Changes in bacterial species composition in enrichment cultures with various dilutions of inoculum as monitored by denaturing gradient gel electrophoresis

Denaturing gradient gel electrophoresis revealed changes in the bacterial species obtained from enrichment cultures with different inoculum dilutions. This inoculum dilution enrichment approach may facilitate the detection and isolation of a greater number of bacterial species than traditional enrichment techniques.     

Effect of progesterone on prostaglandin F2 alpha secretion and outcome of pregnancy during cloprostenol-induced abortion in mares

OBJECTIVES: To determine the role of progesterone in the regulation of endogenous prostaglandin F2 alpha (PGF2 alpha) secretion during cloprostenol-induced abortion and to investigate use of progestins to prevent prostaglandin-associated abortion. ANIMALS: 16 pregnant mares. PROCEDURE: To induce abortion, cloprostenol (250 micrograms/d) was administered daily until fetal expulsion or for up to 5 days. In experiment 1, 8 mares, 98 to 153 days' pregnant, received progesterone (300 mg/d) at 24-hour intervals for 5 days, starting 18 hours after the first cloprostenol administration. In experiment 2, 8 mares, 93 to 115 days' pregnant, received altrenogest (44 mg/d) at 24-hour intervals, starting 12 hours after the first cloprostenol administration. Historic control mares, 82 to 102 days' pregnant, received cloprostenol (250 micrograms/d) daily until fetal expulsion. RESULTS: In control mares, fetal expulsion occurred after 2 to 3 cloprostenol administrations and was associated with significant increases in PGF2 alpha secretion. Abortion did not occur in 5 of 8 progesterone-treated mares and 8 of 8 altrenogest-treated mares, and endogenous PGF2 alpha secretion was inhibited, compared with values in aborting mares. CONCLUSION: Circulating progestogen concentrations may have a role in the outcome of pregnancy during prostaglandin-induced abortion. Altered prostaglandin secretion may be a reflection of a direct effect of progesterone or may be caused by the abortion process. CLINICAL RELEVANCE: Progestogens might be useful for prevention of abortion in mares in which pregnancy is at risk owing to diseases that are associated with excess prostaglandin secretion.     

Heart rate variability: two eras of investigation

Changes in membrane potential are a regulatory signal for voltage-gated ion channels including the family of Ca2+ channels. This regulatory role includes the voltage-dependent opening, closing, and inactivation of ion channels and the control of drug access and affinity for discrete channels states. Membrane potential is both a short- and long-term regulatory signal controlling the number and function of these voltage-gated channels. Depolarization of neuronal and neurosecretory cells produces down-regulation of L-type voltage-gated Ca2+ channels. These processes are described and their relevance to physiologic and pathologic processes of neuronal development and neuroprotection are indicated.     

Physiological response of Lactobacillus plantarum to salt and nonelectrolyte stress

In this report, we compared the effects on the growth of Lactobacillus plantarum of raising the medium molarity by high concentrations of KCl or NaCl and iso-osmotic concentrations of nonionic compounds. Analysis of cellular extracts for organic constituents by nuclear magnetic resonance spectroscopy showed that salt-stressed cells do not contain detectable amounts of organic osmolytes, whereas sugar-stressed cells contain sugar (and some sugar-derived) compounds. The cytoplasmic concentrations of lactose and sucrose in growing cells are always similar to the concentrations in the medium. By using the activity of the glycine betaine transport system as a measure of hyperosmotic conditions, we show that, in contrast to KCl and NaCl, high concentrations of sugars (lactose or sucrose) impose only a transient osmotic stress because external and internal sugars equilibrate after some time. Analysis of lactose (and sucrose) uptake also indicates that the corresponding transport systems are neither significantly induced nor activated directly by hyperosmotic conditions. The systems operate by facilitated diffusion and have very high apparent affinity constants for transport (>50 mM for lactose), which explains why low sugar concentrations do not protect against hyperosmotic conditions. We conclude that the more severe growth inhibition by salt stress than by equiosmolal concentrations of sugars reflects the inability of the cells to accumulate K+ (or Na+) to levels high enough to restore turgor as well as deleterious effects of the electrolytes intracellularly.     

Production of human normal adult and fetal hemoglobins in Escherichia coli

A hemoglobin expression system in Escherichia coli is described. In order to produce authentic human hemoglobin, we need to co-express both methionine aminopeptidase and globin genes under the control of a strong promoter. We have constructed three plasmids, pHE2, pHE4 and pHE7, for the expression of human normal adult hemoglobin and a plasmid, pHE9, for the expression of human fetal hemoglobin, in high yields. The globin genes can be derived from either synthetic genes or human globin cDNAs. The extra amino-terminal methionine residues of the expressed globins can be removed by the co-expressed methionine aminopeptidase. The heme is inserted correctly into the expressed alpha- globin from our expression plasmids. A fraction (approximately 25%) of the heme is not inserted correctly into the expressed beta- or gamma- globin. However, the incorrectly inserted hemes can be converted into the correct conformation by carrying out a simple oxidation-reduction process on the purified hemoglobin molecule. We have investigated the functional properties of the expressed hemoglobins by measuring their oxygen-binding properties and their structural features by obtaining their 1H-NMR spectra. Our results show that authentic human normal adult and fetal hemoglobins can be produced from our expression plasmids in E. coli and in high yields. Our expression system allows us to design and to produce any recombinant hemoglobins needed for our research on the structure-function relationship in hemoglobin.      

Megavoltage imaging with low Z targets: implementation and characterization of an investigational system

The poor quality of stereotactic radiotherapy portal images is a limiting factor in precise image registration. To alleviate this problem, a low atomic number (Z) target was implemented on our Siemens MXE linear accelerator. This investigational system was used to assess the performance of various target materials by filming an aluminum contrast object. Beryllium, carbon and conventional target materials were studied. The bremsstrahlung spectra of these materials were simulated using Monte Carlo techniques. These spectra were used to calculate the dependence of narrow beam contrast on phantom thickness for verification of the data measured from film. A Monte Carlo simulation of the beryllium spectrum in a wide beam geometry was used to evaluate the effect of phantom-to-film distance on contrast. Although the same degree of contrast improvement with distance was not realized in practice, the improvement in image quality rivaled that achieved using a scatter reduction grid. A comparison of conventional localization images of the head and neck of an anthropomorphic phantom with images produced with a beryllium or carbon target and a mammography film and screen system supports earlier suggestions that the technique is clinically useful.     

A two-term MEDLINE search strategy for identifying randomized trials in obstetrics and gynecology

In this study, we examined whether endothelin (ET) plays a role in the short-term increase in mean arterial pressure (MAP) after nitric oxide synthase (NOS) inhibition with N(omega)-nitro-L-arginine methyl ester (L-NAME) in stroke-prone spontaneously hypertensive rats (SHRSPs). Experiments were performed by using Inactin-anesthetized male SHRSPs that were pretreated with chlorisondamine to block reflex autonomic cardiovascular effects. Injection of L-NAME (10 mg/kg, i.v.), but not D-NAME, produced rapid and marked increases (74 +/- 3 mm Hg) in MAP that were sustained for >1 h. In SHRSPs that were treated with the ET(A/B) receptor antagonist, L-754,142 (15 mg/kg + 15 mg/kg/h), L-NAME increased MAP by 45 +/- 4 mm Hg (p < 0.0001 compared with L-NAME alone). L-754,142 blocked pressor responses to big ET-1 by >90% but was without effect on pressor responses to norepinephrine. Plasma levels of ET-1 averaged 5 +/- 1 pg/ml in animals given vehicle and were slightly increased in animals given either L-NAME alone (7 +/- 2 pg/ml) or L-754,142 alone (7 +/- 2 pg/ml) but increased markedly when L-NAME and L-754,142 were given together (114 +/- 18 pg/ml). This may relate to an effect of L-754,142 to block ET-receptor-mediated clearance of ET-1. We conclude that ET plays a role in the short-term pressor response after NOS inhibition in SHRSPs.     

Long-term potentiation in vivo in layers II/III of rat barrel cortex

Long-term potentiation was studied in vivo in the rat barrel cortex. It was found that LTP lasting several hours could be induced in layer II/III by tetanic stimuli applied in layer IV. The probability of inducing LTP at a given site was high (86%) provided that the electrodes were not displaced too far horizontally. LTP was not observed if the stimulating electrode was located on the far side of the neighbouring barrel-column from the recording electrode. The strongest LTP was induced by stimulating layer IV septal locations or the edge of the barrel and recording in the near half of the neighbouring barrel. However, examples were found of LTP from layer IV to II/III within the same barrel, within the same septum and from barrel to adjacent septum. The probability of inducing LTP on a particular occasion was greatly increased by iontophoresis of bicuculline at the recording site during the tetanus (from 20 to 55% judged by a change in peak amplitude). The average increase in the peak amplitude was 29 +/- 3.2% for protocol 1 (urethane anesthesia, monopolar stimulation) and 23 +/- 7% for protocol 2 (barbiturate anesthesia, bipolar stimulation). The probability of inducing LTP was greater if the first tetanus was accompanied by BMI application (67%) than for any subsequent attempts (39%). These results suggest it should be possible to study the effect of LTP on sensory processing in defined positions within the barrel field.