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Biomedical subjects

A P Koretsky

Publications and source records attributed to A P Koretsky.

At least 19 recordsLinked to original sources
PubMed1999

Perfusion analysis using dynamic arterial spin labeling (DASL).

A variety of magnetic resonance (MR) techniques have proved useful to quantify perfusion using endogenous water as a blood flow tracer. Assuming that water is a freely diffusable tracer, the model used for these techniques predicts that the quantitation of perfusion is based on three parameters, all of which can depend on blood flow. These are the longitudinal tissue relaxation time, the transit time from point of labeling to tissue, and the difference in tissue MR signal between an appropriate control and the labeled state. To measure these three parameters in parallel, a dynamic arterial spin labeling (DASL) technique is introduced based on the analysis of the tissue response to a periodic time varying degree of arterial spin labeling, called here the labeling function (LF). The LF frequency can be modulated to overdetermine parameters necessary to define the system. MR schemes are proposed to measure the tissue response to different LF frequencies efficiently. Sprague-Dawley rats were studied by DASL, using various frequencies for the LF and various arterial pCO2 levels. During data processing, the periodic behavior of the tissue response to the LF allowed for frequency filtering of periodic changes in signal intensity unrelated to perfusion and arterial spin labeling. Measures of transit time, tissue longitudinal relaxation time, and perfusion agreed well over a range of LF frequencies and with previous results. DASL shows potential for more accurately quantifying perfusion as well as measuring transit times associated with arterial spin labeling techniques.

Animals↗
PubMed1999

Detection of single mammalian cells by high-resolution magnetic resonance imaging.

This study reports the detection of single mammalian cells, specifically T cells (T lymphocytes) labeled with dextran-coated superparamagnetic iron oxide particles, using magnetic resonance microscopy. Size amplification due to sequestration of the superparamagnetic particles in vacuoles enhances contrast in localized areas in high-resolution magnetic resonance imaging. Magnetic resonance images of samples containing differing concentrations of T cells embedded in 3% gelatin show a number of dark regions due to the superparamagnetic iron oxide particles, consistent with the number predicted by transmission electron microscopy. Colabeling of T cell samples with a fluorescent dye leads to strong correlations between magnetic resonance and fluorescence microscopic images, showing the presence of the superparamagnetic iron oxide particles at the cell site. This result lays the foundation for our approach to tracking the movement of a specific cell type in live animals and humans.

Animals↗
PubMed1999

Creatine and cyclocreatine treatment of human colon adenocarcinoma xenografts: 31P and 1H magnetic resonance spectroscopic studies.

Creatine (Cr) and cyclocreatine (cyCr) have been shown to inhibit the growth of a variety of human and murine tumours. The purpose of this study was to evaluate the anti-tumour effect of these molecules in relation to drug accumulation, energy metabolism, tumour water accumulation and toxicity. Nude mice carrying a human colon adenocarcinoma (LS174T) with a creatine kinase (CK) activity of 2.12 units mg(-1) protein were fed Cr (2.5% or 5%) or cyCr (0.025%, 0.1% or 0.5%) for 2 weeks and compared with controls fed standard diet. Cr concentrations of 2.5% and 5% significantly inhibited tumour growth, as did 0.1% and 0.5% cyCr. In vivo 31P magnetic resonance spectroscopy (MRS) after 2 weeks of treatment showed an increase in [phosphocreatine (PCr)+phosphocyclocreatine (PcyCr)]/nucleoside triphosphate (NTP) with increasing concentrations of dietary Cr and cyCr, without changes in absolute NTP contents. The antiproliferative effect of the substrates of CK was not related to energy deficiency but was associated with acidosis. Intratumoral substrate concentrations (measured by 1H-MRS) of 4.8 micromol g(-1) wet weight Cr (mice fed 2.5% Cr) and 6.2 micromol g(-1) cyCr (mice fed 0.1% cyCr) induced a similar decrease in growth rate, indicating that both substrates were equally potent in tumour growth inhibition. The best correlant of growth inhibition was the total Cr or (cyCr+Cr) concentrations in the tissue. In vivo, these agents did not induce excessive water accumulation and had no systemic effects on the mice (weight loss, hypoglycaemia) that may have caused growth inhibition.

Adenocarcinoma↗
PubMed1998

Cardiac MRI of the normal and hypertrophied mouse heart.

With the development of recent transgenic techniques, studies involving mice offer opportunities to increase understanding of cardiac disease. This provides motivation for the current study to perform noninvasive evaluation of the normal and hypertrophied mouse heart with MRI. By acquiring ECG and respiratory signals, the MR image acquisition was gated to both the cardiac and respiratory cycles. Combining a spin-warp imaging sequence with an RF surface coil resulted in short-axis images that allowed quantification of in vivo cardiac mass. Excellent agreement between MRI-determined (y) and postmortem heart weight (x) was obtained: y = 0.991x + 1.43 (r = 0.996). Isoproterenol, at 282 micromol/kg body weight (BW) and 573 micromol/kg BW, induced a dose-dependent increase in the ratio of heart weight to BW of 16.8 +/- 1.09% and 24.1 +/- 1.71%, respectively, which was accurately measured by MRI. These results demonstrate the ability of MRI to noninvasively monitor cardiac anatomy in the mouse.

Animals↗
PubMed1998

In vivo neuronal tract tracing using manganese-enhanced magnetic resonance imaging.

Development of efficient imaging techniques to trace neuronal connections would be very useful. Manganese ion (Mn2+) is an excellent T1 contrast agent for magnetic resonance imaging (MRI). Four reports utilizing radioactive Mn2+ in fish and rat brain indicate that Mn2+ may be useful for tracing neuronal connections. Therefore, the purpose of this work was to determine if Mn2+ can be used as an in vivo MRI neuronal tract tracer. The results indicate that topical administration of MnCI2 solution to the naris of mice as well as to the retinal ganglion cells via intravitreal injection leads to enhancement of contrast along the respective pathways. Therefore, application of Mn2+ to neurons allows the use of MRI to visualize neuronal connections.

Animals↗
PubMed1998

Comparison of the effects of ORG 30029, dobutamine and high perfusate calcium on function and metabolism in rat heart.

Cardiac contractility may be enhanced via multiple cellular mechanisms resulting in varied effects on cardiac energetics. The mechanisms that account for the varied energetic responses are not well understood. The purpose of this investigation was to compare the effects of the calcium sensitizing agent ORG 30029 (N-hydroxy-5,6-dimethoxy-benzo[b]thiophene-2-carboximidamide hydrochloride, a calcium sensitizing agent which increases contractility without increasing calcium transients significantly), dobutamine and high perfusate calcium on contractility and energetics. Langendorff-perfused rat hearts were stimulated with ORG 30029, dobutamine and high perfusate calcium in graduated concentrations while myocardial oxygen consumption (MVO2) and force-time integral were measured. ORG 30029, dobutamine and high perfusate calcium increased contractility in a dose-dependent manner. Despite an increase of 50% in systolic pressure and a 17% increase in force-time integral from control, ORG 30029 had no significant effect on MVO2 at the lower concentrations (n = 6). However, dobutamine (n = 4) and high perfusate calcium (n = 4) caused a 65% increase in systolic pressure and a 17% increase in force-time integral and a 50% and 41% increase in MVO2 respectively (P < 0.05). High energy phosphates (by 31P NMR), and lactate production were unaltered by these agents, suggesting that metabolism was steady state. Basal metabolism tended to increase slightly with dobutamine but not with ORG 30029 or high perfusate calcium. ORG 30029, dobutamine, and high perfusate calcium increase contractility in perfused rat hearts with disparate effects on energetics. These differences may be accounted for, in part, by differences in energy expenditure for calcium handling.

Animals↗
PubMed1998

Radial echo-planar imaging.

A new ultrafast magnetic resonance imaging pulse sequence named radial echo-planar imaging (rEPI) is introduced. The sequence is based on a modification of the echo-planar imaging (EPI) sequence to scan k-space radially, in an attempt to combine the speed of EPI with the benefits of radial sampling. Like in EPI, all the desired lines in k-space are scanned consecutively in opposite directions. The unique feature of this new sequence, however, is that the orientation of the readout gradient is incrementally rotated, so that all the echoes are refocused through the center of k-space. Therefore, rEPI data are acquired in a polar grid, and image reconstruction can be done either by means of filtered back-projection or by regridding the data to a Cartesian matrix followed by 2D Fourier transform. First results show that rEPI images can be acquired with the same speed and signal-to-noise ratio of EPI images. rEPI images are also shown to be less sensitive to off-resonance effects than EPI images. Further studies are underway to investigate the usefulness of rEPI for spectroscopic imaging and applications affected by motion.

Echo-Planar Imaging↗
PubMed1998

Simultaneous glutamate and perfusion fMRI responses to regional brain stimulation.

Functional magnetic resonance imaging (fMRI) rests on the assumption that regional brain activity is closely coupled to regional cerebral blood flow (rCBF) in vivo. To test the degree of coupling, cortical brain activity was locally stimulated in rats by reversed microdialysis infusion of picrotoxinin, alphagamma-aminobutyric acid-A antagonist. Before and during the first 30 minutes of infusion, simultaneous fMRI (rCBF) and neurochemical (interstitial glutamate concentration) measures of brain activity were highly correlated (r = 0.83). After 30 minutes of picrotoxinin-induced stimulation, glutamate levels decreased but rCBF remained elevated, suggesting that additional factors modulate the relationship between neuronal neurotransmitters and hemodynamics at these later stages.

Animals↗
PubMed1998

Fluorescence measurement of calcium transients in perfused rabbit heart using rhod 2.

Surface fluorescence spectroscopy of the beating heart to measure cytosolic calcium has been limited by the need to use ultraviolet excitation light for many of the commonly used calcium indicators. Ultraviolet light in the heart produces a high level of background fluorescence and is highly absorbed, limiting tissue penetration. Visible wave-length fluorescence dyes such as rhod 2 are available; however, the lack of spectral shift with calcium binding precludes the use of ratio techniques to account for changes in cytosolic dye concentration. We have developed a method for in vivo quantitation of cytosolic rhod 2 concentration that in conjunction with calcium-dependent fluorescence measurements permits estimation of cytosolic calcium levels in perfused rabbit hearts. Reflective absorbance of excitation light by rhod 2 loaded into myocardium was used as an index of dye concentration and the ratio of fluorescence intensity to absorbance as a measure of cytosolic calcium concentration. Endothelial cell loading of rhod 2 was found to be minimal (< 5%), and dye leak rate out of the cytosol was slow, with approximately 5% loss of dye fluorescence occurring between 10 and 30 min after dye loading. Rhod 2 loading into subcellular compartments, determined by manganese quenching, was also minimal (< 5%). The dissociation constant of rhod 2 for calcium was measured in vitro to be 500 nM, and this value increased to 710 nM in the presence of 0.5 mM myoglobin. On the basis of this value and in vivo fluorescence measurements, cytosolic calcium concentration in the rabbit heart was found to be 229 +/- 90 nM at end diastole and 930 +/- 130 nM at peak systole, with peak fluorescence preceding peak ventricular pressure by approximately 40 ms. This technique should facilitate detailed analysis of calcium transients from the whole heart.

Animals↗
PubMed1998

Absence of myofibrillar creatine kinase and diaphragm isometric function during repetitive activation.

Creatine kinase (CK) provides ATP buffering in skeletal muscle and is expressed as 1) cytosolic myofibrillar CK (M-CK) and 2) sarcomeric mitochondrial CK (ScCKmit) isoforms that differ in their subcellular localization. We compared the isometric contractile and fatigue properties of 1) control CK-sufficient (Ctl), 2) M-CK-deficient (M-CK[-/-]), and 3) combined M-CK/ScCKmit-deficient null mutant (CK[-/-]) diaphragm (Dia) to determine the effect of the absence of M-CK activity on Dia performance in vitro. Baseline contractile properties were comparable across groups except for specific force, which was approximately 16% lower in CK[-/-] Dia compared with M-CK[-/-] and Ctl Dia. During repetitive activation (40 Hz, (1)/(3) duty cycle), force declined in all three groups. This decline was significantly greater in CK[-/-] Dia compared with Ctl and M-CK[-/-] Dia. The pattern of force decline did not differ between M-CK[-/-] and Ctl Dia. We conclude that Dia isometric muscle function is not absolutely dependent on the presence of M-CK, whereas the complete absence of CK acutely impairs isometric force generation during repetitive activation.

Adenylate Kinase↗
PubMed1997

The role of creatine kinase in inhibition of mitochondrial permeability transition.

Cyclosporin A sensitive swelling of mitochondria isolated from control mouse livers and from the livers of transgenic mice expressing human ubiquitous mitochondrial creatine kinase occurred in the presence of both 40 microM calcium and 5 microM atractyloside which was accompanied by a 2.5-fold increase over state 4 respiration rates. Creatine and cyclocreatine inhibited the latter only in transgenic liver mitochondria. Protein complexes isolated from detergent solubilised rat brain extracts, containing octameric mitochondrial creatine kinase, porin and the adenine nucleotide translocator, were reconstituted into malate loaded lipid vesicles. Dimerisation of creatine kinase in the complexes and exposure of the reconstituted complexes to >200 microM calcium induced a cyclosporin A sensitive malate release. No malate release occurred with complexes containing octameric creatine kinase under the same conditions.

Animals↗
PubMed1997

Functional equivalence of creatine kinase isoforms in mouse skeletal muscle.

Creatine kinase (CK) is a highly conserved enzyme abundant in skeletal muscle that has a key role in high energy phosphate metabolism. The localization of the muscle isoenzyme of CK (MM-CK) to the M line and the sarcoplasmic reticulum of myofibrils has been suggested to be important for proper force development in skeletal muscle. The importance of this subcellular compartmentation has not been directly tested in vivo. To test the role of myofibrilar localization of CK, the consequences of a complete CK isoform switch from MM-CK to the brain (BB-CK) isoform, which does not localize to the M line, was studied in transgenic mouse skeletal muscle. In MM-CK knockout mice there are large contractile defects. When MM-CK was replaced by BB-CK, the aberrant contractile phenotypes seen in MM-CK knockout mice were returned to normal despite the lack of myofibrillar localization. These results indicate that CK compartmentation to the myofibril of skeletal muscle is not essential for contractile function and that there is functional equivalence of creatine kinase isoforms in supporting cellular energy metabolism.

Animals↗
PubMed1997

Estimation of water extraction fractions in rat brain using magnetic resonance measurement of perfusion with arterial spin labeling.

The model used for calculating perfusion by MRI techniques that use endogenous water as a tracer assumes that arterial water is a freely diffusible tracer. Evidence shows that this assumption is not valid in the brain at high blood flow rates, at which movement of water into and out of the microvasculature becomes limited by diffusion across the blood-brain barrier. In this work, the arterial spin-labeling technique is used to show that fraction of arterial water that is dependent on blood flow rate remains in the vasculature and does not exchange with brain tissue water. By using perfusion measurements without and with magnetization transfer (MT) effects, one can distinguish arterial label that exchanges into tissue because blood has much smaller MT than brain tissue. Using this technique, the extraction fraction for water is measured in the rat brain at various cerebral blood flow rates. At high flow rates (approximately 5 ml/g/min), the extraction fraction for water is found to be about 45% in rat brain. Disruption of the blood-brain barrier with D-mannitol caused an increase in the extraction fraction for water. It was possible to form an image related to the extraction fraction for water. The ability to estimate the amount of vascular water exchanging with tissue water by MRI may represent a noninvasive approach to detect the integrity of the blood-brain barrier.

Animals↗
PubMed1997

Evidence for the exchange of arterial spin-labeled water with tissue water in rat brain from diffusion-sensitized measurements of perfusion.

The extraction fraction of vascular water in rat brain is investigated by means of diffusion measurements of arterial spin labeled water at varying cerebral blood flow (CBF) values. The apparent diffusion coefficient (ADC) of the difference of the proton magnetization signal in the brain acquired with and without continuous arterial spin labeling is modeled to provide a measure of the amount of arterial water in tissue and vasculature and thus of the extraction fraction. The tissue and vascular portion of the arterial spin labeled water are differentiated based on their diffusion characteristics in a manner analogous to the intravoxel incoherent motion (IVIM) method. The amount of labeled arterial water that exchanges with tissue water is determined by estimating the fraction of the total signal that is associated with the slow-decaying component of a biexponential fit to the normalized difference signal between the magnetization of brain tissue acquired with and without arterial spin labeling. The results indicate that, at normal CBF (1.15 +/- 0.21 ml x g(-1) x min[-1]), about 90% of the arterial spin labeled water diffuses with an ADC of (1.21 +/- 0.37) x 10[-3] mm2 s[-1]), which is equal to tissue. At high CBF, an increasing fraction of the labeling water has a fast-pseudo-diffusion coefficient due to a decrease in water extraction fractions. The results also show that the contribution of vascular water to the measurement of perfusion by techniques that use endogenous water as a tracer can be efficiently eliminated by the use of diffusion sensitizing gradients with small effective b values (b approximately 20 s/mm2), enabling these techniques to monitor true changes in tissue perfusion.

Animals↗
PubMed1997

Manganese ion enhances T1-weighted MRI during brain activation: an approach to direct imaging of brain function.

Present techniques for functional MRI rely on detecting changes in hemodynamics that result as a consequence of brain activation. It would be useful if MRI techniques could be developed that enable imaging of a parameter directly related to neuronal activity. Influx of calcium into neurons is necessary for release of neurotransmitters. Divalent manganese ions (Mn2+) can enter cells through voltage-gated calcium channels and Mn2+ is paramagnetic. Mn2+ accumulation in brain due to activation should alter relaxation times offering an approach to sensitize MRI to calcium influx in the brain. To test this idea, T1-weighted MRI was obtained from the rat brain in the presence of a continuous intravenous infusion of 3.6 mumol/min MnCl2. In the anesthetized rat brain, signal enhancement was detected in regions corresponding to ventricles. Activation of the brain with glutamate led to increase in MRI signal intensity in the brain to 238 +/- 23% of the original. This increase in signal was dependent on the presence of MnCl2 and was not due to changes in blood flow. It was necessary to break the blood brain barrier with mannitol to make Mn2+ accessible to the active sites for efficient detection. Enhancement of MRI signal in the brain was also detected with decreasing anesthesia and with somatosensory stimulation. Due to the slow clearance of Mn2+ from the stimulated region of the brain, MRI enhancement could also be detected after stimulation that occurred on awake, behaving rats outside the magnet. These data indicate that MnCl2 shows potential as a MRI contrast agent that is directly sensitive to brain activation.

Animals↗
PubMed1997

Near-simultaneous hemoglobin saturation and oxygen tension maps in mouse brain using an AOTF microscope.

A newly developed microscope using acousto-optic tunable filters (AOTFs) was used to generate in vivo hemoglobin saturation (SO2) and oxygen tension (PO2) maps in the cerebral cortex of mice. SO2 maps were generated from the spectral analysis of reflected absorbance images collected at different wavelengths, and PO2 maps were generated from the phosphorescence lifetimes of an injected palladium-porphyrin compound using a frequency-domain measurement. As the inspiratory O2 was stepped from hypoxia (10% O2), through normoxia (21% O2), to hyperoxia (60% O2), measured SO2 and PO2 levels rose accordingly and predictably throughout. A plot of SO2 versus PO2 in different arterial and venous regions of the pial vessels conformed to the sigmoidal shape of the oxygen-hemoglobin dissociation curve, providing further validation of the two mapping procedures. The study demonstrates the versatility of the AOTF microscope for in vivo physiologic investigation, allowing for the generation of nearly simultaneous SO2 and PO2 maps in the cerebral cortex, and the frequency-domain detection of phosphorescence lifetimes. This class of study opens up exciting new possibilities for investigating the dynamics of hemoglobin and O2 binding during functional activation of neuronal tissues.

Acoustics↗
PubMed1997

Expression of functional mitochondrial creatine kinase in liver of transgenic mice.

The mitochondrial isoform of creatine kinase (MiCK) is localized to the mitochondrial intermembrane space, and its precise role in vivo is still actively being investigated. Here, we report a transgenic mouse model in which MiCK is expressed in liver, a tissue that does not normally express significant levels of CK. Expression of the genomic clone for human, ubiquitous MiCK was controlled by the promoter/enhancer region of the transthyretin gene. Three of seven founder mice were chosen to establish lines and had MiCK activity values ranging from 13 to 269 micromol x min(-1) x g wet wt(-1). Differential centrifugation and histochemical staining demonstrated that >90% of the CK activity is localized to the mitochondrial intermembrane space. An unusual mitochondrial morphology characterized by an angular nature to the membranes was detected using electron microscopy in the transgenic line expressing the highest levels of MiCK. Increasing hepatic total creatine levels led to a return to normal mitochondrial morphology. 31P-nuclear magnetic resonance spectroscopy demonstrated that the expressed MiCK is capable of producing and utilizing phosphocreatine. These mice will be useful for investigating gain of function effects of MiCK in cellular energetics.

Animals↗