Umbilical cord blood coagulability, acidosis and intracranial hemorrhage

Peer Reviewe

Generation of an inducible double-stable cell line expressing MOP-YFP and c-myc-5-HT_2C-Cerulean receptors.

<p>Images obtained by confocal microscopy from living Flp-In HEK293 cells expressing permanently MOP-YFP receptors and c-myc-5-HT_2C-Cerulean receptors in an inducible manner. The right column corresponds to the images resulting after exciting the cells with the YFP light settings whereas in the left column are the same microscopy field illuminated for CFP visualization. When indicated, +DOX corresponds to treatment with doxycycline (0.01 μg/ml) for 24 hours prior to microscope observation.</p

Effect of different doses of DAMGO on the time-course generation of endocytic vesicles containing MOP-YFP receptors.

<p><b>A.</b> Data graph show representative results from time-course MOP-YFP receptor endocytosis experiments performed with DAMGO at different concentrations (see symbol legends). <b>B.</b> Dose-response curve obtained after non-linear analysis of data corresponding to the maximal effect in terms of number of vesicles per cells generated by different concentrations of DAMGO. Each point represents the mean ± SEM of three independent experiments.</p

Kinetic parameters obtained from endocytosis assays performed with cells heterologously co-expressing MOP-YFP and c-myc-5-HT2C-Cerulean receptors.

<p>Cells were treated with DAMGO (10 μM), sufentanyl (1 μM), methadone (10 μM), morphine (10 μM), morphine (10 μM) plus 5-HT (10 μM), morphine (10 μM) plus DOI (10 μM), morphine (10 μM) plus WAY161503 (1 μM), oxycodone (10 μM) plus 5-HT (10 μM) or buprenorphine (10 μM) plus 5-HT (10 μM). Data indicate mean ± SEM of (n) independent experiments. t_1/2: time to reach the 50% of the maximal response (minutes). Emax: maximal response (number of vesicles per cell). n.d.: not detected.</p><p>Kinetic parameters obtained from endocytosis assays performed with cells heterologously co-expressing MOP-YFP and c-myc-5-HT2C-Cerulean receptors.</p

MOP-YFP receptor endocytosis studies in living cells by confocal microscopy.

<p><b>Upper panels:</b> Confocal microscopy images from Flp-In HEK293 cells permanently expressing MOP-YFP receptors without induction of c-myc-5-HT_2C-Cerulean receptor expression. Cells were treated with either DAMGO (10 μM) (left panel) or morphine (10 μM) (right panel) for 30 minutes before picture acquisition. <b>Lower panels:</b> Images from cells treated with doxycycline (0.01 μg/ml) for 24 hours in order to express c-myc-5-HT_2C-Cerulean receptors. Cells were treated with either DAMGO (10 μM), morphine (10 μM), 5-HT (10 μM) or morphine plus 5-HT for 30 minutes before picture acquisition.</p

Effect of different doses of serotonin (5-HT) on the facilitation of the endocytosis of MOP-YFP receptors by morphine.

<p><b>A.</b> Data graph display representative results from time-course MOP-YFP receptor endocytosis experiments conducted by co-treatment with morphine at 10 μM plus 5-HT at different concentrations (see symbol legends). <b>B.</b> Dose-response curve (continuous line) obtained after non-linear analysis of the maximal effect (number of vesicles per cell) generated by morphine (10 μM) plus different concentrations of 5-HT. Each point represents the mean ± SEM of three independent experiments. Dotted line display the characteristic bell shaped profile observed when connecting these points in an increasing dose-dependent manner.</p

Kinetic parameters obtained from endocytosis assays performed with HeLa cells transiently expressing MOP-YFP receptors.

<p>Cells were treated with DAMGO (10 μM), sufentanyl (1 μM) or morphine (10 μM) for 15 minutes. Data indicate mean ± SEM of (n) independent experiments. t_1/2: time to reach the 50% of the maximal response (minutes). Emax: maximal response (number of vesicles per cell). n.d.: not detected.</p><p>Kinetic parameters obtained from endocytosis assays performed with HeLa cells transiently expressing MOP-YFP receptors.</p