title: Role of mitochondrial cytochrome c in cocaine-induced apoptosis in rat testes.
abstract: OBJECTIVE
  We have previously demonstrated that cocaine exposure leads to apoptosis in rat testes. To understand further the mechanism of cocaine-induced testicular damage, we studied the effect of cocaine on cytochrome c release from the mitochondria. We also determined the caspase 3, caspase 8, and caspase 9 activities in rat testes after chronic cocaine exposure.
  
  
  METHODS
  Thirty-day-old male Sprague-Dawley rats received cocaine hydrochloride or equal volumes of normal saline subcutaneously daily for 90 days. The testes were removed at 15, 30, and 90 days of cocaine or saline administration. Mitochondria and cytosolic fractions from testes were isolated. Western blotting was performed in both fractions using anti-cytochrome c antibody. Caspase 3, caspase 8, and caspase 9 activities were determined by fluorometric assay.
  
  
  RESULTS
  The expression of cytochrome c protein in the cytosolic fraction was increased on day 15 and persisted for up to 90 days after cocaine injection compared with controls. However, the expression of cytochrome c in testes was decreased in the mitochondria fraction on days 15, 30, and 90 after cocaine injections compared with the corresponding controls. The caspase activity study showed caspase 3 and caspase 9 activities increased in cocaine-treated testes at each point of the study compared with the corresponding controls. However, the caspase 8 activity in cocaine-treated testes did not change significantly at each point of the study compared with the corresponding controls.
  
  
  CONCLUSIONS
  Our results suggest that the release of cytochrome c from mitochondria and its subsequent activation of caspase 9 and caspase 3 in testes play a key role in cocaine-induced germ cell apoptosis. Our findings also indicate that cocaine-induced testicular germ cell apoptosis in rats is at least initiated through a mitochondria-associated pathway.
authors.affiliation: Department of Urology, Wayne State University School of Medicine, Detroit, Michigan 48201, USA.
authors.forename: Haikun
authors.initials: H
authors.lastname: Li
authors.forename: Liping
authors.initials: L
authors.lastname: Xu
authors.forename: Joseph C
authors.initials: JC
authors.lastname: Dunbar
authors.forename: C B
authors.initials: CB
authors.lastname: Dhabuwala
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases / Q000378:metabolism; D003042:Cocaine / Q000494:pharmacology*; D003574:Cytochrome c Group / Q000187:drug effects / Q000378:metabolism*; D004789:Enzyme Activation / Q000187:drug effects; D005470:Fluorometry; D008297:Male; D008928:Mitochondria / Q000187:drug effects* / Q000201:enzymology* / Q000378:metabolism; D023421:Models, Animal; D051381:Rats; D017207:Rats, Sprague-Dawley; D013094:Spermatozoa / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism; D013737:Testis / Q000187:drug effects* / Q000201:enzymology* / Q000378:metabolism
chemical_list: D003574:Cytochrome c Group; C505410:Casp3 protein, rat; C505528:Casp8 protein, rat; C506435:Casp9 protein, rat; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases; D003042:Cocaine