Environmental enrichment has been shown to induce several changes in the brain including induction of several genes and proteins, neurogenesis and improvement in cognitive performance. I am planning to setup enrichment paradigm for mice to study changes in certain protein levels In hippocampus by western blot and qPCR but I would like to have positive control to make sure. What are good positive controls for western blot and qPCR after environmental enrichment?
I think it really depends on the age of the animal, enrichment duration and the time at which you harvest tissue following enrichment. The following recent paper should give you some good ideas as far as what proteins/genes to analyze:
http://www.ncbi.nlm.nih.gov/pubmed/?term=23473322
Broadly, I would suggest synaptic scaffolding proteins/genes (PSD-95, synapsin, stargazin, shank family, homer family, etc) as a start, as enrichment probably causes synaptogenesis. Immediate early genes (Arc, zif268, c-fos, etc) may also be something to look at. Transcription factors (CREB, SCIRR69, etc) that occupy the promoters of trophic genes, and the tropic factors themselves (BDNF, NT3 etc) , could also be an option.
In any case, it may be necessary to impoverish a subset of animals after enrichment (for a period of 1-2 months) to support the finding that enrichment indeed augments your protein/gene of interest. As a corollary, it may be necessary to show that enrichment downregulates certain behaviors. I believe enrichment has robust effects on anxiety, so an increase in your positive control with a decrease in anxious behavior would be strong.
Hope this helps.
Thank you very much for the suggestions and the article. If you have experience in isolating synaptosomes, please let me know how difficult it is to set it up and how robust the results from them are to work with. I had some experience in trying to isolate microglia from the brain after mechanical dissociation and it was kind of difficult to achieve similar numbers in each trial.
I personally have not isolated pure synaptosomal fractions, but have heard people use a pretty simple Ficoll gradient-based protocol. This might've been the protocol they were following:
http://www.nature.com/nprot/journal/v3/n11/full/nprot.2008.171.html
Looks like this would be easier than screwing around with pH or some other property and it appears to be pure and repeatable. Again, I havent personally used this protocol so I can't do much as far as troubleshooting goes, but this is where I'd probably start. Best of luck.
Thank you very much for your answer. I think it will help me greatly.
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