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Preparation of Synaptoneurosomes from Mouse Cortex using a Discontinuous Percoll-Sucrose Density Gradient
Published on: September 17, 2011
A rapid Percoll gradient procedure for preparation of synaptosomes
Peter R Dunkley1, Paula E Jarvie, Phillip J Robinson
1School of Biomedical Sciences and Hunter Medical Research Institute, Faculty of Health, The University of Newcastle, Callaghan, New South Wales 2308, Australia. peter.dunkley@newcastle.edu.au
Nature Protocols
|October 18, 2008
Summary
This study presents a rapid method to isolate functional synaptosomes from brain tissue using a Percoll gradient. This technique allows for efficient neurotransmitter uptake and release studies in neuroscience research.
Area of Science:
- Neuroscience
- Cell Biology
- Biochemistry
Background:
- Brain tissue homogenization severs nerve terminals, forming synaptosomes.
- Isolating intact, functional synaptosomes is crucial for studying neurotransmission.
Purpose of the Study:
- To develop a rapid and efficient protocol for isolating viable synaptosomes from brain homogenates.
- To enable functional experiments on synaptosomes with minimal delay.
Main Methods:
- Homogenization of fresh brain tissue in an isotonic medium.
- Isolation of synaptosomes using a discontinuous Percoll gradient and medium-speed centrifugation.
- Minimizing mechanically damaging steps and maintaining isotonic conditions.
Main Results:
- The Percoll gradient method isolates synaptosomes quickly (approx. 1 hour).
- Yields 2.5-4 mg synaptosomal protein per gram rat brain.
- Produces relatively homogeneous synaptosomes with minimal contamination.
Conclusions:
- The described protocol provides a fast and effective means to obtain viable, functional synaptosomes.
- This method facilitates efficient neurotransmitter uptake and release studies.
- The protocol offers advantages in speed and purity over existing methods.
