs. sham group, respectively; for both relationships, P < 0.05). This enhanced central gain of the cardiac sympathetic afferent reflex in the heart failure group was significantly attenuated after intravenous and cerebroventricular injection of the angiotensin II receptor antagonist losartan (5 mg/kg iv and 0.125 mg/kg in 0.1 ml icv). These data suggest that the central gain of the cardiac sympathetic afferent reflex is enhanced in dogs with heart failure and central angiotensin II plays an important role in this enhanced response.
Postischemic endothelium-dependent vascular reactivity is preserved in adhesion molecule-deficient mice
Authors: Banda Michael A. , Lefer David J. , Granger D. Neil
Published: December 2017

Previous studies utilizing monoclonal antibodies directed against specific leukocyte-endothelial cell adhesion proteins have suggested that neutrophils mediate endothelial cell injury in a number of vascular beds after ischemia-reperfusion (I/R). In the present study, we investigated superior mesenteric artery (SMA) vascular reactivity to acetylcholine (ACh) and sodium nitroprusside (SNP) after occlusion and reperfusion in wild-type (C57BL/6) mice and in gene-targeted mice that are deficient in either CD11/CD18, intercellular adhesion molecule 1 (ICAM-1), or P-selectin. All mice were 4 wk of age, and the SMA was occluded for 45 min and then reperfused for 45 min. Segments of SMA were isolated and suspended in organ chambers and contracted with phenylephrine (10−5M), and the maximal vasorelaxation to ACh (10−6M) and SNP (10−6M) was measured. SMA from sham-operated C57BL/6 mice relaxed 83.5 ± 3.3% to ACh and 91.7 ± 3.4% to SNP. In contrast, segments of SMA from C57BL/6 mice subjected to I/R demonstrated a marked impairment in vasorelaxation to ACh (51.3 ± 4.7%, P < 0.01 vs. sham) without any impairment in the vasoreactivity to SNP (86.1 ± 5.5%). In CD11/CD18-deficient mice, SMA reactivity to ACh (84.7 ± 2.3%) and SNP (91.2 ± 2.8%) was unaffected by I/R. Similarly, SMA rings from ICAM-1-deficient mice exhibited normal vasorelaxation to ACh and SNP with maximal vasorelaxation of 83.1 ± 2.9 and 87.4 ± 3.0%, respectively. We also observed profound preservation of endothelium-dependent vasorelaxation after I/R in P-selectin-deficient mice. These findings indicate that leukocyte-endothelial cell adhesion molecule deficiency is associated with the preservation of endothelium-dependent vascular responses after I/R.
In vivo diffusion of immunoglobulin G in muscle: effects of binding, solute exclusion, and lymphatic removal
Authors: Flessner Michael F. , Lofthouse Joanne , Zakaria El Rasheid
Published: December 2017

Previously, we demonstrated that immunoglobulin G (IgG), dissolved in an isotonic solution in the peritoneal cavity, transported rapidly into the abdominal wall when the intraperitoneal (ip) pressure was >2 cmH2O. We hypothesized that this was chiefly caused by convection and that diffusion of IgG was negligible. To investigate the role of diffusion, we dialyzed rats with no pressure gradient across the abdominal wall muscle for 2 or 6 h with an ip isotonic solution containing125I-labeled IgG. At the end of the experiment, the animal was euthanized and frozen and abdominal wall tissue was processed to produce cross-sectional autoradiograms. Quantitative densitometric analysis resulted in IgG concentration profiles with far lower magnitude than profiles from experiments in which convection dominated. In other in vivo experiments, we determined the lymph flow rate to be 0.8 × 10−4ml ⋅ min−1⋅ g−1and the fraction of extravascular tissue (θs) available to the IgG to be 0.041 ± 0.001. An in vitro binding assay was used to determine the time-dependent, nonsaturable binding constant: 0.0065 min−1× duration of exposure. A non-steady-state diffusion model that included effects of θs, time-dependent binding, and lymph flow was fitted to the diffusion profile data, and the IgG diffusivity within the tissue void was estimated to be 2 × 10−7cm2/s, a value much higher than that published by other groups. We also demonstrate from our previous data that convection of IgG through tissue dominates over diffusion at ip pressures >2 cmH2O, but diffusion may not be negligible. Furthermore, nonsaturable binding must be accounted for in the interpretation of tissue protein concentration profiles.
Ceruloplasmin impairs endothelium-dependent relaxation of rabbit aorta
Authors: Cappelli-Bigazzi Maurizio , Ambrosio Giuseppe , Musci Giovanni , Battaglia Carmine , Di Patti Maria Carmela Bonaccorsi , Golino Paolo , Ragni Massimo , Chiariello Massimo , Calabrese Lilia
Published: December 2017

This study evaluated the effects of ceruloplasmin, the copper-containing blue oxidase of vertebrate plasma, on the relaxation of rabbit aortic rings after endothelial release of nitric oxide (NO). Ceruloplasmin at physiological, i.e., micromolar, concentrations inhibited relaxation of rabbit aorta induced by endothelium-dependent agonists like acetylcholine or ADP, whereas it was ineffective toward vasodilation due to direct stimulation of smooth muscle cells by nitroglycerin. The effect was reversible and specific for native, fully metalated ceruloplasmin, since relaxation was not impaired by the heat-treated or metal-depleted derivatives. A trapping mechanism, involving a direct interaction of NO or other NO-containing species (like nitrosothiols and iron-dinitrosyls) with the copper sites and/or with the free thiol of ceruloplasmin, could be safely excluded on the basis of spectroscopic and chemical analyses of the protein exposed to authentic NO, nitrosothiols, or iron-dinitrosyls. The data presented in this paper constitute the first evidence of impairment of the endothelium-dependent vasodilatation by a plasma protein and may shed some light on the still uncertain physiological role of ceruloplasmin.
Mechanism of constant contractile efficiency under cooling inotropy of myocardium: simulation
Authors: Mikane Takeshi , Araki Junichi , Kohno Kunihisa , Nakayama Yasunori , Suzuki Shunsuke , Shimizu Juichiro , Matsubara Hiromi , Hirakawa Masahisa , Takaki Miyako , Suga Hiroyuki
Published: December 2017

We have reported that, in canine hearts, cardiac cooling to 29°C enhanced left ventricular contractility but changed neither the contractile efficiency of cross-bridge (CB) cycling nor the excitation-contraction coupling energy. The mechanism of this intriguing energetics remained unknown. To get insights into this mechanism, we simulated myocardial cooling mechanoenergetics using basic Ca2+and CB kinetics. We assumed that both adenosinetriphosphatase (ATPase)-dependent sarcoplasmic reticulum (SR) Ca2+uptake and CB detachment decelerated with cooling. We also assumed that all the ATPase-independent SR Ca2+release, Ca2+binding to and dissociation from troponin, and CB attachment remained unchanged. The simulated cooling shifted the CB force-free Ca2+concentration curve to a lower Ca2+concentration, increasing the Ca2+responsiveness of CB force generation, and increased the maximum Ca2+-activated force. The simulation most importantly showed that these cooling effects combined led to a constant contractile efficiency when Ca2+uptake and CB detachment rate constants changed appropriately. This result seems to account for our experimentally observed constant contractile efficiency under cooling inotropy.
CCD imaging in cryospectrophotometric determination of microvascular oxyhemoglobin saturations
Authors: Måseide Kårstein , Rofstad Einar K.
Published: December 2017

A microspectrophotometric imaging method has been developed for localized measurements of intravascular oxyhemoglobin (HbO2) saturations in microvessels from sections of quick-frozen tissue. HbO2saturation was calculated from the absorption spectrum of red blood cells measured at five selected wavelengths in the 520- to 570-nm range. We combined the use of narrow-bandwidth interference filters and a CCD camera mounted on a microscope to obtain one gray image of the sample at each wavelength. Each pixel is a quantitative measure of transmitted light intensity from the tissue sample at that location. A linear calibration curve for blood frozen in vitro (humans and mice) and in vivo (mice) was obtained using a multicomponent analysis. Oxy- and deoxyhemoglobin were assumed to be the only hemoglobin components present. A constant term compensates for light loss due to scattering on red blood cells and ice crystals. The standard error in single measurements of HbO2saturation was 5%. The present method allows off-line analysis of the HbO2saturation distribution within a microvessel network and offers new possibilities for comparative morphological studies.
Modification of cardiac β-adrenoceptor mechanisms by H2O2
Authors: Persad Sujata , Rupp Heinz , Jindal Rashi , Arneja Jugpal , Dhalla Naranjan S.
Published: December 2017

From the role of oxidative stress in cardiac dysfunction, we investigated the effect of H2O2, an activated species of oxygen, on β-adrenoceptors, G proteins, and adenylyl cyclase activities. Rat heart membranes were incubated with different concentrations of H2O2before the biochemical parameters were measured. Both the affinity and density of β1-adrenoceptors were decreased, whereas the density of the β2-adrenoceptors was decreased and the affinity was increased by 1 mM H2O2. Time- and concentration-dependent biphasic changes in adenylyl cyclase activities in the absence or presence of isoproterenol were observed when membranes were incubated with H2O2; however, activation of the enzyme by isoproterenol was increased or unaltered. The adenylyl cyclase activities in the absence or presence of forskolin, NaF, and Gpp(NH)p were depressed by H2O2. Catalase alone or in combination with mannitol was able to significantly decrease the magnitude of alterations due to H2O2. The cholera toxin-stimulated adenylyl cyclase activity and ADP ribose labeling of Gsproteins were decreased by treatment with 1 mM H2O2, whereas Giprotein activities, as reflected by pertussis toxin-stimulation of adenylyl cyclase and ADP ribosylation, were unaltered. The Gsand Giprotein immunoreactivities, estimated by labeling with respective antibodies, indicate a decrease in binding to the 45-kDa band of Gsprotein, whereas no change in the binding of antibodies to the 52-kDa band of Gsprotein or the 40-kDa subunit of Giprotein was evident when the membranes were treated with 1 mM H2O2. These results suggest that H2O2in high concentrations may attenuate the β-adrenoceptor-linked signal transduction in the heart by changing the functions of Gsproteins and the catalytic subunit of the adenylyl cyclase enzyme.
ANF elicits phosphorylation of the cGMP phosphodiesterase in vascular smooth muscle cells
Authors: Wyatt Todd A. , Naftilan Allen J. , Francis Sharron H. , Corbin Jackie D.
Published: December 2017

Guanosine 3′,5′-cyclic monophosphate (cGMP)-binding, cGMP-specific phosphodiesterase (PDE5) is abundant in vascular smooth muscle, and this enzyme is a potent substrate for cGMP-dependent protein kinase (PKG) in vitro. Binding of cGMP to the allosteric sites of PDE5 is required for this phosphorylation to occur. Vascular smooth muscle cells (VSMC) were used to determine if PDE5 is phosphorylated in intact cells when cGMP is increased. With the use of anti-PDE5 antibodies, a phosphorylated 93-kDa protein band was immunoprecipitated from early passaged primary cultures of VSMC that had been preincubated with32Pito label cellular ATP and then treated with atrial natriuretic factor (ANF). In the absence of ANF, there was no detectable incorporation of radiolabeled phosphate into this band. Phosphorylation of the 93-kDa protein was augmented by pretreating cells with 8-bromoguanosine 3′,5′-cyclic monophosphate (8-BrcGMP) to activate PKG before addition of ANF. 8-BrcGMP, which interacts poorly with the allosteric sites of PDE5, had no effect on PDE5 phosphorylation in the absence of ANF. Phosphorylation of PDE5 in response to treatment of cells with ANF was associated with a two- to fourfold increase in PDE activity in immunoprecipitates. Multiple-passaged VSMC, which are deficient in PKG but retain PDE5, demonstrated no ANF-dependent increase in phosphorylation or catalytic activity of PDE5. However, incubation of immunoprecipitated PDE5 from these cells with purified PKG, cGMP, and a phosphorylation mixture containing [γ-32P]ATP resulted in32Piincorporation into PDE5 that was correlated with increased catalytic activity. These studies are the first to demonstrate phosphorylation of PDE5 in intact cells, thus suggesting a physiological role for this enzyme in smooth muscle regulation.
Voltage-gated sodium channels in cardiac microvascular endothelial cells
Authors: Walsh Kenneth B. , Wolf Matthew B. , Fan Jinping
Published: December 2017

The goal of this study was to determine whether inward Na+ or Ca2+ currents could be measured in cardiac microvascular endothelial cells (CMEC). CMEC were isolated from rat ventricular muscle and studied during days 1–4 in culture. Differential uptake of fluorescently labeled acetylated low-density lipoproteins (LDL) indicated that the primary culture contained >90% CMEC. Membrane currents were measured with the use of the whole cell arrangement of the patch-clamp technique with a Cs+ internal solution to prevent contamination by outward K+currents. Voltage steps positive to −30 mV resulted in the activation of a fast, inward Na+current ( I Na). In 20 cells examined, the peak inward current measured at 0 mV was 2.1 pA/pF. The half-maximal voltage required for inactivation of I Na was −45 mV, and the current recovered from inactivation with a time constant of 10 ms. Inward currents were eliminated by replacement of external sodium with N-methylglucamine and were blocked by both tetrodotoxin (TTX) (dissociation constant = 5 nM) and saxitoxin (50 nM). Stimulation of protein kinase C, through application of phorbol 12,13-dibutyrate, resulted in an increase in the amplitude of I Na without any change in the voltage dependence of current activation. Thus the endothelium of cardiac microvessels may be unique in expressing voltage gated, TTX-sensitive Na+ channels.