I would like to try to block all synaptic transmission. TTX blocks the AP-dependent release at the terminal and so will Cd2+ be effecttive in blocking the VGCC. What about minis?
Hi! Most likely to block it completely is impossible. Try EGTA in Ca-free bath solution/culture medium and as you mentione + TTX and VGCC blockers - that could be the most what you can get. You can not avoide some spontaneous release even in tetanus-toxin treated neurons (which cleaves SNARE comples responcible for vesicle fusion) because in the synapse there is some Tetanus-toxin-insensitive SNARE protein, which also can drive the vesicle fusion. And this is only one protein, but there are a lot of similar and you'd faster kill the cell than _COMPLETELY_ block exocytosis.
If you want just to block the postsynaptic responces, then the coctail of AP5, CNQX and bicucculine or picrotoxine will wipe it out, but the neurotransmitter release will be there. Your choise.
Hi! Most likely to block it completely is impossible. Try EGTA in Ca-free bath solution/culture medium and as you mentione + TTX and VGCC blockers - that could be the most what you can get. You can not avoide some spontaneous release even in tetanus-toxin treated neurons (which cleaves SNARE comples responcible for vesicle fusion) because in the synapse there is some Tetanus-toxin-insensitive SNARE protein, which also can drive the vesicle fusion. And this is only one protein, but there are a lot of similar and you'd faster kill the cell than _COMPLETELY_ block exocytosis.
If you want just to block the postsynaptic responces, then the coctail of AP5, CNQX and bicucculine or picrotoxine will wipe it out, but the neurotransmitter release will be there. Your choise.
Blocking the GABA and glutamate postsynaptic receptors with CNQX, AP-5, and bicuculline is probably the simplest way of blocking all transmission.
we need more information to help you, in which preparation are you working? why do you want to block all synaptic transmission? how much is the mini frequency? I´m totally agree with Yaroslav. Also reducing the temperature the minis could decrease as well, but always some modification could affect other cellular process. So, the strategy will change if you need to block vesicle fusion or if you just want to block the transmission, in the last one Stephen is totally right if there is only gaba and glutamate NT.
I agree with those that suggested a cocktail to block glutamate and GABA receptors. In fact we did just that and looked at the effect on cortical pyramidal cells a number of years ago (Paré D, Shink E, Gaudreau H, Destexhe A, Lang EJ. J Neurophysiol. 1998 Mar;79(3):1450-60). I would avoid using bicuculline because that also blocks SK channels (see Debarbieux F, Brunton J, Charpak S. J Neurophysiol. 1998 Jun;79(6):2911-8. So to block GABA-A receptors I think it would be better to use picrotoxin or GABAzine
You might use Botolinum toxin A. If you work in cell cultures, incubation with the recombinant toxin might be sufficient to cleave most of SNAP25 and abolish minis. We have done it in our 2011 paper and it worked quite nice, but we had the special case to work in autapses, allowing precise control over every single neuron we recorded from. Botulinum toxin C killed our neurons, we never got it to work. The only other option is to use Munc13-1 and -2 double knockouts. They have absolutely no release, but are actually viable in culture (you have to do E18 preparations).
Article Activation of metabotropic GABA receptors increases the ener...
Depending on your tissue, I would probably try a cocktail of picrotoxin (maybe 100micro might be enough for u), APV and cnqx
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