bluuebunny/crossref_metadata_2025 huggingface dataset:
Hypoglycemia-associated autonomic failure in diabetes
Authors: Cryer Philip E.
Published: December 2017

Hypoglycemia is the limiting factor in the glycemic management of diabetes. The concept of hypoglycemia-associated autonomic failure (HAAF) in diabetes posits that recent antecedent iatrogenic hypoglycemia causes both defective glucose counterregulation (by reducing the epinephrine response to falling glucose levels in the setting of an absent glucagon response) and hypoglycemia unawareness (by reducing the autonomic and the resulting neurogenic symptom responses) and thus a vicious cycle of recurrent hypoglycemia. Perhaps the most compelling support for HAAF is the finding that as little as 2–3 wk of scrupulous avoidance of hypoglycemia reverses hypoglycemia unawareness and improves the reduced epinephrine component of defective glucose counterregulation in most affected individuals. Insight into this pathophysiology has led to a broader view of the clinical risk factors for hypoglycemia to include indexes of compromised glucose counterregulation and provided a framework for the study of the mechanisms of iatrogenic hypoglycemia and, ultimately, its elimination from the lives of people with diabetes.
Characterization of the growth hormone receptor in human dermal fibroblasts and liver during development
Authors: Goodyer Cynthia G. , Figueiredo Rilene M. O. , Krackovitch Stephanie , De Souza Li Lilia , Manalo Jennifer A. , Zogopoulos George
Published: December 2017

Human tissues express growth hormone receptors (hGHR) by the 3rd mo of gestation. We assessed developmental changes in hGHR function in fibroblasts and liver, testing binding and hormonal response. Fetal cells showed low but reproducible hGH binding. No age-related changes occurred in fibroblasts (9 wk–34 yr). In contrast, there was a fourfold increase in hGH binding in postnatal liver, with a sixfold increase in hGHR mRNA. Both full-length and truncated hGHR mRNAs were detected in all livers. Cross-linking revealed a larger hGH/receptor complex in fetal liver. Fetal hepatocytes produced 10 times more insulin-like growth factor (IGF)-II than IGF-I, and responded to hGH (150 ng/ml) with a significant increase in IGF-II. Fetal hepatocytes secreted three IGF-binding proteins (IGFBPs), including IGFBP1, but not IGFBP3. hGH did not alter fetal hepatocyte IGFBPs but stimulated glucose uptake. Exposure of fibroblasts to hGH decreased hGH binding only in >1-yr postnatal fibroblasts, whereas treatment with dexamethasone (100–400 nM) increased binding only in postnatal cells. Thus, although fetal hepatocytes and fibroblasts possess functional hGHR, these receptors (and/or their signaling pathways) are immature or have adapted to the in utero environment.
MDCK cells secrete neutral proteases cleaving insulin-like growth factor-binding protein-2 to -6
Authors: Shalamanova Liliana , Kübler Bernd , Scharf Jens-Gerd , Braulke Thomas
Published: December 2017

Proteolysis of insulin-like growth factor-binding proteins (IGFBPs) may be an important mechanism to regulate IGF availability and IGF-independent functions of IGFBPs. We analyzed the secretion of IGFBP proteases in Madin-Darby canine kidney (MDCK) cells. The results showed that several specific proteases were secreted, cleaving IGFBP-2 to -6 at neutral pH. The proteolytic activity against IGFBP-6 differed at least from IGFBP-5 protease activity in its sensitivity both to IGF-II and to the hydroxamic acid-based disintegrin metalloprotease inhibitor, as well as serine protease inhibitors. During partial purification steps, the serine protease inhibitor-sensitive fraction with IGFBP-6 protease activity was separated from fractions characterized by the presence of a 30-kDa disintegrin immunoreactive band. Whereas the IGFBP-4 and -6 proteases are predominantly secreted across the basolateral membrane, the majority of IGFBPs are sorted to the apical medium from filter-grown cells. These studies indicate that the side-specific secretion of several distinct IGFBP proteases with partially overlapping IGFBP specificities may be another level in the regulation of IGF-dependent epithelial functions.
Molecular mechanisms involved in gastrin-mediated regulation of cAMP-responsive promoter elements
Authors: Thommesen Liv , Hofsli Eva , Paulssen Ruth H. , Anthonsen Marit W. , Lægreid Astrid
Published: December 2017

In the present study, we explore the role of cAMP-responsive (CRE) promoter elements in gastrin-mediated gene activation. By using the minimal CRE promoter reporter plasmid, pCRELuc, we show that gastrin can activate CRE. This activation is blocked by H-89 and GF 109203x, which inhibit protein kinases A and C, respectively. Moreover, Ca2+-activated pathways seem to be involved, because the calmodulin inhibitor W-7 reduced gastrin-mediated activation of pCRELuc. Deletion of CRE from the c- fos promoter rendered this promoter completely unresponsive to gastrin, indicating that CRE plays a central role in c- fos transactivation. Interestingly, gastrin-induced expression of the inducible cAMP early repressor (ICER), a gene that is known to be regulated by CRE promoter elements, was not reduced by H-89, W-7, or GF 109203x. Furthermore, bandshift analyses indicated that the region of the ICER promoter containing the CRE-like elements CARE 3–4 binds transcription factors that are not members of the CRE-binding protein-CRE modulator protein-activating transcription factor, or CREB/CREM/ATF-1, family. Our results underline the significance of the CRE promoter element in gastrin-mediated gene regulation and indicate that a variety of signaling mechanisms are involved, depending on the CRE promoter context.
Effect of acute exercise on uncoupling protein 3 is a fat metabolism-mediated effect
Authors: Schrauwen Patrick , Hesselink Matthijs K. C. , Vaartjes Ilonca , Kornips Esther , Saris Wim H. M. , Giacobino Jean-Paul , Russell Aaron
Published: December 2017

Human and rodent uncoupling protein (UCP)3 mRNA is upregulated after acute exercise. Moreover, exercise increases plasma levels of free fatty acid (FFA), which are also known to upregulate UCP3. We investigated whether the upregulation of UCP3 after exercise is an effect of exercise per se or an effect of FFA levels or substrate oxidation. Seven healthy untrained men [age: 22.7 ± 0.6 yr; body mass index: 23.8 ± 1.0 kg/m2; maximal O2 uptake (V˙o 2 max): 3,852 ± 211 ml/min] exercised at 50% V˙o 2 max for 2 h and then rested for 4 h. Muscle biopsies and blood samples were taken before and immediately after 2 h of exercise and 1 and 4 h in the postexercise period. To modulate plasma FFA levels and fat/glucose oxidation, the experiment was performed two times, one time with glucose ingestion and one time while fasting. UCP3 mRNA and UCP3 protein were determined by RT-competitive PCR and Western blot. In the fasted state, plasma FFA levels significantly increased ( P < 0.0001) during exercise (293 ± 25 vs. 1,050 ± 127 μmol/l), whereas they were unchanged after glucose ingestion (335 ± 54 vs. 392 ± 74 μmol/l). Also, fat oxidation was higher after fasting ( P < 0.05), whereas glucose oxidation was higher after glucose ingestion ( P< 0.05). In the fasted state, UCP3L mRNA expression was increased significantly ( P < 0.05) 4 h after exercise (4.6 ± 1.2 vs. 9.6 ± 3.3 amol/μg RNA). This increase in UCP3L mRNA expression was prevented by glucose ingestion. Acute exercise had no effect on UCP3 protein levels. In conclusion, we found that acute exercise had no direct effect on UCP3 mRNA expression. Abolishing the commonly observed increase in plasma FFA levels and/or fatty acid oxidation during and after exercise prevents the upregulation of UCP3 after acute exercise. Therefore, the previously observed increase in UCP3 expression appears to be an effect of prolonged elevation of plasma FFA levels and/or increased fatty acid oxidation.
Adipose tissue-specific increase in angiotensinogen expression and secretion in the obese (fa/fa) Zucker rat
Authors: Hainault Isabelle , Nebout Guillaume , Turban Sophie , Ardouin Bernadette , Ferré Pascal , Quignard-Boulangé Annie
Published: December 2017

We investigated angiotensinogen (AGT) expression in adipose tissue and liver of Zucker rats during the onset of obesity. The developmental pattern of AGT expression (protein and mRNA) in liver was similar in both genotypes. In inguinal adipose tissue, AGT cell content was similar in suckling and weaned pups in lean rats, whereas it continuously increased with age in obese rats. AGT amount in adipocytes was unaffected by the genotype until weaning. Thereafter, adipocytes from obese rats displayed a significant increase in AGT content that was strengthened with age. Compared with the cell content, the amount of secreted AGT over 24 h was higher, and a genotype effect was observed as early as 14 days of age. Using fat cell populations differing by size, we showed that this AGT oversecretion was not solely related to adipocyte hypertrophy. Our results demonstrate that the fa genotype exerts a control on the production of AGT in a tissue-specific manner, suggesting a local role of AGT in the overdevelopment of adipose tissue.
Dynamic changes in G alpha i-2 levels in rat hearts associated with impaired heart function after myocardial infarction
Authors: Shi B. , Heavner J. E. , McMahon K. K. , Spallholz J. E.
Published: December 2017

The objective of this study was to determine if levels and function of Gs alpha and G alpha i-2 in rat hearts change over time following acute myocardial infarction (MI), and if so, whether the changes in G proteins are associated with changes in heart function. As compared with sham-operated controls, the G alpha i-2 level of MI rats did not change at day 1, increased by 64% at day 3 (P < 0.01) and by 55% at day 9 (P < 0.05) accompanied by reduced adenylyl cyclase activity, and returned to control by day 21. By contrast, the Gs alpha level did not change at any time. Cardiac function in MI animals was markedly impaired at days 1, 3, and 9 as evidenced by substantial elevation in LVEDP and reduction in maximum rates of pressure development and relaxation, and was partially restored at day 21. Increased G alpha i-2 level in MI rats correlated significantly to severity of impaired cardiac function. The results show a three-phase dynamic pattern in G alpha i-2 level following acute MI: a lag phase, an increased expression phase associated with marked impairment of heart function, and a late phase in which the expression returns to control level accompanied by partially restored cardiac function.
Overdrive suppression of spontaneously beating chick heart cell aggregates: experiment and theory
Authors: Kunysz A. , Glass L. , Shrier A.
Published: December 2017

In spontaneously beating chick heart cell aggregates, sustained periodic stimulation at a rate faster than the intrinsic frequency is generally followed by a transient slowing of the automatic rhythm called "overdrive suppression." We characterize the qualitative aspects of overdrive suppression using three sets of experimental protocols: 1) stimulation at a fixed frequency with various numbers of stimuli, 2) stimulation at different frequencies, 3) stimulation with different intensities. We develop a mathematical model based on a system of nonlinear ordinary differential equations to account for the experimental observations. The main idea of the model is that overdrive suppression arises as a result of a hyperpolarizing current that is induced by action potentials. This work shows that the frequency of action potentials is the major determinant of overdrive suppression. Consequently, during periodic pacing of spontaneous oscillators at different rates, the fastest frequency where 1:1 entrainment can be maintained is associated with maximal overdrive suppression. This type of model is complementary to the development of a rigorous ionic model and can help provide insight into the physiological mechanisms of overdrive suppression.
Downregulation of atrial natriuretic factor clearance receptors in experimental chronic renal failure rats
Authors: Luk J. K. , Wong E. F. , Wong N. L.
Published: December 2017

Studies were performed to examine the changes of renal ANF second messenger guanosine 3',5'-cyclic monophosphate (cGMP) responses and receptor properties in chronic renal failure (CRF). Five-sixths-nephrectomized and sham-operated Wistar rats were used. The glomerular filtration rate was decreased in the five-sixths-nephrectomized rats, which also had significantly higher plasma blood urea nitrogen and plasma atrial natriuretic factor (ANF) levels (148.5 +/- 10.2 vs. 115.7 +/- 7.3 pg/ml, P = 0.01) than the sham rats. In vitro ANF-stimulated cGMP accumulations in glomeruli of five-sixths-nephrectomized rats were higher than controls. Radioligand-binding experiments showed downregulation of the total ANF receptor in both acid and nonacid wash CRF glomeruli (nonacid wash: 189 +/- 25 vs. 362.8 +/- 52.8 fmol/mg protein, P < 0.05; acid wash: 449.8 +/- 67 vs. 652.7 +/- 52.5 fmol/mg protein, P < 0.05). No change in receptor densities was observed in the des(Gln18,Ser19,Gly20,Leu21)atrial natriuretic peptide-(4--23)-NH2-resistant receptors between sham and CRF rat glomeruli. Therefore, downregulation of ANF clearance receptors exists in CRF rat glomeruli, and this is associated with the exaggerated ANF-stimulated cGMP response in these CRF glomeruli. Hypersensitivity of CRF rat to ANF, together with high plasma ANF levels and downregulation of clearance receptor, may contribute to increased sodium excretion in CRF.
Fibroblast proliferation during myocardial development in rats is regulated by IGF-1 receptors
Authors: Reiss K. , Cheng W. , Kajstura J. , Sonnenblick E. H. , Meggs L. G. , Anversa P.
Published: December 2017

To determine whether the growth of cardiac fibroblasts during development is modulated by the insulin-like growth factor (IGF)-1 receptor (IGF-1R), the expression of IGF-1, IGF-2, and IGF-1R was determined in fibroblasts from fetal and postnatal hearts. The expression of proliferating cell nuclear antigen (PCNA) and DNA polymerase-alpha was also evaluated in combination with the estimation of DNA replication. In comparison with fetal hearts, at postnatal day 21, fibroblast expression of IGF-1R mRNA, IGF-2, PCNA, and DNA polymerase-alpha was reduced by 77, 70, 80, and 86%, respectively. Moreover, IGF-1R protein decreased by 48% at 21 days. Bromodeoxyuridine labeling decreased by 88 and 89% in the left and right ventricle, respectively, at this time. Two different antisense oligodeoxynucleotides to IGF-1R reduced DNA replication by 60 and 44% in fibroblasts in culture. In addition, this intervention markedly attenuated the growth response of fibroblasts to IGF-1 or serum. In conclusion, the IGF-1R system appears to play a major role in the regulation of fibroblast growth in the heart in vivo.
Changes in contractility and afterload have only slight effects on subendocardial systolic flow impediment
Authors: Iwanaga S. , Ewing S. G. , Husseini W. K. , Hoffman J. I.
Published: December 2017

To test the hypothesis that contractility did not greatly influence systolic impediment to subendocardial flow we perfused the coronary artery at 70 mmHg and altered aortic pressure in 7 dogs and contractility in another 7. We measured myocardial systolic flow impediment (SFI) by comparing regional flows while beating and during asystole. Cardiac contraction impeded 29% of subendocardial asystolic flow, which was not affected by either intervention. In subepicardium, contraction increased flow by 25%, but dobutamine impeded systolic flow. Subepicardial SFI was only 16% of subendocardial SFI. Dobutamine slightly decreased estimated percent systolic myocardial blood flow (%SMBF) in subendocardium (+/- 12%) but decreased subepicardial %SMBF (45.5 to 17.4%). Phasic coronary flow pulsatility increased more with dobutamine than increased afterload, and pulsatility and SFI correlated only in subepicardium. Systolic-to-total coronary flow ratio and %SMBF did not correlate closely in subendocardium. SFI was most prominent in the subendocardium, whereas subepicardial SFI mainly determined epicardial coronary flow pulsatility. We conclude that the effects of contractility changes differ when evaluating regional SFI vs. phasic flow pulsatility.
Adhesion of flowing neutrophils to cultured endothelial cells after hypoxia and reoxygenation in vitro
Authors: Rainger G. E. , Fisher A. , Shearman C. , Nash G. B.
Published: December 2017

Using a novel in-line deoxygenating system linked to an in vitro flow-based adhesion assay and video microscopy, we have studied neutrophil recruitment and migration after hypoxia and reoxygenation of cultured human umbilical vein endothelial cells (HUVEC). Unstimulated purified neutrophils were perfused over reoxygenating HUVEC immediately after various periods of endothelial hypoxia. Adhesion to HUVEC was dependent on the duration of hypoxia, with 30, 60, and 100 min of exposure causing graded increments in neutrophil recruitment. The degree of hypoxia also markedly influenced the endothelial response. Severe hypoxia (O2 < 2.5%) induced stationary attachment and then migration of neutrophils, in contrast to rolling adhesion alone under a less intense regime (O2 = 2.5-4.0%). Judged from studies with monoclonal antibodies, P-selectin was essential for adhesion after severe hypoxia, and neutrophil immobilization was attributable to the activation of neutrophil beta 2-integrin. Perfusion of neutrophils with an antibody against interleukin-8 or a platelet-activating factor antagonist reduced levels of adhesion. However, IL-8 appeared to be the dominant agent involved in the immobilization from flow, whereas platelet-activating factor was the more potent agent involved in initiating subendothelial migration. Thus endothelial cells alone can initiate all stages of adhesion and migration of flowing neutrophils after hypoxia and reperfusion.
Serum tumor necrosis factor-alpha does not mediate endotoxin-induced myocardial depression in rabbits
Authors: Nishikawa Y. , Mathison J. , Lew W. Y.
Published: December 2017

Tumor necrosis factor-alpha (TNF-alpha) is an endogenous mediator for several effects of endotoxin. To evaluate whether TNF-alpha mediates endotoxin-induced left ventricular (LV) dysfunction, we measured LV function (sonomicrometers) and serum TNF-alpha (cytolytic assay) in anesthetized rabbits given endotoxin (100 micrograms/kg iv). In the control group (n = 8), systolic depression (defined by a > 10% increase in end-systolic volume at a matched end-systolic pressure) developed in four rabbits and diastolic dilation (> 10% increase in end-diastolic volume at a matched end-diastolic pressure) developed in three rabbits. Neither the increase in end-systolic volume nor the increase in end-diastolic volume correlated with the increase in TNF-alpha, which reached a peak of 2,875 +/- 762 U/ml. In a second group of rabbits (n = 7), a goat polyclonal anti-rabbit antibody to TNF-alpha was given 30-60 min before endotoxin. Anti-TNF-alpha antibody alone did not alter LV function. Although the TNF-alpha response to endotoxin was effectively blunted (peak TNF-alpha remained < 100 U/ml), all seven rabbits developed systolic depression (P = 0.08 compared with control group) and diastolic dilation (P = 0.03). We conclude that serum TNF-alpha does not mediate endotoxin-induced LV systolic depression or diastolic dilation in this model.
Early changes in protein synthesis in epicardial coronary artery of pressure-overloaded heart
Authors: Gerova M. , Pechanova O. , Stoev V. , Kittova M. , Bernatova I. , Barta E.
Published: December 2017

In anesthetized dogs, a 4-h, approximately 30% increase in blood pressure induced by constriction of the abdominal aorta brought about an increase in the total RNA content in the left anterior descending coronary artery (LADCA) and the left ventricular (LV) myocardium (9.05 +/- 1.72 and 11.06 +/- 4.68%, respectively) but not in the left circumflex coronary artery (LCCA). Under the same experimental conditions, [14C]leucine incorporation increased in LADCA and LV myocardium (45.34 +/- 13.54 and 58.07 +/- 11.91%, respectively), but not in LCCA. The data indicate an early shift in protein synthesis in LADCA and simultaneously in the myocardium during a short-term pressor event. The difference in the shift of protein synthesis in the two main branches of the left coronary artery was related to the quantitatively different deformation of the LADCA and LCCA due to different deformation of the underlying myocardium and/or of the annulus fibrosus atrioventricularis during changes in the left or right ventricle volume [M. Gerova, E. Barta, M. Stolarik, and J. Gero, Am. J. Physiol. 262 (Heart Circ. Physiol. 31): H1049-H1053, 1992]. The results support the hypothesis that the deformation and/or rate of deformation of cells in the coronary wall may trigger an increase in protein synthesis. Changes in protein synthesis in the myocardium and LADCA were found to be reversible 2 h after releasing the aortic constriction.
A new technique for study of impact of arterial elasticity on left ventricular mass in rats
Authors: Amin F. , Niederhoffer N. , Tatchum-Talom R. , Makki T. , Guillou J. , Tankosic P. , Atkinson J.
Published: December 2017

We investigated possible links between left ventricular mass and central arterial elasticity in the adult spontaneously hypertensive rat (SHR) and in a subgroup of SHR in which blood pressure was normalized by chronic antihypertensive drug treatment; results were compared with those of age-matched normotensive Wistar-Kyoto rats. Two indexes of arterial elasticity, based on the measurement of aortic pressure pulse wave velocity, were used. First, the slope relating carotidofemoral pulse wave velocity to blood pressure in the phenylephrine-infused pithed preparation was used as a pressure-independent index of wall elasticity. Second, to account for hypertension- and treatment-induced aortic remodeling, elastic modulus was determined from the pulse wave velocity recorded when blood pressure reached that measured in awake animals before anesthesia and pithing, together with values for wall thickness and lumen diameter evaluated by histomorphometric analysis after in situ fixation at the same pressure. In control SHR, regression analysis of variance revealed significant correlations between left ventricular mass and both wave velocity/pressure slope and elastic modulus. Chronic antihypertensive treatment normalized all three parameters. In conclusion, this new technique provides experimental evidence of a link between left ventricular mass and central arterial elasticity.
Nitric oxide synthase does not participate in negative inotropic effect of acetylcholine in frog heart
Authors: Mery P. F. , Hove-Madsen L. , Chesnais J. M. , Hartzell H. C. , Fischmeister R.
Published: December 2017

In the heart, the parasympathetic neurotransmitter acetylcholine (ACh) reduces the force of contraction. Although the effect of ACh can be partly explained by an inhibition of adenylyl cyclase, some of the effects of ACh may also be mediated via stimulation of nitric oxide synthase (NOS) and production of guanosine 3', 5'-cycle monophosphate (cGMP). NOS inhibitors can prevent the negative chronotropic effect of ACh on spontaneously beating cardiomyocytes and suppress the inhibition of the L-type calcium current (ICa) by ACh in sinoatrial myocytes. This pathway may be relevant not only to the chronotropic effect of ACh but also to its inotropic effect, because ACh, NO, and cGMP regulate the force of contraction and ICa in the cardiac ventricle. Here we report the effects of L-arginine (L-Arg), the substrate of NOS, and NG-monomethyl-L-arginine (L-NMMA) and NG-nitro-L-arginine (L-NNA), two NOS inhibitors, on muscarinic effects in the cardiac ventricle. We found that L-Arg, L-NMMA, and L-NNA have no effect on the muscarinic inhibition of ICa in isolated frog myocytes. In addition, these compounds have no significant effects on basal ICa or beta-adrenergic stimulation of ICa. L-Arg and its analogues did not change the negative inotropic effect of ACh in frog ventricular fibers. Basal active tension and the positive inotropic effect of isoproterenol, a beta-adrenergic agonist, also were unaffected. We conclude that NOS in not involved in muscarinic inhibition of ICa in isolated from ventricular myocytes or the negative inotropic effect of ACh in the frog ventricle.
Postischemic vasodilation in human forearm is dependent on endothelium-derived nitric oxide
Authors: Meredith I. T. , Currie K. E. , Anderson T. J. , Roddy M. A. , Ganz P. , Creager M. A.
Published: December 2017

Although endothelium-derived nitric oxide contributes to basal vascular tone, little is known about its role in regulating blood flow during changes in metabolic supply and demand. We examined the contribution of endothelium-derived nitric oxide to reactive hyperemia in the forearm of 20 normal subjects (12 women, 8 men) aged 27 +/- 4 yr (means +/- SD), using the nitric oxide synthase inhibitor, NG-monomethyl-L-arginine (L-NMMA). Forearm ischemia was induced by suprasystolic blood pressure cuff inflation for 5 min, and the subsequent hyperemic flow was recorded for 5 min using venous occlusion strain-gauge plethysmography. The efficacy of nitric oxide blockade was tested by comparing the dose-response relationship to the endothelium-dependent agonist, acetylcholine (3, 10, and 30 mg/min), before and after intra-arterial infusion of up to 2,000 mg/min of L-NMMA. L-NMMA produced a significant downward and rightward shift in the dose-response relationship to acetylcholine and a 39% reduction in response to the maximum dose (P < 0.001). In the presence of L-NMMA, peak hyperemic flow was reduced 16% (26.5 +/- 2.1 to 22.3 +/- 1.5 ml.min-1.100 ml of forearm-1, P < 0.03), and the minimum forearm vascular resistance was increased 22.8% (3.5 +/- 0.3 to 4.3 +/- 0.4 mmHg.ml-1.min.100 ml, P < 0.02). Total hyperemia, calculated from the area under the flow vs. time curve, at 1 and 5 min after cuff release was 17 and 23% less, respectively (13.6 +/- 1.2 vs. 11.3 +/- 1.1 and 31.8 +/- 2.7 vs. 24.6 +/- 1.8 ml/100 ml, P < 0.002), following L-NMMA. These data suggest that endothelium-derived nitric oxide plays a role in both reactive hyperemia and in the maintenance of the hyperemic response following ischemia in the forearm.
Changes in endothelial actin cytoskeleton at leakage sites in the rat mesenteric microvasculature
Authors: Thurston Gavin , Baldwin Ann L. , Wilson Lisa M.
Published: December 2017

Pages H316–H329: Gavin Thurston, Ann L. Baldwin, and Lisa M. Wilson. “Changes in endothelial actin cytoskeleton at leakage sites in the rat mesenteric microvasculature.” The volume number in the abstract line was incorrectly cited and should appear as the following.

Thurston, Gavin, Ann L. Baldwin, and Lisa M. Wilson.Changes in endothelial actin cytoskeleton at leakage sites in the rat mesenteric microvasculature. Am. J. Physiol. 268 ( Heart Circ. Physiol. 37): H316–H329, 1995.
Nitric oxide synthase inhibition partially prevents decreased LV contractility during endotoxemia
Authors: Herbertson M. J. , Werner H. A. , Walley K. R.
Published: December 2017

Decreased contractility of myocytes after cytokine exposure can be prevented by nitric oxide synthase inhibition. Whether this is true in an intact animal model of sepsis is unknown. Anesthetized pigs were pretreated with saline or a nitric oxide synthase inhibitor, N omega-nitro-L-arginine, and then treated with saline or endotoxin. We measured hemodynamics and left ventricular pressures (Millar catheter) and volumes (conductance catheter). Left ventricular contractility was assessed using the slope (E(max)) of the end-systolic pressure-volume relationship. Four hours after endotoxin infusion, E(max) had decreased by 44 +/- 5% (P < 0.05) and mean arterial pressure had decreased by 30 +/- 10% (P < 0.05). Pretreatment with N omega-nitro-L-arginine significantly reduced the decrease in E(max) to 28 +/- 3% (P < 0.05) and prevented the decrease in mean arterial pressure. However, it also raised pulmonary arterial pressure. We conclude that nitric oxide contributes to the early decrease in left ventricular contractility after endotoxin in the intact animal. However, the vascular effects of nitric oxide synthase inhibition increase right and left ventricular afterloads, which were detrimental to cardiac function.
PAF attenuates endothelium-dependent coronary arteriolar vasodilation
Authors: DeFily D. V. , Kuo L. , Chilian W. M.
Published: December 2017

Platelet-activating factor (PAF) has been reported to play a role in neutrophil activation, microvascular permeability, and endothelial dysfunction in a variety of vascular preparations. Although a majority of the effects of PAF are thought to be mediated by the activation of neutrophils, it is unclear the extent to which the deleterious effects of PAF extend to coronary resistance vessels. Therefore, the purpose of this study was to determine whether PAF causes coronary arteriolar endothelial dysfunction in vivo and whether this dysfunction is independent of activated neutrophils. To test these hypotheses, we measured changes in coronary arteriolar diameter to endothelium-dependent and -independent dilators in vivo by measuring coronary microvascular diameters in a beating canine heart using intravital videomicroscopy following intracoronary infusion of PAF (20 ng.kg-1.min-1). Changes in coronary arteriolar diameter following incubation with PAF were also measured in isolated coronary arterioles. In vivo, incubation with PAF resulted in a significant attenuation of endothelium-dependent dilation to intracoronary acetylcholine (0.1 microgram.kg-1.min-1, 39 +/- 7 vs. 20 +/- 3% dilation) and serotonin (1 microgram.kg-1.min-1, 29 +/- 6 vs. 2 +/- 2% dilation). Papaverine-induced relaxation, however, was unchanged. Likewise, in vitro relaxation to serotonin (10 nM) was significantly reduced (38 +/- 4 vs. 3 +/- 5%) following treatment with PAF, whereas nitroprusside (10 nM)-induced relaxation was unchanged. Because PAF impaired endothelium-dependent arteriolar dilation both in vivo and in vitro, we conclude that the presence of activated neutrophils is not required for PAF-induced coronary microvascular dysfunction.
Characterization of a TTX-sensitive Na+ current in pacemaker cells isolated from rabbit sinoatrial node
Authors: Muramatsu H. , Zou A. R. , Berkowitz G. A. , Nathan R. D.
Published: December 2017

A tetrodotoxin (TTX)-sensitive Na+ current (iNa) was investigated in single pacemaker cells after 1-4 days in culture. Ruptured-patch and perforated-patch whole cell recording techniques were used to record iNa and spontaneous electrical activity, respectively. For seven cells exposed to 20 mM Na+ (22-24 degrees C) and held at -98 mV (25% of the channels inactivated), the uncorrected maximum iNa was -39 +/- 10 pA/pF at -29.1 +/- 2.4 (SE) mV, maximum conductance was 0.9 +/- 0.2 nS/pF (1.6 +/- 0.2 mS/cm2). Half-activation and inactivation potentials were -41.4 +/- 2.0 and -90.6 +/- 2.5 mV, and the corresponding slope factors were 6.0 +/- 0.4 and 6.4 +/- 0.6 mV. Inactivation and recovery from inactivation were best fit by sums of two exponentials. During action potential clamp, a TTX-sensitive compensation current accounted for 55% of the upstroke velocity. The results suggest that iNa contributes significantly to the action potential in some nodal pacemaker cells, and the characteristics of iNa are similar to those of atrial and ventricular myocytes.
Low external pH limits cell death of energy-depleted cardiomyocytes by attenuation of Ca2+ overload
Authors: Atsma D. E. , Bastiaanse E. M. , Van der Valk L. , Van der Laarse A.
Published: December 2017

We studied the effect of external pH (pHe) on cell injury, ATP content, and intracellular concentration of Ca2+ ([Ca2+]i), Na+ ([Na+]i), and H+ (pHi) during metabolic inhibition (NaCN + 2-deoxyglucose) in neonatal rat cardiomyocytes. Cell death during metabolic inhibition decreased at pHe < 7.4, with almost no cell death at pHe 6.0. Lowering pHe resulted in only temporary ATP conservation. During metabolic inhibition at pHe 7.4, [Ca2+]i rose from 86 +/- 44 nM to 2.5 +/- 0.4 microM, but at pHe 6.0, [Ca2+]i rose to only 510 +/- 215 nM. During metabolic inhibition at pHe 7.4, pHi decreased from 7.25 +/- 0.06 to 6.82 +/- 0.16, but at pHe 6.0, pHi decreased to 6.34 +/- 0.17. During metabolic inhibition at pHe 7.4, [Na+]i increased from 9.1 +/- 0.86 to 26.1 +/- 4.1 mM. At pHe 6.0, [Na+]i rose more rapidly, to 27.3 +/- 3.5 mM. At pHe < 7.4, sarcolemmal Na+/Ca2+ exchanger activity, involved in the development of Ca2+ overload, was decreased, as assessed during Na(+)-free incubation. We conclude that low pHe protects cardiomyocytes during metabolic inhibition by limiting Ca2+ overload via Na+/Ca2+ exchanger inhibition.
Stability of high-energy phosphates in right ventricle: myocardial energetics during right coronary hypotension
Authors: Itoya M. , Mallet R. T. , Gao Z. P. , Williams A. G. , Downey H. F.
Published: December 2017

This study was conducted to determine if mechanisms that reduce right coronary (RC) blood flow (RCBF) and right ventricular (RV) oxygen consumption (MVO2) during moderate RC hypotension preserve RV high-energy phosphates. RC arteries of anesthetized dogs were cannulated and perfused with arterial blood supplied by a pressurized extracorporeal circuit. RC perfusion pressure (RCPP) was either kept constant at 100 mmHg or reduced to 60 or 30 mmHg for 20 min followed by a freeze-clamp biopsy of RV. Left ventricular (LV) biopsy was also performed to compare energy metabolism between RV and LV.RCBF and MVO2 significantly decreased when RCPP was reduced to 60 mmHg, but RV segment shortening (%SS) was unchanged; ATP, creating phosphate (CrP) and phosphorylation state of CrP ([CrP]/[Cr][Pi]) did not differ from control values. RV %SS, CrP, and phosphorylation state fell markedly at 30 mmHg RCPP. At 100 mmHg RCPP, CrP phosphorylation state in RV was only 35% of that in LV. These results indicate that RV increases its energetic efficiency without significant changes in high-energy phosphates or CrP phosphorylation state during moderate RC hypotension. Furthermore, the RV myocardium maintains a much lower energy level than LV myocardium, commensurate with its lower energy requirements.
In vivo and in vitro vasoactive reactions of coronary arteriolar microvessels to nitroglycerin
Authors: Jones C. J. , Kuo L. , Davis M. J. , Chilian W. M.
Published: December 2017

The actions of nitroglycerin on the coronary microcirculation are controversial, with some laboratories reporting that coronary arterioles dilate to the drug and others reporting that they do not. Our goal was to reconcile these disparate observations. Specifically, we hypothesized that dilation of coronary arterioles by nitroglycerin is overwhelmed by intrinsic autoregulatory escape mechanisms. Accordingly, we projected that coronary arterioles would show transient, but not sustained, dilation to nitroglycerin in vivo. Furthermore, we hypothesized that isolated coronary arterioles would show sustained dilation to the drug, because intrinsic escape mechanisms would be absent under these conditions. To test these hypotheses, we measured diameter changes of canine coronary microvessels in vivo during continuous nitroglycerin administration (intracoronary infusion or epicardial suffusion) using intravital fluorescent microscopy (n = 17 dogs) at two time points: early (1-3 min), when coronary artery blood flow velocity was increased, and late (15-20 min), after blood flow velocity returned to control. Tb study responses of coronary arterioles in the absence of autoregulatory influences, we measured the diameter of isolated canine coronary arterioles to varying doses of nitroglycerin (n = 8 vessels, maximal diameter 81 +/- 4 microns). During the early phase of nitroglycerin infusion (1,3, and 10 micrograms.kg-1.min-1), coronary arterioles dilated by 4 +/- 1, 7 +/- 2, and 13 +/- 2% (all P < 0.05), whereas small arteries dilated by 1 +/- 2, 3 +/- 1, and 4 +/- 1%, respectively (P < 0.05 for the higher doses). Coronary artery blood velocity measured increased by 45 +/- 15% (3 micrograms.kg-1.min-1, P < 0.05). Suffusion of nitroglycerin (10(-5) M) dilated coronary arterioles, but not small arteries, by 17 +/- 5% (P < 0.05) between 1 and 3 min. After 15-20 min of nitroglycerin (3 micrograms.kg-1.min-1 by intracoronary infusion), diameters of coronary arterioles and coronary artery blood velocity returned to control, whereas dilation of small arteries remained significant at 4 +/- 1%. Coronary arteriolar dilation by epicardial suffusion of nitroglycerin also waned to control values by 15-20 min, whereas dilation of small arteries was observed: 5 +/- 2% (P < 0.05). In vitro, nitroglycerin caused dose-dependent dilation of coronary arterioles to their maximal diameter, which was sustained for 20 min. Thus nitroglycerin dilates coronary arterioles and small arteries. The dilation in vivo is transient for arterioles but sustained for arteries. In vitro, the dilation is sustained. Because microvessels in vitro are capable of sustaining dilation for 20 min, we conclude that the waning of arteriolar dilation in vivo is related to autoregulatory escape from dilation by nitroglycerin.
Heat stress protects the perfused rabbit heart from complement-mediated injury
Authors: Gralinski M. R. , Black S. C. , Stancato L. F. , Kilgore K. S. , Campau P. A. , Park J. L. , Ozeck M. J. , Pratt W. B. , Lucchesi B. R.
Published: December 2017

We determined if heat stress induction of heat shock protein (HSP) 70 modulates complement activation in an experimental model of xenograft rejection. Male New Zealand White rabbits were heat stressed (core body temperature to 42 degrees C for 15 min; n = 9). Control rabbits (n = 13) were not exposed to heat stress. Hearts were removed 18 h later and perfused by the Langendorff method. After equilibration, human plasma (source of human complement) was added to the perfusion medium. Hemodynamic variables recorded during perfusion with human plasma were improved in hearts from heat-stressed animals compared with control hearts. Assembly of the soluble membrane attack complex was reduced in the interstitial fluid effluent from the heat-stressed hearts. Electron microscopic evidence of ultrastructural changes was attenuated in the hearts from heat-stressed rabbits. Myocardial tissue from heat-stressed animals exhibited an increase in inducible HSP 70 that was virtually absent in the hearts of control rabbits. Previous whole body hyperthermia protects the rabbit heart against the detrimental effects of heterologous plasma, suggesting that heat-stress induction of HSP 70 limits the extent of complement activation by a discordant vascularized tissue (xenograft). Induction of heat stress proteins by the donor organ might be an important mechanism affecting the outcome of xenograft transplants.
Survival of metabolically inhibited ventricular myocytes is enhanced by inhibition of rigor and SR Ca2+ cycling
Authors: Koyama T. , Boston D. , Ikenouchi H. , Barry W. H.
Published: December 2017

During severe ATP depletion, sarcolemmal rupture resulting from rigor- and/or Ca(2+)-induced myofilament force development is considered to be an important cause of irreversible cell injury. Recent experiments in our laboratory demonstrated that during prolonged metabolic inhibition (MI) in adult rabbit ventricular myocytes, in which rigor was prevented by exposure to 30 mM 2,3-butanedione monoxime (BDM), cyclic uptake and release of cystolic Ca2+ occurred and was associated with strong phasic contractions. To investigate the relative contribution of this sarcoplasmic reticulum Ca2+ cycling and associated force development to energy depletion injury, the effects of BDM together with 7 mM caffeine were examined in isolated rabbit ventricular myocytes subjected to MI with 2 mM NaCN and 20 mM 2-deoxyglucose (2-DG). During 90 min of MI with CN and 2-DG, no cells retained a rod shape in the absence of BDM or caffeine. In the presence of both 30 mM BDM and 7 mM caffeine during MI, preservation of rod morphology was enhanced, and 52 +/- 6.2% of cells retained a rod shape 48 h after metabolic inhibition and had normal ATP content and resting membrane potential. Both systolic and diastolic functions of cells that survived MI, however, were impaired. We conclude that exposure to caffeine together with BDM markedly enhances survival of myocytes during severe prolonged ATP depletion. After recovery, these isolated myocytes show some characteristics of stunning.
Differences between cytokine effects in the microcirculation of the rat
Authors: Baudry N. , Rasetti C. , Vicaut E.
Published: December 2017

We studied by in vivo microscopy in rat cremaster muscle the acute and delayed effects of short exposure to tumor necrosis factor (TNF), interleukin (IL)-1 beta, and IL-6 on basal tone and vascular reactivity of second- to fourth-order arterioles (A2-A4). A 20-min exposure to recombinant human (rh) TNF (0.1-10 ng/ml) induced a significant arteriolar vasodilation, but no significant changes in basal tone were found after exposure to the same doses of IL-1 beta. In contrast, the same exposure to IL-6 (0.1-10 ng/ml) induced a significant dose-dependent vasoconstriction (i.e., 8, 15, and 21% at 10 ng/ml in A2-A4 arterioles, respectively). This vasoconstriction was inhibited by the thromboxane A2 receptor antagonist SQ-29548. We did not find any significant effect of rhTNF or IL-6 on vascular reactivity to norepinephrine immediately after exposure to these two cytokines or 100 min after the end of the exposure. Contrastingly, a large dose-dependent decrease in reactivity to norepinephrine was found immediately after exposure to IL-1 beta and still persisted 100 min after the end of the exposure. Such a decrease was not found for the vasoconstriction in response to KCl. We conclude that, at the microvascular level, large differences exist between the three cytokines generally considered to mediate the harmful cardiovascular effects in sepsis. 1) TNF but not IL-1 beta is responsible for a vasodilatory effect, whereas the effect of IL-6 is a thromboxane A2-mediated vasoconstriction. 2) Short exposure to IL-1 beta but not to rhTNF or IL-6 diminishes the response of the arterioles to norepinephrine but not to KCl.
Modeling and simulation of illumination effects for evaluation of microvessels of the conjunctiva
Authors: Wick C. E. , Loew M. H. , Kurantsin-Mills J.
Published: December 2017

We present the development of a comprehensive model that was undertaken to determine the relationships between the components of an image and the light intensity values present in the image of the microvessels of translucent tissues such as the bulbar conjunctiva. Experiments were conducted during the modeling process by use of a cylindrical microvessel embedded in a diffuse medium (phantom) on a reflecting background to affirm model components and simulations. The three-dimensional model was reduced to a single illumination plane with four regions of interest and modeled as Lambertian radiators and surfaces. The modeling showed that the top of the cylinder and its immediate vicinity are diffuse reflectors of light from the source plus light reflected from the background. The limbus of the cylinder is a diffuse reflector of the source and background illumination and a specular reflector of background reflections that achieve a high grazing angle with the cylinder. The immediate vicinity of the cylinder receives direct illumination from the source, but the light is partially obscured by the cylinder. The region beyond the shadow of the cylinder is a diffuse reflector of the overhead light. The diffuse medium additionally reflects the source and also attenuates the illumination reaching the other compo- rents of the scene. The direct and reflected illumination at each region of the model was calculated by use of specific geometric relationships. To verify those calculations, we analyzed a video simulation for the effects of different illumination conditions and their contributing elements. Intensity values were calculated from the relative reflectivity data determined from the video signals. The illumination values at the points along the line at the meridian of the cylinder were due to its reflectivity and also that of the medium. Similarly, the values of points distant from the shadow of the cylinder were due to the reflectivity of the background and the medium. The excellent agreement between the model and the phantom provides a foundation for the detection and precise measurement of microvessel dimensions within a diffuse medium. The additional ability to compute relative depth, from a single view, also permits discrimination between neighboring microvessels in complex images.
Preconditioning prevents chronic reperfusion-induced coronary endothelial dysfunction in rats
Authors: Kaeffer N. , Richard V. , Francois A. , Lallemand F. , Henry J. P. , Thuillez C.
Published: December 2017

Experiments were designed to test whether preconditioning protects against chronic endothelial injury after ischemia and reperfusion. Coronary arteries were isolated from rats subjected to sham surgery or 20 min of ischemia followed by 1 h, 1 day, 1 wk, or 1 mo of reperfusion without or with preconditioning. The endothelium-dependent relaxations to acetylcholine (ACh; assessed in vitro) were markedly reduced after ischemia and 1 h of reperfusion (31 +/- 6 vs. 57 +/- 6% in sham; P < 0.01) and did not recover after longer durations of reperfusion (1 mo: 32 +/- 5 vs. 56 +/- 2%; P < 0.01). The impaired response to ACh was restored by preconditioning at all time points (1 h: 53 +/- 6; 1 mo: 65 +/- 4%). After 1 mo, the potency of ACh in preconditioned arteries was also increased compared with that in sham animals. Electron microscopy showed marked endothelial damage after 1 h of reperfusion and signs of regenerated endothelium after 1 mo of reperfusion. Both acute and chronic ultrastructural changes were prevented by preconditioning. Thus preconditioning, in addition to protecting myocardial cells, also protects against chronic reperfusion-induced endothelial injury, both in terms of functional and structural changes.
Nitric oxide modulates arteriolar responses to increased sympathetic nerve activity
Authors: Nase G. P. , Boegehold M. A.
Published: December 2017

The purpose of this study was to determine whether arteriolar responses to increased sympathetic nerve activity are limited by the actions of endogenous nitric oxide. Intravital microscopy was used to examine diameter responses of small feed arteries (SFA), first-order arterioles (1A) and second-order arterioles (2A) to perivascular sympathetic nerve stimulation in the superfused rat small intestine. Stimulation induced a frequency-dependent constriction in all vessel types that was completely abolished by the alpha-adrenoceptor antagonist phentolamine (10(-6) M). In SFA and 1A, the magnitude of sympathetic constriction was increased significantly in the presence of the nitric oxide synthase inhibitor NG-monomethyl-L-arginine(L-NMMA, 10(-4) M). In SFA (n = 7), stimulation at 3, 8, and 16 Hz induced constrictions of 11 +/- 1, 28 +/- 4, and 42 +/- 3%, respectively, under the normal superfusate vs. 28 +/- 3, 46 +/- 5, and 76 +/- 3% in the presence of L-NMMA. For 1A (n = 7), stimulation induced constrictions of 10 +/- 1, 27 +/- 4, and 37 +/- 3% under the normal superfusate vs. 24 +/- 2, 47 +/- 3, and 72 +/- 4% in the presence of L-NMMA. The effect of L-NMMA on sympathetic constriction in SFA (n = 7) was completely reversed by the additional presence of 5 x 10(-3) M L-arginine in the superfusate. These results suggest that endogenous nitric oxide activity can attenuate sympathetic neurogenic constriction in the intestinal microvasculature.