TY - JOUR
T1 - 17β-estradiol reduces the effect of metabolic inhibition on gap junction intercellular communication in rat cardiomyocytes via the estrogen receptor
AU - Chung, Tun Hui
AU - Wang, Seu Mei
AU - Wu, Jiahn Chun
N1 - Funding Information:
This work was supported by National Science Council of the Republic of China grants NSC-90-2314-B002-336 and NSC-91-2314-B002-131.
PY - 2004/11
Y1 - 2004/11
N2 - The effects of 17β-estradiol (E2) on gap junction intercellular communication (GJIC) were assessed by Lucifer yellow dye coupling in cultured neonatal rat cardiomyocytes after metabolic inhibition (MI) using potassium cyanide and sodium iodoacetate. MI significantly reduced dye coupling of cardiomyocytes to 8.5% ± 0.6% of control levels, and pretreatment with E2, but not its inactive isomer 17α-estradiol, dose-dependently (EC 50 = 0.41 μM) increased the dye coupling up to 76% ± 15% of control levels. The effect of E2 on MI-induced dye uncoupling was abolished by tamoxifen, a potent estrogen receptor (ER) antagonist. The ligand, E2-BSA-FITC, labeled the cardiomyocyte surface, whereas BSA-FITC did not, suggesting the presence of membrane-associated E2 receptors. Double immunofluorescence microscopy showed that MI-induced the accumulation of non-phosphorylated Cx43 at the gap junction and that this was prevented by E2 pretreatment. Labeling of Lucifer yellow-microinjected cardiomyocytes with antibodies specific for Ser368-phosphorylated Cx43 (Ser368Cx43) or non-phosphorylated Cx43 confirmed that E2 reduced the MI-induced inhibition of dye coupling and accumulation of non-phosphorylated Cx43 concomitant with the reappearance of Ser368Cx43 at the gap junction. MI caused a decrease in Ser368Cx43 protein levels, and pretreatment with E2 significantly increased the levels of Ser368Cx43. Inhibition of protein kinase C (PKC) with chelerythrine blocked the E2-induced increase of Ser368Cx43 levels in MI-treated cardiomyocytes. These results suggest that E2 attenuates the inhibitory effect of MI on GJIC in cardiomyocytes by affecting the phosphorylation of Cx43, possibly mediated by activation of PKC via a membrane-associated signaling mechanism.
AB - The effects of 17β-estradiol (E2) on gap junction intercellular communication (GJIC) were assessed by Lucifer yellow dye coupling in cultured neonatal rat cardiomyocytes after metabolic inhibition (MI) using potassium cyanide and sodium iodoacetate. MI significantly reduced dye coupling of cardiomyocytes to 8.5% ± 0.6% of control levels, and pretreatment with E2, but not its inactive isomer 17α-estradiol, dose-dependently (EC 50 = 0.41 μM) increased the dye coupling up to 76% ± 15% of control levels. The effect of E2 on MI-induced dye uncoupling was abolished by tamoxifen, a potent estrogen receptor (ER) antagonist. The ligand, E2-BSA-FITC, labeled the cardiomyocyte surface, whereas BSA-FITC did not, suggesting the presence of membrane-associated E2 receptors. Double immunofluorescence microscopy showed that MI-induced the accumulation of non-phosphorylated Cx43 at the gap junction and that this was prevented by E2 pretreatment. Labeling of Lucifer yellow-microinjected cardiomyocytes with antibodies specific for Ser368-phosphorylated Cx43 (Ser368Cx43) or non-phosphorylated Cx43 confirmed that E2 reduced the MI-induced inhibition of dye coupling and accumulation of non-phosphorylated Cx43 concomitant with the reappearance of Ser368Cx43 at the gap junction. MI caused a decrease in Ser368Cx43 protein levels, and pretreatment with E2 significantly increased the levels of Ser368Cx43. Inhibition of protein kinase C (PKC) with chelerythrine blocked the E2-induced increase of Ser368Cx43 levels in MI-treated cardiomyocytes. These results suggest that E2 attenuates the inhibitory effect of MI on GJIC in cardiomyocytes by affecting the phosphorylation of Cx43, possibly mediated by activation of PKC via a membrane-associated signaling mechanism.
KW - Connexin43
KW - Membrane estrogen receptor
KW - Metabolic inhibition
KW - Non-genomic effect
KW - Serine368 phosphorylation
KW - Signaling pathway
UR - http://www.scopus.com/inward/record.url?scp=7444226471&partnerID=8YFLogxK
U2 - 10.1016/j.yjmcc.2004.08.003
DO - 10.1016/j.yjmcc.2004.08.003
M3 - Article
C2 - 15522278
AN - SCOPUS:7444226471
SN - 0022-2828
VL - 37
SP - 1013
EP - 1022
JO - Journal of Molecular and Cellular Cardiology
JF - Journal of Molecular and Cellular Cardiology
IS - 5
ER -