Explore the vast range of titles available.

Azithromycin is recommended for the treatment of uncomplicated urogenital chlamydia infection although the standard 1gram dose sometimes fails to eradicate the infection (treatment failure). One hypothesis proposed for treatment failure has been insufficient levels of the antibiotic at the site of infection. We developed an assay using liquid chromatography and tandem mass spectrometry (LC-MS/MS) to measure azithromycin concentration in high-vaginal swabs and monitor how concentration changes over time following routine azithromycin treatment. Azithromycin concentrations were measured in two groups of women either within the first 24h of taking a 1g dose (N = 11) or over 9 days (N = 10). Azithromycin concentrations were normalised to an internal standard (leucine enkephalin), and the bulk lipid species phosphatidylcholine [PC(34:1)], using an Agilent 6490 triple quadrupole instrument in positive ionisation mode. The abundances of azithromycin, PC(34:1), and leu-enkephalin were determined by multiple reaction monitoring and absolute levels of azithromycin estimated using standard curves prepared on vaginal specimens. Vaginal azithromycin concentrations of women were rapidly obtained after 5h post-treatment (mean concentration = 1031mcg/mg of lipid, range = 173-2693mcg/mg). In women followed for 9 days, peak concentrations were highest after day 2 (mean concentration = 2206mcg/mg, range = 721-5791mcg/mg), and remained high for at least 9 days with a mean concentration of 384mcg/mg (range = 139-1024mcg/mg) on day 9. Our study confirmed that a single 1g dose of azithromycin is rapidly absorbed and remains in the vagina at relatively high levels for at least a week, suggesting that poor antibiotic absorption is unlikely to be an explanation for treatment failure.

Enkephalins are opioid peptides that modulate analgesia, reward, and stress. In vivo detection of enkephalins remains difficult due to transient and low endogenous concentrations and inherent sequence similarity. To begin to address this, we previously developed a system combining in vivo optogenetics with microdialysis and a highly sensitive mass spectrometry-based assay to measure opioid peptide release in freely moving rodents (Al-Hasani et al., 2018, eLife). Here, we show improved detection resolution and stabilization of enkephalin detection, which allowed us to investigate enkephalin release during acute stress. We present an analytical method for real-time, simultaneous detection of Met- and Leu-enkephalin (Met-Enk and Leu-Enk) in the mouse nucleus accumbens shell (NAcSh) after acute stress. We confirm that acute stress activates enkephalinergic neurons in the NAcSh using fiber photometry and that this leads to the release of Met- and Leu-Enk. We also demonstrate the dynamics of Met- and Leu-Enk release as well as how they correlate to one another in the ventral NAc shell, which was previously difficult due to the use of approaches that relied on mRNA transcript levels rather than posttranslational products. This approach increases spatiotemporal resolution, optimizes the detection of Met-Enk through methionine oxidation, and provides novel insight into the relationship between Met- and Leu-Enk following stress.

Enkephalins are endogenous opioid pentapeptides that play a role in neurotransmission and pain modulation in vertebrates. However, the distribution pattern of enkephalinergic neurons in the brains of reptiles has been understudied. This study reports the organization of the methionine-enkephalin (M-ENK) and leucine-enkephalin (L-ENK) neuronal systems in the central nervous system of the gecko Hemidactylus frenatus using an immunofluorescence labeling method. Although M-ENK and L-ENK-immunoreactive (ir) fibers extended throughout the pallial and subpallial subdivisions, including the olfactory bulbs, M-ENK and L-ENK-ir cells were found only in the dorsal septal nucleus. Enkephalinergic perikarya and fibers were highly concentrated in the periventricular and lateral preoptic areas, as well as in the anterior and lateral subdivisions of the hypothalamus, while enkephalinergic innervation was observed in the hypothalamic periventricular nucleus, infundibular recess nucleus and median eminence. The dense accumulation of enkephalinergic content was noticed in the pars distalis of the hypophysis. In the thalamus, the nucleus rotundus and the dorsolateral, medial, and medial posterior thalamic nuclei contained M-ENK and L-ENK-ir fibers, whereas clusters of M-ENK and L-ENK-ir neurons were observed in the pretectum, mesencephalon, and rhombencephalon. The enkephalinergic fibers were also seen in the area X around the central canal, as well as the dorsal and ventral horns. The widespread distribution of enkephalin-containing neurons within the central nervous system implies that enkephalins regulate a variety of functions in the gecko, including sensory, behavioral, hypophysiotropic, and neuroendocrine functions.

Intrauterine hypoxia (gestation days 15-19, pO 2 65 mm Hg, duration 4 h) led to an increase in the expression of p53, beclin-1, endothelial NO synthase (eNOS), and caspase-3 proteins in cardiomyocytes and reduced the number of mast cells in the heart of 60-day-old albino rats. Administration of a non-opiate analogue of leu-enkephalin (NALE peptide: Phe–D-Ala–Gly–Phe–Leu–Arg, 100 μg/kg) on days 2-6 of the neonatal period decreased the severity of delayed posthypoxic myocardial reaction. The content of eNOS + cardiomyocytes and the total number of mast cells of these animals did not differ from the control parameters; the content of p53 + cardiomyocytes was significantly lower than in animals exposed to intrauterine hypoxia. The cardioprotective activity of NALE was partially neutralized by co-administration with the NO synthase inhibitor (L-NAME, 50 mg/kg). Correction of the delayed posthypoxic changes, similar to the effects of NALE peptide, was observed after neonatal administration of its arginine-free analogue, G peptide (Phe–D-Ala–Gly–Phe–Leu–Gly; 100 μg/kg). Non-opiate analogues of leu-enkephalin NALE and G peptides can be considered as promising substances capable of preventing long-term cardiac consequences of intrauterine hypoxia.

The δ-opioid receptor (δOR) holds great potential as a therapeutic target. Yet, clinical drug development, which has focused on δOR agonists that mimic the potent and selective tool compound SNC80 have largely failed. It has increasingly become apparent that the SNC80 scaffold carries with it potent and efficacious β-arrestin recruitment. Here, we screened a relatively small (5120 molecules) physical drug library to identify δOR agonists that underrecruit β-arrestin, as it has been suggested that compounds that efficaciously recruit β-arrestin are proconvulsant. The screen identified a hit compound and further characterization using cellular binding and signaling assays revealed that this molecule (R995045, compound 1) exhibited ten-fold selectivity over µ- and κ-opioid receptors. Compound 1 represents a novel chemotype at the δOR. A subsequent characterization of fourteen analogs of compound 1, however did not identify a more potent δOR agonist. Computational modeling and in vitro characterization of compound 1 in the presence of the endogenous agonist leu-enkephalin suggest compound 1 may also bind allosterically and negatively modulate the potency of Leu-enkephalin to inhibit cAMP, acting as a ‘NAM-agonist’ in this assay. The potential physiological utility of such a class of compounds will need to be assessed in future in vivo assays.

The present investigation was conducted to elucidate the influence of an opioid peptide, leucine-enkephalin (L-ENK), on the reproductive axis of the tilapia Oreochromis mossambicus . In the first experiment, administration (i.p.) of 25, 100, and 300 μg L-ENK to the stripped female tilapia, for a period of 22 days, resulted in a significantly higher number of stage I follicles compared to those of initial controls and experimental controls, whereas the mean number of stage II and III follicles and serum levels of E 2 did not significantly differ among different experimental groups. A significant increase in the number of stage V (fully ripened) follicles was concomitant with significant reduction in the follicular diameter in 25 or 100 μg L-ENK-treated fish compared to those of experimental controls. However, significant reduction in the mean number and diameter of these follicles was observed in 300 μg L-ENK-treated fish compared to those of experimental controls and 25 or 100 μg L-ENK-treated fish. In the second experiment, the stimulatory effect of 25 μg L-ENK on the ovary was abolished in combination with gonadotropin-releasing hormone antagonist (GnRH-A). In conclusion, these results suggest that L-ENK exerts stimulatory as well as inhibitory effects on the ovary in a dose-dependent manner, and that these effects are possibly mediated through the GnRH, for the first time in fish.

Dipeptidyl peptides III (DPP III) is a dual-domain zinc exopeptidase that hydrolyzes peptides of varying sequence and size. Despite attempts to elucidate and narrow down the broad substrate-specificity of DPP III, there is no explanation as to why some of them, such as tynorphin (VVYPW), the truncated form of the endogenous heptapeptide spinorphin, are the slow-reacting substrates of DPP III compared to others, such as Leu-enkephalin. Using quantum molecular mechanics calculations followed by various molecular dynamics techniques, we describe for the first time the entire catalytic cycle of human DPP III, providing theoretical insight into the inhibitory mechanism of tynorphin. The chemical step of peptide bond hydrolysis and the substrate binding to the active site of the enzyme and release of the product were described for DPP III in complex with tynorphin and Leu-enkephalin and their products. We found that tynorphin is cleaved by the same reaction mechanism determined for Leu-enkephalin. More importantly, we showed that the product stabilization and regeneration of the enzyme, but not the nucleophilic attack of the catalytic water molecule and inversion at the nitrogen atom of the cleavable peptide bond, correspond to the rate-determining steps of the overall catalytic cycle of the enzyme.

Incubation of primary culture of pulmonary fibroblasts with non-opiate analogue of leuenkephalin (NALE; Phe-D-Ala-Gly-Phe-Leu-Arg, 0.1 μM) reduced generation of superoxide anion-radical (by 20.7%) and decreased the number of p53 + cells (by 40.2%) induced by exposure to H 2 O 2 (60 μM). The cytoprotective effect of NALE was potentiated by NO synthase inhibitor L-NAME (1 mM): the number of p53 + cells decreased by 65.3% and morphometric parameters of the cell nuclei and nucleoli were improved. Incubation of pulmonary fibroblasts culture with peptide G (Phe-D-Ala-Gly-Phe-Leu-Gly, 0.1 μM) also significantly reduced the damaging effect of H 2 O 2 : the number of p53 + cells decreased by 73.5%, the area of cell nuclei returned to normal, and generation of superoxide anion-radical decreased by 18.4%. These results indicate that C-terminal amino acid Arg and activation of NO synthase are not involved in the direct cytoprotective effect of NALE.

Hypobaric hypoxia (pO 2 65 mm Hg, duration 4 h) induced a significant increase in the number of cardiomyocytes expressing р53, beclin-1, endothelial NO synthase and accumulation and degranulation of mast cells in the epicardium in hearts of prepubertal female rats (age 45-47 days); the number of cardiomyocytes with nucleoli decreased, while the number of single-nucleolar cardiomyocytes increased after this exposure. Five-fold administration of non-opiate analogue of leu-enkephalin (NALE peptide: Phe–D-Ala–Gly–Phe–Leu–Arg; 100 μg/kg) during the neonatal period reduced the severity of the post-hypoxic changes in the heart. Neonatal administration of NALE (100 μg/kg) against the background of NO synthase blockade with L-NAME (50 mg/kg) did not abolish the cardioprotective effects of the peptide. A similar correction of posthypoxic changes in the heart was observed after neonatal administration of original peptide G (Phe–D-Ala–Gly–Phe–Leu–Gly; 100 μg/kg). Thus, NO synthase—NO system and C-terminal amino acid Arg in the molecule of non-opiate analogue of leu-enkephalin are not required for the cardioprotective effects of peptides. Non-opiate leu-enkephalin analogs, peptides NALE and G, can be considered as promising substances for increasing heart resistance to hypoxia during later age periods.

In white rats with experimental hypothyroidism, changes in the myeloid compartment of the blood system induced by 6-h immobilization stress and the corrective effect of the analogue of leu-enkephalin (dalargin) on these shifts was analyzed. It was found that in rats with hypothyroidism, stress in the anxiety stage did not cause leukocytosis typical of euthyroid animals, but at the stage of resistance provoked leukopenia at the expense of eosinopenia and neutropenia with depletion of the intramedullary reserve. Dalargin increased white blood cells count, neutrophil count, and the intramedullary depot of these cells.