Cryopreservation of brain mitochondria: A novel methodology for functional studies

Vidya N. Nukala, Indrapal N. Singh, Laurie M. Davis, Patrick G. Sullivan

Research output: Contribution to journalArticlepeer-review

42 Scopus citations

Abstract

Often, comparative studies involving large number of animals or human post-mortem tissue samples are precluded, especially those requiring structurally and functionally intact cells and/or organelles. The ability to 'bank' such samples for storage and restore or 'reanimate' them at a later time without causing damage to the structure and/or function becomes imperative. However, to date, such attempts have produced conflicting results. We here demonstrate for the first time that isolated rat brain mitochondria can be successfully cryopreserved and restored for later use. We added a well characterized cryoprotectant 10% (v/v) dimethyl sulfoxide (DMSO) to purified rat cortical mitochondria and allowed them to cool at a uniform rate of ∼1°C/min and stored them at -80°C. Freshly isolated as well as reanimated brain mitochondria were analyzed for respiration. Structural integrity of cryopreserved mitochondria was also verified by electron microscopy. Mitochondrial membrane marker levels were assessed along with cytochrome c levels. Intact structure and function of the cryopreserved brain mitochondria observed allows us the opportunity to store mitochondria for longer periods of time as well as perform metabolic studies as needed. This will considerably expand the time-frame required for carrying out functional analysis in large comparative studies.

Original languageEnglish
Pages (from-to)48-54
Number of pages7
JournalJournal of Neuroscience Methods
Volume152
Issue number1-2
DOIs
StatePublished - Apr 15 2006

Bibliographical note

Funding Information:
The authors would like to thank Dr. Melanie McEwen, Doug Price, Dr. Steve Scheff and Maile Brown for their expertise and advice. This work was supported by an ADRC pilot grant (A605144-20) and NIH grants (NS48191 and NS046426) to P.G.S.

Keywords

  • Brain mitochondria
  • Cryopreservation
  • Cytochrome c
  • Dimethyl sulfoxide (DMSO)
  • Membrane marker proteins
  • Respiration
  • Ultra-structure

ASJC Scopus subject areas

  • General Neuroscience

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