Role of protein kinase C in estrogen protection against apoptotic cerebellar cell death in ethanol-withdrawn rats.
Results of studies from our laboratory have shown that administration of 17beta-estradiol (E(2)) reduces cerebellar neuronal damage during ethanol withdrawal (EW). In the current study, we examined mechanisms underlying E(2) protection against EW-associated cerebellar damage by assessing apoptotic indicators: DNA fragmentation, caspase-3 activity, and protein kinase C (PKC) activity. Ovariectomized rats, implanted with E(2) or oil pellets, received ethanol [7.5% weight/volume (wt./vol.)] (EW/E(2) group and EW/Oil group, respectively) chronically (for 5 weeks) or control dextrin diet (Dextrin/Oil group). At day 14 of EW, cerebelli were collected for the terminal deoxynucleotidyltransferase (TdT)-mediated dUDP-biotin nick end labeling (TUNEL) assay to detect DNA fragmentation and for immunohistochemistry to detect caspase-3 activation. A separate group of rat cerebelli was prepared to assess for total PKC activity, as well as for activity of a specific PKC isozyme, epsilon (PKCepsilon), by using an in vitro [gamma-(32)P]ATP phosphorylation assay at days 1 and 14 of EW. Results indicated that rats in the EW/Oil group had more DNA fragments and caspase-3-positive neuronal cells than observed for control rats, and these effects were inhibited by E(2) treatment. For total PKC activity at day 1 of EW, rats in the EW/E(2) group had a lower cytosolic PKC activity than observed for either rats in the EW/Oil group or control rats. At day 14 of EW, both EW groups had a lower total PKC activity than observed for control rats. For PKCepsilon activity, rats in the EW/E(2) group had a lower cytosolic PKCepsilon activity than observed for rats in the EW/Oil group or for control rats at day 1, and they had a lower membrane PKCepsilon activity at day 14 of EW than observed for control rats. These findings support the suggestion that E(2) protects against cerebellar neuronal damage in ethanol-withdrawn rats by inhibition of DNA fragmentation and caspase-3 activation, and that reduced PKC activity may be involved in the protection.
Jung
Marianna E
ME
Department of Pharmacology and Neuroscience, University of North Texas HSC at Fort Worth, 3500 Camp Bowie Boulevard, Fort Worth, TX 76107-2699, USA. mjung@hsc.unt.edu
Watson
David G
DG
Wen
Yi
Y
Simpkins
James W
JW
D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology; D016923:Cell Death / Q000187:drug effects / Q000502:physiology; D002531:Cerebellum / Q000187:drug effects* / Q000201:enzymology / Q000473:pathology; D004958:Estradiol / Q000502:physiology* / Q000627:therapeutic use; D000431:Ethanol / Q000494:pharmacology*; D005260:Female; D011493:Protein Kinase C / Q000502:physiology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013375:Substance Withdrawal Syndrome / Q000201:enzymology* / Q000473:pathology / Q000517:prevention & control
D000431:Ethanol; D004958:Estradiol; D011493:Protein Kinase C