Early alterations of actin cytoskeleton in OK cells by opioids.
Recently we identified and characterized opioid binding sites in OK (opossum kidney) cells and observed decreased proliferation of these cells in response to opioids. In the present study we investigated the effects of opioids on the actin cytoskeleton and explored whether their antiproliferative action may relate to alterations in the distribution or the dynamics of actin microfilaments. Exposure of OK cells to the opioids alphaS1 casomorphin and ethylketocyclazocine resulted in a rapid and substantial actin microfilament reorganization. This was documented by a significant dose-dependent decrease in the amounts of F-actin, determined by measurements of quantitative fluorescence, by immunoblot analysis and by a concomitant increase of the G/total-actin ratio measured by the DNase I inhibition assay. These changes were verified by confocal laser scanning microscopy, which showed marked redistribution of the microfilamentous structures in the presence of the opioids without affecting the organization of microtubules or vimentin intermediate filaments. The effect of opioids on actin polymerization dynamics occurred within 15 min and persisted for at least 2 h, while their restoration to control levels was accomplished 6 h later, indicating a reversible phenomenon. Northern blot analysis showed that the concentration of the actin transcript was unaffected. The addition of diprenorphine, a general opioid antagonist, prevented the effects of opioids on the actin cytoskeleton. The inhibition of OK cell proliferation, induced by ethylketocyclazocine and alphaS1 casomorphin was partially prevented in the presence of phallacidin, which stabilizes microfilaments. Our findings demonstrate that opioids, acting via kappa 1 binding sites, induce rapidly modifications in the dynamics of actin polymerization, and in the organization of microfilaments in OK cells, which may relate to their antiproliferative effect on these cells.
Papakonstanti
E A
EA
Department of Biochemistry, University of Crete, School of Medicine, Heraklion, Greece.
Bakogeorgou
E
E
Castanas
E
E
Emmanouel
D S
DS
Hartig
R
R
Stournaras
C
C
D000199:Actins / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D002460:Cell Line; D003599:Cytoskeleton / Q000187:drug effects* / Q000378:metabolism; D007382:Intermediate Filaments / Q000187:drug effects; D007668:Kidney / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D018613:Microscopy, Confocal; D008870:Microtubules / Q000187:drug effects; D009294:Narcotics / Q000494:pharmacology*; D009893:Opossums; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism
D000199:Actins; D009294:Narcotics; D012333:RNA, Messenger