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Published on: March 23, 2011
Using FASS-LTP in postmortem mice brain tissues to assess pathological synaptic function
Joshua Currie1, Shaneilahi Budhwani2, Klarissa H Garza3
1John Sealy School of Medicine, University of Texas Medical Branch at Galveston, USA; Mitchell Center for Neurodegenerative Diseases, Department of Neurology, University of Texas Medical Branch, USA.
Background:
Study of synaptic integrity using conventional electrophysiology is a gold standard for quantitative assessment of neurodegeneration. Fluorescence assisted single-synapse long-term potentiation (FASS-LTP) provides a high throughput method to assess the synaptic integrity of neurotransmission within and between different brain regions as a measure of pharmacological efficacy in translational models.
New Method:
We adapted the existing method to our purpose by adding a step during the thawing of frozen samples, by an extra step of placing them on a rocker at room temperature for 30 minutes immediately following thawing with constant mixing on a shaker. This allowed for gradual, uniform thawing, effectively separating the synaptosomes. Our study demonstrates FASS-LTP on four brain regions at 6- and 12-month periods in the 3xTg-AD mouse model, treating sibling cohorts with VU0155069 (a small molecule inhibitor) or vehicle (0.9 % saline).
Results:
Our findings demonstrate the robust ability of the FASS-LTP technique to characterize the functional synaptic integrity maintained by disease-treatment therapies in multiple brain regions longitudinally using frozen brain tissue.
Comparison With Existing Methods:
By providing a detailed, user-friendly protocol for this well-known analysis and including a recovery step improved the ability to robustly replicate the FASS-LTP between different brain regions. This may be extrapolated to a translational use on human clinical samples to improve understanding of the therapeutic impact on synaptic performance related to glutamate neurotransmission.
Conclusions:
FASS-LTP method offers a robust analysis of synaptosomes isolated from frozen tissue samples, demonstrating greater reproducibility in rodent and human synapses in physiological and pathological states.
