bluuebunny/crossref_metadata_2025 huggingface dataset:
Endogenous heme oxygenase prevents impairment of cerebral vascular functions caused by seizures
Authors: Carratu Pierluigi , Pourcyrous Massroor , Fedinec Alex , Leffler Charles W. , Parfenova Helena
Published: March 2015

In newborn pigs, the mechanism of seizure-induced cerebral hyperemia involves carbon monoxide (CO), the vasodilator product of heme catabolism by heme oxygenase (HO). We hypothesized that seizures cause cerebral vascular dysfunction when HO activity is inhibited. With the use of cranial window techniques, we examined cerebral vascular responses to endothelium-dependent (hypercapnia and bradykinin) and endothelium-independent (isoproterenol and sodium nitroprusside) dilators during the recovery from bicuculline-induced seizures in saline controls and in animals pretreated with a HO inhibitor, tin protoporphyrin (SnPP). SnPP (3 mg/kg iv) blocked dilation to heme and reduced the CO level in cortical periarachnoid cerebrospinal fluid, indicating HO inhibition in the cerebral microcirculation. In saline control piglets, seizures increased the CO level, which correlated with the time-dependent cerebral vasodilation; during the recovery (2 h after seizure induction), responses to all vasodilators were preserved. In SnPP-treated animals, cerebral vasodilation and the CO responses to seizures were greatly reduced, and cerebral vascular reactivity was severely impaired during the recovery. These findings suggest that HO in the cerebral microcirculation is rapidly activated during seizures and provides endogenous protection against seizure-induced vascular injury.
Protein kinase C-δ mediates adenosine A3receptor-induced delayed cardioprotection in mouse
Authors: Zhao Ting Cun , Kukreja Rakesh C.
Published: March 2015

We investigated the role of protein kinase C in adenosine A3receptor (A3AR)-induced delayed cardioprotection in the mouse heart. Mice were treated with selective A3AR agonist N6-(3-iodobenzyl)adenosine-5′- N-methyluronamide (IB-MECA). Twenty-four hours later, hearts were perfused in the Langendorff mode and subjected to 30 min of global ischemia and 30 min of reperfusion. Infarct size was determined by computer morphometry of tetrazolium-stained sections, and ventricular function was monitored by inserting a fluid-filled balloon into the left ventricle (LV). Chelerythrine chloride (CHE, 5.0 mg/kg) and rottlerin (Rot, 0.3 mg/kg) were given 30 min before IB-MECA to block total and PKC-δ isoforms, respectively. IB-MECA caused postischemic reduction in necrosis and improvement in ventricular function, which was abolished by CHE. Western blot analysis demonstrated translocation of the PKC-δ isoform but not the α, ϵ, ξ, η isoform(s) from cytoplasm to the membrane fraction after 30 min of IB-MECA administration. A3AR antagonist MRS-1191 and CHE blocked the translocation of PKC-δ. Furthermore, IB-MECA-induced increase in nuclear factor-κB binding was diminished by CHE. These results provide direct evidence of an essential role of PKC, and more specifically, PKC-δ in A3AR-induced delayed cardioprotection.
Quantitative assessment of tyrosine nitration of manganese superoxide dismutase in angiotensin II-infused rat kidney
Authors: Guo Wei , Adachi Takeshi , Matsui Reiko , Xu Shanqin , Jiang Bingbing , Zou Ming-Hui , Kirber Michael , Lieberthal Wilfred , Cohen Richard A.
Published: March 2015

Hypertension caused by angiotensin II is characterized by an increase in tissue oxidant stress as evidenced by increased quantities of reactive oxygen and nitrogen species. Manganese superoxide dismutase (MnSOD) is a key mitochondrial antioxidant enzyme that is inactivated in conditions of oxidant stress by reacting with peroxynitrite to form 3-nitrotyrosine in its active site. The increase in 3-nitrotyrosine content in MnSOD in the kidney of angiotensin II-infused rats was assessed in this study by immunohistochemistry, Western blotting, immunoprecipitation, and HPLC with UV detection (HPLC-UV). MnSOD activity decreased ∼50% in angiotensin II-infused rat kidneys (24 ± 4.6 vs. 11 ± 5.2 U/mg) without a change in protein expression. Immunohistochemical staining showed 3-nitrotyrosine predominantly in distal tubules and collecting duct cells in the angiotensin II-infused rat kidneys. By two-photon microscopy, 3-nitrotyrosine colocalized with MnSOD. Total 3-nitrotyrosine content in kidney homogenates was increased in angiotensin II-infused rat kidney [3.2 ± 1.9 (sham treated) vs. 9.5 ± 2.3 ng/mg protein by HPLC-UV detection]. With tracer amounts of tyrosine-nitrated recombinant MnSOD, the most sensitive technique to detect tyrosine nitration of MnSOD was immunoprecipitation from tissue with anti-MnSOD antibody, followed by detection of 3-nitrotyrosine by Western blotting or HPLC. By HPLC, 3-nitrotyrosine content of kidney MnSOD increased 13-fold after angiotensin II infusion, representing an increase from approximately one-twentieth to one-fifth of the total 3-nitrotyrosine content in sham-treated and angiotensin II-infused rat kidney, respectively. Angiotensin II-induced hypertension is accompanied by increased tyrosine nitration of MnSOD, which, because it inactivates the enzyme, may contribute to increased oxidant stress in the kidney.
Enhancement of closed-state inactivation in long QT syndrome sodium channel mutation ΔKPQ
Authors: Chen Tiehua , Sheets Michael F.
Published: March 2015

ΔKPQ, a three amino acid [lysine (K), proline (P), glutamine (Q)] deletion mutation of the human cardiac Na channel (hH1), which is one cause of long QT syndrome (LQT3), has impaired inactivation resulting in a late sodium current. To better understand inactivation in ΔKPQ, we applied a site-3 toxin anthopleurin A, which has been shown to inhibit inactivation from the open state with little or no effect on inactivation from the closed state(s) in wild-type hH1. In contrast to the effect of site-3 toxins on wild-type hH1, inactivation from closed state(s) in toxin-modified ΔKPQ demonstrated a large negative shift in the Na channel availability curve of nearly −14 mV. Recovery from inactivation showed that toxin-modified ΔKPQ channels recovered slightly faster than those in control, whereas development of inactivation at potentials negative to −80 mV showed that inactivation developed much more rapidly in toxin-modified ΔKPQ channels compared with control. An explanation for our results is that closed-state inactivation in toxin-modified ΔKPQ is enhanced by the mutated inactivation lid being positioned “closer” to its receptor resulting in an increased rate of association between the inactivation lid and its receptor.
Contractility and ischemic response of hearts from transgenic mice with altered sarcolemmal KATP channels
Authors: Rajashree R. , Koster J. C. , Markova K. P. , Nichols C. G. , Hofmann P. A.
Published: March 2015

The functional significance of ATP-sensitive K+ (KATP) channels is controversial. In the present study, transgenic mice expressing a mutant Kir6.2, with reduced ATP sensitivity, were used to examine the role of sarcolemmal KATP in normal cardiac function and after an ischemic or metabolic challenge. We found left ventricular developed pressure (LVDP) was 15–20% higher in hearts from transgenics in the absence of cardiac hypertrophy. β-Adrenergic stimulation caused a positive inotropic response from nontransgenic hearts that was not observed in transgenic hearts. Decreasing extracellular Ca2+ decreased LVDP in hearts from nontransgenics but not in those from transgenics. These data suggest an increase in intracellular [Ca2+] in transgenic hearts. Additional studies have demonstrated hearts from nontransgenics and transgenics have a similar postischemic LVDP. However, ischemic preconditioning does not improve postischemic recovery in transgenics. Transgenic hearts also demonstrate a poor recovery after metabolic inhibition. These data are consistent with the hypothesis that sarcolemmal KATP channels are required for development of normal myocardial function, and perturbations of KATP channels lead to hearts that respond poorly to ischemic or metabolic challenges.
Role of glycocalyx in leukocyte-endothelial cell adhesion
Authors: Mulivor A. W. , Lipowsky H. H.
Published: March 2015

The binding of fluorescently labeled microspheres (FLMs, 0.1-μm diameter) coated with antibody (1a29) to ICAM-1 was studied in postcapillary venules during topical application of the chemoattractant N-formylmethionyl-leucyl-phenylalanine (fMLP). FLM adhesion to endothelial cells (ECs) increased dramatically from 50 to 150 spheres per 100-μm length of venule after superfusion of the mesentery with fMLP and equaled or exceeded levels of leukocyte (WBC) adhesion. Removal of the EC glycocalyx by micropipette infusion of the venule with heparinase increased FLM-EC adhesion to levels attained with fMLP. Subsequent application of fMLP did not increase FLM adhesion further, suggesting that the FLMs saturated all ICAM-1 binding sites. Perfusion with heparinase after suffusion with fMLP significantly increased FLM-EC adhesion above levels attained with fMLP. However, WBC adhesion fell because of possible removal of selectins necessary to maintain WBC rolling at the wall. It is concluded that the glycocalyx serves as a barrier to adhesion and that its shedding during natural activation of ECs may be an essential part of the inflammatory response.
Biventricular mechanical asynchrony predicts hemodynamic effect of uni- and biventricular pacing
Authors: Yu Yinghong , Kramer Andrew , Spinelli Julio , Ding Jiang , Hoersch Walter , Auricchio Angelo
Published: March 2015

We tested whether biventricular resynchronization explains contractile function changes with univentricular and biventricular pacing in heart failure patients with varying magnitudes of baseline biventricular asynchrony. Thirty patients (New York Hospital Association class ≥ III, QRS duration ≥120 ms) were tested. Contractile function was measured by left ventricular maximum first derivative of pressure over time (dP/d tmax). Biventricular mechanical asynchrony was quantified by the normalized pressure-pressure (NPP) loop area formed by the cross-plot of right and left intraventricular pressure curves from each cardiac cycle. Any ventricular pacing increased dP/d tmax if it decreased baseline NPP loop area and almost always worsened dP/d tmax and asynchrony when baseline NPP loop area <0.3. The quantitative relationship between dP/d tmax and NPP loop area change depended on ventricular pacing site and timing relative to intrinsic activation. For similar NPP loop decreases, dP/d tmax increased 16% more with left and biventricular pacing compared with right ventricular pacing. In conclusion, right, left, or biventricular pacing can improve contractile function only in patients having sufficient baseline biventricular asynchrony. However, biventricular resynchronization is only one of the improvement mechanisms.
A synergistic effect of albumin and fibrinogen on immunoglobulin-induced red blood cell aggregation
Authors: Ben-Ami Ronen , Barshtein Gershon , Mardi Tamar , Deutch Varda , Elkayam Ori , Yedgar Saul , Berliner Shlomo
Published: March 2015

Therapeutic administration of immunoglobulins (Ig) has the potential to precipitate thrombotic events. This phenomenon may be explained by red blood cell (RBC) aggregation, which can be potentiated by Ig. The contribution of plasma albumin and fibrinogen to Ig-induced RBC aggregation is unclear. We examined RBC aggregation in three settings: 1) patients receiving therapeutic infusions of Ig; 2) patients receiving plasma supplemented in vitro with Ig; and 3) patients receiving RBC suspensions in standard buffer with varying concentrations of albumin, Ig, and fibrinogen. Ig infusion augmented aggregation of RBCs from patients with normal or high plasma levels of albumin but decreased aggregation in those with lower plasma albumin concentrations. In vitro, RBC aggregation was significantly increased only when all three components, fibrinogen, albumin, and Ig, were present at or above normal concentrations in the suspension but was unaffected when any one of the components was absent from the suspension. Our results suggest a three-way interaction among fibrinogen, Ig, and albumin that synergistically induces RBC aggregation in plasma. Understanding these interactions may help predict clinically important phenomena related to RBC aggregation, such as thrombotic complications of Ig infusion.
Cardioselective overexpression of HO-1 prevents I/R-induced cardiac dysfunction and apoptosis
Authors: Vulapalli Sreesatya Raju , Chen Zhongyi , Chua Balvin H. L. , Wang Tingchung , Liang Chang-Seng
Published: March 2015

Heme oxygenase (HO)-1 converts heme to bilirubin, carbon monoxide, and iron. Our prior work has suggested a cardioprotective role for HO-1 in heart failure. To test whether HO-1 (heat shock protein 32) prevents cardiomyocyte apoptosis and cardiac dysfunction after ischemia-reperfusion (I/R), we generated transgenic mice overexpressing HO-1 in the heart under the control of the α-myosin heavy chain promoter. HO-1 transcript and protein increased markedly in the heart only. In an isolated heart preparation, we observed an enhanced functional recovery during reperfusion after ischemia in the transgenic hearts compared with nontransgenic controls. I/R injury was also performed in intact animals by coronary ligation and reperfusion to assess the protective role of HO-1 overexpression on heart apoptosis. HO-1 overexpression reduced cardiac apoptosis, as evidenced by fewer terminal deoxynucleodidyl transferase-mediated dUTP nick-end labeling-positive or in situ oligo ligation-positive myocytes, compared with nontransgenic mice. Our results indicate that cardioselective overexpression of HO-1 exerts a cardioprotective effect after myocardial I/R in mice, and this effect is probably mediated via an antiapoptotic action of HO-1.
Developmental differences in Ca2+-activated K+channel activity in ovine basilar artery
Authors: Lin Mike T. , Hessinger David A. , Pearce William J. , Longo Lawrence D.
Published: March 2015

A primary determinant of vascular smooth muscle (VSM) tone and contractility is the resting membrane potential, which, in turn, is influenced heavily by K+channel activity. Previous studies from our laboratory and others have demonstrated differences in the contractility of cerebral arteries from near-term fetal and adult animals. To test the hypothesis that these contractility differences result from maturational changes in voltage-gated K+channel function, we compared this function in VSM myocytes from adult and fetal sheep cerebral arteries. The primary current-carrying, voltage-gated K+channels in VSM myocytes are the large conductance Ca2+-activated K+channels (BKCa) and voltage-activated K+(KV) channels. We observed that at voltage-clamped membrane potentials of +60 mV in perforated whole cell studies, the normalized outward current densities in fetal myocytes were >30% higher than in those of the adult ( P < 0.05) and that these were predominately due to iberiotoxin-sensitive currents from BKCachannels. Excised, insideout membrane patches revealed nearly identical unitary conductances and Hill coefficients for BKCachannels. The plot of log intracellular [Ca2+] ([Ca2+]i) versus voltage for half-maximal activation ( V½) yielded linear and parallel relationships, and the change in V½for a 10-fold change in [Ca2+] was also similar. Channel activity increased e-fold for a 19 ± 2-mV depolarization for adult myocytes and for an 18 ± 1-mV depolarization for fetal myocytes ( P > 0.05). However, the relationship between BKCaopen probability and membrane potential had a relative leftward shift for the fetal compared with adult myocytes at different [Ca2+]i. The [Ca2+] for half-maximal activation (i.e., the calcium set points) at 0 mV were 8.8 and 4.7 μM for adult and fetal myocytes, respectively. Thus the increased BKCacurrent density in fetal myocytes appears to result from a lower calcium set point.
Hyperhomocysteinemia leads to pathological ventricular hypertrophy in normotensive rats
Authors: Joseph Jacob , Joseph Lija , Shekhawat Nawal S. , Devi Sulochana , Wang Junru , Melchert Russell B. , Hauer-Jensen Martin , Kennedy Richard H.
Published: March 2015

A recent report indicated that hyperhomocysteinemia (Hhe), in addition to its atherothrombotic effects, exacerbates the adverse cardiac remodeling seen in response to hypertension, a powerful stimulus for pathological ventricular hypertrophy. The present study was undertaken to determine whether Hhe has a direct effect on ventricular remodeling and function in the absence of other hypertrophic stimuli. Male Wistar-Kyoto rats were fed either an amino acid-defined control diet or an intermediate Hhe-inducing diet. After 10 wk of dietary treatment, rats were subjected to echocardiographic assessment of left ventricular (LV) dimensions and systolic function. Subsequently, blood was collected for plasma homocysteine measurements, and the rats were killed for histomorphometric and biochemical assessment of cardiac remodeling and for in vitro cardiac function studies. Significant LV hypertrophy was detected by echocardiographic measurements, and in vitro results showed hypertrophy with significantly increased myocyte size in the LV and right ventricle (RV). LV and RV remodeling was characterized by a disproportionate increase in perivascular and interstitial collagen, coronary arteriolar wall thickening, and myocardial mast cell infiltration. In vitro study of LV function demonstrated abnormal diastolic function secondary to decreased compliance because the rate of relaxation did not differ between groups. LV systolic function did not vary between groups in vitro. In summary, in the absence of other hypertrophic stimuli short-term intermediate Hhe caused pathological hypertrophy and remodeling of both ventricles with diastolic dysfunction of the LV. These results demonstrate that Hhe has direct adverse effects on cardiac structure and function, which may represent a novel direct link between Hhe and cardiovascular morbidity and mortality, independent of other risk factors.
Estrogen replacement reduces PGHS-2-dependent vasoconstriction in the aged rat
Authors: Armstrong Stephen J. , Zhang Yunlong , Stewart Ken G. , Davidge Sandra T.
Published: March 2015

The reduction in estrogen in postmenopausal women contributes to an increase in vascular dysfunction. Models of aging have shown that this is due, in part, to increased prostaglandin H synthase (PGHS)-dependent vasoconstriction. We showed previously that inducible PGHS-2-dependent vasoconstriction is increased with aging. In the present study, we hypothesized that estrogen suppresses PGHS-2-dependent constriction in the aged rat. Isolated mesenteric arteries from placebo- or estrogen-treated, ovariectomized aged (24 mo) Fisher rats were assessed for endothelium-dependent relaxation in the absence or presence of PGHS inhibitors. PGHS inhibition (meclofenamate, 1 μmol/l) enhanced methacholine-induced relaxation only in the placebo group. Specific PGHS-2 inhibition (NS-398, 10 μmol/l) increased arterial relaxation to a greater extent than PGHS-1 inhibition (valeryl salicylate, 3 mmol/l). Estrogen prevented the PGHS-dependent constrictor effect but did not enhance nitric oxide-dependent relaxation in this model. PGHS-1 and endothelial nitric oxide synthase were not altered by estrogen, whereas PGHS-2 expression was decreased in the estrogen-replaced rats ( P < 0.05). In summary, estrogen replacement improved vasodilation in aged rats by decreasing PGHS-dependent constriction.
Fumonisin-induced blockade of ceramide synthase in sphingolipid biosynthetic pathway alters aortic input impedance spectrum of pigs
Authors: Constable Peter D. , Smith Geoffrey W. , Rottinghaus George E. , Tumbleson Mike E. , Haschek Wanda M.
Published: March 2015

The sphingolipid signaling pathway appears to play an important role in regulating vascular tone. We examined the effect of fumonisin B1, a fungal toxin in corn that blocks ceramide synthase in the sphingolipid signaling pathway, on the ascending aortic impedance spectrum of pigs. Sixteen pigs were fed culture material containing fumonisin B1(20 mg/kg body wt) ( n = 7) or a control diet ( n = 9) daily for 3 days and then instrumented under α-chloralose anesthesia for measurement of ascending aortic pressure and flow. Fumonisin ingestion increased serum sphinganine and sphingosine concentrations. Fumonisin ingestion also decreased cardiac output and characteristic impedance and increased the frequency of the first minimum impedance modulus, systemic vascular resistance, and the terminal, first, and second harmonic reflection coefficients, without changing mean arterial pressure. Thus blockade of ceramide synthase is accompanied by decreased vascular tone in systemic conduit arteries and increased vascular tone in systemic resistance vessels. The results indicate that the sphingolipid signaling pathway influences vascular tone in α-chloralose-anesthetized pigs.
Cardiovascular regulation after destruction of the C1 cell group of the rostral ventrolateral medulla in rats
Authors: Madden Christopher J. , Sved Alan F.
Published: March 2015

To evaluate the role of C1 neurons in the rostral ventrolateral medulla (RVLM) in cardiovascular regulation, we studied rats in which this cell group was destroyed by the injection of anti-dopamine-β-hydroxylase-saporin into the RVLM. These immunotoxin injections resulted in 32–99% depletion of the RVLM-C1 neurons and ∼50% depletion of the A5 cell population. In conscious rats with large (>80%) depletion of the RVLM-C1 cell population, resting arterial pressure was ∼10 mmHg lower than in control injected rats, although heart rate was not significantly different. Similar results were observed when arterial pressure was recorded in urethan-anesthetized rats, although under anesthesia, heart rate was also reduced in rats with large (>80%) depletion of the RVLM-C1 neuronal population. Sympathoexcitatory responses to baroreceptor unloading, chemoreceptor activation, and electrical stimulation of sciatic nerve afferent fibers were attenuated in rats with >80% depletion of the RVLM-C1 cell population. These effects of RVLM-C1 plus A5 cell populations were not mimicked by either smaller lesions of the RVLM-C1 population or by selective destruction of the A5 cell population with 6-hydroxydopamine. Sympathoinhibitory responses such as decreases in arterial pressure and heart rate evoked by injection of GABA into the RVLM or by intravenous phenylbiguanide administration were not altered by RVLM-C1 plus A5 cell depletion. These data suggest that RVLM-C1 cells contribute to the maintenance of baseline arterial pressure and play an integral role in sympathoexcitatory responses.
Prior exercise training produces NO-dependent increases in collateral blood flow after acute arterial occlusion
Authors: Yang H. T. , Ren Jie , Laughlin M. Harold , Terjung Ronald L.
Published: March 2015

We previously reported that prior training improves collateral blood flow (BF) to the calf muscles after acute-onset occlusion of the femoral artery (Yang HT et al. Am J Physiol Heart Circ Physiol 279: H1890–H1897, 2000). The purpose of this study was to test the hypothesis that increased release of nitric oxide (NO) by NO synthase (likely endothelial NOS) contributes to the increased BF to calf muscles of trained rats after acute femoral artery occlusion. Adult male Sprague-Dawley rats (∼325 g) were limited to cage activity and were sedentary (SED; n = 28) or exercise trained (TR; n = 30) for 6 wk by treadmill running. On the day of the investigation, rats were anesthetized with ketamine-acepromazine and instrumented for determination of BF (using 141Ce- and 85Sr-labeled microspheres) and distal limb arterial pressure, and femoral arteries were occluded bilaterally. Four hours after surgery, collateral BF was determined twice during treadmill running: first at a demanding speed (20 m/min, 15% grade) and second, after a brief rest and at a faster running speed (25 m/min, 15% grade). The fact that BF did not increase further at the higher running speed indicated that maximal collateral BF was measured. Approximately half of the rats in each group received 20 mg/kg body wt N G-nitro-l-arginine methyl ester (l-NAME) intra-arterially 30 min before treadmill exercise and BF measurement to block production of NO by NOS. Results indicate that prior training improved collateral-dependent BF to the skeletal muscle of rats after acute femoral artery occlusion due primarily to an increase in the conductance of the upstream collateral circuit. Blockade of NOS with l-NAME produced decreased vascular conductance, both in the upstream collateral circuit and in the distal skeletal muscle microcirculation, and the difference between collateral vascular conductance in TR and SED rats was abolished. Our results indicate that the primary determinant of the increased collateral BF with prior training is the resistance of the upstream collateral circuit and imply that enhanced endothelium-mediated dilation induced by training serves to increase collateral BF following acute arterial occlusion.
Staphylococcal α-toxin provokes neutrophil-dependent cardiac dysfunction: role of ICAM-1 and cys-leukotrienes
Authors: Grandel Ulrich , Reutemann Mathias , Kiss Ladislau , Buerke Michael , Fink Ludger , Bournelis Emmanoyil , Heep Martina , Seeger Werner , Grimminger Friedrich , Sibelius Ulf
Published: March 2015

The role of polymorphonuclear neutrophils (PMN) in septic myocardial dysfunction is presently unknown. Staphylococcus aureus infections are frequently associated with septic sequelae. Therefore, we perfused isolated rat hearts with low doses of α-toxin, the major staphylococcal exotoxin, followed by application of human PMN, N-formyl-methionyl-leucyl-phenylalanine, and arachidonic acid. In contrast to sham-perfused hearts (no α-toxin), a rise in coronary perfusion pressure (CPP) and a reduction of contractile function were noted, and cardiac expression of intercellular adhesion molecule (ICAM)-1 was detected by immunohistochemical methods and real-time PCR. Histological analysis and myeloperoxidase activity indicated cardiac PMN accumulation in α-toxin-challenged hearts. Major quantities of cysteinyl (cys)-leukotrienes (LT), LTB4, and 5-hydroxyeicosatetraenoic acid (5-HETE) were found in the perfusate of α-toxin-exposed hearts. With an anti-ICAM-1 antibody, neutrophil accumulation, leukotriene (LT) synthesis, coronary vasoconstriction, and the accompanying cardiodepression were suppressed. Similarly, the lipoxygenase inhibitor MK-886 blocked LT synthesis and maintained cardiac function. We conclude that low-dose α-toxin provokes coronary endothelial ICAM-1 expression and neutrophil accumulation, with subsequent synthesis of cys-LTs, LTB4, and 5-HETE under conditions of appropriate stimulation. This response is linked with coronary vasoconstriction and contractile dysfunction, with cys-LT synthesis and maldistribution of perfusion offered as likely underlying mechanisms.
α2-Adrenergic receptors in NTS facilitate baroreflex function in adult spontaneously hypertensive rats
Authors: Hayward Linda F. , Riley Alecia P. , Felder Robert B.
Published: March 2015

We examined the effect of α2-adrenoreceptor blockade in the nucleus of the solitary tract (NTS) on baroreflex responses elicited by electrical stimulation of the left aortic depressor nerve (ADN) in urethane-anesthetized spontaneously hypertensive rats (SHR, n = 11) and normotensive Wistar-Kyoto rats (WKY, n = 11). ADN stimulation produced a frequency-dependent decrease in mean arterial pressure (MAP), renal sympathetic nerve activity (RSNA), and heart rate (HR). In SHR, unilateral microinjection of idazoxan into the NTS markedly reduced baroreflex control of MAP, RSNA, and HR and had a disproportionately greater influence on baroreflex control of MAP than of RSNA. In WKY, idazoxan microinjections did not significantly alter baroreflex function relative to control vehicle injections. These results suggest that baroreflex regulation of arterial pressure in SHR is highly dependent on NTS adrenergic mechanisms. The reflex regulation of sympathetic outflow to the kidney is less influenced by the altered α2-adrenoreceptor mechanisms in SHR.
Role of NO in modulating neuronal activity in superficial dorsal horn of spinal cord during exercise pressor reflex
Authors: Li Jianhua , Mitchell Jere H.
Published: March 2015

Static contraction of hindlimb skeletal muscle in cats induces a reflex pressor response. The superficial dorsal horn of the spinal cord is the major site of the first synapse of this reflex. In this study, static contraction of the triceps surae muscle was evoked by electrical stimulation of the tibial nerve for 2 min in anesthetized cats (stimulus parameters: two times motor threshold at 30 Hz, 0.025-ms duration). Ten stimulations were performed and 1-min rest was allowed between stimulations. Muscle contraction caused a maximal increase of 32 ± 5 mmHg in mean arterial pressure (MAP), which was obtained from the first three contractions. Activated neurons in the superficial dorsal horn were identified by c-Fos protein. Distinct c-Fos expression was present in the L6-S1 level of the superficial dorsal horn ipsilateral to the contracting leg (88 ± 14 labeled cells per section at L7), whereas only scattered c-Fos expression was observed in the contralateral superficial dorsal horn (9 ± 2 labeled cells per section, P < 0.05 compared with ipsilateral section). A few c-Fos-labeled cells were found in control animals (12 ± 5 labeled cells per section, P < 0.05 compared with stimulated cats). Furthermore, double-labeling methods demonstrated that c-Fos protein coexisted with nitric oxide (NO) synthase (NOS) positive staining in the superficial dorsal horn. Finally, an intrathecal injection of an inhibitor of NOS, N-nitro-l-arginine methyl ester (5 mM), resulted in fewer c-Fos-labeled cells (58 ± 12 labeled cells per section) and a reduced maximal MAP response (20 ± 3 mmHg, P < 0.05). These results suggest that the exercise pressor reflex induced by static contraction is mediated by activation of neurons in the superficial dorsal horn and that formation of NO in this region is involved in modulating the activated neurons and the pressor response to contraction.
Sympathoexcitation to intravenous interleukin-1β is dependent on forebrain neural circuits
Authors: Kenney Michael J. , Blecha Frank , Wang Yan , McMurphy Rose , Fels Richard J.
Published: March 2015

We investigated the contributions of forebrain, brain stem, and spinal neural circuits to interleukin (IL)-1β-induced sympathetic nerve discharge (SND) responses in α-chloralose-anesthetized rats. Lumbar and splenic SND responses were determined in spinal cord-transected (first cervical vertebra, C1), midbrain-transected (superior colliculus), and sham-transected rats before and for 60 min after intravenous IL-1β (285 ng/kg). The observations made were the following: 1) lumbar and splenic SND were significantly increased after IL-1β in sham C1-transected rats but were unchanged after IL-1β in C1-transected rats; 2) intrathecal administration of dl-homocysteic acid (10 ng) increased SND in C1-transected rats; 3) lumbar and splenic SND were significantly increased after IL-1β in sham- but not midbrain-transected rats; and 4) midbrain transection did not alter the pattern of lumbar and splenic SND, demonstrating the integrity of brain stem sympathetic neural circuits after decerebration. These results demonstrate that an intact forebrain is required for mediating lumbar and splenic sympathoexcitatory responses to intravenous IL-1β, thereby providing new information about the organization of neural circuits responsible for mediating sympathetic-immune interactions.
Gene-environment interactions in wet beriberi: effects of thiamine depletion in CD36-defect rats
Authors: Tanaka Takao , Kono Tatsuji , Terasaki Fumio , Kintaka Taigo , Sohmiya Koichi , Mishima Takayuki , Kitaura Yasushi
Published: March 2015

Selective vulnerability to thiamine deficiency is known to occur between individuals and within different tissues. However, no comprehensive explanation for this has been found, and there are no reports that reproduce the cardiovascular manifestations of human wet beriberi in animals. We hypothesized that the distinction of substrate reliance, namely, the primary dependency on glucose as substrate, could be an underlying factor in the selective vulnerability of thiamine deficiency. In the setting of impaired fatty acid entry, which occurs in CD36-defect rats, substrate reliance shifts from fatty acid to glucose, which would be expected to lead to a susceptibility to thiamine deficiency. Genomic DNA was analyzed for CD36 defects in three cognate strains of rats [spontaneously hypertensive rats (SHR)/NCrj, SHR/Izm, and Wistar-Kyoto (WKY)/NCrj], which identified the presence of a CD36 defect in SHR/NCrj rats but not in SHR/Izm and WKY/NCrj rats. Treatment with 2 wk of thiamine-depleted chow on 4-wk-old rats of each of these strains resulted in increased body and lung weight in the SHR/NCrj rats but not in the SHR/Izm and WKY/NCrj rats. The increased lung weight in the SHR/NCrj rats was accompanied with histological changes of congestive vasculopathy, which were not observed in either the SHR/Izm or the WKY/NCrj rats. Thiamine-deficient 12-wk-old SHR/NCrj rats demonstrated increased body weight (305.6 ± 6.2 g in thiamine-deficient rats vs. 280.8 ± 9.1 g in control; P < 0.0001), lactic acidemia (pH, 7.322 ± 0.026 in thiamine-deficient rats vs. 7.443 ± 0.016 in control; P < 0.0001; lactate, 2.42 ± 0.28 mM in thiamine-deficient rats vs. 1.20 ± 0.11 mM in control; P < 0.0001) and reduced systemic vascular resistance (4.61 ± 0.42 × 104dyn·s·cm–5in thiamine-deficient rats vs. 6.55 ± 1.36 × 104dyn·s·cm–5in control; P < 0.0001) with high cardiac output (186.0 ± 24.7 ml in thiamine-deficient rats vs. 135.4 ± 27.2 ml in control; P < 0.0019). In conclusion, SHR/NCrj rats harboring a genetic defect of long-chain fatty acid uptake present the relevant clinical cardiovascular signs of human wet beriberi, strongly indicating a close gene-environment interaction in wet beriberi.
Hypoxia induces myocyte-dependent COX-2 regulation in endothelial cells: role of VEGF
Authors: Wu Guifu , Mannam Arjuna P. , Wu Jiaping , Kirbis Simona , Shie Jue-Lon , Chen Christopher , Laham Roger J. , Sellke Frank W. , Li Jian
Published: March 2015

There is increasing evidence that cyclooxygenase (COX)-2 possess both angiogenic and cardioprotective properties. We examined the effects of hypoxic cardiac myocytes (H9c2 cells) on COX-2 expression in human umbilical vein endothelial cells (HUVECs) to determine the pathway involved in COX-2 regulation. The medium from hypoxic (<1% O2) cardiac myocytes (HMCM) or normoxic cardiac myocytes (21% O2) was added to HUVEC cultures. HMCM induced a transient increase of COX-2 mRNA expression at 1 and 3 h without affecting the COX-1 mRNA level. A similar effect also observed in HMCM from cultured primary cardiac myocytes (rat neonatal cardiac myocytes). The increased COX-2 mRNA was associated with a time-dependent increase in COX-2 protein expression. COX-2 was significantly induced by VEGF (4.86 ± 1.03-fold) and IL-1β (3.93 ± 0.89-fold) and slightly increased by TNF-α but not by FGF2, IGF-1, or PDGFs. Analysis of proteins secreted in HMCM showed increased levels of VEGF but not IL-1β or TNF-α. The HMCM-induced COX-2 expression was inhibited by the addition of an anti-VEGF neutralizing antibody. VEGF induced endothelial cell COX-2 expression by both increasing COX-2 transcription and prolonging the COX-2 mRNA half-life. Furthermore, staurosporine, a nonselective PKC inhibitor, prevented the induction of VEGF by hypoxia. Both a selective PKC-α and -β inhibitor and an inducible nitric oxide synthase (NOS) inhibitor decreased the induction of COX-2 by HMCM and VEGF. Finally, HMCM-induced upregulation of COX-2 was accompanied by upregulation of PGI2and PGE2. These results suggest that VEGF is one of the principal factors produced by hypoxic myocytes that is responsible for the induction of endothelial cell COX-2 expression. This process likely involves both PKC and NOS pathways. Our findings have important implications regarding the cardiac protection of COX-2 in ischemic heart disease.
Differential ANG II generation in plasma and tissue of mice with decreased expression of the ACE gene
Authors: Wei Chih-Chang , Tian Baohong , Perry Gilbert , Meng Qing Cheng , Chen Yiu-Fai , Oparil Suzanne , Dell'Italia Louis J.
Published: March 2015

We utilized mice with homozygous disruption of angiotensin-converting enzyme (ACE) (−/−), mice with heterozygous deletion of ACE (+/−), and wild-type mice (+/+) to test the hypothesis that genetic variation in ACE modulates tissue and plasma angiotensin (ANG) II concentrations. With the use of ANG I as substrate, kidney, heart, and lung ACE activity was reduced 80% in −/− mice compared with +/+ mice. However, ANG II concentrations and ANG II-to-ANG I ratios in the kidney, heart, and lung did not differ among genotypes. In contrast, plasma ANG II concentrations in −/− mice were <2 fmol/ml, whereas plasma ANG I concentrations were extremely high (765 fmol/ml). Chymase activity was increased 14-fold in the kidney ( P < 0.05) and 1.5-fold in the heart ( P < 0.05) of −/− versus +/+ mice but did not differ among genotypes in the lung. ANG II formation from enzymes other than ACE and chymase contributed <2% of total ANG II formation in all genotypes. These data suggest that ACE is essential to ANG II formation in the vascular space, whereas chymase may provide an important mechanism in maintaining steady-state ANG II levels in tissue.
Oxygen regulation and limitation to cellular respiration in mouse skeletal muscle in vivo
Authors: Marcinek David J. , Ciesielski Wayne A. , Conley Kevin E. , Schenkman Kenneth A.
Published: March 2015

In skeletal muscle, intracellular Po2can fall to as low as 2–3 mmHg. This study tested whether oxygen regulates cellular respiration in this range of oxygen tensions through direct coupling between phosphorylation potential and intracellular Po2. Oxygen may also behave as a simple substrate in cellular respiration that is near saturating levels over most of the physiological range. A novel optical spectroscopic method was used to measure tissue oxygen consumption (Ṁo2) and intracellular Po2using the decline in hemoglobin and myoglobin saturation in the ischemic hindlimb muscle of Swiss-Webster mice.31P magnetic resonance spectroscopic determinations yielded phosphocreatine concentration ([PCr]) and pH in the same muscle volume. Intracellular Po2fell to <2 mmHg during the ischemic period without a change in the muscle [PCr] or pH. The constant phosphorylation state despite the decline in intracellular Po2rejects the hypothesis that direct coupling between these two variables results in a regulatory role for oxygen in cellular respiration. A second set of experiments tested the relationship between intracellular Po2and Ṁo2. In vivo Ṁo2in mouse skeletal muscle was increased by systemic treatment with 2 and 4 mg/kg body wt 2,4-dinitrophenol to partially uncouple mitochondria. Ṁo2was not dependent on intracellular Po2above 3 mmHg in the three groups despite a threefold increase in Ṁo2. These results indicate that Ṁo2and the phosphorylation state of the cell are independent of intracellular Po2throughout the physiological range of oxygen tensions. Therefore, we reject a regulatory role for oxygen in cellular respiration and conclude that oxygen acts as a simple substrate for respiration under physiological conditions.
Responsiveness vs. basal activity of plasma ANG II as a determinant of arterial pressure salt sensitivity
Authors: Osborn John W. , Ariza-Nieto Pilar , Collister John P. , Soucheray Sandra , Zimmerman Benjamin , Katz Stephen
Published: March 2015

Infusion of angiotensin II (ANG II) causes salt-sensitive hypertension. It is unclear whether this is due to the body's inability to suppress ANG II during increased salt intake or, rather, an elevated basal level of plasma ANG II itself. To distinguish between these mechanisms, Sprague-Dawley rats were instrumented with arterial and venous catheters for measurement of arterial pressure and infusion of drugs, respectively. The sensitivity of arterial pressure to salt was measured in four groups with the following treatments: 1) saline control (Con, n = 12); 2) administration of the angiotensin-converting enzyme inhibitor enalapril to block endogenous ANG II (ANG-Lo, n = 10); 3) administration of enalapril and 5 ng · kg–1 · min–1 ANG II to clamp plasma ANG II at normal levels (ANG-Norm, n = 10); and 4) administration of enalapril and 20 ng · kg–1 · min–1 ANG II to clamp ANG II at high levels (ANG-Hi, n = 10). Rats ingested a 0.4% NaCl diet for 3 days and then a 4.0% NaCl diet for 11 days. Arterial pressure of rats fed the 0.4% NaCl diet was lower in ANG-Lo (84 ± 2 mmHg) compared with Con (101 ± 3 mmHg) and ANG-Norm (98 ± 4 mmHg) groups, whereas ANG-Hi rats were hypertensive (145 ± 4 mmHg). Salt sensitivity was expressed as the change in arterial pressure divided by the change in sodium intake on the last day of the 4.0% NaCl diet. Salt sensitivity (in mmHg/meq Na) was lowest in Con rats (0.0 ± 0.1) and progressed from ANG-Lo (0.8 ± 0.2) to ANG-Norm (1.5 ± 0.5) to ANG-Hi (3.5 ± 0.5) rats. We conclude that the major determinant of salt sensitivity of arterial pressure is the basal level of plasma ANG II rather than the responsiveness of the renin-angiotensin system.
Positive inotropic effects of ouabain in isolated cat ventricular myocytes in sodium-free conditions
Authors: Nishio Manabu , Ruch Stuart W. , Wasserstrom J. Andrew
Published: March 2015

The inotropic and toxic effects of cardiac steroids are thought to result from Na+-K+-ATPase inhibition, with elevated intracellular Na+(Na[Formula: see text])causing increased intracellular Ca2+(Ca[Formula: see text]) via Na-Ca exchange. We studied the effects of ouabain on cat ventricular myocytes in Na+-free conditions where the exchanger is inhibited. Cell shortening and Ca[Formula: see text] transients (with fluo 4-AM fluorescence) were measured under voltage clamp during exposure to Na+-free solutions [LiCl or N-methyl-d-glucamine (NMDG) replacement]. Ouabain enhanced contractility by 121 ± 55% at 1 μmol/l ( n = 11) and 476 ± 159% at 3 μmol/l ( n = 8) (means ± SE). Ca[Formula: see text] transient amplitude was also increased. The inotropic effects of ouabain were retained even after pretreatment with saxitoxin (5 μmol/l) or changing the holding potential to −40 mV (to inactivate Na+current). Similar results were obtained with both Li+and NMDG replacement and in the absence of external K+, indicating that ouabain produced positive inotropy in the absence of functional Na-Ca exchange and Na+-K+-ATPase activity. In contrast, ouabain had no inotropic response in rat ventricular myocytes (10–100 μmol/l). Finally, ouabain reversibly increased Ca2+overload toxicity by accelerating the rate of spontaneous aftercontractions ( n = 13). These results suggest that the cellular effects of ouabain on the heart may include actions independent of Na+-K+-ATPase inhibition, Na-Ca exchange, and changes in Na[Formula: see text].
Role of dual-site phospholamban phosphorylation in the stunned heart: insights from phospholamban site-specific mutants
Authors: Said M. , Vittone L. , Mundiña-Weilenmann C. , Ferrero P. , Kranias E. G. , Mattiazzi A.
Published: March 2015

Phosphorylation of phospholamban (PLB) at Ser16(protein kinase A site) and at Thr17[Ca2+/calmodulin kinase II (CaMKII) site] increases sarcoplasmic reticulum Ca2+uptake and myocardial contractility and relaxation. In perfused rat hearts submitted to ischemia-reperfusion, we previously showed an ischemia-induced Ser16phosphorylation that was dependent on β-adrenergic stimulation and an ischemia and reperfusion-induced Thr17phosphorylation that was dependent on Ca2+influx. To elucidate the relationship between these two PLB phosphorylation sites and postischemic mechanical recovery, rat hearts were submitted to ischemia-reperfusion in the absence and presence of the CaMKII inhibitor KN-93 (1 μM) or the β-adrenergic blocker dl-propranolol (1 μM). KN-93 diminished the reperfusion-induced Thr17phosphorylation and depressed the recovery of contraction and relaxation after ischemia. dl-Propranolol decreased the ischemia-induced Ser16phosphorylation but failed to modify the contractile recovery. To obtain further insights into the functional role of the two PLB phosphorylation sites in postischemic mechanical recovery, transgenic mice expressing wild-type PLB (PLB-WT) or PLB mutants in which either Thr17or Ser16were replaced by Ala (PLB-T17A and PLB-S16A, respectively) into the PLB-null background were used. Both PLB mutants showed a lower contractile recovery than PLB-WT. However, this recovery was significantly impaired all along reperfusion in PLB-T17A, whereas it was depressed only at the beginning of reperfusion in PLB-S16A. Moreover, the recovery of relaxation was delayed in PLB-T17A, whereas it did not change in PLB-S16A, compared with PLB-WT. These findings indicate that, although both PLB phosphorylation sites are involved in the mechanical recovery after ischemia, Thr17appears to play a major role.
Inhibition of ERK attenuates force development by lowering myosin light chain phosphorylation
Authors: D'Angelo Gerard , Adam Leonard P.
Published: March 2015

Phosphorylation of the actin-associated protein caldesmon (CaD) by extracellular signal-regulated kinases (ERK1/2) is purported to participate in force maintenance by vascular smooth muscle. We examined the interrelationship among ERK1/2 activity, phosphorylation of the high molecular weight isoform of CaD (h-CaD) and the 20-kDa myosin light chain (LC20), and isometric force in strips of porcine carotid artery stimulated with endothelin-1 (ET-1; 50 nM). After an initial delay, ERK1/2 activity increased in parallel with ET-1-mediated force; h-CaD phosphorylation increased modestly. 2-(2′-Amino-3′-methoxyphenyl)-ox-anaphthalen-4-one (PD-098059; 50 μM), an ERK1/2 kinase inhibitor, significantly reduced basal ERK1/2 activity within 1 h, but only partially attenuated h-CaD phosphorylation at 3 h. The mechanisms underlying the temporal dissociation of ERK1/2 activity from h-CaD phosphorylation are unknown, but include the possibility that a kinase other than ERK1/2 phosphorylates h-CaD or, more likely, that phosphate turnover in h-CaD is very slow. PD-098059 partially inhibited the development of ET-1-stimulated force only in Ca2+-replete physiological saline solution, primarily by reducing LC20phosphorylation, yet had no effect on myosin light chain kinase in vitro. These inhibitory effects were most evident during the early phase of force production. The inhibitory effect of PD-098059 on force could not be correlated with a corresponding effect on ERK1/2-mediated h-CaD phosphorylation because force in arterial strips stimulated with ET-1 in the absence or presence of PD-098059 tended to approximate each other over time despite significant differences in the level of h-CaD phosphorylation. Force and LC20phosphorylation in response to KCl depolarization were unaffected by PD-098059. These results show that ERK1/2 may regulate force in arterial smooth muscle, but suggest that the mechanism for this effect is by inhibiting LC20phosphorylation.