bluuebunny/crossref_metadata_2025 huggingface dataset:
Endothelial vasodilator production by uterine and systemic arteries. VI. Ovarian and pregnancy effects on eNOS and NOx
Authors: Magness Ronald R. , Sullivan Jeremy A. , Li Yun , Phernetton Terrance M. , Bird Ian M.
Published: December 2017

Normal pregnancy and the follicular phase of the ovarian cycle are both estrogen-dominated physiological states that are characterized by elevations in uterine blood flow and endothelial nitric oxide synthase (eNOS) protein expression in the uterine artery (UA) endothelium. It is unknown if elevations in mRNA level account for the changes in protein or eNOS activity. We tested the hypothesis that pregnancy and the follicular phase are associated with increases in eNOS mRNA and the consequent elevated expression of eNOS protein results in increased circulating nitric oxide (NO) levels. UA were obtained from pregnant (PREG; n = 8; 110–130 days gestation; term = 145 ± 3 days), nonpregnant luteal (LUT; n = 6), nonpregnant follicular (FOL; n= 6), and nonpregnant ovariectomized (OVEX; n = 6) sheep. Circulating NO levels were analyzed as total NO2-NO3(NOx). Western analysis performed on UA endothelial-isolated proteins demonstrated that eNOS protein levels were OVEX = LUT ≤ FOL < PREG ( P < 0.05), whereas eNOS mRNA expression (RT-PCR) in UA endothelial cells obtained by limited collagenase digestion was OVEX < LUT < FOL < PREG ( P < 0.05). Pregnancy dramatically elevated eNOS protein (4.1- to 6.9-fold) and mRNA (2.4- to 6.9-fold) over LUT controls ( P < 0.01). Circulating NOxlevels were not altered by ovariectomy or the ovarian cycle but were elevated from 4.4 ± 1.1 μM in LUT to 12 ± 4, 22 ± 3, and 41 ± 3 μM at 110, 120, and 130 days gestation ( P < 0.01). Systemic NOxlevels in singleton (12.5 ± 1.6 μM) were less ( P < 0.01) than in multiple (twin 27.6 ± 6.5 μM; triplet = 46 ± 10 μM) pregnancies. Therefore, the follicular phase and, to a much greater extent, pregnancy are associated with elevations in UA endothelium-derived eNOS expression, although significant increases in systemic NOxlevels were only observed in the PREG group (multiple > singleton). Thus, although UA endothelial increases in eNOS protein and mRNA levels are associated with high estrogen states, increases in local UA NO production may require additional eNOS protein activation to play its important role in the maintenance of uterine blood flow in pregnancy.
Role of leukocyte accumulation and oxygen radicals in ischemia-reperfusion-induced injury in skeletal muscle
Authors: Schlag M. G. , Harris K. A. , Potter R. F.
Published: December 2017

The role of leukocytes and nonleukocyte-derived reactive oxygen metabolites (ROMs) in reperfusion-induced skeletal muscle injury was determined. Male rats received 2 h no-flow hindlimb ischemia-reperfusion (I/R, n = 6) or were rendered neutropenic via antineutrophil serum (ANS) before I/R (I/R + ANS, n = 5). Oxygen radicals in the absence of neutrophils were tested by administration of dimethylthiourea (DMTU) (I/R + ANS + DMTU, n = 5). Perfused capillaries (CDper) and rolling (Lr), adherent (La), and extravasated leukocytes (Le) in the extensor digitorum longus muscle were measured every 15 min during 90 min of reperfusion using intravital microscopy. The vital dyes bisbenzimide (BB) and ethidium bromide (EB) provided direct measures of tissue injury (EB/BB). CDper decreased immediately on reperfusion in the I/R and I/R + ANS groups. CDper in the I/R + ANS + DMTU group remained at baseline throughout reperfusion. La increased in the I/R group; however, EB/BB was the same between I/R and I/R + ANS groups. Injury in the I/R + ANS + DMTU group did not differ from other groups ≥60 min, after which EB/BB became significantly lower. Le did not differ between groups and was highly correlated to tissue injury. The results suggest that Le lead to parenchymal injury, and ROMs lead to perfusion deficits during the early reperfusion period after ischemia.
Sodium-induced rise in blood pressure is suppressed by androgen receptor blockade
Authors: Caplea Ann , Seachrist Darcie , Dunphy Gail , Ely Daniel
Published: December 2017

Our objective was to test the hypothesis that 1) a high Na (HNa, 3%) diet would increase blood pressure (BP) in male Wistar-Kyoto (WKY) and spontaneously hypertensive Y chromosome (SHR/y) rat strains in a territorial colony; 2) sympathetic nervous system (SNS) blockade using clonidine would lower BP on a HNa diet; and 3) prepubertal androgen receptor blockade with flutamide would lower BP on a HNa diet. A 2 × 4 factorial design used rat strains (WKY, SHR/y) and treatment [0.3% normal Na (NNa), 3% HNa, HNa/clonidine, and HNa/flutamide]. BP increased in both strains on the HNa diet ( P < 0.0001). There was no significant decrease in BP in either strain with clonidine treatment. Androgen receptor blockade with flutamide significantly decreased BP in both strains ( P < 0.0001) and normalized BP in the SHR/y colony. Neither heart rate nor activity could explain these BP differences. In conclusion, a Na sensitivity was observed in both strains, which was reduced to normotensive values by androgen blockade but not by SNS blockade.
Regulation of extracellular volume and interstitial fluid pressure in rat bone marrow
Authors: Iversen Per Ole , Berggreen Ellen , Nicolaysen Gunnar , Heyeraas Karin
Published: December 2017

The volume and fluid pressure characteristics of the intact bone marrow is incompletely understood. We used microspheres and lipoproteins for measurements of intravascular volume (IVV) and EDTA for interstitial fluid volume (IFV) within the rat bone marrow. Interstitial fluid pressure (IFP) was determined with micropipettes connected to a servo-controlled counter-pressure system. Both the microspheres and the lipoproteins yielded estimates of IVV of ∼1 ml/100 g. After a brief reactive hyperemia, IVV increased to 2.5 ml/100 g, whereas IFV decreased with ∼1.5 ml/100 g, so that total extracellular volume did not change. Baseline bone marrow IFP was 9.7 mmHg. The hyperemia led to a transient twofold increase in IFP, whereas a marked blood loss decreased IFP by almost one-half. These novel data suggest that extracellular volume and IFP within the bone marrow can be measured with tracer methods and the micropuncture technique. The responses of IVV, IFV, and IFP during changes in blood flow to the bone marrow suggest a tight regulation and are thus compatible with those for a low-compliant tissue.
The role of ANG II and endothelin-1 in exercise-induced diastolic dysfunction in heart failure
Authors: Cheng Che-Ping , Ukai Tomohiko , Onishi Katsuya , Ohte Nobuyuki , Suzuki Makoto , Zhang Zhu-Shan , Cheng Heng-Jie , Tachibana Hideo , Igawa Akihiko , Little William C.
Published: December 2017

The diastolic dysfunction present at rest in congestive heart failure (CHF) is exacerbated during exercise (Ex). Increases in circulating ANG II and endothelin-1 (ET-1) during Ex may contribute to this response. We assessed the effect of Ex on circulating plasma levels of ANG II and ET-1 and left ventricular (LV) dynamics before and after pacing-induced CHF at rest and during Ex in nine conscious, instrumented dogs. Before CHF, there were modest increases in circulating levels of ANG II (but not ET-1) during Ex. LV diastolic performance was enhanced during Ex with decreases in the time constant of LV relaxation (τ), LV end-systolic volume (VES), and LV minimum pressure with a downward shift of the LV early diastolic portion of the pressure-volume (P-V) loop. This produced an increase in peak LV filling rate without an increase in mean left atrial (LA) pressure. After CHF, the resting values of ANG II and ET-1 were elevated and increased to very high levels during Ex. After CHF, mean LA pressure, τ, and LV minimum pressure were elevated at rest and increased further during Ex. Treatment with L-754,142, a potent ET-1 antagonist, or losartan, an ANG II AT1-receptor blocker, decreased these abnormal Ex responses in CHF more effectively than an equally vasodilatory dose of sodium nitroprusside. Combined treatment with both ANG II- and ET-1-receptor blockers was more effective than either agent alone. We conclude that in CHF, circulating ANG II and ET-1 increase to very high levels during Ex and exacerbate the diastolic dysfunction present at rest.
Lack of muscarinic regulation of Ca2+channels in Gi2α gene knockout mouse hearts
Authors: Chen Fuhua , Spicher Karsten , Jiang Meisheng , Birnbaumer Lutz , Wetzel Glenn T.
Published: December 2017

The purpose of the present study was to examine the role of Gi2α in Ca2+channel regulation using Gi2α gene knockout mouse ventricular myocytes. The whole cell voltage-clamp technique was used to study the effects of the muscarinic agonist carbachol (CCh) and the β-adrenergic agonist isoproterenol (Iso) on cardiac L-type Ca2+currents in both 129Sv wild-type (WT) and Gi2α gene knockout (Gi2α−/−) mice. Perfusion with CCh significantly inhibited the Ca2+current in WT cells, and this effect was reversed by adding atropine to the CCh-containing solution. In contrast, CCh did not affect Ca2+currents in Gi2α−/− ventricular myocytes. Addition of CCh to Iso-containing solutions attenuated the Iso-stimulated Ca2+current in WT cardiomyocytes but not in Gi2α−/− cells. These findings demonstrate that, whereas the Iso-Gsα signal pathway is intact in Gi2α gene knockout mouse hearts, these cells lack the inhibitory regulation of Ca2+channels by CCh. Therefore, Gi2α is necessary for the muscarinic regulation of Ca2+channels in the mouse heart. Further studies are needed to delineate the possible interaction of Giand other cell signaling proteins and to clarify the level of interaction of G protein-coupled regulation of L-type Ca2+current in the heart.
Receptor subtypes mediating adenosine-induced dilation of cerebral arterioles
Authors: Ngai Al C. , Coyne Ellicia F. , Meno Joseph R. , West G. Alexander , Winn H. Richard
Published: December 2017

The purpose of this study was to investigate the receptor subtypes that mediate the dilation of rat intracerebral arterioles elicited by adenosine. Penetrating arterioles were isolated from the rat brain, cannulated with the use of a micropipette system, and luminally pressurized to 60 mmHg. Both adenosine and the A2A receptor-selective agonist CGS-21680 induced dose-dependent vasodilation (−logEC50: 6.5 ± 0.2 and 8.6 ± 0.3, respectively). However, adenosine, which is capable of activating both A2A and A2B receptors, caused a greater maximal dilation than CGS-21680. The A2Areceptor-selective antagonist ZM-241385 (0.1 μM) only partially inhibited the dilation induced by adenosine but almost completely blocked CGS-21680-induced dilation. Neither 8-cyclopentyl-1,3-dipropylxanthine (0.1 μM), an A1receptor-selective antagonist, nor MRS-1191 (0.1 μM), an A3 receptor-selective antagonist, attenuated adenosine dose responses. Moreover, ZM-241385 had no effect on the dilation induced by ATP (10 μM) or acidic (pH 6.8) buffer. We concluded that the A2A receptor subtype mediates adenosine-induced dilation of intracerebral arterioles in the rat brain. Furthermore, our results suggest that A2B receptors may also participate in the dilation response to adenosine.
Neutrophils are primary source of O2radicals during reperfusion after prolonged myocardial ischemia
Authors: Duilio Carlo , Ambrosio Giuseppe , Kuppusamy Periannan , DiPaula Anthony , Becker Lewis C. , Zweier Jay L.
Published: December 2017

Although many studies document oxygen radical formation during ischemia-reperfusion, few address the sources of radicals in vivo or examine radical generation in the context of prolonged ischemia. In particular, the contribution of activated neutrophils remains unclear. To investigate this issue, we developed a methodology to detect radicals without interfering with blood-borne mechanisms of radical generation. Dogs underwent aorta and coronary sinus catheterization. No chemicals were infused; instead, blood was drawn into syringes prefilled with a spin trap and analyzed by electron paramagnetic resonance spectroscopy. After 90 min of coronary artery occlusion, transcardiac concentration of oxygen radicals rose severalfold 10 min after reflow and remained significantly elevated for at least 1 h. Radicals were mostly derived from neutrophils, as shown by marked reduction after the administration of 1) neutrophil NADPH oxidase inhibitors and 2) a monoclonal antibody (R15.7) against neutrophil CD18 adhesion molecule. Reduction of radical generation by R15.7 was also associated with a significantly smaller infarct size and no-reflow areas. Thus our data demonstrate that neutrophils are a major source of oxidants in hearts reperfused in vivo after prolonged ischemia and that antineutrophil interventions can effectively prevent the increase in oxygen radical concentration during reperfusion.
Aging, oxidative responses, and proliferative capacity in cultured mouse aortic smooth muscle cells
Authors: Moon Sung-Kwon , Thompson Larry J. , Madamanchi Nageswara , Ballinger Scott , Papaconstantinou John , Horaist Chris , Runge Marschall S. , Patterson Cam
Published: December 2017

The cellular mechanisms that contribute to the acceleration of atherosclerosis in aging populations are poorly understood, although it is hypothesized that changes in the proliferative capacity of vascular smooth muscle cells is contributory. We addressed the relationship among aging, generation of reactive oxygen species (ROS), and proliferation in primary culture smooth muscle cells (SMC) derived from the aortas of young (4 mo old) and aged (16 mo old) mice to understand the phenotypic modulation of these cells as aging occurs. SMC from aged mice had decreased proliferative capacity in response to α-thrombin stimulation, yet generated higher levels of ROS and had constitutively increased mitogen-activated protein kinase activity, in comparison with cells from younger mice. These effects may be explained by dysregulation of cell cycle-associated proteins such as cyclin D1 and p27Kip1 in SMC from aged mice. Increased ROS generation was associated with decreased endogenous antioxidant activity, increased lipid peroxidation, and mitochondrial DNA damage. Accrual of oxidant-induced damage and decreased proliferative capacity in SMC may explain, in part, the age-associated transition to plaque instability in humans with atherosclerosis.
Erythrocyte deformability is a nitric oxide-mediated factor in decreased capillary density during sepsis
Authors: Bateman Ryon M. , Jagger Justin E. , Sharpe Michael D. , Ellsworth Mary L. , Mehta Sanjay , Ellis Christopher G.
Published: December 2017

Erythrocyte deformability has been recognized as a determinant of microvascular perfusion. Because nitric oxide (NO) is implicated in the modulation of red blood cell (RBC) deformability and NO levels increase during sepsis, we tested the hypothesis that a NO-mediated decrease in RBC deformability contributes to decreased functional capillary density (CD) in remote organs. With the use of a peritonitis model of sepsis in the rat [cecal ligation and perforation (CLP)] and aminoguanidine (AG) to prevent increases in NO, we measured CD in skeletal muscle (intravital microscopy), mean erythrocyte membrane deformability ([Formula: see text]; micropipette aspiration), systemic NO production [plasma nitrite/nitrate (NOx) chemiluminescence], and NO accumulation in RBC [NO bound to hemoglobin (HbNO) detected by electron paramagnetic resonance spectroscopy]. In untreated CLP animals relative to sham, NOx increased 254% ( P < 0.05), stopped flow capillaries increased 149% ( P < 0.05), and [Formula: see text] decreased 12.7% ( P < 0.05), with a subpopulation (5%) of RBC with deformabilities below the normal range. AG prevented increases in NOx, accumulation of HbNO, and decreases in both [Formula: see text]and functional CD. We found no evidence of leukocyte plugging postcapillary venules. Our findings suggest that decreased functional CD during sepsis resulted from a NO-mediated decrease in erythrocyte deformability.
Renal sympathetic nerve regulation to heating is altered in rats with heart failure
Authors: Kenney Michael J. , Musch Timothy I. , Weiss Mark L.
Published: December 2017

Heart failure (HF) alters the regulation of basal sympathetic nerve discharge (SND); however, the effect of HF on SND responses to acute stress is not well established. In the present study, renal SND responses to hyperthermia were determined in chloralose-anesthetized HF rats and in sham controls. Whole body heating (colonic temperature increased from 38 to 41°C) was used as an acute stressor because increased internal body temperature provides a potent stimulus to the sympathetic nervous system. Left ventricular end-diastolic pressure and the right ventricular wt-to-body wt ratio were increased ( P < 0.05) in HF compared with sham rats. The following observations were made: 1) renal sympathoexcitatory responses to heating were significantly reduced in HF compared with sham rats, 2) renal blood flow remained unchanged from control levels during heating in HF rats but was significantly reduced in sham rats, and 3) renal SND responses to heating were significantly higher in HF rats with bilateral lesions of the hypothalamic paraventricular nucleus (PVN) compared with sham PVN-lesioned HF rats. These results demonstrate a marked attenuation in the responsiveness of renal SND to heating in HF rats and suggest that HF alters the organization of neural pathways mediating SND responses to heating.
Changes in collagenase and collagen gene expression after induction of aortocaval fistula in rats
Authors: Dolgilevich S. M. , Siri F. M. , Atlas S. A. , Eng C.
Published: December 2017

Progressive ventricular dilatation commonly accompanies the transition to overt failure in chronically overloaded hearts; however, only recently have studies begun to elucidate underlying molecular alterations. In particular, the potential role of altered myocardial expression of the procollagenase gene in this process has not previously been examined. Biventricular hypertrophy and dilatation were produced in rats by creating an abdominal aortocaval fistula. The time courses of changes in expression of collagen I and III genes and of the procollagenase gene (matrix metalloproteinase-1, MMP-1) were assessed by Northern blot hybridization. Expression of all three genes increased promptly; however, collagenase gene expression peaked much earlier (8 h) than did expression of either of the collagen genes (7 days), and all returned to baseline levels by 45 days. These data corroborate earlier reports of increased collagen gene expression in this model, but more importantly, they provide the first evidence of concurrent activation of collagenase gene expression, suggesting that enhancement of collagen degradation may be a prerequisite for structural cardiac dilatation.
Glucocorticoid modulation of protein phosphorylation and sarcoplasmic reticulum function in rat myocardium
Authors: Rao M. K. , Xu A. , Narayanan N.
Published: December 2017

To decipher the mechanism(s) underlying glucocorticoid action on cardiac contractile function, this study investigated the effects of adrenalectomy and dexamethasone treatment on the contents of sarcoplasmic reticulum (SR) Ca2+-cycling proteins, their phosphorylation by endogenous Ca2+/calmodulin-dependent protein kinase II (CaM kinase II), and SR Ca2+sequestration in the rat myocardium. Cardiac SR vesicles from adrenalectomized rats displayed significantly diminished rates of ATP-energized Ca2+uptake in vitro compared with cardiac SR vesicles from control rats; in vivo administration of dexamethasone to adrenalectomized rats prevented the decline in SR function. Western immunoblotting analysis showed that the relative protein amounts of ryanodine receptor/Ca2+-release channel, Ca2+-ATPase, calsequestrin, and phospholamban were neither diminished significantly by adrenalectomy nor elevated by dexamethasone treatment. However, the relative amount of SR-associated CaM kinase II protein was increased 2.5- to 4-fold in dexamethasone-treated rats compared with control and adrenalectomized rats. Endogenous CaM kinase II activity, as judged from phosphorylation of ryanodine receptor, Ca2+-ATPase, and phospholamban protein, was also significantly higher (50–80% increase) in the dexamethasone-treated rats. The stimulatory effect of CaM kinase II activation on Ca2+uptake activity of SR was significantly depressed after adrenalectomy and greatly enhanced after dexamethasone treatment. These findings identify the SR as a major target for glucocorticoid actions in the heart and implicate modification of the SR CaM kinase II system as a component of the mechanisms by which dexamethasone influences SR Ca2+-cycling and myocardial contraction.
P2 purinergic receptor activation enhances cardiac contractility in isolated rat and mouse hearts
Authors: Mei Qibing , Liang Bruce T.
Published: December 2017

Activation of P2 purinergic receptors exerts a potent positive inotropic effect in the cardiac myocyte. However, it is unknown whether its activation can also cause an increased contractility in intact heart. With the use of isolated rat and mouse hearts, the objective of the present study was to investigate the effect of P2 receptor agonist on the function of the intact heart. In both Langendorff rat hearts and working rat and mouse heart models, the P2X receptor agonist 2-methylthio-ATP (2-meSATP) caused dose-dependent increases in left ventricular developed pressure, rate of contraction, and rate of relaxation. The extent of P2X receptor agonist-stimulated increase in contractility was significantly less than that stimulated by the β-adrenergic agonist isoproterenol. However, the increase in contractility occurred without a significant effect on the basal heart rate, in contrast to that caused by isoproterenol. In isolated rat ventricular myocytes, both ATP and the P2X receptor agonist 2-meSATP stimulated large increases in the myocyte contractile amplitude (107 ± 13% and 99 ± 9%, n = 17 cells from 5 rats and n = 19 cells from 6 rats, respectively). 2-meSATP caused only a slight increase in phospholipase C activity and could stimulate myocyte contractility in the presence of phospholipase C inhibitor U-73122, consistent with the role of a phospholipase C-independent P2X receptor in mediating the positive inotropic effect of 2-meSATP. The data provide evidence for a potentially important physiological role of the cardiac P2X receptor and for the concept that agonist at this receptor may be beneficial for the treatment of cardiac dysfunction.
Frequency dependence of endocytosis in aortic baroreceptor neurons and role of group III mGluRs
Authors: Pamidimukkala Jaya , Hay Meredith
Published: December 2017

Synaptic transmission between baroreceptor afferents and the nucleus tractus solitarius (NTS) is known to exhibit frequency-dependent depression. Reductions in neurotransmitter release and alterations in mechanisms regulating synaptic transmission are hypothesized to be involved in the activity-dependent depression observed in baroreceptor afferent neurons. The present study utilized cultured aortic baroreceptor neurons and the fluorescent dyes FM1-43 and FM2-10 to characterize the process of endocytosis or vesicle retrieval and its dependence on 1) frequency of neuronal activation, 2) metabotropic glutamate receptor (mGluR) activation, and 3) calcium concentrations inside and outside the cell. Endocytosis per spike, measured in fluorescence units after a 10-s stimulus applied at frequencies of 0.5 (53 ± 4), 1.0 (23 ± 1), and 10.0 Hz (2.7 ± 0.2), was significantly depressed at higher frequencies. Blockade of group III mGluRs with ( RS)-cyclopropyl-4-phosphonophenylglycine (CPPG) facilitated endocytosis at all frequencies, suggesting that this receptor subtype may be involved in the inhibition of endocytosis. Manipulating the extracellular and intracellular calcium concentrations subsequent to exocytosis had no effect on endocytosis. These results suggest that frequency-dependent depression of endocytosis observed in vitro could contribute to the frequency-dependent depression of baroreceptor afferent neurotransmission and that group III mGluRs inhibit endocytosis.
Inducible HSP70 mediates delayed cardioprotection via U-50488H pretreatment in rat ventricular myocytes
Authors: Zhou Jing-Jun , Pei Jian-Ming , Wang Guan-Ying , Wu Song , Wang Wei-Ping , Cho Chi-Hin , Wong Tak-Ming
Published: December 2017

To test the hypothesis that heat-shock proteins (HSPs) mediate delayed cardioprotection of prior κ-opioid receptor (κ-OR) stimulation, we first correlated cellular injury and viability with the expression of HSP70s in isolated rat ventricular myocytes subjected to prior κ-OR stimulation with the selective agonist trans-(±)-3,4-dichloro- N-methyl- N-[2-(1-pyrrolidinyl)cyclohexyl]benzeneacetamide (U-50488H) and delayed lethal simulated ischemia (LSI). Cell injury and viability were indicated by lactate dehydrogenase release and trypan blue exclusion, respectively. The reduced injury and increased viability after pretreatment with U-50488H were concentration dependent and correlated directly with the expression of both stress-inducible (HSP70) and constitutive (HSC70) proteins. The effects mimic those with metabolic inhibition preconditioning (MIP). The cardioprotection against LSI by pretreatment with U-50488H and MIP was abolished and antagonized, respectively, via blockade of the κ-OR by its selective antagonist, nor-binaltorphimine. We also found that blockade of the production of HSP70 but not HSC70 blocked the inhibitory effect of pretreatment with U-50488H on injury and viability. These observations provide evidence that stress-inducible HSP70 mediates delayed cardioprotection of prior κ-OR stimulation.
Effects on sympathetic activity of 8-OHDPAT and clonidine in cat medullary lateral tegmental field
Authors: Orer Hakan S. , Barman Susan M. , Gebber Gerard L.
Published: December 2017

This study was designed to test the hypothesis that 8-hydroxy-2-(di- n-propylamino)tetralin (8-OHDPAT) and clonidine reduce sympathetic nerve discharge (SND) and mean arterial pressure (MAP), in part by actions in the medullary lateral tegmental field (LTF). We microinjected these drugs bilaterally into the LTF of baroreceptor-innervated and -denervated cats anesthetized with Dial-urethane. Neither drug altered SND (as quantified by using power spectral analysis) or MAP when injected into the LTF of baroreceptor-denervated cats. However, cardiac-related power in SND was significantly increased to 148 ± 12 (mean ± SE) and 149 ± 5% of control by microinjections of 8-OHDPAT ( n = 5) and clonidine ( n = 5), respectively, in baroreceptor-innervated cats whose MAP was kept constant; there was no change in 0- to 6-Hz power or total power. SND was significantly reduced by microinjection of these drugs into the rostral ventrolateral medulla of baroreceptor-innervated and -denervated cats. In conclusion, although 8-OHDPAT and clonidine did not reduce SND when injected into the LTF, they acted in this region to facilitate baroreceptor reflex control of SND, as evidenced by a selective increase in cardiac-related power.
Cross-sectional area and volume compliance of porcine left coronary arteries
Authors: Kassab G. S. , Molloi S.
Published: December 2017

We have determined the cross-sectional area (CSA) compliance of the first several generations of pig coronary arteries and the volume compliance of the coronary arterial tree (vessels >0.5 mm in diameter) using a videodensitometric technique. The coronary arteries of four KCl-arrested maximally vasodilated pig hearts were perfused with iodine and 3% Cab-O-Sil. Because Cab-O-Sil occludes small arteries, the flow can be stopped and the pressure can be maintained while the trunk of the coronary artery and its subbranches are imaged using digital angiography. The coronary arteries were preconditioned several times with cyclic changes in pressure from 0 to 160 mmHg. The pressure was then varied in a triangular pattern, and the absolute CSA of each vessel and the total arterial volume were calculated using videodensitometry in conjunction with digital subtraction angiography. Our results have shown that the pressure-diameter and pressure-volume relationships are linear in the 60–140 mmHg pressure range. Furthermore, the compliance of the coronary arteries is small; i.e., the diameter of the coronary artery changes by <15% in the 80-mmHg pressure range. The compliance data couples the mechanics of the blood vessel wall to the mechanics of blood flow to yield a pressure-flow relationship for each coronary arterial segment.
High-throughput assessment of calcium sensitivity in skinned cardiac myocytes
Authors: Lim Chee Chew , Helmes Michiel H. B. , Sawyer Douglas B. , Jain Mohit , Liao Ronglih
Published: December 2017

Isolated permeabilized cardiac myocytes have been used in the study of myofilament calcium sensitivity through measurement of the isometric force-pCa curve. Determining this force-pCa relationship in skinned myocytes is relatively expensive and carries a high degree of variability. We therefore attempted to establish an alternative high-throughput method to measure calcium sensitivity in cardiac myocytes. With the use of commercially available software that allows for precise measurement of sarcomere spacing, we measured sarcomere length changes in unloaded skinned cardiac myocytes over a range of calcium concentrations. With the use of this technique, we were able to accurately detect acute increases or decreases in myofilament calcium sensitivity after exposure to 10 mM caffeine or 5 mM 2,3-butanedione monoxime, respectively. This technique allows for the simple and rapid determination of myofilament calcium sensitivity in cardiac myocytes in a reproducible and inexpensive manner and could be used for high-throughput screening of pharmacological agents and/or transgenic mouse models for changes in myofilament calcium sensitivity.
LV systolic performance improves with development of hypertrophy after transverse aortic constriction in mice
Authors: Nakamura Akihiro , Rokosh D. Gregg , Paccanaro Mariemma , Yee Rupsa R. , Simpson Paul C. , Grossman William , Foster Elyse
Published: December 2017

Transverse aortic constriction (TAC) is an effective technique for inducing left ventricular (LV) hypertrophy in mice. With the use of transthoracic echocardiography and Doppler measurements, we studied the effects of an acute increase in pressure overload on LV contractile performance and peak systolic wall stress index (WSI) at early time points after TAC and the time course of the development of LV hypertrophy in mice. The LV mass index was similar between TAC and sham-operated mice at postoperative day 1 but progressively increased in TAC mice by day 10. There was no further increase in the LV mass index between postoperative days 10 and 20. On day 1, whereas peak systolic WSI increased significantly, the LV ejection fraction (LVEF) and percent fractional shortening (%FS) decreased in TAC mice compared with sham-operated mice. By day 10, peak systolic WSI, LVEF, and %FS had recovered to baseline levels and were not significantly different between postoperative days 10 and 20. Thus LV systolic performance in mice declines immediately after TAC, associated with increased peak systolic WSI, but recovers to baseline levels with the development of compensatory LV hypertrophy over 10–20 days.
Angiotensin II and VEGF are involved in angiogenesis induced by short-term exercise training
Authors: Amaral Sandra L. , Papanek Paula E. , Greene Andrew S.
Published: December 2017

Results from our laboratory have suggested a pathway involving angiotensin II type 1 (AT1) receptors and vascular endothelial growth factor (VEGF) in angiogenesis induced by electrical stimulation. The present study investigated if similar mechanisms underlie the angiogenesis induced by short-term exercise training. Seven days before training and throughout the training period, male Sprague-Dawley rats received either captopril or losartan in their drinking water. Rats underwent a 3-day treadmill training protocol. The tibialis anterior and gastrocnemius muscles were harvested under anesthesia and lightly fixed in formalin (vessel density) or frozen in liquid nitrogen (VEGF expression). In controls, treadmill training resulted in a significant increase in vessel density in all muscles studied. However, the angiogenesis induced by exercise was completely blocked by either losartan or captopril. Western blot analysis showed that VEGF expression was increased in the exercised control group, and both losartan and captopril blocked this increase. The role of VEGF was directly confirmed using a VEGF-neutralizing antibody. These results confirm the role of angiotensin II and VEGF in angiogenesis induced by exercise.
Characterization of mice with a combined suppression ofI to and I K,slow
Authors: Brunner Michael , Guo Weinong , Mitchell Gary F. , Buckett Peter D. , Nerbonne Jeanne M. , Koren Gideon
Published: December 2017

Cardiac-specific expression of a truncated Kv1.1 polypeptide (Kv1DN) attenuates the slow inactivating outward K+ current ( I K,slow), increases action potential duration (APD) and Q-T intervals, and induces spontaneous ventricular arrhythmias. Expression of the pore mutant of Kv4.2 (Kv4DN) eliminates the fast component of the transient outward current ( I to) and prolongs APDs and Q-T intervals markedly; however, no arrhythmias are seen in Kv4DN mice, suggesting that APD and Q-T prolongation are not per se proarrhythmic. To test this hypothesis, the Kv1DN and Kv4DN lines were crossbred to produce animals (Kv1/Kv4DN) expressing both transgenes in an identical genetic background. Whole cell voltage-clamp recordings from left ventricular apex cells confirmed that in Kv1/Kv4DN left ventricular apex cells, both components (fast and slow) of I to and the 4-aminopyridine-sensitive component of I K,sloware eliminated, resulting in marked APD prolongation compared with wild-type, Kv1DN, or Kv4DN cells. Telemetric electrocardiogram monitoring ( n = 10 mice/group) revealed a significant prolongation of Q-Tc and P-R intervals in Kv1/Kv4DN animals compared with Kv1DN or Kv4DN animals. Spontaneous arrhythmias were observed mainly in Kv1DN mice. Thus the attenuation of fast I to in addition to I K,slow in Kv1/Kv4DN mice causes significant prolongation of APD and Q-T intervals and attenuation of spontaneous arrhythmias.
Effects of chronic alcohol consumption on regulation of myocardial protein synthesis
Authors: Vary Thomas C. , Lynch Christopher J. , Lang Charles H.
Published: December 2017

Heart disease represents an important etiology of mortality in chronic alcoholics. The purpose of the present study was to examine potential mechanisms for the inhibitory effect of chronic alcohol exposure (16 wk) on the regulation of myocardial protein metabolism. Chronic alcohol feeding resulted in a lower heart weight and 25% loss of cardiac protein per heart compared with pair-fed controls. The loss of protein mass resulted in part from a diminished (30%) rate of protein synthesis. Ethanol exerted its inhibition of protein synthesis through diminished translational efficiency rather than lower RNA content. Chronic ethanol administration decreased the abundance of eukaryotic initiation factor (eIF)4G associated with eIF4E in the myocardium by 36% and increased the abundance of the translation response protein (4E-BP1) associated with eIF4E. In addition, chronic alcohol feeding significantly reduced the extent of p70S6 kinase (p70S6K) phosphorylation. The decreases in the phosphorylation of 4E-BP1 and p70S6Kdid not result from a reduced abundance of mammalian target of rapamycin (mTOR). These data suggest that a chronic alcohol-induced impairment in myocardial protein synthesis results in part from inhibition in peptide chain initiation secondary to marked changes in eIF4E availability and p70S6Kphosphorylation.
Mitochondrial ATPase and high-energy phosphates in failing hearts
Authors: Liu Jingbo , Wang Chunsheng , Murakami Yo , Gong Guangrong , Ishibashi Yutaka , Prody Catherine , Ochiai Koichi , Bache Robert J. , Godinot Catherine , Zhang Jianyi
Published: December 2017

This study examined high-energy phosphates (HEP) and mitochondrial ATPase protein expression in hearts in which myocardial infarction resulted in either compensated left ventricular remodeling (LVR) or congestive heart failure (CHF). The response of HEP (measured via31P magnetic resonance spectroscopy) to a modest increase in the cardiac work state produced by dobutamine-dopamine infusion and pacing (if needed) was examined in 17 pigs after left circumflex coronary artery ligation (9 with LVR and 8 with CHF) and compared with 7 normal pigs. In hearts with LVR, the baseline phosphocreatine (PCr)-to-ATP ratio decreased, and calculated ADP increased; these changes were most severe in hearts with CHF. HEP levels did not change in normal or LVR hearts during dobutamine-dopamine infusion. However, in hearts with CHF, the PCr-to-ATP ratio decreased further, and free ADP increased. The mitochondrial protein levels of the F0F1-ATPase subunits were normal in hearts with compensated LVR. However, in failing hearts, the α-subunit decreased by 36%, the β-subunit decreased by 16%, the oligomycin sensitivity-conferring protein subunit decreased by 40%, and the initiation factor 1 subunit decreased by 41%. Thus in failing hearts, reductions in mitochondrial F0F1-ATPase protein expression are associated with increased myocardial free ADP.
Temporal effects of 17β-estradiol on caveolin-1 mRNA and protein in bovine aortic endothelial cells
Authors: Jayachandran Muthuvel , Hayashi Toshio , Sumi Daigo , Iguchi Akihisa , Miller Virginia M.
Published: December 2017

Endothelial nitric oxide synthase (eNOS) is regulated both by caveolin-1 and 17β-estradiol (E2). Temporal relationships between effects of estrogen on caveolin-1 and nitric oxide (NO) are not known. Therefore, this study was designed to determine whether estrogen regulates caveolin-1 and, if so, whether such regulation corresponds to changes in nitrite/nitrate (NOx) production. Bovine aortic endothelial cells (BAECs) were cultured in the absence and presence of 17β-estradiol or 17α-estradiol (10−8and 10−10M) for 12, 24, and 48 h. eNOS protein expression and NOxproduction increased significantly after 24 h but not after 12-h treatment with 17β- and not 17α-estradiol. Both mRNA and protein for caveolin-1 were increased significantly only after 48-h treatment with E2, but eNOS protein and NOxproduction were decreased compared with cells treated for 24 h. These increases in caveolin-1 were inhibited by the estrogen receptor antagonist ICI-182,780 (10−6M). Results of this study suggest that E2stimulates caveolin-1 transcription and translation through estrogen receptor-mediated mechanisms. The results further suggest that estrogen may indirectly regulate NOxthrough caveolin-1 expression, which inhibits eNOS catalytic activity.
A novel method to study contraction characteristics of a single cardiac myocyte using carbon fibers
Authors: Yasuda So-Ichiro , Sugiura Seiryo , Kobayakawa Naoshi , Fujita Hideo , Yamashita Hiroshi , Katoh Kaoru , Saeki Yasutake , Kaneko Hiroko , Suda Yoshihisa , Nagai Ryozo , Sugi Haruo
Published: December 2017

To facilitate cardiac muscle research, we developed a novel method by which the force and length of a single ventricular myocyte can be recorded with a pair of carbon graphite fibers attached firmly to both ends. One fiber was stiff, whereas the other fiber was compliant to allow the recording of force and shortening during twitch contractions. The image of the compliant carbon fiber was projected onto a pair of photodiodes, and their output was fed to a piezoelectric transducer after variable amplifications to alter the effective compliance of the carbon fiber. Thus contraction of the myocyte was induced under virtually isometric conditions as well as under auxotonic conditions. We obtained a bell-shaped relation between the compliance under an auxotonic load and the work output of the myocyte, which was directly related to myocyte performance in the heart. Because it is easy to attach myocytes to the experimental apparatus, the present method would allow us to study cardiac muscle mechanics at the cellular and molecular levels.
Myocardial and interstitial matrix metalloproteinase activity after acute myocardial infarction in pigs
Authors: Etoh Takuma , Joffs Cassandra , Deschamps Anne M. , Davis Jennifer , Dowdy Kathryn , Hendrick Jennifer , Baicu Simona , Mukherjee Rupak , Manhaini Marlina , Spinale Francis G.
Published: December 2017

A structural event during the evolution of a myocardial infarction (MI) is left ventricular (LV) remodeling. The mechanisms that contribute to early changes in LV myocardial remodeling in the post-MI period remain poorly understood. Matrix metalloproteinases (MMPs) contribute to tissue remodeling in several disease states. Whether and to what degree MMP activation occurs within the myocardial interstitium after acute MI remains to be determined. Adult pigs ( n = 15) were instrumented to measure regional myocardial function and interstitial MMP levels within regions served by the circumflex and left anterior descending arteries. Regional function was measured by sonomicrometry, and interstitial MMP levels were determined by selective microdialysis and zymography as well as by MMP interstitial fluorogenic activity. Measurements were performed at baseline and sequentially for up to 3 h after ligation of the obtuse marginals of the circumflex artery. Regional fractional shortening fell by over 50% in the MI region but remained unchanged in the remote region after coronary occlusion. Release of soluble MMPs, as revealed by zymographic activity of myocardial interstitial samples, increased by 2 h post-MI. The increased zymographic activity after MI was consistent with MMP-9. Myocardial interstitial MMP fluorogenic activity became detectable within the ischemic region as early as 10 min after coronary occlusion and significantly increased after 1 h post-MI. MMP fluorogenic activity remained unchanged from baseline values in the remote region. The present study demonstrated that myocardial MMP activation can occur within the MI region in the absence of reperfusion. These unique results suggest that MMP release and activation occurs within the ischemic myocardial interstitium in the early post-MI period.
Blunted nitric oxide-mediated inhibition of renal nerve discharge within PVN of rats with heart failure
Authors: Zhang Kun , Li Yi-Fan , Patel Kaushik P.
Published: December 2017

We have demonstrated a decreased neuronal nitric oxide (NO) synthase (nNOS) message in the hypothalamus of rats with heart failure (HF). Subsequently, we have demonstrated that NADPH diaphorase (a commonly used marker for nNOS activity) positive neurons are decreased in paraventricular nucleus (PVN) of rats with coronary artery ligation model of HF. The goal of the present study was to examine the influence of endogenous NO within the PVN on renal sympathetic nerve discharge (RSND) during HF. In α-chloralose- and urethane-anesthetized rats, an inhibitor of NO synthase, N G-monomethyl-l-arginine (l-NMMA) microinjected into the PVN (50, 100, and 200 pmol in 50–200 nl) produced a dose-dependent increase in RSND, blood pressure, and heart rate in control and HF rats. These responses were attenuated in rats with HF compared with control rats. On the other hand, the NO agonist, sodium nitroprusside, microinjected in PVN produced a dose-dependent decrease in RSND and blood pressure in control and HF rats. These responses were less in rats with HF compared with control rats. These data suggest that the endogenous NO-mediated effect within the PVN of HF rats is less potent in suppressing RSND compared with control rats. These data support the conclusion that the NO system within the PVN involved in controlling autonomic outflow is altered during HF and may contribute to the elevated levels of renal sympathoexcitation commonly observed in HF.
Adenylyl cyclase isoforms and signal integration in models of vascular smooth muscle cells
Authors: Webb Jerry G. , Yates Phillip W. , Yang Qing , Mukhin Yurii V. , Lanier Stephen M.
Published: December 2017

Adenylyl cyclases present a potential focal point for signal integration in vascular smooth muscle cells (VSMC) influencing contractile state and cellular responses to vessel wall injury. In the present study, we examined the influence of the vasoactive peptide arginine vasopressin (AVP) on cAMP regulation in primary cultures of rat aortic VSMC and in the A7r5 arterial smooth muscle cell line. In cultured VSMC and A7r5 cells, AVP had no effect on basal cAMP but differentially affected β-adrenergic receptor-induced activation of adenylyl cyclase. AVP synergistically increased (twofold) isoproterenol-stimulated cAMP production in VSMC but inhibited the effect of isoproterenol (50%) in the A7r5 cell line. The effects of AVP in both preparations were blocked when cells were pretreated with a selective V1vasopressin receptor antagonist. Moreover, the actions of AVP in both models were dependent on release of intracellular Ca2+and were mimicked by elevation of Ca2+with the ionophore A23187 , suggesting that the responses to AVP involve Ca2+-mediated regulation of adenylyl cyclase stimulation. Adenylyl cyclase types I, III, and VIII are stimulated by Ca2+/calmodulin, whereas types V and VI are directly inhibited by Ca2+. RNA blot analysis for effector isotypes indicated that both VSMC and A7r5 cells expressed types III, V, and VI. VSMC also expressed mRNA for type IV and VIII effectors, which could account for the cell-specific responses to peptide hormone and Ca2+.
Acute hibernation decreases myocardial pyruvate carboxylation and citrate release
Authors: Panchal Ashish R. , Comte Blandine , Huang Hazel , Dudar Basil , Roth Bridgette , Chandler Margaret , Des Rosiers Christine , Brunengraber Henri , Stanley William C.
Published: December 2017

In the well-perfused heart, pyruvate carboxylation accounts for 3–6% of the citric acid cycle (CAC) flux, and CAC carbon is lost via citrate release. We investigated the effects of an acute reduction in coronary flow on these processes and on the tissue content of CAC intermediates. Measurements were made in an open-chest anesthetized swine model. Left anterior descending coronary artery blood flow was controlled by a extracorporeal perfusion circuit, and flow was decreased by 40% for 80 min to induce myocardial hibernation ( n = 8). An intracoronary infusion of [U-13C3]lactate and [U-13C3]pyruvate was given to measure the entry of pyruvate into the CAC through pyruvate carboxylation from the13C-labeled isotopomers of CAC intermediates. Compared with normal coronary flow, myocardial hibernation resulted in parallel decreases of 65% and 79% in pyruvate carboxylation and net citrate release by the myocardium, respectively, and maintenance of the CAC intermediate content. Elevation of the arterial pyruvate concentration by 1 mM had no effect. Thus a 40% decrease in coronary blood flow resulted in a concomitant decrease in pyruvate carboxylation and citrate release as well as maintenance of the CAC intermediates.