Glutamate Induced Plasticity in primary hippocampal cultures

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pk2342
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Glutamate Induced Plasticity in primary hippocampal cultures

Hi,
So I'm having trouble inducing plasticity in my primary hippocampal neurons from neonatal mice. At the day of recording my culture is 10 to 15 days old and the culture medium contains Kyneurenic acid.
Here are my solutions:
My bath solution consists of [mM]: 119 NaCl, 20 HEPES, 30 Glucose, 2 CaCl2, 2 MgCl2 - I also add 100 uM Picrotoxin, 1 uM TTX and 1 uM Glycine
My electrode solution consists of [mM]: 130 K-Gluconate, 10 KCl, 5 MgCl2, 0.6 EGTA, 5 HEPES, 0.06 CaCl2, 2 Mg-ATP, 0.2 GTP, 20 Phosphocreatine
The procedure:
- basal mEPSC recording for 15 minutes at a holding of -70 mV
- 1 min of perfusion with Mg-free bath solution
- direct application of Mg-free bath solution containing 200 uM Glutamate (I just pipet drops on the side of my chamber until they enter the bath)
- after 30 seconds I switch back to normal bath solution
For some reason I fail to induce plasticity in 99% of the cells.
Thank you and all the best
Peter

pk2342
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if nobody has experience with

if nobody has experience with glutamate induced LTP, I would also appreciate your alternatives to glutamate application for LTP induction in culture :)

biophysics.tushar
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 I guess you can try some

 I guess you can try some younger neurons (you mentioned 10-15 days old neurons) or dissociated acute neurons.