Concerning the angina of "syndrome X".
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Epinephrine↗
Biomedical subjects
A J Miller
Publications and source records attributed to A J Miller.
Xenopus oocytes as a heterologous expression system for plant proteins.
The Xenopus oocyte is a robust and convenient system for the transient expression of many different animal proteins and it has recently been demonstrated that oocytes can also translate, process, and target plant proteins. This expression system can also be used to clone genes, characterize function, and study posttranslational processing of proteins. Here we describe the methodology for the expression of plant proteins, in particular membrane proteins, in Xenopus oocytes.
Animals↗
Plasticity of craniomandibular muscle function: 31P magnetic resonance spectroscopy of the rabbit masseter muscle.
The masseter muscle was studied during postnatal development of the rabbit from the juvenile to adult stage in which the oral function was altered during maturation by modifying the diet to soft food. The muscle was assessed using phosphate magnetic resonance (31P NMR) spectroscopy with a single-turn copper surface coil to study potential changes in phosphate metabolism. The 31P NMR spectra consisted of five peaks related to unbound forms of inorganic phosphate (Pi), creatine phosphate (PCr), and three peaks related to the adenosine triphosphate (ATP). The masseter was assessed in one group of five rabbits at 8 weeks postnatally (juvenile) and after 4 months of this experimental group masticating on soft food. They were compared with a control group of five rabbits raised on a normal hard diet. The Pi/PCr ratio increased in the adult masseter much higher during twitching, tetany, and periodic contraction than in the juvenile regardless as to whether the adult animal had been raised from the juvenile period on soft or hard diet. There were relatively few differences between the experimental adult animals raised on a soft diet and the normal adult animals despite the soft diet animals demonstrating a significantly lower weight and smaller muscle mass. These findings suggest that chronic underuse of the masseter muscle by decreasing the masticatory loads has a minimal effect on the phosphate metabolism of the maturing masseter.
Adaptation, Physiological↗
In situ characterization of Clostridium botulinum neurotoxin synthesis and export.
A monoclonal antitoxin/colloidal gold probe and sequential centrifugation were used to study synthesis, translocation and export of Clostridium botulinum strain 62A neurotoxin (NT). Exponential growth occurred after 5 h of anaerobic incubation of spores and continued for 15-16 h. NT was detected at 15 h using the probe and transmission electron microscopy (TEM), 2 h earlier than the first detection by the mouse bioassay. During exponential growth, the probe localized NT primarily in the cytoplasm, on the inner side of the cytoplasmic membrane and in the cell wall. During stationary and death phases, the NT was located within the cytoplasm, cell wall and extracellularly. NT was released from the cell during cell wall exfoliation. Cells retained NT after repeated gelatin-phosphate washes and sequential centrifugations, consistent with the TEM observation that the NT is bound to the cell wall. These observations indicate that the process of Cl. botulinum type A NT production follows a sequence of synthesis, translocation across the cytoplasmic membrane and export through the cell wall.
Animals↗
Further analysis of the mechanisms underlying the tracheal relaxant action of SCA40.
1. SCA40 (1nM-10 microM), isoprenaline (1-300 nM) and levcromakalim (100 nM-10 microM) each produced concentration-dependent suppression of the spontaneous tone of guinea-pig isolated trachea. Propranolol (1 microM) markedly (approximately 150 fold) antagonized isoprenaline but did not antagonize SCA40. The tracheal relaxant action of SCA40 was unaffected by suramin (100 microM) or 8-(p)-sulphophenyltheophylline (8-SPT; 140 microM). 2. An isosmolar, K(+)-rich (80 mM) Krebs solution increased tracheal tone, antagonized SCA40 (approximately 60 fold), antagonized isoprenaline (approximately 20 fold) and very profoundly depressed the log concentration-effect curve for levcromakalim. Nifedipine (1 microM) did not itself modify the relaxant actions of SCA40, isoprenaline or levcromakalim. However, nifedipine prevented the rise in tissue tone and the antagonism of SCA40 and isoprenaline induced by the K(+)-rich medium. In contrast, nifedipine did not prevent the equivalent antagonism of levcromakalim. 3. Charybdotoxin (100 nM) increased tracheal tone, antagonized SCA40 (approximately 4 fold) and antagonized isoprenaline (approximately 3 fold). Nifedipine (1 microM) prevented the rise in tissue tone and the antagonism of SCA40 and isoprenaline induced by charybdotoxin. 4. Quinine (30 microM) caused little or no change in tissue tone and did not modify the relaxant action of isoprenaline. However, quinine antagonized SCA40 (approximately 2 fold). Nifedipine (1 microM) prevented the antagonism of SCA40 induced by quinine. 5. Tested on spontaneously-beating guinea-pig isolated atria SCA40 (1 nM-10 microM) increased the rate of beating in a concentration-dependent manner. Over the concentration-range 1 microM-10 microM, SCA40 also caused an increase in the force of atrial contraction. 6. Intracellular electrophysiological recording from guinea-pig isolated trachealis showed that the relaxant effects of SCA40 (1 micro M) were often accompanied by the suppression of spontaneous electrical slow waves but no change in resting membrane potential. When the concentration of SCA40 was raised to 10 micro M, its relaxant activity was accompanied both by slow wave suppression and by plasmalemmal hyperpolarization.7. SCA40 (10 nM- 100 micro M) more potently inhibited the activity of cyclic AMP phosphodiesterase (PDE)than that of cyclic GMP PDE derived from homogenates of guinea-pig trachealis. Theophylline(1 micro M- 1O mM) also inhibited these enzymes but was less potent than SCA40 in each case and did not exhibit selectivity for inhibition of cyclic AMP hydrolysis.8. Tested against the activity of the isoenzymes of cyclic nucleotide PDE derived from human blood cells and lung tissue, SCA40 proved highly potent against the type III isoenzyme. It was markedly less potent against the type IV and type V isoenzymes and even less potent against the isoenzymes types I and II.9. It is concluded that the tracheal relaxant action of SCA40 (1 nM- 1 micro M) does not involve the activation of beta-adrenoceptors or P1 or P2 purinoceptors. Furthermore, this action is unlikely to depend upon the opening of BKca channels with consequent cellular hyperpolarization and voltage-dependent inhibition of Ca2+ influx. The tracheal relaxant action of SCA40 (up to 1 micro M) is more likely to depend upon its selective inhibition of the type III isoenzyme of cyclic nucleotide PDE. At concentrations above 1 micro M, SCA40 exerts more general inhibition of the isoenzymes of cyclic nucleotide PDE and may then promote the opening of BKca channels.
Animals↗
The three Williams--Hunter, Hewson and Cruikshank: their unique contributions to our knowledge of the lymphatics.
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Anatomy↗
31P-magnetic resonance spectroscopy of the rabbit masseter muscle.
Dynamic biochemical changes in the masseter muscle were studied in 14 New Zealand adult male rabbits by 31P-nuclear magnetic resonance (NMR) spectroscopy. NMR spectra were obtained during rest and electrical stimulation of the muscle in the anaesthetized animal at 33 recording sessions. Electrical stimulation was applied by a pair of copper wires placed separately with hypodermic needles into the muscle. NMR spectra were acquired with a 2 x 3 cm, double-turn, copper transmit/receive coil. Sixteen spectra were averaged over 30 s to obtain averaged spectra continuously during a 30-min recording. The spectra were processed automatically using a non-linear 'least-squares' fitting program on the spectrometer. A Lorentzian line shape was assumed for the peaks, and values of peak height, area and chemical shifts were generated. Each averaged spectrum consisted of five peaks: inorganic phosphate (Pi), creatine phosphate (PCr), and three peaks related to ATP. Data were analysed as to absolute changes in Pi and PCr, in the ratio of Pi/PCr, and the shift of Pi to PCr to estimate pH. Several protocols were used in which ranges of frequency, intensity and duration of electrical stimulation were tested. The protocol for detailed studies involved stimulating the muscle twice at 5 Hz for 3 min separated by a 3-min rest period, then stimulating twice at 50 Hz for 3 min separated by a rest period. During contraction of the muscle, there was a significant increase in the Pi/PCr ratio (p < 0.05) as compared to the resting level. The ratio reached a plateau over a 3-min contraction using 5-Hz stimulation, then increased significantly more with the 50-Hz stimulation but decayed during the 3 min. Sustained stimulation with 50 Hz for 15-45 min evoked an initial sharp change in Pi/PCr, which then reached a steady plateau that remained over the entire stimulation. These findings indicate that the rabbit masseter muscle is relatively fatigue resistant in maintaining a steady-state equilibrium in the relation of Pi to PCr.
Adenosine Triphosphate↗
Vancomycin penetration into biofilm covering infected prostheses and effect on bacteria.
To evaluate the adequacy of penetration of antibiotics into biofilm, a novel in vitro model for prosthesis-related infection was developed. Sterile stainless steel orthopedic nuts were incubated with slime-producing Staphylococcus epidermidis. Biofilm-covered nuts were exposed to varying concentrations of vancomycin; then biofilms were harvested. Vancomycin levels in biofilm, as measured by fluorescent polarization immunoassay, far exceeded the MIC and MBC of vancomycin for the organism. Bacterial growth in biofilm was inversely related to vancomycin concentration in biofilm, but even extremely high drug concentrations did not eradicate bacteria embedded in biofilm. The MICs and MBCs for bacteria recovered from biofilm did not differ from those for incubating organisms. Thus, failure of glycopeptide antibiotics to cure prosthesis-related infection is not due to poor penetration of drugs into biofilm but likely due to diminished antimicrobial effect on bacteria in the biofilm environment.
Bioprosthesis↗
Disruptive effects of protein on performance of liquid membrane-based ion-selective microelectrodes.
The effects of addition of protein [bovine serum albumin (BSA)] to the calibration solutions of various liquid membrane-based ion-selective microelectrodes have been assessed. Protein at typically cytosolic levels of 100 mg/ml had no effect on the response of Na(+)-, K(+)-, Mg(2+)-, Ca(2+)-, H(+)-, NO3(-)-, and Cl(-)- selective microelectrodes at ion concentrations well above the respective detection limits. By contrast, this level of protein resulted in a pronounced increase in the detection limits of Na(+)-, K(+)-, Mg(2+)-, Ca2+, and Cl(-)-selective microelectrodes; H+ and NO3- microelectrodes were largely unaffected by the protein. The effect of BSA on detection limit is independent of pH and ionic strength, nor is it due to hysteresis, to binding of ions to the protein, or to the generation of junction potentials. More extensive studies on Na(+)-selective microelectrodes revealed that other proteins and large molecules (e.g., polyethylene glycol) also increase detection limits, with the extent of this effect apparently correlated positively with the molecular weight. Because biological macromolecules at physiological concentrations can influence the properties of alkali- and alkali earth cation-selective microelectrodes, care must be taken if the use of these electrodes entails cytosolic measurements close to their detection limits.
Equipment Failure↗
The efficacy of choline magnesium trisalicylate (CMT) in the management of metastatic bone pain: a pilot study.
Twenty-six patients with painful, bony metastases were recruited into a randomized, double-blind, single dose, two-treatment, three-part crossover study of choline magnesium trisalicylate (CMT) and placebo. Assessments were made prior to and at one, two, three and four hours after dosing. Bone pain caused by metastatic cancer was significantly relieved one hour after the administration of 1500 mg CMT (p = 0.04). At all four time points the pain was less than baseline with CMT and at three time points greater than baseline with placebo but these results did not reach statistical significance. The summed pain intensity difference for patients was greater with CMT than with placebo, but this also did not reach significance. The incidence of volunteered side-effects was similar for both treatments. The results suggest that a nonacetylating, nonsteroidal anti-inflammatory drug may have a role complementary to that of an opioid in the management of metastatic bone pain.
Adolescent↗
The heterologous expression of H(+)-coupled transporters in Xenopus oocytes.
The Xenopus oocyte has become a convenient and robust system for the expression of many different animal transport proteins and it has now been demonstrated that oocytes can also translate, process and target plant membrane proteins. Here we review the background to this expression system and discuss how it can be used to express plant transporters. A H+/hexose cotransporter (STP1) from Arabidopsis thaliana has been successfully expressed in oocytes and further characterization has shown the transporter to have properties similar to the same transporter expressed in yeast. However, the expression of H(+)-coupled transporters may present problems for oocyte intracellular pH regulation, because the influx of protons during their operation may acidify the cell. To investigate this possibility, proton-selective microelectrodes have been used to measure intracellular pH during symport of H+ and hexose in oocytes expressing STP1. These measurements showed that oocyte cytosolic pH is unaltered during H+/glucose cotransport, but is sensitive to changes in external pH.
Animals↗
The cardiac lymphatic system and coronary artery disease.
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Animals↗
The lymphatics of the canine parietal pericardium.
A careful anatomical study of the lymphatic drainage from the parietal pericardium reveals a complicated network that is notably different from that defined by the instillation of markers into the pericardial sac. The pericardial cupola and the posterior (dorsal) area of the parietal pericardium drain directly to the cardiac lymph node in the right upper mediastinum by relatively short lymphatics. The lymphatics of the anterior and lateral areas of the parietal pericardium pass to collecting vessels that travel cranially or caudally along the phrenic nerves. The former traverse specific lymph nodes and then enter the right or left venous angles. The latter pass caudally and then drain to the major collecting systems from the area of the diaphragm.
Animals↗
Functional expression of a plant plasma membrane transporter in Xenopus oocytes.
A full-length cDNA clone for the H+/hexose co-transporter (STP1) from Arabidopsis thaliana has been transcribed in vitro and the mRNA injected into Xenopus oocytes. Under optimized conditions, oocytes injected with the STP1 mRNA accumulated 3-O-[methyl-14C]glucose at rates of more than a 1000-fold greater than water-injected control oocytes. A hexose-elicited depolarization of the oocyte membrane potential was demonstrated, and uptake was shown to be stimulated by low external pH, confirming the activity of a H+/hexose co-transport system. This is the first example of the functional expression of a plant membrane transporter in oocytes.
3-O-Methylglucose↗
Unstable asthma and theophylline.
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Asthma↗
Fixation of fractures of the glenoid rim.
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Adult↗
The role of the lymphatic system in coronary atherosclerosis.
Arguments are presented to support the hypothesis that the integrity of the coronary lymphatic system is an important factor in the development of coronary atherosclerosis. It is proposed that the lymphatic drainage of the epicardial coronary arteries is genetically inadequate, and that the flow of lymph from the heart is further impaired by gravitational factors due to man's upright position. Arguments are presented to support the concept that the intramural coronary arteries are protected from atherosclerosis because of a highly effective pericoronary lymphatic and venous capillary system and that the internal thoracic (internal mammary) artery is similarly protected from atherosclerosis because of its close relationship to an extensive lymphatic chain. It is proposed that the failure of the cardiac lymphatic system in the allogenic transplanted heart is a cause of the accelerated atherosclerosis in both epicardial and intramural coronary arteries.
Animals↗
Inhibition by adrenergic neurone blocking agents of the relaxation induced by BRL 38227 in vascular, intestinal and uterine smooth muscle.
1. The adrenergic neurone blocking agents, guanethidine and bretylium, have been tested for inhibitory activity against the actions of some relaxant drugs (BRL 38227, noradrenaline, sodium nitroprusside, theophylline) in vascular, intestinal and uterine smooth muscle. 2. In guinea-pig isolated taenia caeci pre-contracted with KCl (25 mM), BRL 38227 (0.1-10 microM) and noradrenaline (10 nM-100 microM) each caused concentration-dependent relaxation. Guanethidine and bretylium (50 microM) each antagonized the relaxation to BRL 38227 but not that to noradrenaline. At high concentration (500 microM), the adrenergic neurone blocking agents antagonized the action of BRL 38227 and, to some extent, that of noradrenaline. 3. In rat isolated aorta pre-contracted with noradrenaline (300 nM), BRL 38227 (0.0125-3.2 microM) and sodium nitroprusside (0.3-100 nM) each produced concentration-dependent smooth muscle relaxation. Guanethidine and bretylium (5-500 microM) each antagonized the action of BRL 38227 without antagonizing that of sodium nitroprusside. 4. Rats were pretreated with 17-beta oestradiol benzoate. Tension waves were then induced from segments of isolated, oestrogen-dominated uterus by transmural electrical stimulation or by oxytocin (0.2 nM). These tension waves were inhibited by BRL 38227 (0.025-3.2 microM) or theophylline (0.05-0.8 mM) in a concentration-dependent manner. Guanethidine (50 microM) antagonized the action of BRL 38227 in both the electrically- and oxytocin-driven tissues. In the electrically-driven tissues, guanethidine (50 microM) did not antagonize the inhibition to theophylline. 5. In KCl (25 mM)-treated guinea-pig taenia caeci, guanethidine (50 microM) inhibited the efflux of 86Rb+ evoked by BRL 38227 (10 microM) but not that evoked by noradrenaline (10 microM). In contrast, apamin(100 nM) reduced the efflux of 86Rb+ which was promoted by noradrenaline, but did not affect efflux induced by BRL 38227.6. It is concluded that the adrenergic neurone blocking agents, guanethidine and bretylium (each at 50 microM), selectively inhibit the relaxant action of BRL 38227 in vascular, intestinal and uterine smooth muscle. If this inhibition reflects direct blockade of the K+-channel (KKCO) which is opened by BRL 38227, then the adrenergic neurone blocking agents act as inhibitors selective for KKCO as opposed to the small, apamin-sensitive (SKCa) and large (BKca) conductance, Ca2"-dependent K+-channels.
Animals↗