Studies On Mitochondrial Energy Metabolism And On Adenine Nucleotide Translocator (ANT) Activity And Expression Of Rars Cardium During Hypoxic Exposure

To understand the mechanism of disorder in heart energy metabolism and the contributing role of ANT in mitochondrial respiratory function during rats exposed to hypoxia , we observed the changing aspects of mitochondrial oxidative phosphorylation function and the content of adenine nucleotide and the change of it's transport activity and it's gene expression during hypoxic exposure.Adult male wistar rats were exposed to simulated high altitude at 5000m for 0(control),1(1d),5(5d),15(15d)and 30days(30d) in hypobaric chamber. All animals were sacrificed by decaptation under normoxic(control)and hypoxic(other groups)condition respectively. Rats heart ventricle was removed and mitochondria were isolated by centrifugation. Mitochondria respiratory function was measured by Clark oxygen electrode. The size of adenine acid pool(ATP,ADP,AMP)in muscle or mitochondria were separated and measured by high performance liquid chromatography(HPLC). mRNA copies of ANT1,ANT2 in tissues were determined by FQ-RT-PCR. The protein content of ANT in mitochondria was detected by western blot analysis.The results show that: 1) Compared with that in control, mitochondrial state â…¢ respiration(ST3)and OPR decreased significantly in hypoxia exposure groups. RCR level was decreased significantly in 1d,5d and 15d,then restored control in 30d group.st4 level was increased in 1d,5d,15d,then decrease quickly in 30d, even lower than control.2) Compared with control group, the content of ANT decreased significantly in 5d,15d and 30d,the content was lowest in 30d,only 33.4% compared with that in control. The level of energy charge(EC) in muscle was significantly lower than that of control in 5d,30d,but not decreased in 1d,the EC level was restored in 15d, even higher than that of control.3)The content of ATP decreased significantly in 1d,5d,30d,restored in 15d,then reach the lowest level in 30d.The level of EC decreased in 1d and 5d,then restored in 15d and 30d,there were no significant differences compared with control.4) The decreased transport activity of ANT was observed in 1d and 5d,and then increased to control level, in 30d, ANT activity was decreased again.5) The ANT1 mRNA state level was significantly decreased ,ANT2 mRNA state level was increased significantly in 30d,other groups were no significant differences compared with control. During hypoxia ,the total protein of ANT in mitochondria from heart ventricle muscle did not change.Conclusion: 1) Hypoxic exposure could make decrease of energy reserve in heart ventricle muscle, total anenine nucleotide pool decreased steeply was observed when the hypoxic time prolong. mitochondria respiratory dysfunction .This results indicated that the change of energy status in heart muscle contribute to the disorder of heart dyfunction during hypoxia. 2) Acute hypoxic exposure could lead to decrease significantly of ANT transport activity in mitochondria. The change of ANT activity may have coordination function on mitochondrial oxidative phosphorylation and heart energy ststus.3) chronic hypoxia could change the proportion of ANT protein isoforms ,indicated that ANT2 have great function on require of energy of heart during hypoxia.