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                        from Tocris

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