Electrophysiology

Dear Yassine,

Please find below the details of the electrophysiology solutions used in the 2019 elife paper for both whole-cell and excised patch experiments:

Electrophysiology solutions:

Whole-cell patch experiments:

Intracellular Pipette Solutions:

146mM NaCl 

2mM MgCl₂

5mM EGTA

10mM HEPES (pH 7.4)

pH Adjusted: with either NMDG or methanesulfonate.

Extracellular Pipette Solutions:

0-146mM NaCl - Solutions balanced against pipette for ionic strength with NMDG-Methanesulfonate.

2mM MgCl₂

5mM EGTA

10mM HEPES (pH 7.4)

pH Adjusted: with either NMDG or methanesulfonate.

NaCl Excised inside-out patch experiments:

Extracellular Pipette Solutions:

146mM NaCl

2mM MgCl₂

5mM EGTA

10mM HEPES (pH 7.4)

pH Adjusted: with either NMDG or methanesulfonate.

Intracellular Bath Solutions:

0-146mM NaCl₂- Solutions balanced against pipette for ionic strength with NMDG-Methanesulfonate. 

2mM MgCl₂

5mM EGTA

10mM HEPES (pH 7.4).

Under conditions of greater than 150mM NaCl in the bath solution – NaCl was substituted with KCl to minimize the junction potential. 

Bi-ionic substitution inside-out patch experiments:

Extracellular Pipette Solution:

146mM NaCl

2mM MgCl₂

5mM EGTA

10mM HEPES (pH 7.4)

pH Adjusted: with either NMDG or methanesulfonate.

Intracellular Bath Solutions:

For experiments using Br^-, I^-, NO₃^-, SO₄^2- 150mM sodium salt solutions were prepared with 2mM Mg(OAc)₂ used instead of 2mM MgCl₂:

Example:

150mM SO₄^-2_­

2mM Mg(OAc)₂

5mM EGTA

10mM HEPES (7.4)

pH adjusted with either NMDG or methanesulfonate.

For HCO₃^‑experiments:

165mM NaHCO₃^- was used as [HCO₃^-] / [HCO₃^–] + [H₂CO₃ + CO₂]=0.9 at pH 7.4, due to the pKa of 6.4 for protonation of HCO₃^-.

pH monitoring was regularly implemented to ensure that HCO₃^- never decreased by over 20% due to conversion to CO₂ and fresh solutions were prepared every 1-3 hours.