Electrophysiology Chemicals

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Electrophysiology Chemicals

9/22/2004 Colin
TTX                from Alomone Labs, Israel. TTX w/ citrate.
Na channel toxin
1 mg per ampoule, MW = 319.28, soluble in dH2O
lyophilised = 1 year @ 4 deg C  /  liquid = 6 months @ -20 deg C
desired final concentration = 1 uM
1 uM in 100 ml external solution = 0.031928 mg in 100 ml
                        1 mg TTX in 1566 ul of dH2O => 50 ul aliquots each of 0.031928 mg
                        aliquot solution is concentration of 2 mM    

Picrotoxin        from Tocris Cookson.
GABA A R channel toxin
1 g per ampoule, MW = 602.00, soluble in DMSO
lyophilised = 1 year @ RT  /  liquid = 6 months @ -20 deg C
desired final concentration = 100 uM
100 uM in 100 ml external solution = 6.02 mg in 100 ml
                        500 mg PTX in 8306 ul (8.31 ml) of DMSO => 100 ul aliquots of 6.02 mg                         aliquot solution is concentration of 100 mM

APV                from Tocris Cookson.
                        NMDA R antagonist
10 mg per ampoule, MW = 197.13, soluble in dH2O
lyophilised = 6 months @ RT  /  liquid ~  2 months @ -20 deg C
desired final concentration = 100 uM
100 uM in 100 ml external solution = 1.9713 mg in 100 ml
                        10 mg APV in 508 ul of dH2O => 100 ul aliquots each of 1.9713 mg
                        aliquot solution is concentration of 100 mM

CNQX                        from Tocris Cookson.
AMPA, Kainate antagonist
10 mg per ampoule, MW = 232.16, soluble in DMSO to 100 mM
lyophilised = 6 months @ RT  /  liquid ~  2 months @ -20 deg C
desired final concentration = 20 uM
20 uM in 100 ml external solution = 0.46432 mg in 100 ml
                        10 mg CNQX in 1077 ul of DMSO => 50 ul aliquots each of 0.46432 mg
                        aliquot solution is concentration of 40 mM  

NBQX                        from Tocris Cookson.
AMPA, Kainate antagonist
5 mg per ampoule, MW = 336.28
desired final concentration = 20 uM
                        5 mg NBQX in 743 ul of DMSO => 100 ul aliquots each of 0.6726 mg
Kynurenic       from Sigma
Acid                General glutamate R antagonist (AMPA and NMDA Rs)
1 g per ampoule, MW = 189.2,
desired final concentration = 2 mM
2 mM in 100 ml external solution = 37.84 mg in 100 ml
                        Do not make up stock and freeze. Instead, on day of experiment, dissolve
direct into external solution and stir for 10-20 mins. 
           
SR95531         from Tocris (1262)
Hydrobromide GABAA R Antagonist of choice (no effects on K+ channels)
(GABAzine)   10 mg ampoule, MW = 368.23, soluble in dH2O
desired final concentration = 3 uM
3 uM in 100 ml external solution = 0.1105 mg in 100 ml
                        10 mg of SR95531 in 9ml dH2O => 90 aliquots of 100ul each 0.1105mg
                        1 mg of SR95531 in 900ul dH2O => 9 aliquots of 100ul each 0.1105mg
Each aliquot will give 3 uM in 100 ml external

Bicuculline      from Sigma
Methiodide     GABA A receptor antagonist
100 mg pot, MW = 509.3, soluble in dH2O to 1.8 mg/ml
[lyophilised > 6 months @ 2-8 deg C, liquid ~  2 months @ -20 deg C]
desired final concentration = 100 uM
100 uM in 100 ml external solution = 5.093 mg in 100 ml
100 mg Bicuculine Methiodide in 29452 ul (29.5 ml) of dH2O => 1500 ul aliquots each of 5.093 mg, aliquot solution is concentration of 6.666 mM    

QX-314           from Alomone Labs, Israel.
                        Blocks Na channels from inside cell
500 mg ampoule, MW = 298.85, soluble in dH2O or internal solution
store @ RT
desired final concentration in patch pipette = 2-5 mM
dissolve ~1.5 mg in 100 ul internal solution, take volume equivalent to 0.598 mg and add this to 400 ul of internal solution to give 5 mM,
or take volume equivalent to 0.3 mg and add this to 200 ul of internal solution to give 5 mM

Bungarotoxin   from Calbiochem.
nACHR antagonist
1mg ampoule, soluble in dH2O, store stocks @ -20 deg C
desired final concentration = 1-2 ug per ml (Zhang et al. 1998)
25 ml at this concentration = 50 ug in 100ml
Make 50ul aliquots of 1mg/ml in dH2O and add one 50 ul aliquot to 25 ml external to achieve 2 ug per ml
Tubocurarine   from Sigma.
nACHR antagonist
desired final concentration = 50 uM (Zhang et al. 1998)
50 uM in 50 ml external solution = 3.409 mg in 100 ml
                        100 mg Tubocurarine in 5867ul dH2O => 100ul aliquots of 1.705 mg
each is enough to make 50 uM of 50 ml external

Glutamic         from Tocris.
Acid                Major agonist for EAA receptors in CNS
1 g ampoule, MW = 147.13, soluble to 1 eq NaOH
store @ RT, stock solutions at –20 deg C
desired final concentration = 200 uM
200 uM in 100 ml external solution = 2.9426 mg in 100 ml
58.852 mg Glutamic Acid in 2000 ul (2 ml) of 200mM NaOH => 100ul aliquots of 2.9426 mg, aliquot solution is concentration of 200 mM

Muscimol        from Sigma
Selective GABAA agonist
10 mg pot, MW = 114.1, soluble to 100mM in 1 eq NaOH
desired final concentration = 30 uM
30 uM in 100 ml external solution = 0.3423 mg in 100 ml
30 uM in 1 ml external solution = 0.003423 mg in 100 ml
10 mg Muscimol in 1460ul NaOH => 50ul aliquots for 30 uM in 100ml
                                                        or 1ul in 2ml external to achieve 30uM

NMDA           from Tocris (0114).
NMDA R Agonist
50 mg ampoule, MW = 165.15, soluble in dH2O
desired final concentration for puffer application = 200 uM – 1 mM
desired final concentration for bath application = 20 uM (Ben-Ari)
200 uM in 1 ml external solution = 0.033 mg in 1 ml external
20 uM in 100 ml external solution = 0.33 mg in 100 ml dH2O
5 mg NMDA in 1500 ul dH2O => 100ul aliquots of 0.2943 mg
50 ul of aliquot in 50 ml = 20 uM
10 ul of aliquot in 1 ml = 200 uM
aliquot solution is concentration of 2 mM 

GABA R agonist
1 g per ampoule, MW = 103.12, soluble in dH2O to 100 mM
lyophilised = 6 months @ RT  /  liquid ~  2 months @ -20 deg C
desired final concentration = 50 uM (see Kriegstein ref)
50 uM in 100 ml external solution = 0.516 mg in 100 ml
                        10 mg GABA in 1938 ul of dH2O => 100 ul aliquots of 0.516 mg                                       aliquot solution is concentration of 50 mM  

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