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PubMed1993

Microbial delignification with white rot fungi improves forage digestibility.

Three wild-type white rot fungi and two cellulase-less mutants developed from Phanerochaete chrysosporium K-3 (formerly Sporotrichum pulverulentum) were tested for their ability to delignify grass cell walls and improve biodegradation by rumen microorganisms. Fungal-treated and control stems of Bermuda grass were analyzed for their content of ester- and ether-linked aromatics by using alkali extraction and gas chromatography, for in vitro dry weight digestion and production of volatile fatty acids in in vitro fermentations with mixed ruminal microorganisms, for loss of lignin and other aromatics from specific cell wall types by using microspectrophotometry, and for structural changes before and after in vitro degradation by rumen microorganisms by using transmission electron microscopy. P. chrysosporium K-3 and Ceriporiopsis subvermispora FP 90031-sp produced the greatest losses in lignin and improved the biodegradation of Bermuda grass over that of untreated control substrate. However, C. subvermispora removed the most lignin and significantly improved biodegradation over all other treatments. Phellinus pini RAB-83-19 and cellulase-less mutants 3113 and 85118 developed from P. chrysosporium K-3 did not improve the biodegradation of Bermuda grass lignocellulose. Results indicated that C. subvermispora extensively removed ester-linked p-coumaric and ferulic acids and also removed the greatest amount of non-ester-linked aromatics from plant cell walls. Microscopic observations further indicated that C. subvermispora removed esters from parenchyma cell walls as well as esters and lignin from the more recalcitrant cell walls (i.e., sclerenchyma and vascular tissues). C. subvermispora improved in vitro digestion and volatile fatty acid production by ruminal microorganisms by about 80%, while dry matter loss due to fungi was about 20% greater than loss in untreated control stems. The chemical and structural studies used identified sites of specific fungal attack and suggested mechanisms whereby improvement occurred.

Journal Article↗
PubMed1976

Quantitative cytophotometry evaluated as a method for analyses of herpes simplex viral deoxyribonucleic acid synthesis.

Feulgen deoxyribonucleic acid (F-DNA) microspectrophotometry was evaluated as a potential tool for quantification of herpes simplex virus type 1 (HSV-1) and 2(HSV-2) DNA synthesis in single cells. Since HSV DNA synthesis has been extensively studied using incorporation of radioactive precursors into viral and cellular DNA, microspectrophotometric measures were correlated with biochemical data obtained using tritium-labeled thymidine ([3H]TdR). It was established that: (i) viral-induced increaae in F-DNA can be cytophotometrically detected between 1 and 6 h postinjection (p.i.), which corresponds to the initial incorporation of [3H) TdR into viral DNA; (ii) peak F-DNA levels occurred 8 h p.i., which supported cytological observations of a more rapid development of an inclusion body in HSV-2-infected nuclei. Despite the fact that F-DNA cytophotometry is unable to distinguish between cell and viral DNA, the overall study supports the existence of a good correlation between data obtained using microspectrophotometric and conventional isotope methods. Furthermore, cytophotometry complements biochemical evaluations in that it permits analyses of DNA changes on a single-cell basis or the detection of infection in very small numbers of cells.

Cell Nucleus↗
PubMed1992

Electrophoretic karyotypes of clinical isolates of Coccidioides immitis.

Chromosomes of the fungal respiratory pathogen, Coccidioides immitis, were separated by contour-clamped homogeneous electric field gel electrophoresis. Twelve isolates were examined, the majority of which showed four chromosomes with a range of molecular size from 11.5 to 3.2 Mb. Three isolates (C634, C735, and L) revealed three chromosomal bands under the conditions employed for electrophoretic separation. However, in two of these isolates (C634 and C735), four chromosomes were visible on membrane transfers of pulsed-field gels after Southern hybridization between the chromosomal DNA and selected DNA probes. The probes included a conserved ribosomal gene and three previously described cDNAs isolated from C. immitis expression libraries. The L isolate was determined to have the same genome size as a typical four-chromosome isolate on the basis of microspectrophotometric comparison of fluorescence intensity of the ethidium bromide-stained nuclear DNA. The genome size of C. immitis determined by microspectrophotometry was approximately 28.2 +/- 2.6 Mb. The calculated genome size based on addition of the average molecular weights of chromosomal bands separated by contour-clamped homogeneous electric field gel electrophoresis was approximately equal to the estimate derived from the spectrophotometric analyses. This is the first report of the electrophoretic karyotype of C. immitis.

Blotting, Southern↗
PubMed1974

Interactions of polyoma and mouse DNAs. I. Lytic infection of bromodeoxyuridine-prelabeled mouse embryo cells.

On CsCl isopycnic centrifugation of the DNA extracted from secondary mouse embryo (ME) cultures grown in the presence of 5-bromodeoxyuridine (BUdR) and 5-fluorodeoxyuridine (FUdR) for 40 h, 10 to 25% of the DNA was found to be unsubstituted, 70 to 80% was bromouracil-hybrid DNA, and 5 to 10% was heavy DNA. These results together with cell number determinations, autoradiography, and Feulgen microspectrophotometry revealed three types of cells in these cultures: (i) 60 to 80% of the cells replicated their DNA once, divided, and then stopped mitotic activity, (ii) 5 to 10% were going through a second round of DNA replication; whereas (iii) 10 to 30% did not replicate DNA during the BUdR-FUdR exposure. After the transfer of these cultures to normal medium (without BUdR-FUdR), up to 20% of the cells resumed DNA synthesis asynchronously within 60 h, but no increase in cell number was observed. BUdR-FUdR-treated cultures, which were infected with polyoma virus in the absence of the thymidine analogues, supported a lytic infection to the same extent as did untreated ME cultures. This was concluded from the similar number of cells, which were induced to synthesize DNA, from the similar replication rate of the viral DNA, from the similar number of cells containing polyoma capsid proteins, and from the similar yields of progeny virus determined by hemagglutination and plaque formation. Thus, BUdR-prelabeled ME cultures are suitable for the investigation of interactions of the polyoma and mouse genomes during the lytic infection.

Animals↗
PubMed1987

Quantitative DNA analysis of low grade cervical intraepithelial neoplasia and human papillomavirus infection by static and flow cytometry.

Quantitative deoxyribonucleic acid (DNA) analysis of cervical biopsy specimens from 26 women with cytological, colposcopic, and histological evidence of mild cervical atypia consistent with cervical intraepithelial neoplasia grade I, reactive atypia, or human papillomavirus infection alone or in combination was performed in a comparative evaluation of Feulgen microspectrophotometry, the fast interval processor image analysis system, and flow cytometry. The fast interval processor image analysis system showed a distinct advantage over the other methods, being faster and allowing the operator to see the cells that were selected for measurement. The three methods of measurement together showed that the DNA content of at least 2% of the cells measured exceeded 5C (C being the haploid amount of DNA in a normal cell and 2C representing the diploid complement of a normal cell) in all cases of cervical intraepithelial neoplasia grade I and reactive atypia and in 87% of those reported as showing human papillomavirus infection alone. In contrast, the DNA content of cervical biopsy specimens from the transformation zone of 11 normal controls did not exceed 4C. This study shows the value of using a DNA threshold--that is, the "5C exceeding rate"--to distinguish between normal and neoplastic appearances of the cervix. These results support the view that cervical infection by human papillomavirus is a true precursor of neoplasia.

Aneuploidy↗
PubMed1986

Studies of erythroblast function in congenital dyserythropoietic anaemia, type I: evidence of impaired DNA, RNA, and protein synthesis and unbalanced globin chain synthesis in ultrastructurally abnormal cells.

Two patients with congenital dyserythropoietic anaemia, type I (CDA) were studied. Their blood reticulocytes showed unbalanced globin chain synthesis with increased alpha:beta globin chain synthesis ratios. A high proportion of the erythroblasts displayed the characteristic "Swiss cheese" abnormality of the nuclear chromatin and some also showed cytoplasmic intrusions lined with nuclear membrane within the nucleus. Occasional erythroblast profiles contained intracytoplasmic inclusions that were ultrastructurally indistinguishable from precipitated alpha chains. The technique of combined Feulgen microspectrophotometry and 3H-thymidine autoradiography showed gross abnormalities of proliferation in the early polychromatic erythroblasts. The proliferative abnormalities included an arrest of DNA synthesis after the progress of cells through part of the S phase and the formation of several mononucleate and binucleate cells with hypertetraploid total DNA contents. The bone marrow cells gave a normal deoxyuridine suppressed value, indicating that there was no impairment of the methylation of deoxyuridylate. Electron microscope autoradiographic studies showed that a high proportion of the erythroblasts with the "Swiss cheese" nuclear abnormality suffered from a severe impairment, or arrest of DNA, RNA, and protein synthesis.

Adult↗
PubMed2002

Increases in myocardial cyclic GMP attenuate contractile delay in myocardial stunning.

We tested the hypothesis that the effects of myocardial stunning would be reduced by cyclic GMP in rabbit hearts. In three groups of anesthetized open-chest New Zealand white rabbits, myocardial stunning was produced by 15 min of occlusion of the left anterior descending coronary artery followed by 15 min of reperfusion repeated twice. Either control vehicle (saline plus 1% dimethyl sulfoxide) or 8-bromo-cyclic GMP (8-Br-cGMP (10(-4) and 10(-3) M)) was topically applied to the left ventricular surface. Hemodynamic (left ventricular and aortic pressures) and functional parameters (wall thickening, delay in onset of wall thickening, and rate of wall thickening) were determined. Coronary blood flow (microspheres) and O2 extraction (microspectrophotometry) were used to determine myocardial O2 consumption (VO2). Myocardial stunning was observed in the control group through an increased delay in onset of myocardial wall thickening (29 +/- 7 versus 55 +/- 16 ms) and decreased maximal rate of wall thickening (20 +/- 8 versus 11 +/- 3 mm x s(-1)). After treatment with 8-Br-cGMP 10(-4) and 10(-3) M, stunning did not increase the delay (37 +/- 5 versus 39 +/- 7 and 39 +/- 7 versus 28 +/- 8 ms). Myocardial stunning did not significantly alter VO2. 8-Br-cGMP 10(-3) M significantly decreased subepicardial VO2 (6.2 +/- 0.8 versus 3.7 +/- 0.6 mL O2 x min(-1) 100 g(-1)) and insignificantly decreased subendocardial VO2 (8.6 +/- 0.9 versus 6.3 +/- 1.2 mL O2 x min(-1) x 100 g(-1)) when compared with the vehicle-treated rabbits. We conclude that increasing cyclic GMP reduced the effects of myocardial stunning in the rabbit heart by ameliorating the delay in onset of wall thickening and decreasing the local O2 costs in the stunned region.

Animals↗
PubMed1982

Fluorescence and differential light absorption recordings with calcium probes and potential-sensitive dyes in skinned cardiac cells.

This report describes an optical system for microspectrophotometry in a single cardiac cell from which the sarcolemma has been removed by microdissection (skinned cardiac cell). This system is attached to the high power inverted microscope used for the microdissection and includes (a) a single variable wavelength microspectrophotometer used to define the spectrum of a given dye or Ca2+ probe; and (b) a dual wavelength, differential microspectrophotometer used to record differentially between the optimum wavelength and a wavelength separated by 25--30 nm. Results are presented using the following optical methods: (a) fluorescence measurements with chlorotetracycline to monitor the amount of Ca2+ bound to the inner face of the sarcoplasmic reticulum (SR) membrane; (b) differential absorption measurements with arsenazo III to measure changes of myoplasmic [Ca2+]free resulting from Ca2+ release from the SR; (c)fluorescence and (or) differential absorption measurements with the potential-sensitive dyes merocyanine 540, NK 2367, and di-S-C3(5) to monitor changes of charge distribution on the SR membrane during Ca2+ accumulation in the SR, as well as before and during Ca2+-induced release of Ca2+ from the SR. A small and rapid signal is observed which precedes the Ca2+-induced release of Ca2+ from the SR. It is detected as an increase of CA2+ binding inside the SR with chlorotetracycline and as a "hyperpolarization" with potential-sensitive dyes, while no transient change of myoplasmic [Ca2+]free is detected with arsenazo III. This small and rapid signal preceding the Ca2+ release may be a first hint to an understanding of the mechanism whereby a small increase of [Ca2+]free outside the SR triggers Ca2+ release from the SR.

Animals↗
PubMed1995

Myocardial metabolic and functional responses to acetylcholine are altered in thyroxine-induced cardiac hypertrophy.

We tested the hypothesis that acetylcholine would reduce myocardial O2 consumption and function, and that thyroxine (T4, 0.5 mg/kg for 16 days) induced cardiac hypertrophy would change this relationship. Anesthetized open-chest New Zealand white rabbits were divided into four groups: control-vehicle (CV, n = 8), control-acetylcholine (CA, n = 10), T4-vehicle (T4V, n = 9), and T4-acetylcholine (T4A, n = 10). Either vehicle or acetylcholine (10(-3) M) was topically applied to the left ventricular surface. Coronary blood flow (radioactive microspheres) and O2 extraction (microspectrophotometry) were used to determine O2 consumption, and muscarinic receptor density and affinity were also determined. T4 increased the heart weight/body weight ratio from 2.6 +/- 0.1 to 3.4 +/- 0.1. T4-treated animals had higher heart rates, blood pressures, and left ventricular dP/dtmax than control rabbits. Topical acetylcholine depressed hemodynamic parameters with a greater decrement in pressures and cardiac output in the T4A group (CA, -25%, T4A, -40%). Myocardial O2 consumption and coronary blood flow were higher in the T4-treated hearts. Myocardial O2 consumption significantly declined in both groups during acetylcholine, but the reduction was greater in the T4-treated hearts (CV 7.9 +/- 0.4 to CA 5.8 +/- 0.6 and T4V 18.8 +/- 3.0 to T4A 7.3 +/- 1.0 mL O2.min-1.100 g-1). Muscarinic receptor density (Bmax) was elevated by 41% in the T4-treated hearts, but affinity (Kd) was not altered. Thus, the T4-treated hearts responded to acetylcholine to a greater extent than control hearts in terms of functional and O2 consumption decrements.(ABSTRACT TRUNCATED AT 250 WORDS)

Acetylcholine↗
PubMed1984

Albumin influences sulfobromophthalein transport by hepatocytes of each acinar zone.

To determine whether profiles of decreasing concentration were generated among hepatocytes of the liver acinus during the transport of sulfobromophthalein sodium (BSP), rat livers were perfused with various concentrations of this dye (10 microM to 1 mM) in the presence and absence of albumin. After steady-state conditions for the biliary secretion of BSP had been attained, pieces of liver were rapidly frozen. Following the alkalinization of cryostat-cut sections, the relative concentration of BSP in hepatocytes of each zone and the effect of albumin on this localization were quantitated by microspectrophotometry. The results showed that BSP, perfused in the absence of albumin, was efficiently extracted by the liver (95% on a single pass), generating distinct profiles of decreasing cellular concentration from zone 1 to zone 3 at every concentration of BSP. However, the addition of albumin to the perfusate greatly reduced the extraction of BSP from the sinusoidal compartment and resulted in the abolition of the differences in BSP content between hepatocytes of zone 1 and zone 3. These results represent a direct demonstration that, as predicted by mathematical modeling, binding of BSP to albumin indeed results in a more homogeneous distribution of BSP within the liver acinus. A simple and direct microspectrophotometric method is therefore available to follow the changes in the relative concentration of BSP among the hepatocytes of the various acinar zones.

Animals↗
PubMed1985

Effect of pacing on oxygen supply-to-consumption ratio in ischemic myocardium.

This study was performed to determine if the partially occluded myocardium could maintain its O2 supply-to-consumption ratio during atrial pacing. In 14 open-chest anesthetized dogs, the left anterior descending coronary artery was occluded to 50% of base-line flow, and in half of the dogs the hearts were paced to 50% above base-line heart rate. Blood flows were determined by use of microspheres before and after occlusion, and O2 supply-consumption variables were determined by use of microspectrophotometry. After occlusion in control and paced groups, blood flow was depressed in the ischemic region compared with the nonischemic region. In paced animals, blood flows in all regions were significantly higher compared with their respective values in nonpaced animals. After pacing, mean O2 consumption was 34% higher (53% in the occluded region) compared with control animals. The O2 supply-to-consumption ratio was similar in the occluded region of paced animals compared with the same region in controls. This ratio was lower in ischemic regions of both groups compared with their respective nonischemic regions. This indicates that, despite a reduced O2 supply-to-consumption ratio in the ischemic area, an unutilized reserve of flow exists.

Animals↗
PubMed1987

Regional differences in pleural lymphatic albumin concentration in sheep.

We used quantitative reflectance autoradiography to compare the concentration of albumin in visceral pleural lymphatics at the cranial and caudal ends of the sheep's lung in the vertical (60 degrees head-up) and horizontal (supine) positions. Twelve to fourteen hours after injecting 125I-albumin intravenously we placed four anesthetized sheep in the vertical position to establish a microvascular hydrostatic pressure gradient along the vertical height of the lung. We placed two anesthetized sheep in the horizontal position. Four hours later, we fixed the left lung and removed visceral pleural tissue blocks from the cranial and caudal ends, separated by a 15-cm distance, along the costovertebral margin. We measured the silver grain density in the pleural lymphatic autoradiograms by dark-field reflectance microspectrophotometry. In the vertical position, the lymph albumin concentration at the cranial end (top) of the lung averaged 2.5 +/- 0.4 g/dl compared with the caudal end (bottom), which averaged 1.8 +/- 0.3 g/dl. The difference (42% greater at the top than the bottom) is significant (P less than 0.05). The computed gradient in perimicrovascular interstitial albumin osmotic pressure was 0.26 +/- 0.13 cmH2O/cm lung height. There were no differences between the cranial and caudal lymphatic groups in the two horizontal sheep. We conclude that in the sheep lung there is a gradient in perimicrovascular albumin concentration due to the vertical gradient in microvascular hydrostatic pressure.

Albumins↗
PubMed1990

Isoproterenol and myocardial O2 supply/consumption in hypertension-induced myocardial hypertrophy.

Myocardial oxygen supply, O2 consumption (MVO2) and function were determined during base-line and isoproterenol infusion (0.5 microgram.kg-1.min-1) 30 days after New Zealand White rabbits were prepared as one-kidney, one-clip (1K,1C) Goldblatt hypertensive or uninephrectomized (sham) controls. Coronary blood flow and cardiac output, using radioactive microspheres, and small vessel O2 saturations, using microspectrophotometry, were measured in hypertrophied and nonhypertrophied hearts. After 30 days, 33% myocardial hypertrophy was evident in the 1K,1C animals. Base-line blood pressure was significantly higher in the 1K,1C animals compared with sham controls and remained higher during isoproterenol infusion. Base-line heart rate was not different between animal groups, and heart rate increased in both groups during isoproterenol. Cardiac output was similar between sham and 1K,1C animals during base-line conditions (316 +/- 40 vs. 365 +/- 51 ml/min, respectively). After isoproterenol, cardiac output increased in the sham animals (494 +/- 119 ml/min) but not in the 1K,1C group (317 +/- 85 ml/min). Isoproterenol elevated MVO2 threefold in both the sham and 1K,1C animals. Coronary blood flow was significantly increased to similar levels in both animal groups. O2 extraction significantly increased during isoproterenol in both groups. Therefore, impaired cardiac function was evident during isoproterenol stress in the hypertrophied myocardium independent of myocardial O2 supply or consumption restrictions.

Animals↗
PubMed1990

Pacing during reperfusion elevates regional myocardial oxygen consumption.

Reperfusion after 2 h of coronary artery occlusion has been shown to result in depressed coronary blood flow to the reperfused region and elevated regional myocardial extraction. This suggests that reperfused myocardium, even after 4 h of reperfusion, possesses limited flow and O2 consumption reserves. We studied the capacity of reperfused myocardium to elevate regional blood flow and regional O2 consumption in response to sustained increased O2 demand, produced by atrial pacing. Two groups of anesthetized open-chest dogs were subjected to 2 h of left anterior descending coronary artery occlusion followed by 4 h of reperfusion. One group was subjected to atrial pacing (40% increase in heart rate) during the entire 4-h reperfusion period. Regional O2 saturation was measured by microspectrophotometry in samples of reperfused and nonoccluded subepicardium and subendocardium, which were taken at the end of the reperfusion period. In the paced group, regional blood flow (radiolabeled microspheres) to reperfused myocardium was significantly higher than to corresponding regions of unpaced hearts (110 +/- 22 vs. 40 +/- 9 ml.min-1 x 100 g-1 in the subendocardium). In the control group, O2 extraction of reperfused subendocardium was significantly higher than that measured in the corresponding nonoccluded region (11.0 +/- 0.9 vs. 8.0 +/- 0.6 ml O2/100 ml). Pacing did not elevate O2 extraction of reperfused myocardium (8.7 +/- 0.6 vs. 8.3 +/- 0.7 ml O2/100 ml). Myocardial O2 consumption was significantly elevated in all regions of the paced heart. It is concluded that reperfused myocardium possesses significant unutilized O2 supply and consumption reserves.

Animals↗
PubMed1994

Effect of dopamine on local segment work and O2 consumption in collateral-dependent myocardium.

To test the hypothesis that dopamine-stimulated work and myocardial O2 consumption (MVO2) in collateral-dependent myocardium would be reduced, Ameroid constrictors were implanted around the circumflex coronary artery (CFX) in nine dogs. Four weeks later, in an anesthetized open-chest preparation, segment length (ultrasonic dimension gauge) and force (miniature force transducer) were measured in myocardium supplied by the CFX and left anterior descending (LAD) coronary arteries. Work in each region was calculated as the systolic area under the force-length curve. Corresponding regional MVO2 was calculated from local O2 extraction (microspectrophotometry) and flow (radiolabeled microspheres). Dopamine infusion (15 micrograms.kg-1.min-1) increased regional work from 262 +/- 56 to 733 +/- 171 mm.g.min-1 in the control (LAD) region, but to a much smaller extent in the collateral-dependent (CFX) region (from 249 +/- 82 to 414 +/- 81 mm.g.min-1). However, regional MVO2 increased to about the same extent in the CFX (from 6.0 +/- 0.7 to 12.4 +/- 0.9 ml O2.min-1 times 100 g-1) and the LAD region (from 7.0 +/- 0.6 to 14.5 +/- 1.3 ml O2.min-1 times 100 g-1). O2 extraction was not elevated in the CFX region. Therefore, the functional impairment was not secondary to O2 supply or consumption limitations.

Animals↗
PubMed1995

Relationship between cGMP and myocardial O2 consumption is altered in T4-induced cardiac hypertrophy.

We tested the hypothesis that increases in guanosine 3',5'-cyclic monophosphate (cGMP) would reduce myocardial O2 consumption and that thyroxine (T4)-induced (0.5 mg/kg for 16 days) cardiac hypertrophy would change this relationship. Anesthetized open-chest New Zealand White rabbits were divided into four groups: control vehicle (CV, n = 7), control nitroprusside (CN, n = 6), T4 vehicle (T4V, n = 8), and T4 nitroprusside (T4N, n = 8). Vehicle or sodium nitroprusside (10(-4) M) was topically applied to the left ventricular subepicardium for 15 min. Coronary blood flow (radioactive microspheres) and O2 extraction (microspectrophotometry) were used to determine O2 consumption. Guanylate cyclase activity and cGMP were determined by radioimmunoassay. T4 increased the heart weight-to-body weight ratio from 2.7 +/- 0.1 to 3.4 +/- 0.2. Topical application of nitroprusside had no significant hemodynamic effects. Nitroprusside significantly increased myocardial cGMP in control hearts (CV = 4.1 +/- 0.3 to CN = 12.4 +/- 5.0 pmol/g) and T4 hearts (T4V = 3.9 +/- 0.3 to T4N = 5.2 +/- 0.4). The increase in the level of myocardial cGMP was significantly greater in CN (+202%) than in T4N (+33%). There were no significant differences in basal or total guanylate cyclase activity between control and T4 rabbits. Myocardial O2 consumption significantly declined in both groups during nitroprusside (10.8 +/- 1.4 for CV to 7.3 +/- 1.0 for CN (-32%) and 13.6 +/- 1.2 for T4V to 9.9 +/- 1.4 ml O2.min-1.100 g-1 for T4N (-27%).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1995

Reduced effect of phenylephrine on regional myocardial function and O2 consumption in experimental LVH.

In a dog model of left ventricular hypertrophy (LVH) created by aortic valve plication, we examined the hypothesis that regional myocardial inotropic and metabolic responses to alpha-adrenergic stimulation would be diminished due to decreased alpha-adrenoceptor number. After systemic beta-adrenergic blockade, phenylephrine (PE, 5 micrograms.kg-1.min-1) was infused into the left anterior descending artery in eight LVH and nine control open-chest anesthetized dogs. The circumflex region served as control. In both regions, local segment work was calculated as the integrated products of force (miniature transducer) and segment shortening (ultrasonic crystals). Local myocardial O2 consumption was calculated from regional blood flow (microspheres) and O2 saturation (microspectrophotometry). A saturation radioligand binding assay was used to determine adrenoceptor number and affinity. In control animals in the treated region, PE increased work from 815 +/- 140 to 1,493 +/- 149 g.mm.min-1. In LVH, work was not significantly altered (688 +/- 142 vs. 730 +/- 149 g.mm.min-1). Regional blood flow was elevated in controls (81 +/- 10 to 141 +/- 24 ml.min-1.100 g-1) but was not changed in LVH (105 +/- 12 vs. 123 +/- 18 ml.min-1.100 g-1). In controls, but not in LVH, myocardial O2 consumption was almost doubled during PE infusion (6.2 +/- 0.9 vs. 12.0 +/- 2.1 ml O2.min-1.100 g-1). alpha-Adrenoceptor number and dissociation constants values were not different between control and LVH (15.7 +/- 2.8 vs. 16.4 +/- 2.7 fmol/mg protein; 13.2 +/- 3.4 vs. 16.9 +/- 4.3 nm, respectively).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1995

Visualization of oxygen level inside a single cardiac myocyte.

The present paper describes theoretical and experimental bases for quantitation of the oxygen level inside a single isolated ventricular rat myocyte. We applied three-wavelength microspectrophotometry to a single cardiomyocyte, where the sensitivity of the spectrophotometry was augmented by a digital image processing technique known as video-enhanced microscopy. Oxygen-dependent changes in light absorption of the intracellular pigments were detected and represented with the use of a newly defined variable, Z, where one pixel of the reconstructed cell image corresponded to 0.2 microns. Theoretically, Z is a single-valued function of the fractional oxygen saturation of the intracellular pigment. We demonstrated in quiescent cardiomyocytes that Z changed according to extracellular PO2, where the relationship was sigmoidal with the extracellular PO2 for a half-maximal Z of 3.2 Torr. In the aerobic cell, conversion of cytochromes to the reduced forms by NaCN did not affect Z value. Therefore, we conclude that Z mainly represents the fractional oxygen saturation of myoglobin, thus reporting cytosolic PO2. We also demonstrated that for extracellular PO2 of 1.3 Torr, abolition of mitochondrial oxygen consumption by NaCN significantly elevated the intracellular oxygen level, suggesting existence of oxygen pressure gradients in a quiescent single cardiomyocyte that were proportional to oxygen flux.

Animals↗