Localized and non-contact mechanical stimulation of dorsal root ganglion sensory neurons using scanning ion conductance microscopy.
Mechanosensitive ion channels convert external mechanical force into electrical and chemical signals in cells, but their physiological function in different tissues is not clearly understood. One reason for this is that there is as yet no satisfactory physiological method to stimulate these channels in living cells. Using the nanopipette-probe of the Scanning Ion Conductance Microscope (SICM), we have developed a new technique to apply local mechanical stimulus to living cells to an area of about 0.385 microm2, determined by the pipette diameter. Our method prevents any physical contact and damage to the cell membrane by use of a pressure jet applied via the nanopipette. The study used whole-cell patch-clamp recordings and measurements of intracellular Ca2+ concentration to validate the application of the mechanical stimulation protocols in human and rat dorsal root ganglia (DRG) sensory neurons. We were able, for the first time, to produce a non-contact, controlled mechanical stimulation on living neurites of human DRG neurons. Our methods will enable the identification and characterisation of compounds being developed for the treatment of clinical mechanical hypersensitivity states.
Sánchez
Daniel
D
Imperial College Faculty of Medicine, London Centre for Nanotechnology, Division of Medicine, 5th floor MRC Clinical Sciences Centre, Hammersmith Hospital Campus, Du Cane Road, London W12 0NN, UK.
Anand
Uma
U
Gorelik
Julia
J
Benham
Christopher D
CD
Bountra
Chas
C
Lab
Max
M
Klenerman
David
D
Birch
Rolfe
R
Anand
Praveen
P
Korchev
Yuri
Y
D000328:Adult; D000818:Animals; D020013:Calcium Signaling / Q000502:physiology; D002462:Cell Membrane / Q000502:physiology / Q000648:ultrastructure; D002478:Cells, Cultured; D003712:Dendrites / Q000502:physiology; D004594:Electrophysiology; D005727:Ganglia, Spinal / Q000166:cytology / Q000502:physiology*; D006801:Humans; D066298:In Vitro Techniques; D008297:Male; D008855:Microscopy, Electron, Scanning; D008856:Microscopy, Fluorescence; D016501:Neurites / Q000502:physiology; D009475:Neurons, Afferent / Q000502:physiology*; D010812:Physical Stimulation / Q000295:instrumentation* / Q000379:methods; D017729:Presynaptic Terminals / Q000502:physiology; D051381:Rats; D017208:Rats, Wistar