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Slide1
Tweak Your TechniqueElectrophysiology 101
August 19, 2011
Kevin Lee
Slide2What can be studied with electrophysiology?
Single channel behaviour
in expression systems: HEK293, CHO’s, X.
Laevis
oocytes
Intrinsic properties of excitable cells
neurons,
myocytes
Synaptic transmission
neuromuscular junction, neuronal synapses
Slide3Slide4The hippocampus
Slide5Field recordings
Slide6LTP/LTD
Slide7Whole-cell Recordings
Slide8Patch-Recording Methods
Slide9Whole-cell recordings
Current-clamp vs Voltage-clamp
“EPSP”
“EPSC”
Slide10Whole-cell recordings
Current-clamp vs Voltage-clamp
Slide11Voltage-clamp
Slide12mEPSCs
(minis)
mEPSC
amplitude reflects synaptic AMPAR content
Beique
et al. (PNAS, 2006)
Slide13Beique
et al. (PNAS, 2006)
mEPSCs
(minis)
mEPSC
frequency can reflect:
- total number of synapses
-
presynaptic
function
Slide14Release probability (Pr) of glutamate
Changes in frequency of mEPSCs is generally believed to reflect changes in Pr.“Paired-Pulse” electrical stimulation can also provide information about Pr.
Beique
et al. (PNAS, 2006)
Slide15What about Molecular interventions?
Slide16Biolistic transfection of cortical neurons in organotypic slices
Beique
& Andrade (J.
Physiol
, 2002; cover photo)
Slide17Biolistic transfection of cortical neurons in organotypic slices
Slide18Biolistic transfection of cortical neurons in organotypic slices
Barria
& Malinow (Neuron, 2002)
Cull-Candy et al. (Science STKE, 2004)
Slide19Drawbacks to electrical stimulation:
Difficult to isolate individual synaptic events
Spatial Parameters left unknown
Slide202P-Uncaging of MNI-Glutamate
Slide212P-Uncaging of MNI-Glutamate
Béïque et al., PNAS 2006
Uncaging
of MNI-
Glu activates both AMPA and NMDA receptors
High spatial resolution of glutamate
uncaging
Slide22Single Synapse Ca
2+
Imaging
Time
1
2
1
2
Uncage
Simultaneous Glutamate
Uncaging
and Calcium Imaging
Slide23Fundamental dendritic properties probed by glutamate uncaging and calcium imaging
Losonczy
& Magee (Neuron, 2006)
Slide24Fundamental
dendritic properties probed by glutamate uncaging and calcium imaging
Losonczy
& Magee (Neuron, 2006)
Slide25Experimental Wizardry to Envy
Kravitz
et al. (Nature, 2009)
Slide26Experimental Wizardry to Envy
Kravitz
et al. (Nature, 2009)
Slide27Multiple Simultaneous Recordings in Slices
Olah
et al. (Nature, 2009)
Slide28The
End
Tweak Your Technique
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