Mitochondrial function and mitochondria-induced apoptosis in an overstimulated rat ovarian cycle

1 Department of Biochemistry and Molecular Biology and 2 Department of Obstetrics and Gynecology, University Hospital of Puerto Real; 3 Department of Cell Biology and Histology, School of Medicine, University of Cádiz, Cadiz, Spain; and 4 School of Pharmacy and Biochemistry, University of Buenos Aires, Buenos Aires, Argentina Submitted 16 May 2005 ; accepted in final form 3 July 2005 Female rats were treated with FSH (40 IU/kg) on the first and second diestrus days (D1 and D2) and with LH (40 IU/kg) on the proestrus (P) day to synchronize and maximize ovarian changes. Follicle area increased by 50% from D1 to P, and the estrus (E) phase showed multiple corpora lutea and massive apoptosis. Increased oxygen uptakes (42–102%) were determined in ovary slices and in isolated mitochondria in active state 3 along the proliferation phase (D1-D2-P) that returned to initial values in the E phase. Mitochondrial content and the electron transfer activities of complexes I and IV were also maximal in the P phase (20–79% higher than in D1). Production of NO by mitochondrial nitric oxide synthase (mtNOS), biochemically determined, and the mtNOS functional activity in regulating state 3 oxygen uptake were also maximal at P and 79–88% higher than at D1. The moderately increased rate of NO in the proliferative phase is associated with mitochondrial biogenesis, whereas the high rate of NO generation by mtNOS at phase P appears to trigger mitochondria-dependent apoptosis. The calculated fraction of ovary mitochondria in state 3 was at a minimal value at the P phase. Mitochondrial oxidative damage, with increased thiobarbituric acid-reactive substances and protein carbonyls, indicates progressive mitochondrial dysfunction between phases P and E. The roles of mitochondria as ATP provider, as a source of NO to signal for mitochondrial proliferation and mitochondria-dependent apoptosis, and as a source of O 2 – and H 2 O 2 appear well adapted to serve the proliferation-apoptosis sequence of the ovarian cycle. ovarian follicle; oxygen uptake; mitochondrial nitric oxide synthase; respiratory chain; oxidative damage Address for reprint requests and other correspondence: A. Navarro, Departamento de Bioquímica y Biología Molecular, Facultad de Medicina, Plaza Fragela 9, 11003 Cádiz, Spain (e-mail: ana.navarro{at}uca.es )