Evidence of a source of HIV type 1 within the central nervous system by ultraintensive sampling of cerebrospinal fluid and plasma.

Defining the source of HIV-1 RNA in cerebrospinal fluid (CSF) will facilitate studies of treatment efficacy in the brain. Four antiretroviral drug-naive adults underwent two 48-hr ultraintensive CSF sampling procedures, once at baseline and again beginning on day 4 after initiating three-drug therapy with stavudine, lamivudine, and nelfinavir. At baseline, constant CSF HIV-1 RNA concentrations were maintained by daily entry of at least 10(4) to 10(6) HIV-1 RNA copies into CSF. Change from baseline to day 5 ranged from -0.38 to -1.18 log(10) HIV-1 RNA copies/ml in CSF, and from -0.80 to -1.33 log(10) HIV-1 RNA copies/ml in plasma, with no correlation between CSF and plasma changes. There was no evidence of genotypic or phenotypic viral resistance in either CSF or plasma. With regard to pharmacokinetics, mean CSF-to-plasma area-under-the-curve (AUC) ratios were 38.9% for stavudine and 15.3% for lamivudine. Nelfinavir and its active M8 metabolite could not be accurately quantified in CSF, although plasma M8 peak level and AUC(0-8hr) correlated with CSF HIV-1 RNA decline. This study supports the utility of ultraintensive CSF sampling for studying HIV-1 pathogenesis and therapy in the CNS, and provides strong evidence that HIV-1 RNA in CSF arises, at least in part, from a source other than plasma.

A human extrastriate area functionally homologous to macaque V4.

Extrastriate area V4 is crucial for intermediate form vision and visual attention in nonhuman primates. Human neuroimaging suggests that an area in the lingual sulcus/fusiform gyrus may correspond to ventral V4 (V4v). We studied a human neurological patient, AR, with a putative V4v lesion. The lesion does not affect early visual processing (luminance, orientation, and motion perception). However, it does impair hue perception, intermediate form vision, and visual attention in the upper contralateral visual field. Form deficits occur during discrimination of illusory borders, Glass patterns, curvature, and non-Cartesian patterns. Attention deficits occur during discrimination of the relative positions of object parts, detection of low-salience targets, and orientation discrimination in the presence of distractors. This pattern of deficits is consistent with the known properties of area V4 in nonhuman primates, indicating that AR's lesion affects a cortical region functionally homologous to macaque V4.

Prevention of experimental Haemophilus ducreyi infection: a randomized, controlled clinical trial.

Human subjects were infected with Haemophilus ducreyi. All subjects developed papules and were randomized to treatment with a single dose of azithromycin (1 g) or ciprofloxacin (500 mg). At weekly intervals, volunteers were reinoculated with H. ducreyi, and drug concentrations were measured in peripheral blood mononuclear cells (PBMC). When papules developed, the subjects were treated with antibiotics and dismissed from the study. Eight of the ciprofloxacin-treated subjects developed papules 1 week after the initial treatment, and the ninth subject developed disease 2 weeks after treatment. The 9 azithromycin-treated subjects developed papules 4-10 weeks (mean, 6.8) after the initial treatment (P < .001). Azithromycin was detected in PBMC for 3-6 weeks (mean, 4). Pre- and posttreatment lesions had histology typical of experimental chancroid or were culture positive. Azithromycin prevents experimental chancroid for nearly 2 months. These findings have implications for strategies to prevent chancroid.

Serum concentrations of erythromycin after intravenous infusion in preterm neonates treated for Ureaplasma urealyticum infection.

Erythromycin is receiving renewed attention as an alternative for treatment of neonatal infections caused by Ureaplasma urealyticum because of recently proved abilities of this organism to produce systemic disease in this population. Although erythromycin has been used clinically for almost 40 years, very little is known about its activity in the preterm neonate. Fourteen neonates, birth weights < or = 1500 g and < or = 15 days of age, from whom U. urealyticum was isolated from the lower respiratory tract were randomized to receive erythromycin lactobionate either 25 or 40 mg/kg/day in four divided doses at 6-hour intervals scheduled for a total of 10 days. Blood samples collected at multiple time points after initial and steady state doses were assayed for erythromycin by liquid chromatography. Minimal inhibitory concentrations (MICs) of erythromycin for the U. urealyticum isolates were determined. MICs ranged from 0.031 to 2 micrograms/ml; MIC90 = 2 micrograms/ml. Serum erythromycin concentrations met or exceeded most MICs, with peak values of 3.05 to 3.69 and 1.92 to 2.9 micrograms/ml for the 40- and 25-mg/kg/day dosage groups, respectively. Pharmacokinetic parameters were calculated after the initial dose and at steady state for both dosage groups and compared. No adverse effects thought to be related to administration of erythromycin were observed. These preliminary findings showed that erythromycin is well-tolerated, has favorable pharmacokinetic activity in the preterm neonate and should be further investigated for treatment of ureaplasmal infections.

Orientation asymmetry in the flanker task.

Experiments 1 and 2 of this study show that when the target is either a vertical or a horizontal line, diagonal-line flankers tilted 45 degrees either to the right or to the left have the same effect as do incongruent flankers. When the target is a diagonal line tilted either to the right or to the left, vertical- or horizontal-line flankers do not have the same effect as do incongruent flankers. Experiment 3 demonstrates that this asymmetry is not caused by the temporal-dynamic aspects of the processing. Together, these experiments suggest that there is an asymmetrical relation between diagonal lines and either vertical or horizontal lines outside of the central focus of attention. Experiment 4 shows that despite this asymmetry in the flanker task, visual search for a vertical- or a horizontal-line target among diagonal-line distractors is not affected by the number of distractors. Possible explanations of this phenomenon are discussed.

Simultaneous detection of thiol- and disulfide-containing peptides by electrochemical high-performance liquid chromatography with identification by mass spectrometry.

A new HPLC electrochemical detector that can be used to detect selectively and simultaneously both thiol- and disulfide-containing peptides is described. One electrode responds only to thiol-containing peptides, while a second electrode located downstream from a third electrode responds to thiol- as well as disulfide-containing peptides. All three electrodes are located in a single block and sample the HPLC effluent simultaneously. Since the electrochemical detector responds only to peptides that contain thiol or disulfide functionalities, it is ideal for detecting these types of peptides when they are present in complex mixtures. Peptides can be identified by their retention times if reference peptides are available, or by their molecular weights if the appropriate chromatographic fractions are collected and analyzed by mass spectrometry. The utility of the three-electrode electrochemical detector for detecting thiol- and disulfide-containing peptides in a single chromatographic analysis is illustrated with proteolytic digests of bovine alpha A-crystallin and partially reduced bovine insulin.

Optimization of LC apparatus for determinations in neurochemistry with an emphasis on microdialysis samples.

The popularity of in vivo microdialysis sampling of low-molecular-weight substances has focused attention on improved liquid chromatography procedures for such studies. The complexity of the in vivo experiment, coupled with the complexity of LC, has discouraged some workers from developing this capability in their laboratory. Many small-volume dialysate samples are collected over hours of experiments. Proper handling of the animal (anaesthesia, temperature control, probe size and placement, choice of perfusion media) is critical. Simultaneously giving equal attention to sample collection and storage, derivatization, analysis precision, calibration for accuracy, and preventive maintenance of LC pumps, columns, and detectors is difficult for many laboratories. For reliable LC-based assays for microliter volumes of dialysates minimum human interaction with the samples is desirable. Automation strategies and selection of LC components which we have adopted are described for routine analytes such as biogenic amines, amino acids, choline/acetylcholine, and glucose.

Simultaneous multiple electrode liquid chromatographic-electrochemical assay for catecholamines, indole-amines and metabolites in brain tissue.

To enhance the selectivity of liquid chromatographic-electrochemical assays for biogenic amines and metabolites in brain tissue, multiple electrode transducers were investigated. Two configurations of dual working electrodes were examined: parallel-adjacent and series arrangements. Using the raw detector currents with each configuration, peak-height ratios from simultaneously generated chromatograms were calculated to assess the selectivity of the instrument for direct injections of brain tissue supernatant. Ratios were consistent with injections of standards. Nearly coeluting peaks such as norepinephrine and 3-methoxy-4-hydroxyphenylglycol were resolved by using dual detector electrodes in series; only the catecholamines were detected at the downstream electrode owing to their electrochemical reversibility. The scheme was applicable to the assay of norepinephrine, 3,4-dihydroxyphenylacetic acid, dopamine, homovanillic acid, serotonin, 5-hydroxytryptophan and 5-hydroxyindole-3-acetic acid in brain tissue in less than 15 min.

Recent developments in the clinical assessment of the metabolism of aromatics by high-performance, reversed-phase chromatography with amperometric detection.

High-performance liquid chromatography with electrochemical detection has developed into a tool with excellent capability to monitor picomole amounts of individual metabolites in both tissue and body-fluid specimens. Significant technical advances in the area of microparticle chemically-bonded stationary phases have led to dramatic improvements in both sensitivity and resolution. Reversed-phase systems can be modified to include charged exchange sites by addition of detergents to the mobile phase. Adjustment of the surface charge in this manner permits capacity factors for ionic sample components to be increased or decreased. This concept is quite compatible with electrochemical detection and has provided the foundation for several clinical assays now in routine use. Examples of additional applications are presented for the determination of catecholamines in tissue, 3-methoxy-4-hydroxyphenylglycol in urine, and dopamine-beta-hydroxylase activity in human serum.

Determination of urinary normetanephrine, metanephrine, and 3-methoxytyramine by liquid chromatography, with amperometric detection.

We describe a sensitive procedure for measuring the basic catecholamine metabolites--normetanephrine, metanephrine, and 3-methoxytyramine--in urine by use of liquid chromatography, with electrochemical detection. These substances are isolated from hydrolyzed urine by passage through small ion-exchange columns and then pre-concentrated by a rapid set of solvent extractions before being injected onto the liquid-chromatographic column. Concentration and instrumental response are linearly related to 2.0 mg/liter for all three compounds, and practical lower detection limits are about 20 micrograms/liter for actual urine samples. The high selectivity that accrues from the combination of the extraction procedure, high-performance liquid chromatography, and electrochemical detection makes this procedure suitable for quantitative studies of catecholamine metabolism. A set of six samples can be taken through the extraction procedure in parallel in less than 2 h; once prepared, the extracts can be analyzed at a rate of three per hour.