bluuebunny/crossref_metadata_2025 huggingface dataset:
Effects of Rapamycin on Ryanodine Receptor/Ca
2+
-Release Channels From Cardiac Muscle
Authors: Kaftan Edward , Marks Andrew R. , Ehrlich Barbara E.
Published: June 2012

Abstract
Ryanodine receptors (RyRs) are intracellular channels that regulate the release of Ca
2+
from the endoplasmic reticulum of many cell types. The RyRs are physically associated with FK506-binding proteins (FKBPs); immunophilins, with
cis
-
trans
peptidyl-prolyl isomerase activity. FKBP12 copurifies with RyR1 (skeletal isoform) and modulates its gating. A different form of FKBP with a slightly higher molecular weight copurifies with RyR2 (cardiac isoform). Previous studies have demonstrated that FKBP stabilizes gating of the skeletal Ca
2+
-release channel. In the present study, we measured the activity of cardiac RyRs incorporated into planar lipid bilayers to show that rapamycin, a drug that inhibits the prolyl isomerase activity of FKBP and dissociates FKBP from the RyR, increases the open probability and reduces the current amplitude of cardiac muscle Ca
2+
-release channels. These experiments show for the first time that submicromolar concentrations of rapamycin can alter channel function. Our results provide support for the hypotheses that FKBP functionally associates with the RyR and that the immunosuppressant drug, rapamycin, alters the function of both cardiac and skeletal muscle isoforms of the Ca
2+
-release channel. Our findings suggest that FKBP-dependent modulation of channel function may be generally applicable to all members of the intracellular Ca
2+
-release channel family and that FKBPs may play important regulatory roles in many cell processes, ranging from long-term depression in neurons to contractility in cardiomyocytes.
Anisotropic Activation Spread in Heart Cell Monolayers Assessed by High-Resolution Optical Mapping
Authors: Fast Vladimir G. , Darrow Bruce J. , Saffitz Jeffrey E. , Kle´ber Andre´ G.
Published: June 2012

The role of tissue discontinuities in anisotropic impulse propagation was assessed in two-dimensional anisotropic monolayers of neonatal rat myocytes cultured on a growth-directing substrate of collagen. Activation spread and distribution of maximal upstroke rate of rise (V˙
max
) of the action potential were measured with an optical system using a voltage-sensitive fluorescent dye (RH-327) and a 10×10 photodiode array with a spatial resolution ranging from 7 to 15 μm. Activation maps were compared with the cellular architecture and the distribution of gap junctions obtained from immunostaining the gap junction protein connexin43 (Cx43). Four types of structures were studied: (1) dense cell cultures, (2) cultures with anisotropic intercellular clefts of variable size, (3) discontinuities created by inclusion of nonmyocyte cells, and (4) discontinuities resulting from nonuniform expression of gap junctions. In dense monolayers, activation spread was continuous with microinhomogeneities in both longitudinal and transverse directions. The average cell dimensions in such monolayers were smaller than in adult canine myocardium. However, the degree of cellular anisotropy (length-to-width ratio of 5.3±1.4) and connectivity were similar. The presence of small intercellular clefts (less than one cell in length) did not disturb the general pattern of transverse or longitudinal activation spread, but it was associated with wave front microcollisions during transverse propagation and a concomitant increase of V˙
max
beyond the cleft. Long intercellular clefts caused discontinuous transverse propagation. Conduction velocity and V˙
max
decreased significantly at narrow isthmuses formed by closely apposed clefts, rendering such sites susceptible for conduction block. In contrast, V˙
max
increased when the wave front faced the borders of the clefts. Nonmyocyte cells were electrically connected to myocytes and served as sinks for electrotonic currents, thereby producing localized conduction slowing and a decrease in V˙
max
. Localized inhomogeneity in Cx43 distribution correlated accurately with circumscribed conduction block and changes in V˙
max
. Our results provide direct experimental evidence that the cellular structure and gap junction distribution correlate with action potential propagation and distribution of V˙
max
. We suggest that in tissue with a nonuniform anisotropy, connective tissue separating fiber bundles or sites of inhomogeneous connexin distribution may represent predilective sites for block in transverse direction.
Expression of Inducible Nitric Oxide Synthase Inhibits Platelet Adhesion and Restores Blood Flow in the Injured Artery
Authors: Yan Zhong-qun , Yokota Tasuku , Zhang Weiguo , Hansson Go¨ran K.
Published: June 2012

NO generated by endothelial cells is vasoprotective by antagonizing platelet adhesion and smooth muscle contraction. Since vascular smooth muscle cells (VSMCs) produce NO in response to cytokine stimulation and after arterial injury, we speculated that NO produced by VSMCs could compensate for the loss of endothelium. Using balloon catheter denudation of the rat carotid artery as a model for arterial injury and restenosis, we have evaluated the time course of expression of inducible NO synthase (iNOS) by in situ hybridization and immunohistochemistry and studied the effect of iNOS on platelet adhesion and blood flow of the injured vessel. iNOS mRNA and protein were expressed in the innermost layer of the media from day 1 and were subsequently detected in the neointima, whereas no expression was detectable in the uninjured artery. Systemic administration of
N

ω

-nitro-
l
-arginine methyl ester (L-NAME) resulted in a twofold to threefold increase in the adhesion of
111
In-labeled platelets to the injured vessel wall. Platelet adhesion was also enhanced threefold by local delivery of L-NAME from a gel surrounding the injured vessel, whereas the stereoisomer, D-NAME, had no effect. Finally, inhibition of NO synthase led to a 24% reduction of the blood flow in the injured carotid artery. These results demonstrate that arterial injury triggers the expression of iNOS in the lesion and that NO produced by iNOS inhibits platelet adhesion and restores blood flow. This could explain the disappearance of platelet thrombi from deendothelialized arterial surfaces within a few days after injury and indicates the importance of NO generated by iNOS for the maintenance of vascular tone. Thus, expression of iNOS in lesions may represent a protective mechanism that compensates for the loss of endothelium.
Factors Involved in Cardiogenesis and the Regulation of Cardiac-Specific Gene Expression
Authors: Mably John D. , Liew Choong-Chin
Published: June 2012

The delineation of the mechanisms that regulate cardiac gene expression is central to our understanding of cardiac growth and development. Much progress has been made toward the identification of factors involved in tissue-restricted gene expression, especially in skeletal muscle cells. However, the mechanisms regulating the expression of cardiac-specific genes remain less well understood. Certain homeodomain proteins have been implicated in commitment to the cardiac phenotype. Among the best characterized are the murine proteins Csx, Nkx-2.5, and Nkx-2.6, related to the protein tinman, which is essential for heart formation in
Drosophila
. The expression of these genes precedes that of cardiac-specific genes and is therefore believed to play a critical role in the development of the heart. The GATA proteins are a family of zinc finger proteins that are also expressed early in cardiac development and may act separately from, or in concert with, the homeodomain proteins as crucial regulators of heart development. The myosin heavy and light chain genes, the actin genes, the troponin genes, and the atrial natriuretic factor and muscle creatine kinase genes have served as excellent paradigms for the study of cardiac gene expression. Although differences in
cis
-acting elements and their behavior in binding assays have been observed between different genes, there exist similarities that are noteworthy. In this review, we will discuss the factors involved in the regulation of cardiac-specific gene expression in an attempt to provide a better understanding of the process of cardiogenesis.
Identification of a Putative Microvascular Oxygen Sensor
Authors: Harder David R. , Narayanan Jayashree , Birks Eric K. , Liard Jean Francois , Imig John D. , Lombard Julian H. , Lange Andrew R. , Roman Richard J.
Published: June 2012

The vascular response to changes in oxygen levels in the blood and tissue is a highly adaptive physiological response that functions to match tissue oxygen supply to metabolic demand. Defining the cellular mechanisms that can sense physiologically relevant changes in P
o
2
and adjust vascular diameter are vital to our understanding of this process. A cytochrome P450 (P450) enzyme of the 4A family of ω-hydroxylases was localized in renal microvessels, renal cortex, and a striated muscle microvascular bed (cremaster) of the rat. In the presence of molecular oxygen, this P450 enzyme catalyzes formation of 20-HETE from arachidonic acid (AA). Prior studies have shown that 20-HETE potently contracts renal and cerebral arteries and arterioles. The present study demonstrates that 20-HETE constricts striated muscle arterioles as well. In both intact renal microvessels and enriched renal cortical microsomal enzyme preparations, the formation of 20-HETE was linearly dependent on P
o
2
between 20 and 140 mm Hg. Homogenates of cremaster tissue produced 20-HETE when incubated with AA. They also expressed message for P450 4A enzyme, as determined by Southern and Western blots. Administration of 17-octadecynoic acid (17-ODYA), which is a P450 4A inhibitor, attenuated the constriction of third-order cremasteric arterioles in response to elevation of superfusion solution P
o
2
from ≈3 to 5 mm Hg to ≈35 mm Hg. 17-ODYA had no effect on basal vascular tone or response of cremaster arterioles to vasoactive compounds. These results demonstrate the existence of P450 ω-hydroxylase activity and 20-HETE formation in the vasculature and parenchyma of at least two microvascular beds. Our data suggest that a P450 enzyme of the 4A family has the potential to function as an oxygen sensor in mammalian microcirculatory beds and to regulate arteriolar caliber by generating 20-HETE in an oxygen-dependent manner.
Pressure and Angiotensin II Synergistically Induce Aortic Fibronectin Expression in Organ Culture Model of Rabbit Aorta
Authors: Bardy Nathalie , Merval Re´gine , Benessiano Joe¨lle , Samuel Jane-Lyse , Tedgui Alain
Published: June 2012

Aortic fibronectin (FN) expression is augmented in hypertension. Increasing evidence suggests that both angiotensin II (Ang II) and mechanical factors may induce vascular remodeling in response to hypertension. We have previously shown that, in vitro, increased transmural pressure enhances FN expression in rabbit aortic media. To investigate the existence of a link between the effects of pressure and Ang II and to explore the mechanisms underlying such a relationship, we quantified the effect of Ang II and Ang II inhibitors on the pressure-dependent FN expression in a 3-day organ culture model of rabbit aorta using immunolabeling analysis and detected FN mRNAs by in situ hybridization. A dose-dependent effect of Ang II on FN expression was observed at both 80 and 150 mm Hg but not at 0 mm Hg (relaxed vessels). One μmol/L Ang II increased the media cross-sectional surface, showing FN expression from 7.9±0.7% (n=9) to 18.9±1.1% (n=4) at 80 mm Hg (
P
<.01) and from 17.4±1.8% (n=9) to 56.6%±3.6 (n=4) at 150 mm Hg (
P
<.001). In situ hybridization revealed that Ang II and pressure upregulated FN mRNA expression. Losartan, an AT
1
antagonist, not only blocked the Ang II effect but also inhibited the transmural pressure effect. Angiotensin-converting enzyme inhibition abolished the pressure-dependent FN expression and significantly diminished the effect of pressure in the presence of Ang II. The effect of renin-angiotensin system inhibitors was specific for FN, since neither bFGF nor laminin expression was affected by these agents. Taken together, the results demonstrate that (1) the effect of transmural pressure is mediated by the stimulation of a local renin-angiotensin system, resulting in a net Ang II production in the culture medium, (2) transmural pressure and Ang II act synergistically to enhance vascular FN expression, (3) AT
1
receptors mediate both the effects of pressure and of exogenous Ang II, and (4) the effect of Ang II on FN expression is regulated at a pretranslational level.
Functional and Structural Assessment of Intercellular Communication
Authors: Darrow Bruce J. , Fast Vladimir G. , Kléber André G. , Beyer Eric C. , Saffitz Jeffrey E.
Published: June 2012

Remodeling of conduction pathways in the hypertrophic response to myocardial injury is a potential mechanism leading to the development of anatomic substrates of lethal arrhythmias. To delineate the responsible mechanisms and to directly relate changes in intercellular coupling at gap junctions with electrophysiological alterations, we studied the effects of cAMP, a mediator of cardiac hypertrophy, on action potential conduction velocity and connexin expression in neonatal rat ventricular myocyte cultures. Conduction velocity was measured with an optical activation mapping technique in cells loaded with the voltage-sensitive dye RH-237. Action potentials were conducted 24% to 29% more rapidly (
P
<.005) after incubating cultures for 24 hours with the cAMP analogue dibutyryl cAMP (db-cAMP, 1 mmol/L). However, db-cAMP caused no change in the maximum rate of rise of the action potential upstroke, V̇
max
. Electron and immunofluorescence microscopy revealed a significant increase in the number and size of gap junctions in db-cAMP–treated cells. Immunoblotting showed that the total amounts of the ventricular gap junction proteins connexin43 and connexin45 (Cx43 and Cx45, respectively) increased 2- to 4-fold. Immunoprecipitation of metabolically labeled connexin proteins revealed a dose-dependent increase in the rate of Cx45 protein synthesis in myocytes exposed to db-cAMP (>2-fold after a 4-hour exposure) but no change in the Cx43 synthesis rate. Northern blot analysis demonstrated a time-dependent increase in the amount of Cx43 mRNA, with a maximum 3.3-fold increase after 4 hours of exposure to 1 mmol/L db-cAMP; cycloheximide did not block this effect. In contrast, Cx45 mRNA levels were not altered significantly after db-cAMP treatment. Thus, cAMP causes a significant increase in conduction velocity that appears to be attributable largely to enhanced expression of proteins responsible for intercellular communication. Cx43 and Cx45 levels appear to be upregulated by cAMP by disparate molecular mechanisms.
Rapid Microvessel Rarefaction With Elevated Salt Intake and Reduced Renal Mass Hypertension in Rats
Authors: Hansen-Smith Fay M. , Morris Laura W. , Greene Andrew S. , Lombard Julian H.
Published: June 2012

To identify the sequence of events associated with the development of reduced vessel density (rarefaction) in hypertension, microvessel density and ultrastructure were assessed in the cremaster muscle of rats subjected to a 75% surgical reduction of renal mass and normotensive sham-operated control rats. Rats with reduced renal mass (RRM rats) and sham-operated rats were then maintained on either a high salt (4.0% NaCl) or a low salt (0.4% NaCl) diet for 3 days. Acute exposure to the high salt diet significantly increased mean arterial pressure in RRM rats but did not affect sham-operated control rats. Quantitative fluorescence microscopy of cremaster muscle whole mounts using rhodamine-labeled
Griffonia simplicifolia
I lectin revealed substantial rarefaction of microvessels in both RRM hypertensive rats and normotensive sham-operated rats on a high salt diet relative to corresponding control rats on a low salt diet. Confocal microscopy revealed a heterogeneous distribution of microvessels in RRM rats on a high salt diet, with some areas largely devoid of vessels. RRM and sham-operated rats on a high salt diet both exhibited changes in arteriolar ultrastructure, which included a loss of basement membranes and a dissociation of the endothelial and smooth muscle components of the vascular wall, resulting in a loss of vessel integrity. These observations demonstrate that a rapid loss of microvessels can occur not only in rats with RRM hypertension but also in normotensive rats on a high salt diet. This loss of microvessels results from structural alterations, which differ from the degenerative processes associated with microvascular rarefaction in rats with chronic RRM hypertension.
Differential Expression of α
1
Type VIII Collagen in Injured Platelet-Derived Growth Factor-BB–Stimulated Rat Carotid Arteries
Authors: Bendeck Michelle P. , Regenass Stephan , Tom W. David , Giachelli Cecilia M. , Schwartz Stephen M. , Hart Charles , Reidy Michael A.
Published: June 2012

Migration of smooth muscle cells from media to intima is critical for the development of neointimal thickening after balloon catheter injury of the rat carotid artery. The present experiments were designed to identify molecules expressed by smooth muscle cells migrating in vivo in the injured artery. Cell migration was maximized by infusing recombinant platelet-derived growth factor-BB (PDGF-BB) after a minimal filament denudation of the rat carotid artery, whereas cell proliferation was minimized by injecting an antibody against basic fibroblast growth factor (bFGF). This treatment caused an eightfold increase in smooth muscle cell migration into the intima but only a twofold increase in intimal smooth muscle cell replication rates. Differential display screening was used to isolate cDNAs that were overexpressed in the injured PDGF-BB–treated versus unmanipulated rat carotids. One of the clones isolated hybridized to a 4.2-kb mRNA species and shared 90% sequence homology to mouse α
1
type VIII collagen. Northern and Western blots confirmed overexpression of type VIII collagen in the injured PDGF-BB–treated vessels. In a separate series of experiments, we performed filament denudation injury and administered antibodies to inhibit the actions of endogenous bFGF and PDGF-BB, thereby decreasing smooth muscle cell migration, and found that type VIII collagen mRNA expression varied with migration. Using a different arterial injury model (balloon catheter injury), we showed that expression of type VIII collagen was maximal 2 to 4 days after injury, in coincidence with cell migration from the media to the intima. This molecule constitutes an important component of smooth muscle cell response to vessel injury and may play an important functional role in mediating migration.
Muscle Length Directs Sympathetic Nerve Activity and Vasomotor Tone in Resistance Vessels of Hamster Retractor
Authors: Welsh Donald G. , Segal Steven S.
Published: June 2012

Increased resistance to blood flow with muscle extension has been explained by the deformation of vessels within the muscle. In the present study, we developed a novel preparation of the hamster retractor muscle to investigate whether passive changes in skeletal muscle length elicit active vasomotor responses through a range of motion (85% to 130% of in vivo length; sarcomere length, 2.69±0.02 to 4.05±0.01 μm) encompassing the classic length-tension relationship. Arterioles (diameter, 32±3 μm) and feed arteries (diameter, 75±4 μm) were observed to progressively constrict (by 8±1 and 17±2 μm, respectively) with muscle lengthening, reducing blood flow by >50%; reciprocal changes occurred with passive shortening. Sodium nitroprusside (10 μmol/L) dilated vessels (to 47±2 and 98±4 μm, respectively) and abolished vasomotor responses to changing muscle length. The coordination of vasomotor responses between arterioles and feed arteries maintained wall shear rate (control, 1764±200 s
−1
) and perfusion pressure (60±5 mm Hg) into the arteriolar network. Tetrodotoxin (TTX, 1 μmol/L), phentolamine (1 μmol/L), prazosin (0.1 μmol/L), or 6-hydroxydopamine (1 mmol/L) inhibited vasoconstrictor responses, indicating that action potentials initiated by muscle lengthening give rise to norepinephrine release from sympathetic nerves. As shown with glyoxylic acid staining, sympathetic nerves formed a plexus encompassing arterioles and feed arteries. To test for a reflexive response initiated by intramuscular mechanoreceptors, TTX was applied with micropipettes to proximal segments of feed arteries, thereby neurally “isolating” the muscle from the hamster. Whereas lengthening-induced vasoconstriction persisted in arterioles and in feed artery segments distal to TTX, there was no vasomotor response central to the block. We conclude that passive lengthening stimulates the activity of periarteriolar sympathetic nerves; this activity propagates antidromically along nerve fibers into the feed arteries. These findings identify a mechanotransduction sequence by which the length of skeletal muscle actively governs vasomotor tone and the supply of oxygen to muscle fibers.
Role of the Kv4.3 K
+
Channel in Ventricular Muscle
Authors: Dixon Jane E. , Shi Wenmei , Wang Hong-Sheng , McDonald Christine , Yu Hangang , Wymore Randy S. , Cohen Ira S. , McKinnon David
Published: June 2012

The expression of 15 different K
+
channels in canine heart was examined, and a new K
+
channel gene (Kv4.3), which encodes a rapidly inactivating K
+
current, is described. The Kv4.3 channel was found to have biophysical and pharmacological properties similar to the native canine transient outward current (I
to
). The Kv4.3 gene is also expressed in human and rat heart. It is concluded that the Kv4.3 channel underlies the bulk of the I
to
in canine ventricular myocytes, and probably in human myocytes. Both the Kv4.3 and Kv4.2 channels are likely to contribute to the I
to
in rat heart, and differential expression of these two channels can account for observed differences in the kinetic properties of the I
to
in different regions of rat ventricle. There are significant differences in the pattern of K
+
channel expression in canine heart, compared with rat heart, and these differences may be an adaptation to the different requirements for cardiac function in mammals of markedly different sizes. It is possible that the much longer ventricular action potential duration observed in canine heart compared with rat heart is due, in part, to the lower levels of Kv1.2, Kv2.1, and Kv4.2 gene expression in canine heart.
Spatial Changes in Transmembrane Potential During Extracellular Electrical Shocks in Cultured Monolayers of Neonatal Rat Ventricular Myocytes
Authors: Gillis Anne M. , Fast Vladimir G. , Rohr Stephan , Kléber André G.
Published: June 2012

This study investigated the role of different types of discontinuities in tissue architecture on the spatial distribution of the transmembrane potential. Specifically, we tested the occurrence of so-called “secondary sources,” ie, localized hyperpolarizations and depolarizations during the application of extracellular electrical shocks (EESs). Changes in transmembrane potential relative to action potential amplitude (ΔV
m
/APA) were measured in patterned cultures of neonatal rat myocytes, stained with voltage-sensitive dye (RH-237), by optical mapping (96-channel photodiode array, 6- to 30-μm resolution) during the application of EES (field strength, 8 to 22 V/cm; duration, 6 ms). Across narrow cell strands (width, 218±59 [mean±SD] μm), EES applied during the relative refractory period produced a linear and symmetrical profile of ΔV
m
/APA (−65±23% maximal hyperpolarization versus +64±15% maximal depolarization). In contrast, the profile of ΔV
m
/APA was asymmetrical when EESs were applied during the action potential plateau (−95±32% versus +37±14%). At high magnification, no secondary sources were observed at the borders between cells. In dense isotropic cell monolayers or in monolayers and strands showing intercellular clefts, secondary sources were frequently observed. Intercellular clefts of the size of one to several myocytes were sufficient to produce secondary sources of the same magnitude as those that elicited action potentials in dense cell strands. There was a close correlation between the location of secondary sources during EES and localized conduction slowing during propagation. Thus, densely packed cultured cell strands behave as an electrical continuum with no secondary sources occurring at cell borders. Small intercellular clefts can create secondary sources of sufficient magnitude to exert a stimulatory effect.
Overexpression of Tissue Inhibitor of Matrix Metalloproteinase-1 Inhibits Vascular Smooth Muscle Cell Functions In Vitro and In Vivo
Authors: Forough Reza , Koyama Noriyuki , Hasenstab David , Lea Holly , Clowes Monika , Nikkari Seppo T. , Clowes Alexander W.
Published: June 2012

Arterial smooth muscle cells (SMCs) are in a quiescent growth state under normal physiological conditions, but they can be stimulated to proliferate and migrate from one tissue compartment to another if the vessel is injured. This response might require a selective and focal increase in tissue degradation, which might be mediated through the increased production of matrix metalloproteinases (MMPs). Blockade of MMP activity might therefore inhibit the SMC response to injury. To test this hypothesis, we developed clones of rat SMCs that overexpress baboon tissue inhibitor of matrix metalloproteinase-1 (TIMP-1), using retrovirally mediated gene transfer, and characterized the functional capacity of these cells in vitro and in vivo. SMCs transduced with the TIMP-1 vector (LTSN) grew more slowly and also migrated through a gel matrix in a Boyden chamber assay more slowly than the vector alone (LXSN) cells. The conditioned medium from LTSN cells completely inhibited the platelet-derived growth factor-BB–induced migration of normal SMCs across a matrix-coated filter, while the LXSN cell conditioned medium had no effect. The inhibitory activity in the LTSN conditioned medium could be neutralized with an antibody to TIMP-1. In vivo, local overexpression of TIMP-1 using LTSN cells implanted onto balloon-injured rat carotid artery inhibited intimal hyperplasia. Neutralizing antibodies against TIMP-1 suppressed the effect of LTSN cell seeding on intimal thickening. These data support the conclusion that the process of SMC activation leading to a thickened intima is dependent on MMP activity and that TIMP-1 could be utilized to inhibit this process.
Linearity of Pulsatile Pressure-Flow Relations in the Embryonic Chick Vascular System
Authors: Yoshigi Masaaki , Keller Bradley B.
Published: June 2012

The calculation and modeling of vascular input impedance are based on the assumption that pressure and flow are linearly related in the frequency domain. However, this assumption has not been proven for the embryonic circulation. Therefore, we investigated the linearity of pulsatile pressure-flow relations in vivo with acute alterations in cycle length. We simultaneously measured dorsal aortic pressure with a servonull system and flow velocity with a 20-MHz pulsed-Doppler system in stage 24 chick embryos (n=38). Cycle length was acutely altered using thermal probe(s) applied to the sinus venosus. We determined the impedance spectra at several cycle lengths for each embryo and a reference curve from a three-element Windkessel model with the use of nonlinear curve fitting. We then assessed the scatter of experimental impedance along the reference curve as a measure of linearity in the frequency domain. We found that mean vascular resistance did not change after thermal probe applications (
P
>.20 for each), indicating that acute alterations in cycle length did not alter peripheral vascular properties. Superpositioned impedance spectra showed minimal scatter along the model impedance from 0 to 6 Hz. Goodness of fit values (
R
2
) were near unity (.94 to .97) and were similar for all interventions (
P
>.07 for Fisher's
z
, by F test). Above 6 Hz, both modulus and phase spectra exhibited significant scatter (
P
<.05, by F test). Experimental impedance spectra tended to have a fluctuation and a phase-zero crossover, indicating significant wave reflection in the embryonic circulation. Thus, the embryonic vascular system can be approximated as a linear system from 0 to 6 Hz, the range in which the majority (96.0±0.18%) of hydraulic energy is dissipated.
Role of Ca
2+
-Activated K
+
Channels in the Regulation of Membrane Potential and Tone of Smooth Muscle in Human Pial Arteries
Authors: Gokina Natalia I. , Wellman Theresa D. , Bevan Rosemary D. , Walters Carrie L. , Penar Paul L. , Bevan John A.
Published: June 2012

Smooth muscle cells (SMCs) in 58% of human pial arteries obtained during surgery showed no spontaneous contractions and displayed a stable resting membrane potential (MP) of −54.7±1.5 mV. Those that exhibited periodic spontaneous contractions associated with periodic depolarization and generation of spontaneous action potentials (APs) had a less negative MP of −43.1±0.5 mV (42%). Inhibition of calcium-activated potassium (K
Ca
) channels in the silent arteries by charybdotoxin (CTX) and tetraethylammonium ions (TEA) induced dose-dependent depolarization, AP generation, and contraction. TEA and CTX enhanced the spontaneous depolarization and force in arteries that exhibited spontaneous activity. They also prolonged the spontaneous APs up to several times and increased their upstroke amplitude. Both TEA and CTX failed to produce significant depolarization in arteries treated with nifedipine. It is concluded that K
Ca
channels are important regulators of human pial artery SMC resting MP and tone. They are also involved in the control of AP amplitude and duration and the associated contractions. These data suggest that alterations in the activity of SMC K
Ca
channels could be responsible for the appearance of spontaneous activity in human pial arteries in vitro and that impaired function of these channels might be related to vasospastic phenomena in human cerebral circulation.
Mechanical Strain Increases Smooth Muscle and Decreases Nonmuscle Myosin Expression in Rat Vascular Smooth Muscle Cells
Authors: Reusch Peter , Wagdy Hisham , Reusch Regina , Wilson Emily , Ives Harlan E.
Published: June 2012

The effect of cyclic (1-Hz) mechanical strain on expression of myosin heavy chain isoforms was examined in neonatal rat vascular smooth muscle cells cultured on silicone elastomer plates. Myosin heavy chain isoforms were identified by immunoblot using antibodies recognizing (1) smooth muscle myosin heavy chain isoforms SM-1 and SM-2, (2) SM-1 exclusively, and (3) nonmuscle myosin heavy chains A and B. In response to 36 to 72 hours of strain, SM-1 and SM-2 increased by fourfold to sixfold, whereas nonmuscle myosin A decreased to 30% of control. Nonmuscle myosin B was unaffected by strain. SM-1 mRNA increased by twofold to threefold after 12 hours of strain but decreased toward control levels thereafter. SM-2 mRNA was only barely detectable. Nonmuscle myosin A mRNA decreased to 50% of control after 3 hours of strain and then returned to the control level. Since these cells secrete platelet-derived growth factor (PDGF) in response to strain, we assessed the effects of PDGF on myosin isoform expression. Exogenous PDGF (10 ng/mL) decreased SM-1 expression by 35% and increased nonmuscle myosin expression twofold, opposite the effect of strain. In cells exposed to strain with neutralizing antibodies to PDGF-AB, the strain-induced increase in SM-1 was enhanced 10-fold, and nonmuscle myosin A was reduced to 40% of control. Finally, the effect of extracellular matrix on transduction of the strain signal was studied. Forty-eight hours of cyclic strain increased SM-1 by twofold in cells cultured on collagen type I and threefold in cells cultured on laminin. In fibronectin-cultured cells, strain elicited no increase in SM-1. Thus, mechanical strain, sensed through specific interactions with the matrix, can alter myosin isoform expression toward that found in a more differentiated state.
Apoptosis in Ischemic and Reperfused Rat Myocardium
Authors: Fliss Henry , Gattinger Deborah
Published: June 2012

Apoptosis has been observed previously in hearts subjected to either continuous ischemia or ischemia followed by reperfusion. The purpose of this study was to compare the timing and extent of apoptosis in both continuously ischemic and reperfused myocardium. We show that rats subjected to continuous coronary artery occlusion display characteristic signs of apoptosis solely in the ischemic myocardium after only 2.25 hours of ischemia, as illustrated by positive in situ end labeling (ISEL) of apoptotic cardiomyocyte nuclei in tissue sections and/or the presence of DNA “ladders” in agarose gels. In contrast, reperfusion after a 45-minute occlusion accelerated the process, with apoptosis becoming evident solely in the reperfused myocardium after only 1 hour of reperfusion. ISEL and DNA ladder intensity increased with duration of ischemia or reperfusion. The volume of myocardium in which ISEL was observed was smaller in the reperfused hearts, and the ISEL-stained nuclei represented 23% and 33% of the total nuclei in the reperfused and permanently occluded myocardium, respectively. Therefore, the data suggest that reperfusion lowers the extent of apoptosis in ischemic myocardium but, paradoxically, accelerates the residual apoptosis, possibly because of reperfusion injury. A large accumulation of neutrophils was observed in both the permanently occluded and reperfused myocardium, suggesting that the inflammatory response may have contributed to apoptosis in both settings. This study therefore confirms that both ischemic and reperfused rat myocardium can undergo apoptotic cell death. However, the data suggest that although reperfusion lowers the number of myocytes undergoing apoptosis, it accelerates apoptosis in the nonsalvageable cells.
Upregulation of Basement Membrane–Degrading Metalloproteinase Secretion After Balloon Injury of Pig Carotid Arteries
Authors: Southgate Kay M. , Fisher Michael , Banning Adrian P. , Thurston Valerie J. , Baker Andrew H. , Fabunmi Rosalind P. , Groves Peter H. , Davies Malcolm , Newby Andrew C.
Published: June 2012

Basement membrane–degrading metalloproteinases (gelatinases) appear necessary for vascular smooth muscle cell migration and proliferation in culture and for intimal migration of cells after balloon injury to the rat carotid artery. We investigated in the present study the secretion of gelatinases from pig carotid artery tissue after balloon injury. Segments of injured artery and segments proximal and distal to the area of injury were removed 3, 7, and 21 days after balloon dilatation. Medial explants from these segments were then cultured for 3 days, and the serum-free conditioned media were subjected to gelatin zymography. Production of 72- and 95-kD gelatinases was quantified by densitometry. Balloon-injured segments secreted significantly more 72- and 95-kD gelatinase than did paired distal segments at all time points. Release of both gelatinase activities was increased at 3 and 7 days relative to segments from uninjured arteries but declined again by 21 days after balloon injury. Similar results were found for gelatinase levels in extracts of arterial tissue. Consistent with the protein secretion data, in situ hybridization demonstrated that the mRNAs for both gelatinases were upregulated after balloon injury. Expression was prominent in medial smooth muscle cells, particularly around foci of necrosis, and in neointimal cells 3 and 7 days after balloon injury; 72-kD gelatinase mRNA persisted after 21 days and was prominent in regrown endothelial cells. The upregulation of gelatinase activity paralleled the time course of smooth muscle cell migration and proliferation in this model. We conclude that increased gelatinase production occurs in response to balloon injury and may play a role in permitting migration and proliferation of vascular smooth muscle cells.
The Influence of Pancreatic Secretions on the Fat Tolerance of Humans
Authors: TIETZ NORBERT W. , BORDEN TOM A. , HAMILTON JAMES C. , MURPHEY NORMA L.
Published: June 2012

A high fat meal was given to 78 volunteers. Total esterifled fatty acid, cholesterol, and lipase levels were determined in a fasting blood specimen and in specimens obtained 3 and 6 hours after the meal. Attempts were made to correlate the fat tolerance of the individual with the level of fat splitting enzymes in the various serum specimens. The influence of Pancreatic Hormone, whole pancreatic preparations, secretin, and heparin on the fat tolerance as well as on the lipase level was determined. Attempts were made to correlate the findings of fat tolerance and glucose tolerance tests done on the same individuals.
The Plateau of the Action Potential of the Frog Ventricle
Authors: MACFARLANE W. V.
Published: June 2012

A study was made of the changés produced in action potentials of frog ventricular cells by variation in ion concentrations, by administration of NaN
3
, 2:4 DNP, NaCN, NaIA and N
2
which shortened the potentials. The exposure of the perfused tissue to ionized acridines, barbiturates and desoxyglucose caused a prolongation of the action potentials. These actions as obtained were reversible. The evidence from the time-course and differential effects of inhibitors suggests that an active ion transport, modulated by K

o

+
may be important in maintaining the plateau phase of the action potential.
Further Studies on Experimental Atherosclerosis and Dietary Pyrimidines: Orotic Acid, Thiouracil, and Uracil in Male and Female Rats
Authors: FILLIOS LOUIS C. , NAITO CHIKAYUKI , ANDREWS STEPHEN B. , ROACH ALICE M.
Published: June 2012

The comparative potencies of uracil nnd thiouracil in terms of thyroidal changes, hypercholesteremia, azotemia, and incipient atherosclerosis were determined in rats fed cholesterol and cholic acid. Higher dietary levels of uracil were required to produce changes similar to those seen with thiouracil. Certain dietary components related to uracil were also studied. Orotic acid was found to have a dramatic effect on female rats: it favored a markedly lower serum cholesterol level and less cardiovascular sudanophilia. When orotic acid was combined with uracil, the uracil-induced hypercholesteremia and atherogenesis were significantly inhibited, particularly in the females. Orotic acid also protected against the uracil-induced thyroid hyperplasia in these females.
Behavior of the Right Ventricle Following Acute Constriction of the Pulmonary Artery
Authors: TAQUINI A. C. , FERMOSO J. D. , ARAMENDIA P.
Published: June 2012

In 15 anesthetized dogs, constriction of the lumen of the pulmonary artery by less than 50 per cent produced no significant change in intraventricular pressure or cardiac output. Additional constriction caused a progressive increase of the right ventricular pressure; up to a pressure of 62.3±3./min. Hg, there was no change in cardiac output. With higher right ventricular systolic pressure there was a reduction in cardiac output, gradual decrease in systolic pressure but increase in diastolic pressure of both ventricles, and ultimately dilatation of the heart and ventricular arrest or fibrillation.
Effect of Increased Fibrinogen Concentration on the Lysis of In Vivo Thrombi
Authors: BANG N. U. , FREIMAN A. H. , CLIFFTON E. E.
Published: June 2012

The dissolution of clots formed at normal fibrinogen levels and in hyperfibrinogenemia was studied in 10 dogs receiving a standard course of fibrinolytic treatment. A significantly increased resistance of clots formed at high fibrinogen levels to fibrinolytic therapy was found.
In Vitro Effects of Estradiol on the Aorta of Chickens
Authors: MALINOW MANUEL RENÉ
Published: June 2012

Aortas of chickens were incubated during 72 hours with 17α- or with 17β-estradiol and the alkaline phosphomonoesterase activity determined. Paired experiments demonstrated that both isomers are able to enhance the activity of the enzyme in vitro. The possible relation of these results with atherogenesis is discussed.
Arterial Lesions in the Kenya Baboon
Authors: MCGILL HENRY C. , STRONG JACK P. , HOLMAN RUSSELL L. , WERTHESSEN NICHOLAS T.
Published: June 2012

One hundred sixty-three baboons (
Papio doguera
were sacrificed and autopsied immediately after being trapped in their natural habitat in Kenya, British East Africa. Approximately three-fourths of the 67 adults had some degree of aortic intimal lipid deposition, as indicated by gross Sudan IV staining, and a few animals had extensive fatty streaks. These fatty streaks were more frequent with advancing age, but there was no sex difference. Electron microscopy disclosed most of the intimal lipid droplets to be intracellular. Fibrous plaques were infrequent and had a variable but usually low lipid content. One elderly male had fibrous plaques with hemorrhage into their bases. Histologic sections of the coronary arteries showed many small musculo-elastic intimal plaques in which lipid could only rarely be demonstrated. The aortic intimal lipid deposits cannot be attributed to the consumption of excessive animal fat or to hypercholesterolemia. These results indicate that the baboon, like man, is highly susceptible to arterial intimal lipid deposition and that it promises to be an excellent animal for the experimental investigation of atherosclerosis. They further more caution that adequate controls be used in all experimental work involving arterial lesions in primates.
Distensibility and Water Content of Heart Muscle Before and After Injury
Authors: SALISBURY PETER F. , CROSS CECIL E. , RIEBEN P. ANDRE
Published: June 2012

In open-chest dogs the circulation was carried on a heart-lung machine. The right heart and the left atrium were drained. A balloon was placed in the virtually bloodless left ventricle and filled with a known volume of air at the beginning of an experiment; left ventricular pressure was recorded after a steady state had become established. After this control observation, the preparation was exposed to experimental maneuvers, such as variation of the systemic flow rate or changes of the air volume in the balloon. At one or more later stages of the same experiment, the balloon was emptied, the heart was allowed to beat empty for several minutes, and the standard observation was again repeated. Comparison of left ventricular pressures from the initial and later observations showed that the diastolic pressure in the left ventricle increased in the course of the experiment even though other conditions were comparable. From this we conclude that experimental maneuvers injuring the heart muscle reduce its distensibility. The water content of injured hearts was abnormally elevated; myocardial edema is therefore not excluded as one of the mechanisms explaining the reduced distensibility of injured heart muscle.
Homeometric Autoregulation in the Heart
Authors: SARNOFF S. J. , MITCHELL J. H. , GILMORE J. P. , REMENSNYDER J. P.
Published: June 2012

The effect of increasing the activity of the ventricle on its contractility was investigated. Several beats after the ventricle increases the amount of tension it develops per unit of time, it exhibits an increased contractility as shown by the increase in work and the more rapid development of pressure from a given end-diastolic pressure or fiber length. This has been termed homeometric autoregulation in contradistinction to the Frank-Starling or heterometric type of autoregulation. It was found that changes in coronary flow are not essential to the exhibition of this phenomenon. Possible mechanisms and the physiologic significance of the findings are discussed.
Ventricular Endomyocardial Pathology Produced by Chronic Cardiac Lymphatic Obstruction in the Dog
Authors: MILLER ALBERT J. , PICK RUTH , KATZ LOUIS N. , Jones Charles , Ellis Anthony , Malasan John
Published: June 2012

Chronic impairment of cardiac lymph flow was successfully accomplished in 22 dogs after preliminary anatomic studies revealed a feasible surgical technic. The anatomic and surgical details are presented. Gross and microscopic studies were possible in 19 of the operated animals, 3 of which died spontaneously, and 16 of which were sacrificed at varying time intervals between 2 and 16 weeks after surgery. Two of the 19 dogs were completely normal on gross and histologic examination. Abnormalities found in the remaining 17 animals included left and right ventricular subendocardial hemorrhages, increased elastic and fibrous tissue in the left ventricular endocardium, and opacification of the mitral valve leaflets.

It is concluded that chronic impairment of cardiac lymph drainage is productive of significant endomyocardial pathology. These observations are considered to be important as possible etiologic mechanisms in endomyocardial fibrosis and endocardial fibroelastosis. The cardiac lymphatics merit continued intensive investigation, inasmuch as they surely have an important role in cardiac physiology and pathology. Their importance may encompass a far broader area than suggested above. They may be related to the so-called nutritional cardiopathies. The fibroplastic effects of serotonin might occur through an effect on the cardiac lymphatics. Impairment of heart lymph flow may be important in endocarditis and myocarditis of certain types, for it is known that chronic lymphatic obstruction predisposes to recurrent inflammation and infection in the affected parts.
Effects of Metabolic Acidosis on Ventricular Isometric Systolic Tension and the Response to Epinephrine and Levarterenol
Authors: DARBY THOMAS D. , ALDINGER EARL E. , GADSDEN RICHARD H. , THROWER WENDELL B. , Westbroo Sidney H. , Tenneii James M.
Published: June 2012

Hyperglycemia and hypoxia acidosis are known to occur during periods of shock. The plasma levels of catechol amines have been reported to be elevated during periods of shock. Injection of epinephrine or levarterenol can produce increments in blood glucose and have been shown to elicit a metabolic acidosis. In the present studies the effects of reducing circulating blood volume or infusion of lactic acid and of forced CO
2
inhalation on ventricular isometric systolic tension (VCF) and the cardiovascular response to test injections of levarterenol were studied. The experiments with a reduction in circulating blood volume and with lactic acid infusions resulted in a decrease in arterial pH. whole blood total CO
2
and whole blood CO
2
combining power. Concomitant with these changes there was a marked depression of VCF and the response to test injections of levarterenol. Correction of the acid-base changes with 2-amino-2-hydroxymethyl, 1, 3-propane diol resulted in an immediate improvement in VCF and in the response to levarterenol. This change occurred despite a greater increase in blood lactate levels. Anaerobic metabolism with lactate production leads to a decrease in total blood CO
2
and an increase in dissolved CO
2
. This decrease in the CO
2
-carrying capacity would lead to an increase in tissue CO
2
. Since the heart is markedly depressed by CO
2
, this increment could be largely responsible for the myocardial depression. The oxygen debt which the heart can incur is thought to be limited. Therefore, the build-up of CO
2
one of the end-products of aerobic metabolism, would depress oxidative metabolism according to the law of mass action. This hypothesis was further tested by forced CO
2
ventilation which resulted in a marked depression of VCF and the response to levarterenol. Correction of the respiratory acidosis by the administration of THAM which increased the CO
2
-carrying capacity of the blood resulted in an immediate improvement in VCF and the levarterenol response.