I would like to measure a possible change in synaptic coverage by astrocytes upon memory formation. Do you think there is some way to measure it by IHC and optic microscopy (mainly 2-photon microscopy) or one has to do it via EM?
And how would you measure the degree of synaptic coverage? Just by overlapping signals of astrocytes glutamate transporters and neuronal synaptic proteins?
I think you can use the confocal microscopy for this kind of experiment...you can see the live uptake process...I think best way to do electrophysiology...because in memory formation you can see LTP and LDP type of currents on glutamate exposure...so better way to do also electrophysiology using memory paradigm in astrocytes..
Thanks Mouli. However I would not need to measure LTP because I have already a strong and precise behavioural read out for learning. I just need to see if after the memory formation, the astrocytes show a better synaptic coverage in that area. Almost all the papers I ve read measure it by analysis of EM images, but I was wondering if somehow it can be done with optical imaging.
Marco, the definitive is going to be EM but not sure if that works for your system. With immuno, you could potentially do something like STED imaging with glutamate transporters and pre or postsynaptic markers to get high resolution of the numbers of synapses contacted by glia. Electrophysiology will also work with looking at the increase in transporter currents, but that gets complicated a little with the increased synaptic strength due to ltp. You could do just transporter recordings from subjects with and without the behavior training, and then compare immunos between them to make a reasonable case.
I think the definitive method for synaptic coverage would be EM, but if you're stuck or would rather do optical methods, then the next best option would probably be 2PLSM in slices.
If you think you want to go down the EM route, then there are some emerging companies out there that will take your samples and do the EM imaging and reconstruction of astrocytic-synaptic interactions for you...
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