Studies were conducted to determine relationships between feed efficiency and mitochondrial function and biochemistry. After feed efficiency (FE; gain:feed) was determined in broiler breeder males between 6 and 7 wk of age, mitochondria were isolated from breast and leg muscle from birds with high FE (0.83+/-0.01, n = 6) and low FE (0.64+/-0.01, n = 7). Respiratory chain coupling, assessed by the respiratory control ratio (RCR), was greater in high FE breast, and leg mitochondria provided NADH-linked, but not FADH-linked, energy substrates. There were no differences, however, in the adenosine diphosphate to oxygen (ADP:O) ratio (an index of oxidative phosphorylation) when mitochondria were provided either energy substrate. Electron leak, as determined by generation of H202, was greater in the low FE than in high FE breast mitochondria. Electron leak increased following inhibition of electron transport at Complex I (with rotenone) and Complex III (with antimycin A) in low FE but not in high FE breast mitochondria. There were no differences in basal electron leak in leg mitochondria between groups, but H202 generation was elevated (P < 0.07) compared to basal values in low FE leg mitochondria after Complex I inhibition. The activities of Complexes I and II were greater in high FE breast and leg muscle mitochondria compared to those in low FE mitochondria. The results indicate that lower respiratory chain coupling in low FE muscle mitochondria may be due to lower activities of Complexes I and II and defects in electron leak and provide insight into cellular mechanisms associated with the phenotypic expression of feed efficiency in broilers.

Duodenal mitochondria were isolated from broiler breeder males with high (0.79+/-0.01, n = 9) and low (0.63+/-0.02, n = 9) feed efficiency (FE) to assess relationships of FE with duodenal mitochondrial function and site-specific defects in electron transport. Sequential additions of adenosine diphosphate (ADP) resulted in 1) higher respiratory control ratio (RCR; an index of respiratory chain coupling) in high FE mitochondria provided succinate, and 2) higher ADP to oxygen ratio (ADP:O; an index of oxidative phosphorylation) in low FE mitochondria provided NADH-linked substrates (malate, pyruvate, or both). Basal electron leak, measured as H2O2 production, was greater in low FE mitochondria provided succinate (P = 0.08) or NADH-linked substrates. As H2O2 levels were elevated in low FE compared with high FE mitochondria by complex I (P+/-0.07) and complex II inhibition, the higher basal electron leak in low FE mitochondria was apparently due to site-specific defects in electron transport at complexes I and II. Elevations in H2O2 above basal levels indicated that high FE mitochondria may also exhibit electron transport defects at complexes I and III. Despite an ability to produce adenosine triphosphate (ATP) that was equal or superior to that demonstrated in high FE duodenal mitochondria, low FE mitochondria exhibited a greater inherent degree of electron leak. The results provide insight into the role that duodenal mitochondria play in the phenotypic expression of FE in broilers.

1-methyl-4-phenyl-1,2,4,6,-tetrahydropyridine (MPTP) is a selective neurotoxin that produces striatal dopamine depletion resulting in parkinsonism like symptoms in humans and is, therefore, used to generate animal models for Parkinson's disease (PD). In this study, C57BL/6N mice were treated with MPTP acutely (3x20 mg/kg, 2-hour interval, one day injection). Mice were then sacrificed 24 hours after the last injection and brain tissue was collected for analysis. Significant decrease of striatal dopamine (DA) and the metabolites (DOPAC, HVA) was observed after MPTP treatment. MPTP also reduced protein expression of tyrosine hydroxylase (TH) in the striatum. Real time RT-PCR was used to examine selective genes of the dopaminergic system in the substantia nigra. Our data demonstrated that MPTP significantly decreased gene expression of TH, dopamine transporter (DAT), and vesicle monoamine transporter (VMAT), coinciding with the pattern of dopamine concentration changes and protein expression after MPTP treatment. Although a significant decrease of DA metabolites was observed in striatum, there was no change in the expression of monoamine oxidases (MAO-A, MAO-B) or catechol O-methyltransferase (COMT), indicating that these changes might be simply a consequence of reduced monoamine levels. In addition, gene expression of alpha-synuclein was also decreased with MPTP treatment, but there was no change in beta-synuclein and parkin. This is the first study using real-time PCR to indicate that MPTP selectively alters gene expression and provides information for clinical studies in PD. Future studies will focus on gene expression of other pathways that may be affected by MPTP treatment and investigation of gene expression in specific cell types in vivo using LCM technology.

By using a series of chemical inhibitors of mitochondrial respiration, a site-specific defect in the electron transport chain was identified in mitochondria obtained from broilers with pulmonary hypertension syndrome (PHS; ascites). Located at the succinate:ubiquinone oxido-reductase (Complex II:CoQ) interface, this defect would allow electrons to leak from the respiratory chain and consume oxygen by forming reactive oxygen species at a greater rate than in control mitochondria. Lower levels of the primary antioxidants, alpha- and beta-tocopherol, and glutathione (GSH) in PHS mitochondria confirmed the presence of oxidative stress. Respiration studies of PHS liver mitochondria also revealed disease-associated decreases in the respiratory control ratio (RCR, an index of electron transport chain coupling). Differences in the RCR as well as the adenosine diphosphate (ADP) to O ratio (an index of oxidative phosphorylation) between control and PHS mitochondria were accentuated by sequential additions of ADP to isolated mitochondria. In a second experiment, similar improvements in functional indices following sequential additions of ADP and responses to respiratory chain inhibitors were observed in liver mitochondria isolated from Single Comb White Leghorn (SCWL) males (resistant to PHS) similar to that observed in control broiler mitochondria in Experiment 1. The combined results indicate the presence of a site-specific defect at either Complex II, ubiquinone, or both in liver mitochondria obtained from broilers with PHS that may be responsible for the oxidative stress and mitochondrial dysfunction observed in this costly metabolic disease.

The purpose of this study was to assess mitochondrial function and glutathione (a mitochondrial antioxidant) in response to oxidative stress in mitochondria in vitro obtained from broilers with and without pulmonary hypertension syndrome (PHS). Liver mitochondria from Control and PHS broilers were incubated with 0, 1, and 5-mM tertiary-butyl hydroperoxide (tBH). Indices of mitochondrial function [the respiratory control ratio (RCR) and the adenosine diphosphate to oxygen ratio (ADP:O)], and levels of mitochondrial and extra-mitochondrial reduced (GSH) and oxidized (GSSG) glutathione, cysteine, cystine, glutamate and cysteinyl-glycine were determined following tBH treatment. Lower RCR and ADP:O values were observed in PHS mitochondria than in controls. Whereas control mitochondria remained coupled (RCR > 2.0), only 3 PHS preparations remained coupled after 60 min of incubation with 5 mM tBH, indicating a greater susceptibility to oxidative stress in PHS mitochondria. The lower RCR in PHS mitochondria was due to increased oxygen consumption during State IV respiration. Oxidative stress following tBH treatment (decreased GSH and increased GSSG) was observed, but there were no differences in GSH or GSSG between control and PHS mitochondria. The PHS mitochondria did exhibit elevated mitochondrial and extramitochondrial cystine than controls, however. The results indicate that PHS mitochondria do not lack antioxidant protection from GSH, but lower RCR and ADP:O ratios in PHS mitochondria indicate a dysfunction that may contribute to the pathophysiology of this metabolic disease in broilers.