bluuebunny/crossref_metadata_2025 huggingface dataset:
bFGF ameliorates focal ischemic injury by blood flow-independent mechanisms in eNOS mutant mice
Authors: Huang Z. , Chen K. , Huang P. L. , Finklestein S. P. , Moskowitz M. A.
Published: December 2017

Genetically engineered mice deficient in the expression of type III nitric oxide synthase (NOS) [endothelial NOS (eNOS)] were used to decipher the importance of nitric oxide (NO)-dependent augmentation of regional cerebral blood flow (rCBF) to infarct volume reduction following basic fibroblast growth factor (bFGF) infusion during acute middle cerebral artery (MCA) occlusion. We have shown previously that intravenously administered bFGF reduces infarct volume following MCA occlusion in rats and that bFGF dilates cerebral pial arterioles by NO-dependent mechanisms. Halothane-anesthetized eNOS knockout and wild-type mice were subjected to permanent MCA occlusion by intraluminal filament for 24 h. bFGF (100 microg x kg(-1) x h(-1)) was infused intravenously for 2 h, beginning 15 min after the onset of occlusion. Infarct volume was reduced from 119 +/- 8 to 93 +/- 4 mm3 (22% reduction, P < 0.05) or from 102 +/- 9 to 77 +/- 6 mm3 (24% reduction, P < 0.05) in eNOS knockout or wild-type mice, respectively (means +/- SE; n = 10 per group), and neurological deficits were also significantly reduced. Although bFGF infusion caused a 27% increase in rCBF and a 17% reduction in vascular resistance in the infarct margin of wild-type animals as measured by laser Doppler flowmetry, bFGF did not enhance rCBF in the infarct margin of eNOS mutant mice. These data indicate that intravenous bFGF reduces infarct volume following focal ischemia by mechanisms that are largely blood flow independent.
Kinetic analysis of rubidium and thallium as deposited myocardial blood flow tracers in isolated rabbit heart
Authors: Marshall R. C. , Taylor S. E. , Powers-Risius P. , Reutter B. W. , Kuruc A. , Coxson P. G. , Huesman R. H. , Budinger T. F.
Published: December 2017

Evaluation of myocardial perfusion with tracers such as thallium and rubidium is based on the assumption that tissue tracer content is proportional to flow. The purpose of this study was to evaluate the relationship between flow and tissue tracer content of 201Tl and 83Rb in the isolated perfused rabbit heart. 83Rb (86-day half-life), an isotope that is not used clinically, was used as a subsitute for 82Rb (76-s half-life) to improve the accuracy and precision of data acquisition. The multiple indicator-dilution technique was employed with two independent computational approaches. The first approach explicitly deconvolved 201Tl and 83Rb venous concentration curves by the intravascular reference tracer curve. The second approach used a conventional analysis. Both approaches showed that there was more early washout of 83Rb than 201Tl and that the heart retained 201Tl better than 83Rb within 2 min after isotope introduction. These data indicate that 201Tl is a better perfusion tracer than 83Rb in the isolated rabbit heart.
PKC-lambda is the atypical protein kinase C isoform expressed by immature ventricle
Authors: Rybin V. O. , Buttrick P. M. , Steinberg S. F.
Published: December 2017

We recently identified a developmental decline in protein kinase C (PKC) isoform expression, at the level of the protein, in rat ventricular myocardium. To investigate mechanisms regulating PKC isoform expression in cardiac tissue, this study uses Northern blot analysis to compare the abundance of PKC isoform mRNAs in neonatal and adult rat ventricular myocardium. PKC-epsilon protein and mRNA were detected in both neonatal and adult rat ventricular myocardial preparations. In contrast, coordinate postnatal declines in the abundance of PKC-alpha and PKC-delta proteins and transcripts were identified. An antiserum raised against the COOH-terminal sequence of PKC-zeta detected abundant immunoreactivity in neonatal, but not adult, ventricular myocytes. However, PKC-zeta transcripts were not detectable in the heart either by Northern blot analysis or a reverse transcriptase-polymerase chain reaction approach, indicating that neither the myocytes nor the contaminating cellular elements in the heart express PKC-zeta. Rather, PKC-lambda, another atypical PKC isoform that is structurally highly homologous to PKC-zeta, was detected at the protein and mRNA level in neonatal, but not adult, ventricular myocardium. Taken together, these results establish that developmental declines in calcium-sensitive, novel, and atypical PKC isoforms are paralleled by changes in the levels of the mRNAs encoding these proteins, suggesting transcriptional regulation of PKC during normal cardiac development. The results of this study further identify PKC-lambda as the atypical PKC isoform expressed by the immature ventricle.
Alterations in ATP-sensitive potassium channel sensitivity to ATP in failing human hearts
Authors: Koumi S. I. , Martin R. L. , Sato R.
Published: December 2017

Little is known about the involvement of preexisting heart disease on characteristics of ATP-sensitive K+ channels [I[K(ATP)]] in human heart. We have characterized I[K(ATP)] in isolated cardiac myocytes from patients with congestive heart failure (HF) and compared these channel characteristics with those from donor hearts (healthy control) using the patch-clamp technique. During metabolic inhibition induced by treatment with cyanide (1 mM) and 2-deoxyglucose (10 mM), action potential shortening occurred in atrial myocytes isolated from both HF and donors, but this response was significantly greater and sooner in HF than in donors. The action potential duration at 90% repolarization was 24.7 +/- 4.1% (n = 15) of control in HF, whereas it was 58.7 +/- 5.9% (n = 10, P < 0.001) of control in donors measured at 30-min metabolic inhibition. The shortening of the action potential was partially reversed by glibenclamide (0.5 microM) in both groups. Consistent with the action potential measurements, the whole cell membrane current response to metabolic inhibition, evaluated by the differential current measurement, was sooner and greater in HF than in donors. Single-channel atrial I[K(ATP)] from both HF and donors, recorded in the excised inside-out patch configuration, exhibited bursting opening, conductance, and gating behaviors that did not differ between the two groups. However, the concentration of ATP at half-maximal inhibition of the channel in HF was greater (131.0 microM) than in donors (26.1 microM). We conclude that I[K(ATP)] in cardiac myocytes from patients with HF has channel characteristics substantially similar to those in donors, but that the channel is less sensitive to ATP inhibition in HF than in donors.
Beta-blockade reduces effects of adenosine and carbachol by transregulation of inhibitory receptors and Gi proteins
Authors: Borst M. M. , Marquetant R. , Kubler W. , Strasser R. H.
Published: December 2017

Chronic blockade of stimulatory beta-adrenergic receptors may decrease inhibitory receptors of the adrenergic signal transduction system. This transregulation process might reduce the negative inotropic response of the myocardium to inhibitory receptor stimulation. Rats were treated for 6 days with the beta-blocker atenolol (2 mg/day). beta-Adrenergic receptors in cardiac plasma membranes increased from 49 +/- 6 to 75 +/- 9 fmol/mg protein (means +/- SE; P = 0.053), whereas muscarinic M2 receptors decreased (155 +/- 15 vs. 105 +/- 10 fmol/mg protein; P < or = 0.05). Moreover, inhibitory G alpha(i) proteins were reduced by 36%. The functional responses of isolated hearts to inhibitory agonists after prestimulation with isoproterenol (3 nmol/l) were significantly blunted. The Ki value for the negative inotropic response of the maximal rise in developed left ventricular pressure (dP/dt(max)) to adenosine (0.1-100 micromol/l) increased from 5.9 +/- 1.7 to 24.0 +/- 2.5 micromol/l (P < or = 0.001). A similar rightward shift of the dose-response curve was observed for the effects of adenosine on developed left ventricular pressure (LVP) and of carbachol (0.01-10 micromol/l) on LVP and dP/dt. Thus chronic beta-blockade leads to a coordinate transregulation of inhibitory receptors and Gi proteins, reducing the effects of inhibitory receptor activation of the heart. This mechanism may contribute to the beneficial effects of beta-blocker therapy in heart failure.
Endothelial vasodilator production by uterine and systemic arteries. II. Pregnancy effects on NO synthase expression
Authors: Magness R. R. , Shaw C. E. , Phernetton T. M. , Zheng J. , Bird I. M.
Published: December 2017

Pregnancy is characterized by elevations in uterine but not omental artery nitric oxide synthase (NOS)-specific activity. We hypothesized that increases in NO production during pregnancy are associated with elevations in protein expression of the constitutive isoform, endothelial cell NOS (ecNOS), in uterine but not systemic arteries. Arterial NOS-specific activity and guanosine 3',5'-cyclic monophosphate (cGMP) production were tested in pregnant sheep in the presence or absence [+5 mM ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid] of Ca2+. With the use of Western analysis, ecNOS and neuronal NOS (nNOS) constitutive isoform expressions were evaluated in intact and denuded [vascular smooth muscle (VSM)] uterine and systemic (omental and renal) arteries as well as in isolated endothelium-derived proteins from nonpregnant and pregnant sheep. Uterine and omental artery NOS activity and cGMP production were inhibited 75-85% by Ca2+ removal. ecNOS was localized only in uterine and systemic artery endothelium (not VSM) by immunohistochemistry and Western analysis; nNOS was not detected. Compared with nonpregnant ewes, pregnancy increased expression of ecNOS in uterine [2.1- to 4.2-fold (P < 0.0001)] and omental [1.3- to 2.2-fold (P = 0.032)] but not renal (P = 0.1367) artery endothelium. Increases in uterine were greater than in omental artery endothelium. Levels of plasma and urinary cGMP were elevated (P < 0.01) proportionally (1.8- to 2.0-fold) in pregnant versus nonpregnant ewes. During pregnancy, expression of uterine artery endothelium-derived (not VSM) ecNOS constitutive isoform is increased, whereas expression in systemic vessels shows little or no change.
Modulation of propagation from an ectopic focus by electrical load and by extracellular potassium
Authors: Wagner M. B. , Golod D. , Wilders R. , Verheijck E. E. , Joyner R. W. , Kumar R. , Jongsma H. J. , Van Ginneken A. C. , Goolsby W. N.
Published: December 2017

We previously developed a technique (R. Kumar, R. Wilders, R. W. Joyner, H. J. Jongsma, E. E. Verheijck, D. A. Golod, A. C. G. van Ginneken, and W. N. Goolsby. Circulation 94: 833-841, 1996) for study of a mathematical model cell with spontaneous activity, viz. a "real-time" simulation of a rabbit sinoatrial node cell (SAN model cell; R. Wilders, H. J. Jongsma, and A. C. van Ginneken. Biophys. J. 60: 1202-1216, 1991) simultaneously being electrically coupled via our "coupling clamp" [H. Sugiura and R. W. Joyner. Am. J. Physiol. 263 (Heart Circ. Physiol. 32): H1591-H1604, 1992] circuit to a real, isolated ventricular myocyte. We now apply this technique to investigate effects of coupling conductance (Gc), cell size, and the modulation of membrane potential by elevated extracellular potassium concentration on the ability of an ectopic focus, represented by the SAN model cell, to successfully drive a ventricular cell. Values of Gc and the relative sizes of the two cells define three possible outcomes: 1) spontaneous pacing of the SAN model cell but not driving of the ventricular cell, 2) cessation of spontaneous pacing, or 3) pacing of the SAN model cell and driving of the ventricular cell. Below a critical size of the SAN model cell only the first two of these outcomes is possible. Above this critical size there is a range of Gc that allows successful operation of the system as an ectopic focus. Elevation of extracellular potassium concentration from 4 to 8 mM increases both the lower bound and upper bound of Gc for this range. Elevation of extracellular potassium concentration, as commonly observed in myocardial ischemia, may have effects on either inhibiting or releasing from inhibition an ectopic focus.
Cell cycle effects of nitric oxide on vascular smooth muscle cells
Authors: Sarkar R. , Gordon D. , Stanley J. C. , Webb R. C.
Published: December 2017

We characterized the cell cycle block induced by nitric oxide (NO) on smooth muscle cells (SMC). We hypothesized that the inhibition of SMC proliferation by NO was due to a specific block in cell cycle progression. Treatment of cultured rat aortic SMC with the NO donors S-nitroso-N-acetylpenicillamine or S-nitrosoglutathione (0.1 mM for 48 h) resulted in a 50% decrease (P < 0.05) in the fraction of cells in the S and G2 + M phases and a corresponding increase in the G1 fraction, suggesting that NO inhibits entry into S phase, causing accumulation of cells in G1 phase. Application of both NO donors to cycling SMC resulted in a short-term, concentration-dependent (0.06-0.3 mM) inhibition of ongoing DNA synthesis as measured by radiothymidine incorporation, demonstrating that NO causes an S-phase arrest. The S-phase arrest by NO was not mimicked by exogenous guanosine 3',5'-cyclic monophosphate (cGMP, 10 mM) and was associated with, but not due to, a 20% inhibition of RNA synthesis. The S-phase block was completely reversed within 2 h of removal of the NO donors, similar to inhibition by the ribonucleotide reductase inhibitor hydroxyurea. Prolonged treatment of SMC with either NO donor (0.1 mM) did not synchronize cells at the G1-S boundary as expected after a prolonged S-phase arrest, but instead induced a quiescent G0-like state characterized by a 12- to 18-h lag before DNA synthesis returned to normal levels after NO removal. These findings demonstrate that NO inhibition of SMC proliferation is associated with two distinct and reversible cell cycle arrests, an immediate cGMP-independent S-phase block followed by a shift back in the cell cycle from the G1-S boundary to a quiescent G0-like state.
Spatiotemporal aspects of sympathetic C-fiber afferent activity in pressor reflex during abdominal ischemia
Authors: Pan H. L. , Zeisse Z. B. , Pitsillides K. F. , Longhurst J. C.
Published: December 2017

Activation of abdominal sympathetic visceral afferents during ischemia elicits excitatory cardiovascular reflexes. The present study examined the time course and discharge patterns of activation of ischemically sensitive sympathetic C-fiber afferents, and then the relationship between summated afferent activity and the pressor reflex induced by prolonged abdominal ischemia was determined. Single-unit activity of abdominal C-fiber afferents was recorded from the right thoracic sympathetic chain of anesthetized cats during 30 min of ischemia. The reflex pressor response to abdominal ischemia was induced by occlusion of celiac and superior mesenteric arteries. Of 68 C-fiber afferents studied, 36 (approximately 53%) were activated during 30 min of ischemia, whereas the activity of the remaining 32 were not altered. Onset latencies of 36 C-fiber afferents activated by ischemia ranged from 1.0 to 17.4 min with an average of 6.1 +/- 0.8 min. The majority of activated afferents manifested a bursting pattern of discharge activity as ischemia was prolonged beyond 10 min. Summated response of activated afferents, but not individual afferent activity, was related closely to the reflex pressor response during 30 min of ischemia. These results suggest that both recruitment of sufficient numbers of C-fiber afferents and adequate discharge frequency of afferents constitute an encoding mechanism for the pressor reflex during abdominal ischemia.
Temporary acidosis during reperfusion limits myocardial infarct size in dogs
Authors: Kitakaze M. , Takashima S. , Funaya H. , Minamino T. , Node K. , Shinozaki Y. , Mori H. , Hori M.
Published: December 2017

We tested the hypothesis that myocardial extracellular acidosis during early reperfusion limits infarct size. The left anterior descending coronary artery was perfused with blood through a bypass tube in dogs. We occluded the bypass tube for 40 (protocol I; n = 24 hearts) and 90 min (protocol II; n = 36 hearts). In protocols I and II, we infused one group of hearts with HCl (60 micrograms.kg-1.min-1) for 60 min after the onset of reperfusion (the metabolic acidosis group), and another group of hearts were ventilated with 3 liters of 70% O2-30% CO2 mixed with room air 10 min before the onset of reperfusion for 70 min (the respiratory acidosis group). pH in the coronary venous blood and myocardial pH during reperfusion in the metabolic and respiratory acidosis groups were lower than those in the control groups. Infarct sizes in the metabolic (16.4 +/- 2.5 and 22.3 +/- 2.5%) and respiratory (16.7 +/- 2.6 and 22.3 +/- 2.5%) acidosis groups in protocols I and II, respectively, were smaller than those in the control groups (33.1 +/- 3.0 and 40.6 +/- 4.1%, respectively). Thus we conclude that temporary acidosis during reperfusion limits infarct size.
Arteriolar constriction in skeletal muscle during vascular stunning: role of mast cells
Authors: Keller M. W.
Published: December 2017

Striated muscle becomes stunned during reperfusion after sublethal ischemia. Resistance vessel tone and reactivity are altered in stunned muscle tissues. The hypothesis that adenosine-regulated mast cell degranulation occurs during reperfusion and leads to constriction of resistance arterioles was tested. The hamster cremaster muscle was subjected to 1 h of ischemia followed by reperfusion. Resistance arterioles constricted during reperfusion (74% of maximal diameter at baseline vs. 42% of maximal diameter after 30 min of reperfusion; P < 0.01). Mast cells degranulated in reperfusion concomitant with arteriolar constriction. Stimulation of mast cell degranulation in control animals with compound 48/80 or cold superfusate (21 degrees C) caused vasoconstriction that mimicked that seen in reperfusion. The mast cell stabilizer cromolyn blocked degranulation and constriction. If mast cell granules were depleted by applying compound 48/80 before inducing ischemia, then arterioles failed to constrict during reperfusion. Adenosine A3-antagonist BW-A1433 abolished constriction. These findings suggest that arterioles constrict in reperfusion due to adenosine-regulated mast cell degranulation. Vasodilation in response to sodium nitroprusside and acetylcholine was normal in stunned, constricted arterioles. However, the dose-response curves to adenosine were shifted to the left in arterioles constricted by either stunning, compound 48/80, exposure to cold superfusate, or cromolyn compared with control vessels. Depletion of granular components via stunning, compound 48/80, cold superfusate, or inhibition of secretion with cromolyn results in unopposed A1- or A2-mediated vasodilation in response to adenosine, whereas the dilatory effects of adenosine are blunted by simultaneous release of vasoconstrictors from mast cells in control animals. In summary, it was found that mast cell degranulation occurs during reperfusion and leads to constriction of resistance arterioles and altered vascular reactivity to adenosine. Adenosine is released in ischemia and stimulates mast cell degranulation via the A3 receptor located on mast cells during reperfusion.
Effect of phasic vagal stimulation and atrial pacing site on atrioventricular conduction time
Authors: Igel D. A. , Wallick D. W. , Martin P. J. , Levy M. N.
Published: December 2017

We tested the hypothesis that the effect of phasic vagal stimulation on atrioventricular (AV) conduction time is affected by the site of atrial pacing in anesthetized dogs. We paced the right atrium at a constant cycle length from the interatrial septum (IAS), superior coronary sinus (SCS), or inferior coronary sinus (ICS) regions, and we evaluated the time-dependent effects of vagal stimulation on AV conduction at each pacing site. When we stimulated the vagi at stimulus (St)-A phases greater than 136 +/- 40 ms and less than the phase that blocked AV conduction (182 +/- 70 ms), IAS pacing prolonged A-V intervals by 8.6 +/- 8.2 ms more than ICS pacing. A change in pacing site affected the A-V intervals by up to 30 ms when we stimulated the vagus at those times that caused the A-V intervals to prolong maximally. Furthermore, the effect of atrial pacing site on A-V intervals was modulated by AV nodal recovery times during the second or third cycles after the vagal stimulus.
Modeling of arterial and cardiopulmonary baroreflex control of heart rate
Authors: Desai T. H. , Collins J. C. , Snell M. , Mosqueda-Garcia R.
Published: December 2017

We evaluated R-R interval changes (delta R-R interval) in 13 subjects (27 +/- 6 yr; 7 men and 6 women) as a function of blood pressure changes at the carotid sinus and aortic arch and central venous pressure changes at the cardiopulmonary receptors. Neck chamber pressure and suction were used to change pressure at the carotid sinus while lower body negative pressure, phenylephrine infusion, and nitroprusside infusion were used to change pressure at the carotid sinus (delta CSP), aortic arch (delta AAP), and cardiopulmonary receptors (delta CPP). Random effects regression analysis showed a significant linear relationship for delta R-R interval (-1.75 + 1.64 delta CSP + 15.40 delta AAP + 29.02 delta CPP + error), and the correlation (r) between the observed and predicted delta R-R interval was 0.82 (P < 0.00001). Sixty-seven percent of the delta R-R interval variability observed in the study is explained by the model. delta AAP accounts for approximately 63%, delta CSP for 14%, and delta CPP for 23% of the explained delta R-R interval.
Effect of chronic myocardial infarction on in vivo reactivity of skeletal muscle arterioles
Authors: Didion S. P. , Mayhan W. G.
Published: December 2017

The first goal of this study was to test the hypothesis that chronic myocardial infarction alters reactivity of rat skeletal muscle arterioles in vivo. At 4, 8, and 16 wk after induction of chronic myocardial infarction or sham (control) surgery, the spinotrapezius muscle was prepared for direct visualization of the microcirculation. Responses of third-order arterioles (37.9 +/- 0.9 microns) were measured after topical suffusion of acetylcholine (ACh; 0.1, 1.0, and 10 microM), calcitonin gene-related peptide (CGRP; 0.01, 0.1, and 1.0 nM), substance P (SP; 0.01, 0.1, and 1.0 microM), and sodium nitroprusside (SNP; 1.0, 10, and 100 microM). Arteriolar reactivity was impaired after chronic myocardial infarction in response to ACh and CGRP at all time periods examined. In contrast, vasodilatation in response to SP and SNP was preserved after 4, 8, and 16-wk of chronic myocardial infarction. The second goal of this study was to explore the possibility that impaired arteriolar reactivity during chronic myocardial infarction may be related to an altered availability of L-arginine (L-Arg). Suffusion of L-Arg (1.0 mM) partially restored impaired ACh- and CGRP-induced responses in myocardial-infarcted animals toward that observed in controls. Thus the present study demonstrates that impaired reactivity of skeletal muscle arterioles during chronic myocardial infarction appears to be partially related to an alteration in L-Arg availability.
Chronic hypoxia inhibits postnatal maturation of porcine intrapulmonary artery relaxation
Authors: Tulloh R. M. , Hislop A. A. , Boels P. J. , Deutsch J. , Haworth S. G.
Published: December 2017

Neonatal pulmonary hypertension is associated with increased pulmonary vascular reactivity. We studied the responses of isolated porcine intrapulmonary arteries after exposure of piglets to chronic hypobaric hypoxia (CHH) from 0 to 2.5, 3 to 6, or 14 to 17 days of age. CHH inhibited the postnatal development of endothelium-dependent vasorelaxation to acetylcholine (ACh) and the calcium ionophore A-23187. Basal accumulation of guanosine 3', 5'-cyclic monophosphate (cGMP) was unaffected, but cGMP response to ACh was inhibited. Endothelium-independent relaxation to nitric oxide (NO) and zaprinast (a phosphodiesterase inhibitor) was also inhibited, but cGMP accumulation in response to these agonists was normal. The ability of sodium nitroprusside (SNP) to cause vasorelaxation and increase cGMP accumulation was unaffected. Contractile responses to potassium chloride and prostaglandin F2 alpha (PGF2 alpha) were similar to normal after exposure from birth and 3 days and were decreased in the older group, but the ability of NG-monomethyl-L-arginine acetate to increase PGF2 alpha-induced contractions decreased. Thus exposure of newborn piglets to CHH causes 1) no increase in contractile responses and 2) impairment of endothelium-dependent and -independent relaxation by impairing signal transduction mechanisms involved in the release of NO and the effectiveness of cGMP.
Noninvasive analysis of renal artery blood flow dynamics with MR cine phase-contrast flow measurements
Authors: Schoenberg S. O. , Just A. , Bock M. , Knopp M. V. , Persson P. B. , Kirchheim H. R.
Published: December 2017

It was the aim of this study to quantify the measurement of pulsatile flow in the renal artery with the noninvasive magnetic resonance cine phase-contrast (MRCPC) method and combine it with the simultaneous assessment of pulsatile flow with a transit-time ultrasound (TTUS) flowmeter. In seven foxhounds with a chronically implanted precalibrated TTUS flow probe, MRCPC flow measurements were made in the renal artery with a temporal resolution of 32 ms. Mean and pulsatile flow signal were compared by the simultaneous ipsi- or contralateral measurement of the renal blood flow signal by both methods (TTUS and MRCPC). In addition, comparative MRCPC and TTUS flow measurements were made with artificial renal artery stenosis and after the administration of angiotensin II. The mean flow data assessed by the noninvasive MRCPC flow measurements showed an excellent correlation with corresponding TTUS recording (r = 0.99). The MRCPC flow signal displayed a waveform of the renal artery flow profile that was very similar to the TTUS flow pulse. The hemodynamic changes induced by angiotensin II or due to renal artery stenosis were also reliably detected by MRCPC. MRCPC provides a reliable noninvasive method for the quantification of mean blood flow and the assessment of the pulsatile flow signal in the renal artery and proves to be sensitive to hemodynamic changes of pathophysiological importance. Alternatively, the method may be used for studies in physiology that demand a noninvasive approach.
Characterization of ATP-sensitive potassium channels in freshly dissociated rabbit aortic endothelial cells
Authors: Katnik C. , Adams D. J.
Published: December 2017

ATP-sensitive potassium (KATP) channels represent a class of K+ channel regulated by intracellular ATP and serve to transduce changes in cell metabolism into changes in membrane potential. The presence of an KATP conductance has recently been demonstrated in freshly dissociated endothelial cells from rabbit arteries. In the present study, the single-channel activity underlying the KATP conductance in rabbit aortic endothelial cells was examined. Unitary currents were evoked in response to lowering intracellular ATP concentration or application of the K(+)-channel activator levcromakalim and were inhibited by the sulfonylurea drug glibenclamide. Exposure of the cytoplasmic face of an inside-out membrane patch to a solution containing 0.1 mM ATP produced single-channel events with unitary conductances of approximately 150 and approximately 25 pS that were inhibited by either 6 mM ATP or 10 microM glibenclamide. A small conductance channel was also activated in cell-attached patches by bath-applied levcromakalim (25 microM). Activation of endothelial cell KATP channels, and subsequent membrane hyperpolarization, may contribute to endothelium-dependent regulation of vascular smooth muscle tone in response to changes in levels of intracellular metabolites.
Alkalemia reduces recovery from global cerebral ischemia by NMDA receptor-mediated mechanism
Authors: Hurn P. D. , Koehler R. C. , Traystman R. J.
Published: December 2017

In vitro data suggest that low tissue pH reduces, whereas extracellular alkalosis potentiates, cerebral anoxic injury via excitotoxic mechanisms. We tested the hypothesis that in vivo metabolic alkalemia potentiates defects in energy metabolism after global incomplete cerebral ischemia (12 min) and reperfusion (180 min) by an N-methyl-D-aspartate (NMDA) receptor-mediated mechanism. Brain ATP, phosphocreatine, and intracellular pH (pHi) were measured by 31P magnetic resonance spectroscopy in anesthetized dogs treated with 1) preischemic intravenous carbicarb buffer (NaHCO3+Na2CO3, Carb, n = 7); 2) carbicarb infusion plus NMDA receptor antagonist MK-801 (MK-801 + Carb, n = 7); 3) an osmotically equivalent volume of 5% NaCl (NaCl, n = 8); or 4) equivalent volume of 0.9% NaCl (Sal, n = 3). Sagittal sinus pH was raised to 7.82 +/- 0.04 before and 7.65 +/- 0.03 during ischemia in Carb vs. 7.72 +/- 0.01 and 7.60 +/- 0.01 in MK-801+Carb, 7.25 +/- 0.02 and 7.15 +/- 0.03 in NaCl, and 7.31 +/- 0.00 and 7.26 +/- 0.01 in Sal, respectively. Ischemic cerebral blood flow (CBF, radiolabeled microspheres), pHi, and ATP reduction were similar among groups. By 180 min of reperfusion, recovery of ATP was greater in MK-801+Carb (104 +/- 6% of baseline), NaCl (93 +/- 6%), and Sal (94 +/- 6%) than in Carb (47 +/- 6%). Intraischemic pHi was similar among groups, and pHi recovery did not vary among groups despite differences in sagittal sinus pH. In Carb, CBF was restored but with delayed hypoperfusion. We conclude that extracellular alkalosis is deleterious to postischemic CBF and energy metabolism, acting by NMDA receptor-mediated mechanisms.
Upregulation of expression of sarcoplasmic reticulum by TGF-beta 1 in cultured rat cardiac myocytes
Authors: Kimura H. , Takemura H. , Imoto K. , Ohshika H. , Mochizuki Y.
Published: December 2017

The effects of transforming growth factor-beta 1 (TGF-beta 1) on the function and structure of sarcoplasmic reticulum (SR) were studied in cultured neonatal rat cardiac myocytes. The cardiac myocytes at days 2 and 6 of culture exhibited spontaneous contraction; however, the rate of contraction increased and became regular, depending on culture day. Ryanodine and norepinephrine (NE) increased the rate of contraction and frequency of Ca2+ oscillations in myocytes at day 2 of culture. Ryanodine did not affect the spontaneous contraction in nontreated and TGF-beta 1-treated myocytes. On the other hand, NE caused negative and positive chronotropic responses in nontreated and TGF-beta 1-treated cells, respectively. In the absence of extracellular Ca2+, ryanodine and NE did not affect the cytoplasmic Ca2+ concentration ([Ca2+]i) in the nontreated cells, whereas NE increased [Ca2+]i but ryanodine did not in the TGF-beta 1-treated cells. SR structures in TGF-beta 1-treated cells developed more than those in nontreated cells. The results indicate that TGF-beta 1 plays an important role in the upregulation of SR function and structure of cultured neonatal rat cardiac myocytes.
Regulation of hepatic blood flow during resuscitation from hemorrhagic shock: role of NO and endothelins
Authors: Pannen B. H. , Bauer M. , Noldge-Schomburg G. F. , Zhang J. X. , Robotham J. L. , Clemens M. G. , Geiger K. K.
Published: December 2017

We determined the role of nitric oxide (NO) and endothelins (ETs) in the regulation of hepatic blood flow during resuscitation from hemorrhagic shock (HS) in anesthetized rats. Volume resuscitation restored systemic hemodynamics and increased hepatic arterial and portal venous flow above baseline in the vehicle group. Presence of N omega-nitro-L-arginine methyl ester (L-NAME, 1 mg/kg) during resuscitation increased systemic vascular resistance (SVR) above baseline, prevented the restoration of hepatic arterial flow, and abolished portal hyperemia. Although the ETA+B-receptor antagonist bosentan (10 mg/kg) did not alter the systemic hemodynamic response, it abolished the hepatic arterial and portal hyperemia. The ETA-receptor antagonist BQ-610 (150 micrograms/kg) reduced SVR below baseline, allowed hepatic arterial hyperemia to occur, and further enhanced the portal venous hyperemia. This indicates that 1) NO reduces SVR and acts to preserve hepatic blood flow during resuscitation from HS; 2) ETA-receptor-mediated vasoconstriction counteracts the systemic and portal hemodynamic effects of NO; and 3) simultaneous ETB-receptor stimulation enhances blood flow to the liver and may serve to modulate the ETA-receptor-mediated vasoconstrictive effects of ETs.
Capillary red blood cell flow and activation of white blood cells in chronic muscle ischemia in the rat
Authors: Anderson S. I. , Hudlicka O. , Brown M. D.
Published: December 2017

Increased activity of ischemic skeletal muscles in which functional hyperemia is impaired has been linked with capillary endothelial swelling postcapillary white blood cell (WBC) adherence. The perfusion pattern of capillaries under these conditions and time course of WBC activation is not known. Capillary microcirculation was studied by videomicroscopy at rest and after muscle contractions (1 Hz, 10 min) in extensor digi-torum longus muscles of pentobarbital sodium-anesthetized rat during the early stages of chronic ischemia (unilateral ligation of the common iliac artery for 3 days) and in ischemic muscles subjected to increased activity (7 days of ischemia or 3 days of ischemia plus indirect electrical stimulation via planted electrodes, 10 Hz, 7 x 10 min on-90 min off/day) to investigate how perfusion was affected. All ischemic muscles had more intermittently flowing capillaries than did unoperated control) muscles. Temporal heterogeneity of perfusion at rest, assessed by velocity, time spent stationary, and stop/start frequency of red blood cells, was similar to control values in ischemic muscles but greater in ischemic muscles subjected to additional activity. Hyperemic responses to contractions were severely blunted in all ischemic groups. The portion of morphologically nonspherical WBCs, taken to indicate activation, was 24 +/- 3% in venous blood after 3 days of ischemia vs. 14 +/- 1% in control muscles and increased further by 7 days (42 +/- 2%) when activated cells were also found in arterial blood. Thus increased muscular activity may exacerbate the adverse effects of ischemia on capillary perfusion, and WBC activation, evident before endothelial swelling is apparent, provides the potential as a circulating signal for capillary swelling in the ischemic and other muscles.
Regulation of adult cardiocyte growth: effects of active and passive mechanical loading
Authors: Decker M. L. , Janes D. M. , Barclay M. M. , Harger L. , Decker R. S.
Published: December 2017

Fluctuations in hemodynamic load have been documented to modulate contractile protein turnover and myofibrillar structure in the heart; however, the relative importance of active and passive loading in regulating adult cardiocyte growth remains unresolved. To address this issue at the cellular level, adult feline cardiocytes were cultured either on Silastic membranes or plastic surfaces. Cardiocyte-laden membranes were stretched 10% of their rest length to enhance passive loading, whereas heart cells cultured on plastic or Silastic were field stimulated at 1 Hz to mimic active loading. Turnover of contractile proteins and structural integrity of the contractile-cytoskeletal apparatus were monitored for periods ranging from 4 to 72 h. Active and passive loading elevated contractile protein synthesis nearly equally (approximately 50%) and promoted the attachment of remodeled myofibrils to vinculin-positive focal contacts and/or costameres during the first 24 h of loading. Thereafter, rates of contractile protein synthesis returned to control values in passively stretched heart cells but remained elevated in field-stimulated cultures. The fractional rate of growth was increased significantly (approximately 8%/day) in electrically paced cells, whereas in passively stretched cardiocytes the growth rate rose only modestly (approximately 2%/day). Changes in the rate of myocyte growth appeared more closely correlated with the development of focal contacts and myofibril remodeling than with changes in myofibrillar protein turnover per se. 2,3-Butanedione monoxime, nifedipine, and, to a lesser extent, ryanodine blocked field-stimulated contractile protein synthesis and myofibrillar remodeling but had no impact on protein turnover or myofibril reassembly in passively loaded cardiocytes. The results of these experiments imply that both active and passive loading stimulate contractile protein turnover and myofibril remodeling, but the generation of active tension accelerates cardiocyte growth to a greater extent than passive loading. Furthermore, pharmacological interventions suggest that unique pathways may mediate these cellular events in actively and passively loaded adult cardiocytes.
Calcium handling by sarcoplasmic reticulum of neonatal swine cardiac myocytes
Authors: Hohl C. M. , Livingston B. , Hensley J. , Altschuld R. A.
Published: December 2017

In recent years, because of similarities to human infants, neonatal piglets have increasingly become the model of choice for studying neonatal heart function. However, the cardiac sarcoplasmic reticulum (SR) has not been thoroughly characterized in this species. Accordingly, Ca2+ pump kinetics, efflux channel characteristics, Ca2+ transients, and contractile movements were examined in isolated newborn piglet cardiac ventricular myocytes. Maximum uptake rate (Vmax) and concentration required to produce a half-maximal effect (K0.5) for oxalate-supported, ATP-dependent 45Ca2+ uptake by the SR of digitonin-lysed myocytes were 285 +/- 17 nmol 45Ca2+.min-1.mg-1 and 0.69 +/- 0.07 microM, respectively. In the absence of phospholamban phosphorylation, Vmax was reduced to 195 +/- 26 nmol 45Ca2+.min-1.mg-1 (P < 0.05 vs. control) and K0.5 increased to 1.28 +/- 0.13 microM (P < 0.05 vs. control). [3H]ryanodine binding studies yielded a maximum binding capacity of 181 +/- 12 fmol/mg and a dissociation constant of 1.7 +/- 0.2 nM. Raising extracellular Ca2+ (0.5-5 mM) increased peak amplitude and decreased the duration of electrically stimulated fura 2 Ca2+ transients and recordings of cell length changes. Both ryanodine and 2,5-di-tert-butylhydroquinone, an inhibitor of SR calcium adenosinetriphosphatase, completely abolished Ca2+ transients in piglet myocytes. These studies indicate that the SR has a significant role in excitation-contraction coupling in neonatal piglet myocytes.
Inhibition of the voltage-dependent calcium current by extracellular ATP in hamster ventricular cardiomyocytes
Authors: Von zur Muhlen F. , Gonska B. D. , Kreuzer H.
Published: December 2017

The modulation of the high-voltage-activated calcium current (ICa) by external ATP was examined in single ventricular cardiomyocytes of the hamster using the whole-cell configuration of the patch-clamp technique. Extracellular application of ATP (0.1-100 microM) was found to inhibit ICa reversibly. The inhibition followed a slow time course (half time approximately 25 s) and was accompanied by very small changes of the holding current and no shift in the current-voltage relationship. With 100 microM ATP, peak ICa was reduced by approximately 30%. This response was not blocked by the P1 inhibitor 8-cyclopentyl-1,3-dipropylxanthine. The nonhydrolyzable ATP analogs adenosine 5'-O-(3-thiotriphosphate) and AMP-adenosine 5'-[beta,gamma-imido]triphosphate also reduced ICa. The ATP analog alpha,beta-methylene-ATP was about equipotent with ATP at 50 microM. Internal guanosine 5'-O-(3-thiotriphosphate) (200 microM) rendered the ATP-mediated inhibition of ICa poorly reversible, whereas internal guanosine 5'-O-(2-thiodiphosphate) (200-500 microM) had no effect. Holding the intracellular adenosine 3',5'-cyclic monophosphate concentration at a constant high level did not alter the ATP response. We conclude that external ATP inhibits ICa via a P2 purinergic receptor in hamster ventricular myocytes. Our results suggest the involvement of a G protein not coupled to adenylate cyclase. The inhibition of ICa by extracellular ATP might have pathophysiological relevance under conditions of myocardial injury.
Endothelium-derived nitric oxide limits sympathetic neurogenic constriction in intestinal microcirculation
Authors: Nase G. P. , Boegehold M. A.
Published: December 2017

We have recently shown that endogenous nitric oxide (NO) activity can attenuate the sympathetic neurogenic constriction of intestinal arterioles. The purpose of this study was to determine whether the microvascular endothelium is an important site of NO production under these conditions. In the superfused small intestine of the rat, intravital microscopy was used to study the responses of first-order arterioles (1A) to perivascular sympathetic nerve stimulation and directly applied norepinephrine before and then after passage of a CO2 embolus through the 1A lumen to inhibit endothelial function. CO2 embolization did not significantly alter resting arteriolar diameter (50 +/- 4 microns before vs. 51 +/- 4 microns after embolization) but abolished the dilator response to acetylcholine without altering the dilator response to sodium nitroprusside. Stimulation at 3, 8, and 16 Hz caused respective constrictions of 4 +/- 1, 11 +/- 1, and 18 +/- 2 microns, and after CO2 these responses were significantly increased to 9 +/- 1, 18 +/- 1, and 29 +/- 3 microns, respectively. Exposure to the nitric oxide synthase inhibitor NG-monomethyl-L-arginine(10(-4) M in superfusate) after CO2 embolization had no further effect on the magnitude of neurogenic constriction. Similar results were seen when embolization was achieved with N2, and CO2 embolization had the same effect on norepinephrine-induced constriction as it did on neurogenic constriction. These results suggest that nitric oxide of endothelial origin can attenuate sympathetic neurogenic constriction in the intestinal microvasculature.
Effect of long-term food restriction on cardiac mechanics
Authors: Klebanov Simon , Herlihy Jeremiah T. , Freeman Gregory L.
Published: December 2017

Food restriction (FR) is the only known intervention capable of increasing mammalian life span. It not only increases longevity, but reduces the incidence of a broad spectrum of age-related pathologies, including cardiomyopathy, and retards the physiological decline associated with aging. Previous work from this laboratory has shown that long-term FR affects the contractile machinery of the heart, shifting the cardiac myosin profile from the fast, V1 isoform to the slow, V3 isoform. The aim of the present study was to determine whether FR also induces changes in cardiac mechanics. Isolated, isovolumically beating hearts were examined from four groups of rats: 1) ad libitum-fed rats killed at 10–13 mo of age, 2) FR rats offered only 60% of the calories consumed by ad libitum-fed rats and killed at the same age, 3) young ad libitum-fed rats having the same heart weights as the FR rats, and 4) ad libitum-fed rats subjected to short-term FR, i.e., for the last 3 wk of life, and also killed at 10–13 mo of age. Both short- and long-term FR profoundly and to approximately the same extent affected cardiac mechanics. Hearts from FR rats developed much higher pressures than hearts from the ad libitum-fed rats under conditions of low-calcium perfusate. This difference disappeared, however, when contractility was enhanced by either calcium or isoproterenol. FR prolonged both contraction and relaxation times. Long-term ad libitum-fed rats (adult, 10–13 mo of age) had a lower isoproterenol sensitivity than the young ad libitum-fed rats (10 wk of age). Both short- and long-term FR restored the sensitivity to isoproterenol. In summary, FR profoundly affects many aspects of cardiac mechanics, enhancing some age-related changes (prolongation of the contraction and relaxation times), attenuating another (increasing the isoproterenol sensitivity), and, finally, inducing some unique changes unrelated to age (increased pressure development under low-calcium perfusate).
Influence of sympathetic blockade on the acute hypertensive response to aortic constriction
Authors: Fazan R. , Castania J. A. , Ballejo G. , Salgado M. C. O. , Salgado H. C.
Published: December 2017

The objective of the present study was to determine the contribution of the sympathetic nervous system to the hypertensive response to acute (45-min) aortic coarctation in conscious intact or sinoaortic-denervated (SAD) rats. Rats were treated chronically (5 wk) with guanethidine (50 mg ⋅ kg−1 ⋅ day−1ip) to induce sympathetic nerve degeneration or acutely with the α1-adrenergic receptor antagonist prazosin (1 mg/kg iv). Aortic constriction elicited a prompt and sustained rise in mean carotid pressure that was significantly greater in SAD than in intact rats. The increase in pressure was associated with reflex bradycardia only in the intact rats, whereas the heart rate of SAD rats did not change. Guanethidine treatment did not affect the arterial pressure or heart rate responses to aortic coarctation of intact rats but blunted the hypertensive response of SAD rats to the same values exhibited by intact rats. Prazosin administered 10 min after the beginning of aortic coarctation reduced the hypertensive response of SAD rats to the same level as that of intact rats. In conclusion, the data obtained by means of the association of sinoaortic deafferentation with chronic sympathectomy with guanethidine or acute α1-adrenergic receptor blockade with prazosin indicate that the greater hypertensive response of SAD rats involves a lack of suppression of the sympathetic activity in the maintenance of the rise in pressure elicited by aortic coarctation.
Left ventricular function and remodeling after myocardial infarction in aging rats
Authors: Raya Thomas E. , Gaballa Mohamed , Anderson Peter , Goldman Steven
Published: December 2017

— Adaptations of the aging left ventricle (LV) to hemodynamic overload are functionally and structurally distinct from those of the young organism. This study describes the influence of aging on LV hemodynamics and remodeling late after myocardial infarction (MI) in Fischer 344 Brown Norway rats. In sham rats at 23 mo, LV weight, myocyte cross-sectional area (CSA), and myocardial fibrosis were increased, whereas LV dP/d t, LV relaxation, and maximal LV systolic function declined with respect to younger rats (7, 12, and 18 mo of age). Isometric myocardial function was evaluated in papillary muscles of 12- and 23-mo-old sham rats. Myocardial systolic function was decreased in older rats. To determine how aging affects LV function and remodeling after MI, rats were infarcted at 7 and 18 mo of age and were studied 5 mo later. Infarct size was similar in each group. Right ventricular weight, LV end-diastolic pressure, and volume index were increased, whereas LV dP/d t, peak cardiac index, and peak developed LV pressure declined after MI. However, there were no significant differences between young and older rats in any variable of LV systolic function or remodeling after MI. Myocyte CSA increased in younger rats after MI but was unchanged in 23-mo-old rats. After MI, myocardial fibrosis was significantly increased from baseline only in younger rats. The negative interaction of aging and MI on myocyte hypertrophy and fibrosis was highly significant. The findings indicate that baseline LV and myocardial function decline with age. In the aging rat after MI, despite limited compensatory hypertrophy and more advanced baseline myocardial fibrosis, the long-term functional and structural adaptations to MI are similar to those of the mature adult heart.