Targeted axon-attached recording with fluorescent patch-clamp pipettes in brain slices.
Understanding the physiology of axons in the central nervous system requires experimental access to intact axons. This protocol describes how to perform cell-attached recordings from narrow axon fibers (ϕ <1 μm) in acute and cultured brain slice preparations (with a success rate of ∼50%). By using fluorophore-coated glass pipettes and Nipkow disk confocal microscopy, fluorescently labeled axons can be visually targeted under online optical control. In the cell-attached configuration, axonal action potentials are extracellularly recorded as unit-like, sharp negative currents. The axon morphology labeling and cell-attached recordings of axons can be completed within 1-2 h. The recordings are stable for at least 30 min.
Laboratory of Chemical Pharmacology, Graduate School of Pharmaceutical Sciences, University of Tokyo, Tokyo, Japan. t.sasaki.0224@gmail.com
Takuya
T
Sasaki
Norio
N
Matsuki
Yuji
Y
Ikegaya
D000200:Action Potentials; D000818:Animals; D001369:Axons / Q000502:physiology*; D001921:Brain; D005456:Fluorescent Dyes / Q000032:analysis / Q000378:metabolism; D007091:Image Processing, Computer-Assisted; D066298:In Vitro Techniques; D051379:Mice; D008813:Mice, Inbred ICR; D018613:Microscopy, Confocal; D018408:Patch-Clamp Techniques / Q000295:instrumentation* / Q000379:methods*; D051381:Rats; D017208:Rats, Wistar; D012984:Software
D005456:Fluorescent Dyes