bluuebunny/crossref_metadata_2025 huggingface dataset:
Action of cAMP on expression and release of adhesion molecules in human endothelial cells
Authors: Morandini R. , Ghanem G. , Portier-Lemarie A. , Robaye B. , Renaud A. , Boeynaems J. M.
Published: December 2017

The expression of E-selectin induced by tumor necrosis factor (TNF) on the surface of human umbilical vein endothelial cells (HUVEC) was partially inhibited by an increase in the level of adenosine 3',5'-cyclic monophosphate (cAMP), produced by forskolin or cholera toxin combined with the type IV phosphodiesterase inhibitor rolipram and the protein kinase A agonist phosphorothioate analogue of cAMP SpcAMPS. The same agents had no significant effect on the constitutive and TNF-stimulated expression of intercellular adhesion molecule 1 (ICAM-1), whereas the effect on vascular cell adhesion molecule 1 (VCAM-1) expression was variable depending on cell culture conditions. The stimulatory effects of phorbol 12-myristate 13-acetate and bacterial lipopolysaccharide (LPS) on E-selectin expression were also downregulated by the forskolin-rolipram combination and by SpcAMPS. Inhibition of the surface expression of E-selectin was associated with a decrease of the total amount of the protein in the cell lysate and a reduced mRNA level, with no significant effect on mRNA stability. In anesthetized rats, the terbutaline-rolipram combination reduced the rolling of leukocytes induced by LPS in the mesenteric microcirculation. In addition to their partial inhibitory effect on the TNF-induced surface expression of E-selectin on HUVEC, the forskolin-rolipram combination and SpcAMPS strongly inhibited the release of soluble E-selectin from these cells; the release of soluble ICAM-1 and VCAM-1 was unaffected by these agents. Isoproterenol reduced the release of soluble E-selectin, whereas it had no significant effect on the cell surface expression of the protein. This study underscores the potential anti-inflammatory effect of a rise in the endothelial cAMP level.
Chronic hypoxia selectively augments endothelium-dependent pulmonary arterial vasodilation
Authors: Resta T. C. , Walker B. R.
Published: December 2017

We have previously demonstrated that chronic hypoxia (CH) augments pulmonary arterial dilation to the endothelium-derived nitric oxide (EDNO)-dependent pulmonary vasodilator arginine vasopressin (AVP). The present study examined 1) whether this enhanced vasoreactivity is observed with other agents that act by stimulating constitutive NO synthase (cNOS), 2) whether CH increases arterial vascular smooth muscle sensitivity to NO, and 3) whether endogenous endothelin (ET) or an endothelium-derived hyperpolarizing factor (EDHF) contributes to this altered arterial reactivity following CH. We examined responses to the receptor-mediated EDNO-dependent dilators histamine and ET-1, the nonreceptor-mediated EDNO-dependent dilator ionomycin, and the NO donors 1, 3-propanediamine, N- inverted question mark4-[1-(3-aminopropyl)-2-hydroxy-2-nitrosohydrazino] butyl inverted question mark (spermine NONOate) and S-nitroso-N-acetylpenicillamine (SNAP) in U-46619-constricted, isolated perfused lungs from control and CH rats. Additional experiments examined responses to AVP in the presence of the ET-receptor antagonist PD-145065 or the K+ channel blockers glibenclamide or tetraethylammonium (TEA) in lungs from each group. Microvascular pressure was assessed by double occlusion, allowing calculation of segmental resistances. Total and arterial vasodilatory responses to histamine, ET-1, and ionomycin were augmented in lungs from CH vs. control animals. However, CH did not alter the vasodilation to spermine NONOate or SNAP. PD-145065, glibenclamide, and TEA had no effect on responses to AVP in either group. We conclude that increased activity of arterial cNOS may be responsible for the augmented pulmonary arterial dilation to EDNO-dependent vasodilators following CH.
IGF-I increases bFGF-induced mitogenesis and upregulates FGFR-1 in rabbit vascular smooth muscle cells
Authors: Reape T. J. , Kanczler J. M. , Ward J. P. , Thomas C. R.
Published: December 2017

Insulin-like growth factor-I (IGF-I) and basic fibroblast growth factor (bFGF) have both been implicated in the abnormal proliferation of vascular smooth muscle cells (VSMC) that occurs after injury to the arterial wall in vivo. We have investigated the effects of these growth factors on proliferation of rabbit aortic smooth muscle cells (RASMC) in vitro. IGF-I, in contrast to bFGF, is a weak mitogen for RASMC. However, when IGF-I (10 ng/ml) was added in combination with bFGF for 24 h, the effect of the two growth factors on DNA synthesis was synergistic at all concentrations tested (P > 0.001 compared with summed values of bFGF alone plus IGF-I alone), and this synergy was also observed at the level of RASMC proliferation (P < 0.001). Time-course experiments indicated that although bFGF was able to stimulate DNA synthesis after 16 h, activity peaked at 24 h, and a synergistic response with IGF-I was not observed before 24 h. Northern blot analysis demonstrated that IGF-I (10 ng/ml) could selectively upregulate fibroblast growth factor receptor-1 (FGFR-1) mRNA 4.0 +/- 0.24-fold (P < 0.001) without a significant effect on FGFR-2, and this induction in FGFR-1 mRNA occurs in a time- and dose-dependent manner. In addition, IGF-I increases FGFR-1 protein levels in RASMC 2.7 +/- 0.12-fold (P < 0.01), as demonstrated by Western blotting, and this upregulation occurs before the peak in DNA synthesis. These results suggest that IGF-I may be capable of increasing the responsiveness of VSMC to bFGF through modulation of FGFR-1.
Left ventricular diastolic suction with zero left atrial pressure in open-chest dogs
Authors: Ingels N. B. , Daughters G. T. , Nikolic S. D. , DeAnda A. , Moon M. R. , Bolger A. F. , Komeda M. , Derby G. C. , Yellin E. L. , Miller D. C.
Published: December 2017

We investigated left ventricular (LV) diastolic volume changes (suction inflows) with left atrial pressure (LAP) clamped to ambient pressure in six open-chest, anesthetized dogs. The left atrium was cannulated and connected to a servo pump, and LAP was clamped to a set point near 0 mmHg for four beats by withdrawing blood. LAP averaged 5.88 +/- 1.44 mmHg before the clamp and fell to 0.74 +/- 0.61 mmHg (P < 0.0001) after the clamp. During the first clamped beat a transmitral pressure gradient of 1.0 +/- 0.6 mmHg was observed, resulting in LV filling of 2.6 +/- 1.8 ml. Subsequent beats developed suction-driven (mean negative LV pressure: -1.5 +/- 1.3 mmHg; P < 0.005 vs. zero) LV filling of 4.5 +/- 2.8 ml/beat with a peak transmitral pressure gradient of 1.7 +/- 0.6 mmHg. These data are consistent with the hypothesis that LV suction can be an important filling mechanism under condition in which LV end-systolic volume is reduced, e.g., reduced filling pressures, high heart rates, exercise, or increased inotropic drive.
Vagal withdrawal as a function of audience
Authors: De Meersman R. , Reisman S. , Daum M. , Zorowitz R.
Published: December 2017

This investigation examined the effects of psychosocial influences upon vagal cardiac activity. In this crossover, counter-balanced study, 15 subjects were assessed for vagal cardiac activity before and during a presentation in the presence and/or absence of an audience. Electrocardiograms (ECG) were collected throughout the epochs of interest, using a portable holter monitor system. Power spectral density analyses were used to decompose autonomic rhythmicities of heart rate variability. Significantly diminished vagal power was noted before and during presentation episodes with an audience compared with vagal power during a presentation without an audience (P < 0.05). Because respiration modulates autonomic outflow, ECG-derived respiration was derived and compared for all epochs, and no significant differences were noted. The real-life findings in the current investigation are strongly suggestive of the modulating effects of psychosocial interactions upon vagal cardiac electrophysiology and should be considered when assessing autonomic status.
Cyclical strain increases monocyte chemotactic protein-1 secretion in human endothelial cells
Authors: Wung B. S. , Cheng J. J. , Chao Y. J. , Lin J. , Shyy Y. J. , Wang D. L.
Published: December 2017

The effects of mechanical strain on monocyte chemotactic protein-1 (MCP-1) secretion were examined on human endothelial cells (ECs) grown on a flexible membrane base. MCP-1 release into culture medium from strained ECs was demonstrated to be time and strain dose dependent. Northern blot analysis demonstrated a mainly serum-independent 1.8-fold induction of MCP-1 mRNA levels in ECs strained at 15 kPa compared with unstrained controls. ECs treated with actinomycin D abolished this strain-induced expression. Strained ECs at the periphery of wells showed higher MCP-1 gene expression than ECs at the center. Pretreatment of ECs with either cytochalasin D or phalloidin did not abolish strain-induced gene expression. ECs pretreated with stretch-activated ion channel blocker gadolinium or with ryanodine to deplete intracellular stored Ca2+ strongly inhibited the strain-induced MCP-1 levels. We conclude that 1) cyclical strain can modulate the secretion of MCP-1 in a dose-dependent manner, 2) strain-induced MCP-1 production is mediated by increasing MCP-1 mRNA levels via transcription, 3) cytoskeletal rearrangement is not essential for this strain-induced MCP-1 expression, and 4) both Ca2+ influx via stretch-activated ion channels and intracellular Ca2+ release contribute to the strain-induced effect. Such strain-induced MCP-1 secretion might contribute to the trapping of monocytes in the subendothelial space to initiate atherogenesis.
Effects of nitric oxide/EDRF on platelet surface glycoproteins
Authors: Michelson A. D. , Benoit S. E. , Furman M. I. , Breckwoldt W. L. , Rohrer M. J. , Barnard M. R. , Loscalzo J.
Published: December 2017

We examined the effects of nitric oxide (NO)/endothelium-derived relaxing factor (EDRF) on platelet surface glycoproteins (GP). As determined by flow cytometry, in both a washed platelet system and platelet-rich plasma, the EDRF congener (S-nitroso-N-acetylcysteine) markedly inhibited both the thrombin-induced and the (stable thromboxane A2 analogue) U-46619-induced upregulation of P-selectin (alpha-granule protein), CD63 (lysosomal protein), and the GPIIb-IIIa complex (fibrinogen receptor) but minimally inhibited downregulation of the GPIb-IX complex (von Willebrand factor receptor). The inhibitory effects of EDRF were markedly reduced in whole blood or by the addition of washed erythrocytes. Platelets in whole blood were still responsive to guanosine 3',5'-cyclic monophosphate (cGMP), as shown by complete inhibition of P-selectin upregulation by the stable analogue N6,2'-O dibutyryl cGMP. These data suggests that 1) cGMP negatively regulates the platelet surface expression of P-selectin, CD63, and the GPIIb-IIIa complex but not the platelet surface expression of the GPIb-IX complex and 2) hemoglobin within erythrocytes inhibits the effects of EDRF/NO on platelet surface glycoproteins.
NF-kappa B and GTP cyclohydrolase regulate cytokine-induced nitric oxide production by cardiac myocytes
Authors: Oddis C. V. , Finkel M. S.
Published: December 2017

We previously reported that interleukin-1 beta (IL-1) alone stimulated nitric oxide (NO) production by neonatal rat cardiac myocytes (CM) in culture. The present studies were undertaken to explore the signal transduction pathways involved in IL-1-induced NO production by CM. Translocation from the cytosol to the nucleus of nuclear factor-kappa B (NF-kappa B) and activation of guanosine 5'-triphosphate (GTP) cyclohydrolase [rate-limiting enzyme in tetrahydrobiopterin (BH4) synthesis] have been implicated in IL-1 signaling. Accordingly, the effects of the NF-kappa B inhibitor pyrolidine dithiocarbamate (PDTC) and the GTP cyclohydrolase inhibitor 2,4-diamino-6-hydroxypyrimidine (DAHP) on IL-1-induced NO production by CM were studied. PDTC and DAHP inhibited IL-1-induced NO2-production by CM (6.7 +/- 0.6 vs. 0.9 +/- 0.3 and 0.3 +/- 0.1 nmol. 1.25 x 10(5) cells(-1).48 h-1, respectively, P < 0.01, n = 12 for each). Immunohistochemical staining revealed that PDTC blocked IL-1-stimulated nuclear translocation of NF-kappa B. The membrane-permeable analogue of the NO synthase cofactor BH4, methyl-BH4 (mBH4), only partially reversed DAHP inhibition of NO2- formation (6.7 +/- 0.6 vs. 2.4 +/- 0.3 nmol. 1.25 x 10(5) cells-1.48 h-1, P < 0.01, n = 12). Semiquantitative reverse transcription polymerase chain reaction revealed no inducible NO synthase (iNOS) mRNA production in cells treated with IL-1 + PDTC.CM treated with IL-1 + DAHP did express iNOS mRNA. We report for the first time that nuclear translocation of NF-kappa B is essential for II-1-induced iNOS mRNA expression and GTP cyclohydrolase activity is required in addition in addition to BH4 for optimal NO production by CM.
Coronary arteriolar flow-induced vasodilation signals through tyrosine kinase
Authors: Muller J. M. , Davis M. J. , Chilian W. M.
Published: December 2017

Coronary arterioles demonstrate flow-dependent vasodilation that is mediated by endothelial release of nitric oxide. The signaling mechanisms for this response remain unknown. Because tyrosine kinases are an enzyme family linked to many signaling pathways, including some for mechanosensitive transduction, we hypothesized that tyrosine kinase activation is a critical step in flow-induced vasodilation. To test this hypothesis, coronary arterioles were isolated, cannulated with micropipettes, and perfused by two independent reservoir systems. Intraluminal pressure was set at 60 cmH2O, and flow was generated by changing the heights of the reservoirs in equal and opposite directions, thus establishing a pressure difference across the arteriole without altering intraluminal pressure. Vasodilatory responses to intraluminal flow and substance P (1 x 10(-12) to 1 x 10(-7) M) were evaluated before and after intraluminal application of the tyrosine kinase inhibitors genistein (5 microM) and piceatannol (10 microM). Exposure to these inhibitors did not alter spontaneous tone. Substance P caused dose-dependent vasodilation that was not affected by genistein or piceatannol. Increases in intraluminal flow (generated by pressure differences ranging from 4 to 60 cmH2O) elicited graded increases in diameter. Both genistein and piceatannol inhibited the vasodilatory responses to flow. Treatment with daidzein, an inactive analogue of genistein, had no effect on either the flow-induced responses or substance P-induced vasodilation. To further confirm that tyrosine kinase activation is involved in flow-induced vasodilation, vessels were exposed to flow in the absence or presence of genistein and subsequently stained with a fluorescein isothiocyanate-labeled phosphotyrosine antibody. Exposure to flow significantly increased fluorescence of endothelial cells. Genistein treatment reversed the flow-induced increase in tyrosine phosphorylation. These results indicate that endothelium-dependent, flow-induced vasodilation in isolated porcine coronary arterioles is accompanied by an increase in tyrosine kinase activity. We conclude that endothelium-dependent, nitroxidergic, flow-induced vasodilation is mediated, at least in part, by a signaling pathway involving a tyrosine kinase.
Comparison of biaxial mechanical properties of excised endocardium and epicardium
Authors: Kang T. , Humphrey J. D. , Yin F. C.
Published: December 2017

A complete understanding of cardiac mechanics requires knowledge of the mechanical properties of each of the tissues that comprise the heart, Data and constitutive relations are available for the nonlinear multiaxial behavior of epicardium and noncontracting myocardium, but there have been no comparable results for endocardium. In this paper, we present biaxial mechanical data for endocardium and epicardium excised from the same bovine hearts. The data reveal that these two membranes behave differently; endocardium exhibits a greater stiffness in the low-strain range. Moreover, quantification of endocardial behavior requires a seven-parameter, polynomial-exponential pseudostrain-energy function w, whereas epicardium can be described by a four-parameter exponential w. Comparison of our current findings with previous results on canine epicardium reveals further that canine and bovine epicardium behave similarly, although the latter is more extensible. Thus there appear to be marked species differences.
Different responses of epicardium and endocardium to KATP channel modulators during regional ischemia
Authors: Miyoshi S. , Miyazaki T. , Moritani K. , Ogawa S.
Published: December 2017

We examined the responses of epicardial (Epi) and endocardial (Endo) layers to ATP-sensitive K+ (KATP) channel modulators during regional ischemia in anesthetized dogs. Five-minute occlusion of the left anterior descending coronary artery was repeated at 30-min interval. Monophasic action potentials (MAPs) and extracellular K+ concentrations ([K+]o) were measured at Epi and Endo layers. 5-Hydroxydecanoate (5-HD, 30 mg/kg iv), a KATP channel blocker, or nicorandil (NCR, 0.2-0.5 mg/kg iv), an opener, was administered before the third or fourth occlusion. Shortening rate of action potential duration at 90% repolarization (APD90) was greater at the Epi layer than at the Endo layer during the first 4 min after the second control occlusion (19.7 +/- 1.5 vs. 13.1 +/- 2.4%, n = 14, P < 0.05). 5-HD suppressed the shortening preferentially at the Epi layer and reduced the difference between the two layers (11.0 +/- 3.5 vs. 11.5 +/- 3.7%, n = 6, NS). In contrast, NCR augmented the shortening preferentially at the Epi layer and increased the difference between the two layers at 4 min (29.0 +/- 2.0 vs. 5.9 +/- 3.0%, n = 6, P < 0.05). The time differentiation of [K+]o rise was similar at the two layers during the control occlusion (0.44 vs. 0.50 mM/min, n = 12). 5-HD reduced the rate of [K+]o rise at both layers (0.34 vs. 0.40 mM/min), whereas NCR augmented the rate at the Epi layer (0.82 vs. 0.50 mM/min). Activation of KATP channels appears to be involved in ischemia-induced APD shortening and [K+]o rise. The different responses of the two layers suggest a lower threshold for activation and/or a denser distribution of KATP channels or other K+ channels at the Epi layer.
Muscle reflex stimulates sympathetic postganglionic efferents innervating triceps surae muscles of cats
Authors: Hill J. M. , Adreani C. M. , Kaufman M. P.
Published: December 2017

Two neural mechanisms contribute to the cardiovascular responses to exercise. The first, central command, proposes a parallel activation of central locomotor and brain stem circuits controlling cardiovascular function. The second, the muscle reflex, proposes that contraction-activated group III and IV afferents increase cardiovascular function. In humans, whole nerve recordings of sympathetic discharge suggest that central command increases sympathetic outflow to skin but not to skeletal muscle and that the muscle reflex increases sympathetic outflow to skeletal muscle but not to skin. We therefore tested the hypothesis that the muscle reflex, but not central command, increases the discharge of single sympathetic postganglionic efferents innervating the triceps surae muscles of decerebrate unanesthetized cats. Central command was evoked by electrical stimulation of the mesencephalic locomotor region. The reflex was evoked by electrical stimulation of the tibial nerve, which in turn contracted the triceps surae muscles. Hexamethonium abolished spontaneous and evoked activity, verifying that the recordings were from sympathetic postganglionic fibers. The discharge of 13 efferents was increased by static contraction (from 0.6 +/- 0.2 to 1.0 +/- 0.3 imp/s; P < 0.05) but was not increased by central command (from 0.6 +/- 0.2 to 0.8 +/- 0.2 imp/s; P > 0.05). Nevertheless, the discharge of nine efferents, not increased by central command before alpha-adrenergic blockade (from 0.5 +/- 0.2 to 0.9 +/- 0.4 imp/s; P > 0.05), was increased after blockade (from 1.3 +/- 0.2 to 3.2 +/- 0.8 imp/s; P < 0.05). We conclude that the muscle reflex stimulates sympathetic postganglionic efferents innervating the vasculature of skeletal muscle. Furthermore, baroreceptors appear to buffer the central command-induced increases in the discharge of these efferents.
Effect of hypoxia on nitric oxide production in neonatal pig lung
Authors: Nelin L. D. , Thomas C. J. , Dawson C. A.
Published: December 2017

Nitric oxide (NO) synthase inhibitors potentiate hypoxic vasoconstriction (HV), suggesting that NO production during hypoxia normally acts to attenuate HV. To begin to examine the effect of hypoxia on lung NO production, we studied four groups of isolated neonatal pig lungs. In three groups of lungs, the accumulation of nitrite/nitrate (NOx-) was measured in the recirculating perfusate during ventilation with a control gas mixture (Cont), a hypoxic gas mixture (Hyp), or the control gas mixture with N omega-nitro-L-arginine methyl ester (L-NAME) added to the perfusate. Both hypoxia and L-NAME significantly increased perfusion pressure [pulmonary arterial pressure (Pa)-pulmonary venous pressure (Pv)] compared with control. NOx- accumulated in the perfusate at an average rate of 9.1 +/- 2.3 (SE) nmol/min in Cont, 3.7 +/- 0.8 nmol/min (P < 0.05 vs. control) in Hyp, and 3.7 +/- 0.6 nmol/min (P < 0.05 vs. control) in L-NAME. In the fourth group of lungs, exhaled NO output was measured during ventilation with the control gas mixture, the hypoxic gas mixture, and the control gas mixture with L-NAME added to the perfusate. Pa-Pv increased significantly with both hypoxia and L-NAME in these lungs. The exhaled NO output also decreased significantly with both hypoxia and L-NAME. These results suggest that in this preparation there was continuous production of NO that was decreased by hypoxia or L-NAME. It is not clear how the potentiation of HV by NO inhibitors and inhibition of NO production by hypoxia are linked.
Peripheral artery structure and endothelial function in heart failure: effect of ACE inhibition
Authors: Mulder P. , Elfertak L. , Richard V. , Compagnon P. , Devaux B. , Henry J. P. , Scalbert E. , Desche P. , Mace B. , ThuilleZ C.
Published: December 2017

Chronic heart failure (CHF) induces peripheral vasoconstriction and impairs endothelium-dependent relaxation of large arteries. We investigated in a rat model of CHF (coronary artery ligation) 1) whether endothelial dysfunction also exists in resistance arteries, 2) whether this is associated with vascular morphological changes, and 3) the effect of angiotensin-converting enzyme (ACE) inhibition on these parameters. After 1 mo or 1 yr, CHF reduced the vasodilatory response to acetylcholine of isolated, perfused femoral and mesenteric artery segments. This impairment was more marked in femoral than in mesenteric arteries. However, CHF did not induce any arterial remodeling. Chronic treatment with the ACE inhibitor perindopril improved the response to acetylcholine and reduced media cross-sectional area and collagen density. Thus at the level of small peripheral arteries, CHF induces an endothelial dysfunction but does not affect vascular structure. ACE inhibition prevents the CHF-induced endothelial dysfunction and induces vascular remodeling. These changes could contribute to the observed beneficial effects of ACE inhibitors on hemodynamics and survival in CHF.
Effects of 17 beta-estradiol on coronary microvascular responses to endothelin-1
Authors: Lamping K. G. , Nuno D. W.
Published: December 2017

The objective of this study was to examine the effects of 17 beta-estradiol on responses of coronary microvessels to endothelin-1 (ET-1). With the use of isolated pressurized coronary microvessels from the left ventricle of male or female dogs, constrictions to ET-1 were similar in vessels from male and female dogs. 17 beta-Estradiol (1 microM) attenuated constriction to ET-1 of small arteries from both male (percent constriction at 10 microM control: 39 +/- 9%, estradiol: 3 +/- 2%; P < 0.05) and female (percent constriction at 10 microM control: 39 +/- 8%, estradiol: 6 +/- 3%; P < 0.05) dogs similarly. In contrast, testosterone (1 microM) had no effect on constriction to ET-1. Constrictions to ET-1 were completely abolished by BQ-123 (1 microM), a selective ETA-receptor antagonist, and enhanced by BQ-788 (1 microM), a selective ETB-receptor antagonist. Constrictions to ET-1 alone were not altered by indomethacin (Indo, 10 microM) or NG-nitro-L-arginine (L-NNA, 100 microM). 17 beta-Estradiol produced dose-dependent relaxation of coronary microvessels preconstricted with ET-1 that was similar to the response to testosterone and progesterone. Indo or L-NNA alone had no effect on relaxation to 17 beta-estradiol. However, the combination of Indo and L-NNA attenuated Taxation to 17 beta-estradiol (percent dilation at 1 microM control: 64 +/- 13%; Indo plus L-NNA: 21 +/- 6%; P < 0.05) but did not affect relaxation to testosterone. Thus 17 beta-estradiol attenuated constrictions of coronary microvessels to ET-1 more than did similar concentrations of testosterone. The ability of 17 beta-estradiol to modulate responses to endothelin may involve release of vasodilator prostaglandins and/or nitric oxide by 17 beta-estradiol.
Relation between baroreflex sensitivity and carotid artery elasticity in healthy humans
Authors: Bonyhay I. , Jokkel G. , Kollai M.
Published: December 2017

We investigated whether physiological variability in arterial baroreflex sensitivity (BRS) was related to differences in carotid elastic behavior among 19 young healthy subjects (age 18-26 yr). The diameter of the carotid artery (D) and its change during the arterial pulse (delta D) were monitored by a phase-locked echotracking device (UT-4 Hokanson), and pulse pressure (delta P) was measured in the brachial artery by sphygmomanometry. Distensibility coefficient (DC) for the common carotid artery was calculated using the formula DC = (2 x delta D/D)/delta P. Dynamic elastic parameters such as the maximum and mean rate of carotid artery expansion and the dominant harmonic frequency of the diameter curve were also determined. BRS was assessed by regressing R-F.intervals against systolic blood pressure, monitored by finger arterial pressure (FINAPRES), during an elevation of pressure, induced by intravenous bolus injection of phenylephrine. Using correlation and stepwise regression analysis, we found that BRS was significantly related to carotid artery distensibility (r = 0.778, P < 0.001) but was not related to any of the dynamic parameters of carotid pulsation.
ACh dilates pial arterioles in endothelial and neuronal NOS knockout mice by NO-dependent mechanisms
Authors: Meng W. , Ma J. , Ayata C. , Hara H. , Huang P. L. , Fishman M. C. , Moskowitz M. A.
Published: December 2017

We used mice with deletions in either the endothelial nitric oxide synthase (eNOS) or neuronal NOS (nNOS) gene to investigate the role of eNOS and nNOS in acetylcholine (ACh)-induced relaxation of pial arterioles (20-30 microns). Pial arteriolar diameter was measured by intravital microscopy through a closed cranial window, and NOS activity was determined by the conversion of [3H]arginine to [3H]citrulline in subjacent cortex. ACh superfusion (1, 10 microM) caused atropine-sensitive dose-dependent arteriolar dilation in all three mouse strains. At 10 microM, increases of 20 +/- 2, 31 +/- 3, and 23 +/- 3% were recorded in wild-type (n = 25), nNOS mutant (n = 15), and eNOS mutant (n = 20) mice, respectively. NG-nitro-L-arginine (L-NNA, 1 mM) superfusion inhibited cortical NOS activity by > 70% and abrogated the response in wild-type mice while blocking the dilation by approximately 50% in eNOS mutant and nNOS mutant mice. Only in the eNOS mutant did tetrodotoxin (TTX) superfusion (1 microM) attenuate ACh-induced dilation (n = 6). The residual dilation after L-NNA in eNOS mutant mice could be blocked completely by TTX-plus L-NNA. Our findings indicate that 1) ACh dilates pial arterioles of wild-type mice by NOS-dependent mechanisms as reported in other species, 2) the response in nNOS mutant mice resembles the wild-type response except for enhanced dilation to ACh and reduced L-NNA sensitivity, and 3) surprisingly, the response in eNOS mutant mice is partially NOS dependent and attenuated by both TTX and L-NNA. Because nNOS is constitutively expressed in eNOS mutants, these findings coupled with the TTX results suggest that an nNOS-dependent mechanism may compensate for the chronic loss of eNOS activity after targeted gene disruption.
Method for isolation of adult mouse cardiac myocytes for studies of contraction and microfluorimetry
Authors: Wolska B. M. , Solaro R. J.
Published: December 2017

We describe techniques for the isolation of Ca(2+)-tolerant myocytes from mouse (2 to 6 mo old) ventricle for measurements of mechanics of contraction and microfluorimetry. Our approach involved special modifications of existing methods that had been developed for other species but were not successful when applied to the mouse heart. Important features of the method are 1) a requirement for careful timing (< 5 min) of perfusion with nominally Ca(2+)-free solution; 2) perfusion with a solution containing a specially selected batch of collagenase in the presence of a low Ca2+ concentration; and 3) meticulous attention to water quality. Using this method, we could consistently isolate durable, Ca(2+)-tolerant myocytes from adult mouse hearts with a yield of approximately 50%. With slight modifications, the method should enable other investigators to isolate mouse cardiomyocytes for their specific experimental applications.
Coronary hyperperfusion augments myocardial oxygen consumption
Authors: Ishibashi Y. , Zhang J. , Duncker D. J. , Klassen C. , Pavek T. , Bache R. J.
Published: December 2017

This study was performed to test the hypothesis that increases in myocardial oxygen consumption (MVo2) and myocardial contractile function during exercise are flow limited. Studies were performed in 15 chronically instrumented normal dogs. MVo2 and regional percent systolic wall thickening were measured during control conditions and during maximal vasodilation produced by infusion of adenosine (20-75 micrograms.kg-1.min-1) or adenosine combined with nitroglycerin (0.4 micrograms.kg-1.min-1; TNG) into the left anterior descending coronary artery during a three-stage graded treadmill exercise protocol. Adenosine and adenosine plus TNG significantly increased coronary blood flow by 298 +/- 26 and 306 +/- 24%, respectively, at rest and by 134 +/- 7 and 145 +/- 9%, respectively, during the heaviest level of exercise (each P < 0.01). Adenosine and adenosine plus TNG increased MVo2 at rest, but this was associated with a parallel increase in heart rate, so that MVo2 per beat was not significantly changed. Systolic wall thickening was also not changed by hyperperfusion during resting conditions. However, MVo2 per beat was increased by 12 +/- 4% with adenosine and by 13 +/- 5% with adenosine plus TNG during moderate exercise and by 23 +/- 5% with adenosine and by 27 +/- 4% with adenosine plus TNG during the heaviest level of exercise (each P < 0.05). Systolic thickening of the full left ventricular wall did not change during hyperperfusion, but thickening in the subepicardial layer was increased by 14 +/- 3% with adenosine and 18 +/- 3% with adenosine plus TNG during the heaviest level of exercise (each P < 0.05). There was no difference in wall thickening between adenosine and adenosine plus TNG. These findings imply that the increases in MVo2 which occur during exercise are limited by coronary blood flow.
Inotropic agent EMD-53998 weakens nucleotide and phosphate binding to cross bridges in porcine myocardium
Authors: Zhao Y. , Kawai M.
Published: December 2017

The effect of EMD-53998 (EMD) on elementary steps of the cross-bridge cycle in skinned porcine myocardium was studied to understand the positive inotropic mechanisms of EMD. The kinetic constants of the elementary steps were obtained by sinusoidal analysis and compared in the presence and absence of 50 microM EMD. In the presence of 50 microM EMD, the equilibrium constant of the cross-bridge detachment step decreases three times, and the equilibrium constant of the cross-bridge attachment (force generation) step increases two times. Our results further show that, in the presence of 50 microM EMD, the association constants of MgATP and MgADP decrease to one-ninth and one-third, respectively, and the association constant of phosphate decreases to one-third. These results indicate that EMD suppresses the nucleotide binding to cross bridges and increases cross-bridge resistance to phosphate accumulation in myocardium. These results predict that EMD decreases the number of detached cross bridges and increases the number of attached cross bridges. This prediction is consistent with the twofold increase of isometric tension with 50 microM EMD.
Tissue oxygenation and perfusion in endotoxemia
Authors: Sair M. , Etherington P. J. , Curzen N. P. , Winlove C. P. , Evans T. W.
Published: December 2017

Sepsis is believed to induce disturbances in microcirculatory flow and nutrient exchange, which may result in impaired tissue oxygenation. With the use of an established rat model of endotoxemia, voltametric measurements were made of skeletal muscle (tissue) oxygen tension (PtO2) and its response to inspired oxygen concentration (FIO2). Steady-state nutritive flow and the response of endotoxemic muscle to ischemia-reperfusion were also measured. In the presence of a normal arterial PO2, mean muscle PtO2 in the endotoxemic group was significantly lower than controls (52 +/- 9 vs. 24 +/- 4 Torr, P < 0.01; +/- SE). Endotoxemic muscle PtO2 values showed less heterogeneity than control groups and significant attenuation of the response to increasing FIO2 to 0.95 (mean rise in PtO2 +/- SE; 27 +/- 7 vs. 80 +/- 11 Torr for endotoxemic and control groups, respectively; P < 0.01). No steady-state differences in tissue perfusion or response to ligation-induced ischemia-reperfusion could be demonstrated between endotoxemic and control rats. These data suggest that there is significant tissue hypoxia and abnormal microvascular control of oxygenation in endotoxemia, even in the presence of normal microcirculatory perfusion.
Measurement of regional myocardial blood flow in rats by unlabeled microspheres and Coulter channelyzer
Authors: Wang X. , Li F. , Said S. , Capasso J. M. , Gerdes A. M.
Published: December 2017

A relatively inexpensive, expeditious, new nonradioactive microsphere method for measuring regional myocardial blood flow (RMBF) was developed with unlabeled microspheres and a Coulter Channelyzer. To validate the efficiency of this method, hearts from rats were perfused ex vivo by retrograde aortic cannulation. Unlabeled microspheres of varying size were injected into a side arm in the aortic cannula or added to blood samples collected from the rats. Microspheres were then recovered from the cardiac tissue and blood samples. It was found that > 97% of perfused microspheres (diam > 9.4 microns) were retained in the myocardium and that 94.8 +/- 2.2% of the trapped microspheres were recovered and counted successfully using a Counter Channelyzer. The percent recovery of microspheres from 2- and 0.5-ml blood samples were 95.4 +/- 2.3 and 95.3 +/- 3.1%, respectively. Blood flow to the anterior and posterior halves of the ventricular free walls and septum were measured in six rats; excellent agreements were found between the results yielded by 10-, 15-, and 20-microns unlabeled microspheres injected simultaneously. The transmural flow gradients in the left ventricular free wall estimated by 10- and 15-microns spheres did not significantly differ from each other. Thus the method developed here provides a new alternative for measurement of RMBF, which currently allows at least three measurements for nontransmural gradient RMBF and at least two measurements for transmural gradient RMBF.
Lysophosphatidylcholine stimulates phospholipase D in human coronary endothelial cells: role of PKC
Authors: Cox D. A. , Cohen M. L.
Published: December 2017

Lysophosphatidylcholine (lyso PC) mediates multiple potentially atherogenic effects on endothelial cells, although the cellular mechanism of these effects remains unclear. Phospholipase D (PLD) has been recognized as a novel second-messenger system that may regulate cellular function. The purpose of this study was to determine the effect of lyso PC on PLD activity in human coronary artery endothelial cells (HCAEC) by measuring [3H]phosphatidylethanol production in cells labeled with [3H]myristic acid. After incubation with lyso PC (20 microM) for 40 min, PLD activity was markedly stimulated from five- to sixfold. Stimulation of PLD activity by lyso PC was concentration dependent (half-maximum effective concentration of 7.6 microM) and was not mimicked by phosphatidylcholine (20 microM). Because PLD can be regulated by protein kinases, the effect of several protein kinase inhibitors on lyso PC-stimulated PLD activity was tested. The protein kinase A inhibitor H-89 (300 nM) and the tyrosine kinase inhibitors genistein (30 microM) and tyrphostin A25 (100 microM) had no effect on the stimulation of PLD by lyso PC (20 microM). The protein kinase C (PKC) inhibitor calphostin C (10-300 nM) affected neither lyso PC (20 microM)-nor 4 beta-phorbol 12,13-dibutyrate (PDBu, 300 nM)-stimulated PLD activity, suggesting that this agent may not inhibit PKC in these cells. In contrast, the selective PKC inhibitors GF-109203X (0.3-10 microM) and chelerythrine (1-30 microM) concentration dependently inhibited lyso PC (20 microM)-stimulated PLD activity and blocked PDBu (300 nM)-stimulated PLD activity. Together, these data document that lyso PC stimulated PLD in human endothelial cells, possibly by a PKC-dependent mechanism, and provide evidence that PLD activation in human endothelium is a novel and important mechanism by which lyso PC mediates its cellular and possibly atherogenic effects.
L-NNA-sensitive regional cerebral blood flow augmentation during hypercapnia in type III NOS mutant mice
Authors: Ma J. , Meng W. , Ayata C. , Huang P. L. , Fishman M. C. , Moskowitz M. A.
Published: December 2017

The effect of NG-nitro-L-arginine (L-NNA) on regional cerebral blood flow (rCBF) response to hypercapnia (5% CO2 inhalation) was studied in urethan-anesthetized wild-type (SV-129) and type III nitric oxide (NO) synthase (NOS)-deficient mice, using laser-Doppler flowmetry and the closed cranial window technique. Resting rCBF during normocapnia decreased by approximately 25% after L-NNA superfusion in wild-type mice only (n = 18), suggesting a role for type III NOS in baseline blood flow. Hypercapnia augmented rCBF approximately 50% in both wild-type and type III NOS mutant mice. L-NNA superfusion (1 mM) inhibited this increase by approximately 60% in both strains. Hence, synthesis of NO by the constitutively expressed type I NOS contributes to blood flow augmentation during hypercapnia.
Regulation of alpha 1-, beta 1-, and beta 2-adrenergic receptors in rat heart by norepinephrine
Authors: Zhao M. , Hagler H. K. , Muntz K. H.
Published: December 2017

Previous studies suggest that the desensitization and downregulation of beta 1-adrenergic receptors (beta 1-AR) in the failing heart are the result of the elevated plasma catecholamine levels associated with this disease. To examine norepinephrine (NE)-induced regulation of cardiac adrenergic receptors, rats were infused with l-NE (200 micrograms.kg-1.h-1 for 7 days) or vehicle (0.001 N HCl) by implantation of osmotic minipumps. The technique of coverslip autoradiography was used to quantify alpha 1-adrenergic receptors (alpha 1-AR), beta 1-AR, and beta 2-AR in different tissue compartments of rat hearts. For measurement of beta-AR binding, sections were incubated with 70 pM [125I]iodocyanopindolol (ICYP) alone or in the presence of 5 microM dl-propranolol or 5 x 10(-7) M CGP-20712A (a beta 1-antagonist) and then set up for autoradiography. [3H]prazosin (1 nM) with or without phentolamine was used to study alpha-AR binding. Chronic infusion of NE induced a greater downregulation of beta 2-AR compared with beta 1-AR in all regions studied, including atrial and ventricular myocytes, coronary arterioles, and connective tissue. An 18% loss of beta 1-AR was seen only in atrial myocytes; beta 1-AR density actually increased 28% in ventricular myocytes following NE infusion. There was a 15% decrease in alpha 1-AR in ventricular myocytes, whereas no change in alpha 1-AR density was seen in myocardial arterioles. Our study demonstrates that beta 2-AR are more susceptible to NE-induced downregulation than beta 1-AR. Thus other mechanisms may be involved in the selective downregulation of beta 1-AR in certain forms of heart failure.
Participation of galanin in baroreflex inhibition of heart rate by hypothalamic PVN in rat
Authors: Chen Y. L. , Chan S. H. , Chan J. Y.
Published: December 2017

We investigated the role of galanin (Gal) in the suppression of baroreceptor reflex (BRR) response by the paraventricular nucleus of the hypothalamus (PVN) in adult male Sprague-Dawley rats that were anesthetized with pentobarbital sodium. Electrical stimulation (10-s train of 1-ms rectangular pulses at 20-40 microA and 10-20 Hz) of, and microinjection of L-glutamate (1 nmol) into, the PVN significantly inhibited BRR response to transient hypertension induced by phenylephrine (5 micrograms/kg iv). Such a PVN-induced BRR suppression was appreciably antagonized by local administration of Gal antiserum (1:20), but not heat-inactivated Gal antiserum (1:20), to the nucleus tractus solitarius (NTS) bilaterally. Microinjection of Gal (100 pmol) into the NTS bilaterally also resulted in a Gal antiserum-reversible inhibition of the BRR response. Immunohistochemical results demonstrated that the distribution of Gal-containing neurons in the parvocellular subnucleus of the PVN overlapped substantially with the hypothalamic loci on which electrical or chemical activation elicited suppression of the BRR response that was significantly blunted by microinjection of Gal antiserum into the NTS. These results suggest that the PVN may participate in central cardiovascular regulation by suppressing the BRR response via galaninergic neurotransmission at the NTS.
Blocking L-selectin function attenuates reperfusion injury following hemorrhagic shock in rabbits
Authors: Ramamoorthy C. , Sharar S. R. , Harlan J. M. , Tedder T. F. , Winn R. K.
Published: December 2017

Leukocyte adhesion molecule (LAM) blockade reduces ischemia-reperfusion injury. We tested the hypothesis that a monoclonal antibody (MAb) that recognizes a functional epitope of L-selectin would decrease hemorrhagic shock-induced reperfusion injury. Anesthetized rabbits were subjected to 2 h of hemorrhagic shock (cardiac output reduced to 30% of baseline), then given one of the following treatments: MAbs that recognize functional domains of L-selectin (LAM1-3), CD18 (60.3), MAbs that recognize a nonfunctional domain on L-selectin (LAM1-14), or saline, immediately before resuscitation with shed blood. Additional fluids were administered as needed to maintain cardiac output at baseline levels for 6 h. The cumulative fluid resuscitation after MAb LAM1-3 (58 +/- 34 ml/kg) was not significantly different from after MAb 60.3 (21 +/- 24 ml/kg) or MAb LAM1-14 (66 +/- 51 ml/kg), but it was significantly less than saline-treated controls (142 +/- 142 ml/kg). However, two animals treated with MAb LAM1-14 died before 6 h. If their resuscitation volumes are projected to 6 h by linear regression, then the LAM1-14-treated group required significantly greater volume (101 +/- 99 ml/kg) than the MAb LAM1-3-treated group. We conclude that MAbs to a functional domain on L-selectin are protective against reperfusion-injury following hemorrhagic shock.
Control of arteriovenous anastomoses in rabbit ear model of digital perfusion
Authors: Pollock D. C. , Li Z. , Koman L. A. , Gordon E. S. , Smith T. L.
Published: December 2017

The arteriovenous anastomoses (AVA) of the cutaneous microcirculation of the hands and feet are fundamental determinants of thermoregulatory blood flow and may be involved in cold intolerance. These direct microvascular studies are an initial characterization of adrenergic receptor subtypes participating in control of AVA in the ears of anesthetized male New Zealand White rabbits. Adrenergic alpha 1-stimulation with phenylephrine produced AVA constriction, whereas terazosin (an alpha 1-antagonist) produced dilation and attenuated the responses to phenylephrine. Adrenergic alpha 2-stimulation with UK-14304 produced constriction of the AVA, whereas atipamezole (an alpha 2-antagonist) produced dilation and attenuated the responses to UK-14304. When equimolar concentrations of antagonists were studied, the AVA dilation produced by alpha 2-blockade was greater than that produced by alpha 1-blockade. Norepinephrine (a mixed alpha 1- and alpha 2-agonist) also produced vasoconstriction, which was attenuated by both prazosin (an alpha 1-antagonist) and atipamezole. In summary, 1) AVA contain a heterogeneous mixture of both alpha 1- and alpha 2-receptors, and 2) alpha 2-receptors may have a greater influence than alpha 1-receptors on overall tone in AVA.
Habitual exercise attenuates myocardial stiffness in diabetes mellitus in rats
Authors: Woodiwiss A. J. , Kalk W. J. , Norton G. R.
Published: December 2017

The effects of regular voluntary (habitual) exercise on left ventricular (LV) diastolic performance, as well as on those hemodynamic, LV geometric and myocardial interstitial changes that influence myocardial diastolic function, were examined in streptozocin (50 mg/kg)-induced diabetes mellitus (DM) in male rats. After 16 wk of regular exercise on voluntary running wheels, cardiac performance was measured in anesthetized open-chest ventilated rats. Rats with DM developed a decrease in LV end diastolic (LVED) chamber and myocardial wall compliance associated with an increase in myocardial collagen fluorescence (a measure of collagen advanced glycosylation end product concentrations), an increase in systemic blood pressure, and a dilated LV. However, LV end-systolic elastance and total myocardial hydroxyproline concentration were unchanged. Habitual exercise prevented the augmented LVED chamber stiffness (slope of linearized LVED pressure-strain relation, DM vs. exercise DM, P < 0.01) and myocardial wall stiffness (slope of linearized LVED stress-strain relation, DM vs. exercise DM, P < 0.001) produced by DM. However, neither markers of blood glucose control, blood pressure, LV geometry, myocardial hydroxyproline concentration, nor collagen fluorescence were altered by the exercise program. A reduced LVED chamber and myocardial wall stiffness, as noted in euglycemic control rats in response to regular exercise, was also not associated with hemodynamic or myocardial interstitial alterations. Therefore, regular voluntary exercise opposes the decrement in myocardial diastolic performance produced by DM in male rats without influencing the accumulation of myocardial advanced glycosylation end products thought to be partly responsible for the stiff myocardium after chronic hyperglycemia. The beneficial effects of regular exercise on myocardial compliance are likely to be a consequence of changes in active as opposed to passive properties of the myocardial wall.