Presence and localization of connexins 43 and 26 in cell cultures derived from myometrial tissues from nonpregnant and pregnant women and from leiomyomas.

OBJECTIVE: Our objective was to study the appearance and distribution of connexins 43 and 26 in various human myometrial cell cultures. STUDY DESIGN: Scrape loading, Western blotting, and immunohistochemical techniques were applied to cultured cells derived from myometrial tissues obtained from nonpregnant and pregnant women (upper and lower uterine segments) and from leiomyomas (tumor and analogous myometrial tissues). RESULTS: Scrape loading revealed the presence of metabolic coupling in all tissues. Indirect immunohistochemical studies showed membrane localization of connexin 43 in all myometrial cultures. Western blots and indirect immunohistochemical studies showed the presence and localization of the connexin 26 protein and associated gap junctions in tissues from myomas and from nonpregnant and pregnant women except for those derived from the upper segment of the pregnant uterus. CONCLUSION: These results show that human myometrial cultures express various gap junction proteins and that there are regional differences in expression of connexins in tissues from pregnant women.

Ineffectiveness of oxytocin on intercellular communication between term pregnant human myometrial cells before labor.

OBJECTIVE: The effects of the uterotonic agent oxytocin on intercellular communication between term pregnant human myometrial cells before labor were studied to investigate its contribution to the synchronicity of uterine contractions. STUDY DESIGN: The membrane potential and the input resistance of myometrial cells were measured with intracellular micropipettes, and dye-coupling assays were concomitantly performed while the tissues were perfused with three concentrations of oxytocin (10(-8) mol/L, 10(-7) mol/L, and 10(-6) mol/L). The results were compared with those obtained from tissues perfused with gap junctional uncoupling agent octanol and those with Tyrode's solution (control). RESULTS: Octanol increased input resistance and inhibited dye coupling but did not affect membrane potential compared with controls. Oxytocin depolarized membrane potential at all studied concentrations. The mean input resistance values and detectable dye propagation with oxytocin were similar to controls. CONCLUSION: Oxytocin does not contribute to gap junctional intercellular communication in term pregnant human myometrium before labor but may affect contractions by depolarizing membrane potential.

Steroid hormone effects on intercellular communication between term pregnant human myometrial cells before labor.

The appearance of gap junctions (GJs) between myometrial smooth muscle cells is one of the major events associated with the onset of labor. We have employed dye-coupling and electrical-current injection techniques to study the mechanisms by which steroid hormones regulate GJs in term pregnant myometrium of women before labor. Progesterone (P4) did not alter the input resistance (Ro) of the tissues when added to Tyrode's solution, which was used as control treatment. Octanol, the putative gap junctional uncoupling agent, increased the Ro of the cells compared to the control and P4-treated groups. The membrane potential (Em) did not differ between these groups. However, when P4 was applied after the tissue was perfused with estradiol (E2), the results changed dramatically: the Em hyperpolarized, and the Ro increased. Octanol increased the Ro in E2-treated tissues, but did not affect the Em. Consecutive application of E2, octanol, E2, and P4 resulted in rapid changes in the Ro of the cells. Dye-coupling was mostly detected between cells from controls and E2-treated tissues. These results indicate that P4 exerts its effects in the presence of E2 and that P4 has rapid effects on the intercellular communication between human myometrial cells.

Morphometric analysis of gap junctions in nonpregnant and term pregnant human myometrium.

BACKGROUND AND METHODS: Some structures in the human myometrium, namely; caveolae, extracellular space and gap junctions were quantified in tissues from nonpregnant women, women who were not in labor, and women in spontaneous labor or oxytocin-induced labor, in order to reveal morphologic differences that would contribute to explaining the events leading to labor. Transmission electron micrographs taken from tissues from twenty-four pregnant and ten nonpregnant women were studied. RESULTS: Differences were observed in cell growth which was accompanied by enlargement of the extracellular space, and in gap junctions. Gap junctions were demonstrated in nonpregnant women, and even in one postmenopausal woman. Their frequency and size increased gradually between groups of pregnant women in the order of non-labor, oxytocin-induced labor and spontaneous labor. The increase in size of the junctions was less than the increase in number. CONCLUSIONS: Gap junctions are most frequently found in human myometrial cells in spontaneous labor, which strongly suggests a role for these structures in the termination of pregnancy. The effect of oxytocin on the appearance of gap junctions was negligible; hence, the contribution of oxytocin to establish myometrial contractile synchronicity is thought to be minor. Caveolae are probably not relevant for the initiation of labor as their distribution did not differ between the groups.

Dye-coupling between term pregnant human myometrial cells before labor: carboxyfluorescein versus lucifer yellow.

Term pregnant human myometrial cells in whole mounts were microinjected by pressure with the fluorescent probes Lucifer Yellow and carboxyfluorescein. Tissues obtained from acute and elective sections displayed weak dye-coupling when injected with Lucifer Yellow. Injection of carboxyfluorescein into cells from the elective sections resulted in a more extensive dye-coupling than that observed with Lucifer Yellow. These results indicate that term pregnant human myometrial cells are metabolically coupled before labor and carboxyfluorescein is superior to Lucifer Yellow in detecting the coupling.

Direct intracellular injections for studying human myometrial gap junctions prior to labor.

Direct intracellular microinjection of a fluorescent dye (Lucifer Yellow) was performed on ex-situ muscle strips from term pregnant women not in labor. The aim was to characterize the intracellular distribution of LY to obtain criteria for an intracellular injection in whole mounts and ultimately to study the gap junctional communication between myometrial cells in those tissues. Fifteen injections performed on biopsies from ten cases showed a well-bordered fusiform shape and were considered to be intracellular. The intercellular spread of the dye into an adjacent cell was observed in three injections (three cases). The average cell dimension was 284 +/- 95 microns for length and 5 +/- 1.5 microns for width (n = 17). The intracellular injection was confirmed by light microscopy of cross sections. It is concluded that limited coupling exists between myometrial cells of women prior to labor.