Biomedical subjects

Kieran Clarke

Publications and source records attributed to Kieran Clarke.

43 records · Page 3Linked to original sources

PubMed2003

The mechanism underlying the positive inotropic effect of angiotensin II in the isolated perfused rabbit heart: a 31P NMR study.

Activation of the Na(+)/H(+) exchanger may play an important role in the development of cardiac hypertrophy. Isolated ventricular myocyte studies have suggested that angiotensin II (AII) has direct positive inotropic effect caused by intracellular alkalinization due to increased Na(+)/H(+) exchange, but whether this occurs in the whole heart is unknown. Consequently, we have used non-invasive 31P NMR spectroscopy to determine whether AII stimulation alters energetics or intracellular pH (pH(i)) in the intact beating rabbit heart. Heart rate (HR) and developed pressure (DP) were recorded continuously in isolated perfused rabbit hearts, simultaneously with pH(i) and high energy phosphate metabolite levels measured using 31P NMR spectroscopy. AII (11 nM) increased developed pressure by 14+/-2 mmHg (P<0.05) and increased pH(i) by 0.08+/-0.03 pH units (P<0.05, n=6). There were no significant changes in myocardial phosphocreatine (PCr), ATP or Pi concentrations throughout the protocol. Inhibition of Na(+)/H(+) exchange with 1 microM Hoe642 (n=7) abolished the increase in pH(i), but did not prevent the increase in developed pressure, caused by AII. Inhibition of protein kinase C (PKC) using 25 microM chelerythrine chloride prevented the positive inotropic and alkalinizing effects of AII (n=5). We conclude that the positive inotropic effect of AII is associated with, but not caused by, a decreased proton concentration due to stimulation of Na(+)/H(+) exchange in the whole rabbit heart.

Angiotensin II↗

PubMed2003

High-resolution imaging of normal anatomy, and neural and adrenal malformations in mouse embryos using magnetic resonance microscopy.

An efficient investigation of the effects of genetic or environmental manipulation on mouse development relies on the rapid and accurate screening of a substantial number of embryos for congenital malformations. Here we demonstrate that it is possible to examine normal organ development and identify malformations in mouse embryos by magnetic resonance microscopy in a substantially shorter time than by conventional histology. We imaged embryos in overnight runs of under 9 h, with an operator time of less than 1 h. In normal embryos we visualized the brain, spinal cord, ganglia, eyes, inner ear, pituitary, thyroid, thymus, trachea, bronchi, lungs, heart, kidneys, gonads, adrenals, oesophagus, stomach, intestines, spleen, liver and pancreas. Examination of the brain in embryos lacking the transcriptional coactivator Cited2 showed cerebellar and midbrain roof agenesis, in addition to exencephaly. In these embryos we were also able to detect agenesis of the adrenal gland. We confirmed all malformations by histological sectioning. Thus magnetic resonance microscopy can be used to rapidly identify developmental and organ malformations in mutant mouse embryos generated by transgenic techniques, in high-throughput mutagenesis screens, or in screens to identify teratogenic compounds and environmental factors contributing to developmental malformations.

Adrenal Glands↗

PubMed2003

Creatine transporter activity and content in the rat heart supplemented by and depleted of creatine.

The intracellular creatine concentration is an important bioenergetic parameter in cardiac muscle. Although creatine uptake is known to be via a NaCl-dependent creatine transporter (CrT), its localization and regulation are poorly understood. We investigated CrT kinetics in isolated perfused hearts and, by using cardiomyocytes, measured CrT content at the plasma membrane or in total lysates. Rats were fed control diet or diet supplemented with creatine or the creatine analog beta-guanidinopropionic acid (beta-GPA). Creatine transport in control hearts followed saturation kinetics with a K(m) of 70 +/- 13 mM and a V(max) of 3.7 +/- 0.07 nmol x min(-1) x g wet wt(-1). Creatine supplementation significantly decreased the V(max) of the CrT (2.7 +/- 0.17 nmol x min(-1) x g wet wt(-1)). This was matched by an approximately 35% decrease in the plasma membrane CrT; the total CrT pool was unchanged. Rats fed beta-GPA exhibited a >80% decrease in tissue creatine and increase in beta-GPA(total). The V(max) of the CrT was increased (6.0 +/- 0.25 nmol x min(-1) x g wet wt(-1)) and the K(m) decreased (39.8 +/- 3.0 mM). The plasma membrane CrT increased about fivefold, whereas the total CrT pool remained unchanged. We conclude that, in heart, creatine transport is determined by the content of a plasma membrane isoform of the CrT but not by the total cellular CrT pool.

Adenosine Triphosphate↗

PubMed2002

Simultaneous determination of low free Mg2+ and pH in human sickle cells using 31P NMR spectroscopy.

The concentrations of free magnesium, [Mg(2+)](free), [H(+)], and [ATP] are important in the dehydration of red blood cells from patients with sickle cell anemia, but they are not easily measured. Consequently, we have developed a rapid, noninvasive NMR spectroscopic method using the phosphorus chemical shifts of ATP and 2,3-diphosphoglycerate (DPG) to determine [Mg(2+)](free) and pH(i) simultaneously in fully oxygenated whole blood. The method employs theoretical equations expressing the observed chemical shift as a function of pH, K(+), and [Mg(2+)](free), over a pH range of 5.75-8.5 and [Mg(2+)](free) range 0-5 mm. The equations were adjusted to allow for the binding of hemoglobin to ATP and DPG, which required knowledge of the intracellular concentrations of ATP, DPG, K(+), and hemoglobin. Normal oxygenated whole blood (n = 33) had a pH(i) of 7.20 +/- 0.02, a [Mg(2+)](free) of 0.41 +/- 0.03 mm, and [DPG] of 7.69 +/- 0.47 mm. Under the same conditions, whole sickle blood (n = 9) had normal [ATP] but significantly lower pH(i) (7.10 +/- 0.03) and [Mg(2+)](free) (0.32 +/- 0.05 mm) than normal red cells, whereas [DPG] (10.8 +/- 1.2 mm) was significantly higher. Because total magnesium was normal in sickle cells, the lower [Mg(2+)](free) could be attributed to increased [DPG] and therefore greater magnesium binding capacity of sickle cells.

2,3-Diphosphoglycerate↗

PubMed2002

The energetics of ion distribution: the origin of the resting electric potential of cells.

The relation between the energies of ion movement and ATP hydrolysis is unknown in tissues with widely varying electric potentials. Consequently, we measured the concentration of the nine major inorganic ions in the extra- and intracellular phases in heart, liver, and red cells with resting electrical potentials, E(N), of -86, -28, and -6 mV, respectively, under six different physiological conditions. We calculated the Nernst electric potential and the energy of ion movement between the phases. We found that the energy of ATP hydrolysis was essentially constant, between -54 and -58 kJ/mol, in all tissues and conditions. In contrast, as E(N) decreased, the energies of the Na+ and K+ gradients decreased, with slopes approximating their valence. The difference between the energies of Na+ and K+ gradients remained constant at 17 kJ/mol, which is approximately one third of the energy of ATP hydrolysis, demonstrating near-equilibrium of the Na+/K+ ATPase in all tissues under all conditions. All cations, except K+, were pumped out of cells and all anions, except Cl- in liver and red cell, were pumped into cells. We conclude that the energy of ATP was expressed in Na+/K+ ATPase and its linked inorganic ion transporters to create a Gibbs-Donnan near-equilibrium system, an inherent part of which was the electric potential.

Adenosine Triphosphate↗

PubMed2002

Thiazolidinedione treatment normalizes insulin resistance and ischemic injury in the zucker Fatty rat heart.

Obesity is associated with risk factors for cardiovascular disease, including insulin resistance, and can lead to cardiac hypertrophy and congestive heart failure. Here, we used the insulin-sensitizing agent rosiglitazone to investigate the cellular mechanisms linking insulin resistance in the obese Zucker rat heart with increased susceptibility to ischemic injury. Rats were treated for 7 or 14 days with 3 mg/kg per os rosiglitazone. Hearts were isolated and perfused before and during insulin stimulation or during 32 min low-flow ischemia at 0.3 ml small middle dot min(-1) small middle dot grams wet wt(-1) and reperfusion. D[2-(3)H]glucose was used as a tracer of glucose uptake, and phosphorus-31 nuclear magnetic resonance spectroscopy was used to follow energetics during ischemia. At 12 months of age, obese rat hearts were insulin resistant with decreased GLUT4 protein expression. During ischemia, glucose uptake was lower and depletion of ATP was greater in obese rat hearts, thereby significantly impairing recovery of contractile function during reperfusion. Rosiglitazone treatment normalized the insulin resistance and restored GLUT4 protein levels in obese rat hearts. Glucose uptake during ischemia was also normalized by rosiglitazone treatment, thereby preventing the greater loss of ATP and restoring recovery of contractile function to that of lean rat hearts. We conclude that rosiglitazone treatment, by normalizing glucose uptake, protected obese rat hearts from ischemic injury.

Animals↗

PubMedDate not supplied

Uncoupling proteins in human heart.

Abnormal energetic activity in heart failure correlates inversely with plasma free-fatty-acid concentrations. However, the link between energetic and metabolic abnormalities is unknown. To investigate this association, we obtained blood samples from 39 patients undergoing coronary artery bypass graft surgery. Patients fasted overnight before samples were taken. When plasma free-fatty-acid concentrations were raised, cardiac mitochondrial uncoupling proteins (UCP) increased (isoform UCP2, p<0.0001; isoform UCP3, p=0.0036) and those of glucose transporter (GLUT4) protein decreased (cardiac, p=0.0001; skeletal muscle, p=0.0006). Consequently, energy deficiency in heart failure might result from increased mitochondrial UCPs (ie, less efficient ATP synthesis) and depleted GLUT4 (ie, reduced glucose uptake). New treatment to correct these energy defects would be to simultaneously lower plasma free fatty acids and provide an alternative energy source.

Adenosine Triphosphate↗