A laboratory-based hard x-ray monochromator for high-resolution x-ray emission spectroscopy and x-ray absorption near edge structure measurements.

We report the development of a laboratory-based Rowland-circle monochromator that incorporates a low power x-ray (bremsstrahlung) tube source, a spherically bent crystal analyzer, and an energy-resolving solid-state detector. This relatively inexpensive, introductory level instrument achieves 1-eV energy resolution for photon energies of ∼5 keV to ∼10 keV while also demonstrating a net efficiency previously seen only in laboratory monochromators having much coarser energy resolution. Despite the use of only a compact, air-cooled 10 W x-ray tube, we find count rates for nonresonant x-ray emission spectroscopy comparable to those achieved at monochromatized spectroscopy beamlines at synchrotron light sources. For x-ray absorption near edge structure, the monochromatized flux is small (due to the use of a low-powered x-ray generator) but still useful for routine transmission-mode studies of concentrated samples. These results indicate that upgrading to a standard commercial high-power line-focused x-ray tube or rotating anode x-ray generator would result in monochromatized fluxes of order 10(6)-10(7) photons/s with no loss in energy resolution. This work establishes core technical capabilities for a rejuvenation of laboratory-based hard x-ray spectroscopies that could have special relevance for contemporary research on catalytic or electrical energy storage systems using transition-metal, lanthanide, or noble-metal active species.

Changes in Ca(2+) cycling proteins underlie cardiac action potential prolongation in a pressure-overloaded guinea pig model with cardiac hypertrophy and failure.

Ventricular arrhythmias are common in both cardiac hypertrophy and failure; cardiac failure in particular is associated with a significant increase in the risk of sudden cardiac death. We studied the electrophysiologic changes in a guinea pig model with aortic banding resulting in cardiac hypertrophy at 4 weeks and progressing to cardiac failure at 8 weeks using whole-cell patch-clamp and biochemical techniques. Action potential durations (APDs) were significantly prolonged in banded animals at 4 and 8 weeks compared with age-matched sham-operated animals. APDs at 50% and 90% repolarization (APD(50) and APD(90) in ms) were the following: 4 week, banded, 208+/-51 and 248+/-49 (n = 15); 4 week, sham, 189+/-68 and 213+/-69 (n = 16); 8 week, banded, 197+/-40 and 226+/-40 (n = 21); and 8 week, sham, 156+/-42 and 189+/-45 (n = 22), respectively; P<0.05 comparing banded versus sham-operated animals. We observed no significant differences in the K(+) currents between the 2 groups of animals at 4 and 8 weeks. However, banded animals exhibited a significant increase in Na(+) and Na(+)-Ca(2+) exchange current densities compared with controls. Furthermore, we have found a significant attenuation in the Ca(2+)-dependent inactivation of the L-type Ca(2+) current in the banded compared with sham-operated animals, likely as a result of the significant downregulation of the sarcoplasmic reticulum Ca(2+) ATPase, which has been documented previously in the heart failure animals. Our data provide an alternate mechanism for APD prolongation in cardiac hypertrophy and failure and support the notion that there is close interaction between Ca(2+) handling and action potential profile.

PKC translocation without changes in Galphaq and PLC-beta protein abundance in cardiac hypertrophy and failure.

Activation of protein kinase C (PKC) has been implicated as playing a key role in the pathogenesis of cardiac hypertrophy. This study investigates the response of several signal transduction proteins responsible for PKC activation during the transition from compensated pressure-overload hypertrophy (POH) to congestive heart failure (CHF). Pressure overload was produced on male, adult, Hartley strain guinea pigs using a ligature around the descending thoracic aorta. Sham-operated controls, POH, and CHF groups were identified based on left ventricular hypertrophy, pulmonary congestion, and isolated heart Langendorff mechanics. Quantitative immunoblotting revealed phospholipase C (PLC)-betaI and Galphaq were unchanged during POH and CHF, as were RGS2, RGS3, and RGS4 (regulators of G protein signaling, which are activators of intrinsic GTPase activity). Translocation of PKC-alpha, -epsilon, and -gamma from cytosolic to membranous fractions were significantly increased during POH and CHF. Cytosolic PKC activity was also elevated during POH. We conclude that differential PKC activation may be mediated by increases in Galphaq and PLC-betaI activity rather than upregulation of expression.

The effects of levosimendan on the left ventricular function and protein phosphorylation in post-ischemic guinea pig hearts.

The widely accepted theories for the decreased function in the stunned myocardium relate to Ca2+ desensitization and free radical-mediated tissue damage of the myofilaments. The aim of the present study was to examine whether the depressed contractile function and Ca2+ responsiveness of the stunned myocardium may be restored by a new Ca2+ sensitizer (levosimendan), which has been shown to improve the Ca2+ response of the myofilaments. The effects of levosimendan on the left ventricular function and the in vivo protein phosphorylation were examined in both the non-ischemic and the stunned myocardium. Myocardial stunning was induced in Langendorff-perfused guinea pig hearts by suspending the circulation for 8 min, followed by a 20-min reperfusion period. Perfusion of post-ischemic guinea pig hearts with levosimendan (0.03-0.48 microM, 6 min) was associated with dose- and time-dependent increases in both dP/dtmax (contractility) and dP/dtmin (speed of relaxation). When the effectiveness of levosimendan was compared in non-ischemic and post-ischemic hearts, no significant differences were noted in the relative stimulatory effects on contractility and relaxation, at any given time point (time-response curve) or concentration (dose-response curve). Perfusion of the guinea pig hearts with a high (0.3 microM) levosimendan concentration did not reveal any qualitative or quantitative difference in the phosphodiesterase inhibitory potential of the compound (elevation of tissue cyclic AMP levels and characteristics of protein phosphorylation) between the non-ischemic and the post-ischemic myocardium. However, when isoproterenol was administered to induce maximal in vivo phosphorylation of cardiac phosphoproteins, an attenuation of the 32P-incorporation into troponin I was noted in the post-ischemic hearts. The decrease in isoproterenol-induced 32P-incorporation into troponin I was associated with similar alterations in the tissue level of this protein. We conclude that the Ca2+ sensitizer levosimendan exerts dose- and time-dependent positive inotropic and lusitropic effects on the post-ischemic myocardium, lending support to the hypothesis tha Ca2+ desensitization of the myofibrils is involved in myocardial stunning.

Responses of cardiac protein kinase C isoforms to distinct pathological stimuli are differentially regulated.

Currently at least 11 protein kinase C (PKC) isoforms have been identified and may play different roles in cell signaling pathways leading to changes in cardiac contractility, the hypertrophic response, and tolerance to myocardial ischemia. The purpose of the present study was to test the hypothesis that responses of individual PKC isoforms to distinct pathological stimuli were differentially regulated in the adult guinea pig heart. Isolated hearts were perfused by the Langendorff method and were exposed to ischemia, hypoxia, H(2)O(2), or angiotensin II. Hypoxia and ischemia induced translocation of PKC isoforms alpha, beta(2), gamma, and zeta, and H(2)O(2) translocated PKC isoforms alpha, beta(2), and zeta. Angiotensin II produced translocation of alpha, beta(2), epsilon, gamma, and zeta isoforms. Inhibition of phospholipase C with tricyclodecan-9-yl-xanthogenate (D609) blocked hypoxia-induced (alpha, beta(2), and zeta) and angiotensin II-induced (alpha, beta(2), gamma, and zeta) translocation of PKC isoforms. Inhibition of tyrosine kinase with genistein blocked translocation of PKC isoforms by hypoxia (beta(2) and zeta) and by angiotensin II (beta(2)). By contrast, neither D609 nor genistein blocked H(2)O(2)-induced translocation of any PKC isoform. We conclude that hypoxia-induced activation of PKC isoforms is mediated through pathways involving phospholipase C and tyrosine kinase, but oxidative stress may activate PKC isoforms independently of Galphaq-phospholipase C coupling and tyrosine kinase signaling. Because oxidative stress may directly activate PKC, and PKC activation appears to be involved in human heart failure, selective inhibition of the PKC isoforms may provide a novel therapeutic strategy for the prevention and treatment of this pathological process.

Effect of angiotensin-converting enzyme inhibition on protein kinase C and SR proteins in heart failure.

We tested the hypothesis that activation of protein kinase C (PKC) isoforms in pressure-overload heart failure was prevented by angiotensin-converting enzyme (ACE) inhibition, resulting in normalization of cardiac sarcoplasmic reticulum (SR) Ca2+ ATPase (SERCA) 2a and phospholamban protein levels and improvement in intracellular Ca2+ handling. Aortic-banded and control guinea pigs were given ramipril (5 mg. kg-1. day-1) or placebo for 8 wk. Ramipril-treated banded animals had lower left ventricular (LV) and lung weight, improved survival, increased isovolumic LV mechanics, and improved cardiomyocyte Ca2+ transients compared with placebo-treated banded animals. This was associated with maintenance of SERCA2a and phospholamban protein expression. Translocation of PKC-alpha and -epsilon was increased in placebo-treated banded guinea pigs compared with controls and was attenuated significantly by treatment with ramipril. We conclude that ACE inhibition attenuates PKC translocation and prevents downregulation of Ca2+ cycling protein expression in pressure-overload hypertrophy. This represents a mechanism for the beneficial effects of this therapy on LV function and survival in heart failure.

Left ventricular stretch stimulates angiotensin II--mediated phosphatidylinositol hydrolysis and protein kinase C epsilon isoform translocation in adult guinea pig hearts.

Stretch of neonatal cardiomyocytes activates phospholipase C with production of inositol trisphosphate and diacylglycerol in part by formation of angiotensin II (Ang II). However, the response of this pathway to physical stimuli in the adult heart is poorly understood. Thus, in isovolumic perfused guinea pig hearts, we characterized stretch-mediated phosphatidylinositol (PI) hydrolysis and protein kinase C (PKC) isoform translocation using elevated diastolic pressure. Balloon dilatation (minimum diastolic pressure, 25 mm Hg) of the left ventricle (LV) stimulated PI hydrolysis. Pretreatment of stretched hearts with the specific angiotensin (AT1) receptor antagonist losartan abolished stretch-mediated accumulation of inositol phosphates. To examine PKC isoform expression and activation under these conditions, whole-heart extracts were examined by immunoblot analysis. Ang II translocated PKC epsilon to the particulate fraction. 4 beta-Phorbol 12-myristate 13-acetate but not an inactive congener translocated PKC epsilon to the particulate fraction and produced a decrease in myocardial contractile function. Mechanical stretch also translocated PKC epsilon to the particulate fraction; however, this was attenuated but not abolished by losartan. We conclude that in the adult heart, LV dilation produced stretch-mediated activation of phospholipase C, which resulted in PI hydrolysis and PKC epsilon activation in part by stimulation of the local renin angiotensin system. In contrast to stretch-mediated inositol phosphate accumulation, PKC epsilon translocation is not prevented by AT1 receptor blockade, indicating that this PKC isoform can be activated in response to mechanical deformation by an Ang II-independent mechanism in the adult myocardium.

Differential changes in cardiac phospholamban and sarcoplasmic reticular Ca(2+)-ATPase protein levels. Effects on Ca2+ transport and mechanics in compensated pressure-overload hypertrophy and congestive heart failure.

The objective of this study was to elucidate the role of the sarcoplasmic reticulum (SR) in the transition from compensated pressure-overload hypertrophy (increased left ventricular [LV] mass, normal LV function, and no pulmonary congestion) to congestive heart failure (increased LV mass, depressed LV function, and pulmonary congestion). To address this issue, the descending thoracic aorta was banded for 4 and 8 weeks in adult guinea pigs, and the changes in isovolumic LV mechanics, SR Ca2+ transport, and SR protein levels were determined and compared with age-matched sham-operated control animals. A subgroup of the 8-week banded animals manifested the congestive heart failure phenotype with diminished developed LV pressure normalized by LV mass, reduced rates of LV pressure development and relaxation, and markedly increased lung weight-to-body weight ratios. The cardiac mechanical and morphometric changes were associated with depressed protein levels of the SR Ca(2+)-ATPase (85% of the control) and phospholamban (65% of the control) assessed by quantitative immunoblotting. Resultant rates of SR Ca2+ uptake (Vmax) and the affinity of SR Ca(2+)-ATPase for Ca2+ (EC50) were significantly depressed [32 +/- 6 nmol Ca2+.min-1.mg-1 and 0.59 +/- 0.12 (mumol/L)/L, respectively] compared with the 8-week sham-operated control animals [40 +/- 1 nmol Ca2+.min-1.mg-1 and 0.40 +/- 0.05 (mumol/L)/L, respectively]. We conclude that this model of pressure overload-induced cardiac failure is associated with (1) diminished LV force development, rates of pressure development, and decay; (2) depressed protein expression of the Ca(2+)-cycling proteins SR Ca(2+)-ATPase and phospholamban; and (3) decreased Vmax and affinity of the SR Ca(2+)-ATPase for Ca2+.(ABSTRACT TRUNCATED AT 250 WORDS)

Radial nerve palsy: a complication of walker usage.

A patient with diabetic peripheral neuropathy experienced the acute onset of a proximal radial nerve palsy after prolonged use of a walker. Nerve conduction and electromyographic studies confirmed an isolated, severe neurapraxic lesion distal to branches innervating the triceps and anconeus muscles. The acute onset and severity of this lesion suggests that it was caused by mechanical compression of the radial nerve as it exits the spiral groove. Radial mononeuropathy has been reported in conjunction with muscular effort of the triceps muscle. Previous case studies and a review of the literature are discussed. Awareness of this complication in patients using walkers and wheelchairs is important for prevention and diagnosis in rehabilitation.

Contribution of AV3V region in anephric NaCl-induced hypertension in the rat.

The role of the anteroventral third ventricle (AV3V) region in mediating hypertonic sodium chloride-induced pressor responses was investigated in conscious rats. Sham- and AV3V-lesioned rats were prepared with femoral artery and vein catheters and subjected to bilateral nephrectomy under gaseous anesthesia. After recovery, animals were infused intravenously with isotonic (0.5 meq/kg) or hypertonic (10 meq/kg) saline at a rate of 0.0103 ml/min over 2 h. Isotonic saline infusion did not affect arterial pressure or heart rate in either group of rats. Hypertonic saline increased arterial pressure 35 +/- 3 mmHg in sham-lesioned rats and only 10 +/- 4 mmHg in AV3V-lesioned animals (P less than 0.0005). Sham-lesioned rats infused with hypertonic saline had a greater vasopressin-dependent component maintaining arterial pressure than the other groups of rats. Conversely, the sympathetic nervous system-dependent component of blood pressure was suppressed in the hypertonic saline-infused sham-lesioned animals compared with the other animals. However, when the vasopressin receptors were blocked, the neurally mediated portion of blood pressure was similar in all four groups of rats. These results emphasize that circulating vasopressin is important for the rise in arterial pressure accompanying the osmotic stimulus. Furthermore, this study demonstrates that the AV3V region is necessary for vasopressin-dependent pressor responses caused by an osmotic stimulus.

Inability of dorsal spinal rhizotomy to prevent renal wrap hypertension in rats.

Renal hypertension has been shown to be prevented and reversed by renal denervation. It has been postulated that the afferent nerves from the kidney are responsible for mediating the hypertensive stimulus that activates the sympathetic nervous system and increases arterial pressure. This study was designed to directly test the hypothesis that the afferent renal nerves are necessary for the development and maintenance of renal hypertension. In the first experiment, dorsal spinal rhizotomies or sham rhizotomies were performed in rats between T9 and L1, through which afferent renal nerves have been shown to traverse. After the one-kidney, one-wrap procedure, the increase in systolic arterial pressure and water intake was similar in the two groups of rats. To determine whether the removal of afferent renal nerves reversed the hypertensive process, animals with established renal hypertension were subjected to dorsal rhizotomy or the sham-rhizotomy procedure. Again, there was no significant effect on systolic arterial pressure and water intake. Although combined dorsal and ventral rhizotomy and subdiaphragmatic vagotomy did not affect the onset of hypertension, spinal transection at the level of C8 effectively prevented the rise in arterial pressure. Although efferent neural mechanisms contribute to the hypertensive process, these studies suggest that afferent renal nerves are not directly involved in the development and maintenance of one-kidney, one-wrap renal hypertension.

Contribution of sodium to the mechanism of one-kidney, renal-wrap hypertension.

The early and chronic stages of one-kidney, figure-8 renal-wrap hypertension were studied during low or normal dietary sodium intake in rats. The renal-wrap procedure caused a significant elevation in arterial pressure at 3 and 28 days postwrap with normal sodium diet. Sodium-depleted rats did not experience an increased arterial pressure following renal wrapping. Blockade of angiotensin II receptors with [Sar1-Ala8]angiotensin II caused a greater decrease in arterial pressure in the sodium-depleted, renal-wrapped animals compared with sham-operated rats. In sodium-replete rats, angiotensin receptor blockade did not lower arterial pressure. Total ganglionic blockade decreased arterial pressure significantly more in the wrapped animals than in the sodium-replete sham-operated rats and the wrapped or sham-wrapped, sodium-deplete animals. Thus the early and chronic phases of normal sodium, one-kidney, figure-8 renal hypertension are supported by an increased sympathetic nervous system function. These observations suggest that sodium is necessary for the development of one-kidney, figure-8 renal-wrap hypertension and that the presence of sodium in the diet permits a functional activation of the sympathetic nervous system in response to the renal-wrap procedure.

Determination of ethylene oxide, ethylene chlorohydrin, and ethylene glycol in aqueous solutions and ethylene oxide residues in associated plastics.

A gas chromatographic (GC) method was developed for the determination of ethylene oxide and its two reaction products, ethylene chlorohydrin and ethylene glycol, in aqueous ophthalmic solutions. Propylene oxide was used as an internal standard. All three components were determined in one isothermal chromatographic analysis in less than 15 min. An extraction method for the determination of ethylene oxide residues in plastic components was also developed, and certain plastics with different ethylene oxide retention characteristics were identified.

Urinary screening for abnormalities of amino acid or mucopolysaccharide metabolism in patients in a hospital for the mentally handicapped in Wessex.

Urine from 348 patients of a hospital for the mentally handicapped in Wessex was screened for abnormalities of aminoacid and mucopolysaccharide excretion. Persistent aminoacid abnormalities were found in 16 patients: two had phenylketonuria and one a severe variant form of hyperphenylalaninaemia (dihydropteridine reductase deficiency). In at least nine of the other patients, the observed abnormality was probably a secondary phenomenon. Two patients excreted excessive amounts of a glycosaminoglycan identified tentatively as hyaluronic acid. The study also uncovered one undiagnosed diabetic and three patients with urinary tract infection.