gms | German Medical Science

Many theories have been postulated in an attempt to explain the biological basis of aging. One of the theories which has received a growing interest is “the mitochondrial theory of aging”, which links aging to mitochondrial dysfunction, oxidative stress and apoptosis (Physiol Rev, 78:547-81,1998). Skeletal muscle mitochondrial function is a proper target for testing the aforementioned theory, since it is a postmitotic tissue with large energy demands and highly vulnerable to mitochondrial diseases (Science,283:1482-88,1999). Considering that data reported in the literature are not consensual, the main objective of this study was to characterize in vitro skeletal muscle mitochondria functionality from mices of different ages.

Twelve male C57BL/6 strain mice were divided into two groups (GI-1 month;GII-14 months). Skeletal muscle mitochondria were isolated from hindlimb muscles (soleus, gastrocnemius, tibialis anterior, quadriceps) by conventional methods of differential centrifugation (Acta Physiol Scand,161:345-53,1997). Mitochondrial respiratory function was polarographically measured using a Clark-type oxygen electrode (Hansatech DW1, Norford UK). Mitochondrial respiration was initiated by adding pir+mal or suc+rot. State 3 and state 4 respiration, respiratory control ratio (RCR) and ADP/O were determined. Mitochondrial inner membrane permeability and the maximal rate of uncoupled oxidative phosphorylation were determined by adding oligomycin and CCCP, respectively during state 3 respiration with saturated amounts of ADP. Mitochondrial in vitro stimulation was assayed by adding pir+mal and after 30’’ stabilization by adding 3 consecutive pulses of ADP interspersed by 1’30’’ each. State 3 and state 4 respiration, RCR and ADP/O were also determined in the three respiratory cycles.

In both complex I and complex II linked-substrates it was verified that state 3 respiration and RCR were lower in the older animals. State 4 and ADP/O showed no alterations with age. However, the time spent in state 3 (sec.) was higher in the GII group. Respiratory rates in the presence of oligomycin were unaltered. In the presence of CCCP, respiratory rates were lower in the older animals. Considering the in vitro mitochondrial stimulation, a decrease was evident in state 3 along the 3 ADP pulses in both groups studied. State 4, RCR, and ADP/O, were unchanged. When considering the total O2 consumed during the period of stimulation (4’30’’), mitochondria isolated from young animals consumed less oxygen to phosphorylate the same amount of ADP (3x150 nmol ADP).

The results suggest that mitochondria from older animals evidenced a slightly diminished capacity for electron carrying and/or H+ pumping, and an increase susceptibility to protein dysfunction induced by oxidative damage. Concerning inner membrane mitochondrial permeability and mitochondrial phosphorylation efficiency, it appears that aging doesn’t affect the results obtained in both groups.

This work was co-financed by POCI 2010 and FSE