Control of myocardial respiration changes during postnatal development. ADP loses prominence as a stimulator of mitochondrial oxidative phosphorylation as efficency of ADP/ATP exchange increases. Changes in the respiratory control pattern to non-ADP regulation after birth are paralleled by increases in both mRNA and protein expression for the adenine nucletoide translocator (ANT). We tested the hypothesis that mitochondrial maturation is regulated by the postnatal surge in thyroid hormone. Methods: Newborn sheep(age<36hrs; n = 5) underwent thyroidectomy(TH), while preserving parathyoid function. Marked decreases in thyroid hormones were documented. At 30 days TH and control sheep (C), were anesthetized,open-chested, and had coronary sinus shunts placed for measurement of coronary flow and myocardial oxygen consumption (MVO). Using a previously published protocol, we stimulated stimulated MVO with epinephrine, while 31P MRS monitored high energy phosphates. Myocardial biopsies were obtained for Northern and Western analyses for ANT, and the β-F1-ATPase (mitochondrial). Results: Baseline MVO was lower (P<0.05) in TH(1.0±.07μmol/gm/min) than C((2.4±.09). Despite 3-4 fold higher epinephrine doses, MVO in TH(2.6±.3), remained well below C(4.9±0.15). The ΔMVO in C was not accompanied by change in PCr/ATP or calculated ADP. However, a two-fold increase in calculated ADP accompanied the much smaller ΔMVO in TH. Steadystate mRNA levels were near twofold higher for ANT(heart/muscle isoform 1) in C, although there were no differences for β-F1-ATPase. Similarly, ANT protein levels by Western analysis were greater than two-fold higher in C, with no difference in β-F1-ATPase. Conclusions: Thyroid specifically regulates coordinate ANT mRNA and protein expression postnatally. Mitochondrial ATPase does not appear to be closely regulated by thyroid. ANT deficit due to lack of thyroid hormone promotion reduces efficiency of ADP/ATP exchange at the mitochondrial membrane. This creates a bottleneck phenomenon as ADP accumulates in the cytosol awaiting transport into the mitochondria in exchange for ATP. Though, cytosolic ADP appears to stimulate respiration at the mitochondrial membrane, the oxidative capacity is limited by poor ADP availability within the mitochondria. Thus, these studies highlight the importance of thyroid in regulation of mitochondrial respiration and the adenine nucleotide translocator.