Sites of alcohol and volatile anaesthetic action on GABA(A) and glycine receptors

Volatile anaesthetics have historically been considered to act in a nonspecific manner on the central nervous system. More recent studies, however, have revealed that the receptors for inhibitory neurotransmitters such as gamma-aminobutyric acid (GABA) and glycine are sensitive to clinically relevant concentrations of inhaled anaesthetics. The function of GABA(A) and glycine receptors is enhanced by a number of anaesthetics and alcohols, whereas activity of the related GABA rho1 receptor is reduced. We have used this difference in pharmacology to investigate the molecular basis for modulation of these receptors by anaesthetics and alcohols. By using chimaeric receptor constructs, we have identified a region of 45 amino-acid residues that is both necessary and sufficient for the enhancement of receptor function. Within this region, two specific amino-acid residues in transmembrane domains 2 and 3 are critical for allosteric modulation of both GABA(A) and glycine receptors by alcohols and two volatile anaesthetics. These observations support the idea that anaesthetics exert a specific effect on these ion-channel proteins, and allow for the future testing of specific hypotheses of the action of anaesthetics.     

A monitoring, feedback, and triggering system for reproducible breath-hold MR imaging

OBJECTIVE: To evaluate the diagnostic accuracy and clinical usefulness of high-resolution transvaginal duplex Doppler ultrasound in postpartum and post-abortion patients with excessive hemorrhage who are suspected of having residual trophoblast. METHODS: Forty-eight women with excessive hemorrhage referred for possible residual trophoblastic tissue were evaluated by transvaginal duplex Doppler ultrasonography. Based on two-dimensional imaging, the patients were divided prospectively into groups: women who had an empty uterus with a normal uterine cavity, those with a pure endometrial fluid collection and no echogenic foci, those who had a mixed endometrial fluid collection with foci of echogenicity, and those with intracavitary heterogeneous material with mixed echo patterns of fluid and solid components. In each group, Doppler studies were performed and the resistance index (RI) was calculated. The two-dimensional patterns and Doppler results were correlated with clinical and pathologic follow-up. RESULTS: Twenty-eight subjects had a normal uterine cavity and seven had a pure endometrial fluid collection; all were treated conservatively and none showed later clinical evidence of residual trophoblastic tissue. In 13 women, residual trophoblast was strongly suggested from the images of two-dimensional ultrasonography: Five showed an endometrial fluid collection with some echogenic foci, and eight exhibited intracavitary mixed echogenic material. All underwent curettage, and residual trophoblastic tissue was found in ten of the 13. The mean (+/- standard deviation) RI to flow in the myometrial arteries was 0.54 +/- 0.15 in women without residual trophoblast and 0.35 +/- 0.1 in those with residual trophoblastic tissue (P < .01). CONCLUSION: Our experience suggests that transvaginal duplex Doppler ultrasonography is an effective noninvasive method for evaluating patients with excessive postpartum and post-abortion hemorrhage who are suspected of having residual trophoblastic tissue. Its use enhances the positive preoperative diagnosis of residual trophoblastic tissue and may reduce unnecessary curettage procedures.     

Retroviral display of antibody fragments; interdomain spacing strongly influences vector infectivity

Five different single-chain antibody fragments (scFv) against human cell-surface antigens were displayed on murine ecotropic retroviral vectors by fusing them to the Moloney SU envelope glycoprotein. The spacing between the scFv and the SU glycoprotein was varied by fusing the scFv to residue +7 or to residue +1 of Moloney SU and by inserting linker sequences of different lengths between the domains. All of the chimeric envelopes were efficiently incorporated into vector particles and could bind to human cells through their displayed antibody fragments, but did not infect them. The spacing between the scFvs and the SU glycoproteins had no significant effect on the efficiency of envelope expression or viral incorporation and did not affect the binding properties of the chimeric envelopes, nor did it influence the efficiency of targeted gene delivery to human cells by scFv-displaying vectors. However, on murine fibroblasts the infectivity of vectors incorporating the chimeric envelopes was strongly influenced by the length of the interdomain spacer. The titers were very low when the single-chain antibodies were fused through a tripeptide linker to SU residue +7 and were greatly enhanced (up to 10(5)-fold) when they were fused to SU residue +1 through a heptapeptide linker. These results point to the importance of steric interactions between the domains of chimeric envelope glycoproteins and may have implications for retroviral vector design for human gene therapy.     

Polymorphism of bacteriophage T7

For viruses made of nucleic acid and protein, the structure of the protein outer shell has, in the past, been found to be uniquely determined by the viral genome. However, here, non-denaturing agarose gel electrophoresis of bacteriophage T7 reveals two states of the mature T7 capsid; the conditions of growth are found to alter the population by T7 of these two electrophoretically defined states. Both states have been previously observed for a genetically altered T7 and they are observed here for wild-type T7. The average electrical surface charge density of a bacteriophage particle (delta) determines its state; the delta of particles in both states is negative. For a given condition of growth, the population of these two states is influenced by the extent to which the major T7 outer shell protein, p10A, is accompanied by its minor readthrough variant, p10B. Comparison of the two electrophoretic states reveals the following. (1) No difference in radius is present in the outer shell (+/-2%). (2) As the pH of electrophoresis is either increased or decreased from neutrality, the state becomes more highly populated for which delta is greater in magnitude (state 1). By changing the pH, some T7 particles are made to change state. (3) Particles in state 1 adsorb less quickly to host cells than do the particles in the alternative state (state 2). This latter observation suggests the hypothesis that state 1 evolved to reduce the probability of re-initiating an infection when conditions are not favorable for growth. This hypothesis is supported by the observation that, as conditions of growth become apparently more unfavorable, progeny increasingly populate state 1.     

Maternal and neonatal outcomes after prolonged latent phase

Glycol ethers such as 2-ethoxyethanol (EE) are widely used as solvents because they are miscible in aqueous and organic solutions. Toxic effects of EE in rodents include teratogenicity, fetotoxicity, hematotoxicity, and testicular atrophy. The purpose of this study was to determine the effect of dose on the absorption, metabolism, and excretion of 2-ethoxy [U-14C]ethanol by F344/N rats after inhalation exposure. Rats were exposed to either 5 ppm EE for 5 hr 40 min or 46 ppm EE for 6 hr. The uptake and metabolism of EE were linear in the concentration range studied. Significant percentages of the retained doses were exhaled during (22%) and after exposure (16%) as 14CO2. Forty-six percent of the retained dose was excreted in the urine. Approximately 10% of the retained dose was detected in the carcass 66 hr after exposure. The major urinary metabolite was ethoxyacetic acid (EAA), the toxic metabolite of EE. The amount of EAA excreted was linearly related to exposure concentration. Ethylene glycol and N-ethoxyacetyl glycinate were identified as minor metabolites excreted in the urine. The results of this study suggest that the toxicity of inhaled EE should be directly proportional to the exposure concentration up to 46 ppm if the toxicity of EE is due to EAA.