Abstract
It is generally accepted that mitochondria are able to proliferate even in postmitotic cells due to their natural turnover and also to satisfy increased cell energy requirements. However, no detailed studies are available, particularly with respect to specific cell types. Since [3H]-thymidine is incorporated not only into nuclear (n) DNA but also into the DNA of cytoplasmic mitochondria, an autoradiographic approach was developed at the light microscopy level in order to study basic questions of mitochondrial (mt) proliferation in organs of rodents in situ via the cytoplasmic incorporation of [3H]-thymidine injected into the animals 1 h before sacrifice. Experiments carried out on mice after X-irradiation showed that cytoplasmic labeling was not due to a process such as unscheduled nuclear DNA synthesis (nUDS). Furthermore, half-lives of mitochondria between 8-23 days were deduced specifically in relation to cell types. The phase of mtDNA synthesis was about 75 min. Finally, mt proliferation was measured in brain cells of mice as a function of age. While all neurons showed a decreasing extent of mtDNA synthesis during old age, nUDS decreased only in distinct cell types of the cortex and hippocampus. We conclude that the leading theories explaining the phenomenon of aging are closely related, i.e., aging is due to a decreasing capacity of nDNA repair, which leads to unrepaired nDNA damage, or to an accumulation of mitochondria with damaged mtDNA, which leads to a deficit of cellular energy production.
Highlights
Mitochondria, which generate energy for cellular processes through oxidative phosphorylation, contain their own DNA in contrast to other organelles with the exception of the nucleus
MtDNA like nDNA can be labeled with [3H]thymidine (TdR) [4,5,6]. This permitted the study of mitochondria within tissue cells using electron microscopic autoradiography (EM ARG) which up to now - in addition to cells in culture - had been applied mostly to rat adrenal glands [7,8,9,10,11]
In order to answer questions related to the kinetics of mt reduplication within different cell types of an organ, EM ARG studies are much too tedious
Summary
Mitochondria, which generate energy for cellular processes through oxidative phosphorylation, contain their own DNA in contrast to other organelles with the exception of the nucleus. Many questions concerning details remained unanswered, those related to specific cell types within an organ. This may be due to the fact that mostly biochemical or - more recently molecular biology techniques were used which depend on isolated mtDNA from larger pieces of tissue. A very long exposure time of several months was used for the demonstration of nUDS due to the general autoradiographic rule that grains per structure correspond to the product of radioactivity present in the structure times exposure time (provided that the autoradiographic emulsion used works linearly with increasing time) Using such a long exposure time, mtDNA synthesis was demonstrable by light microscopy in the perikaryal cytoplasm of some types of neurons (Figure 2a). Since results regarding mtDNA synthesis were obtained from experiments in which nUDS was measured, distinct results related to nUDS will be reported in this article
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