Tweak Your Technique Electrophysiology 101 August 19, 2011 Kevin Lee.

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Presentation transcript:

Tweak Your Technique Electrophysiology 101 August 19, 2011 Kevin Lee

What 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

The hippocampus

Field recordings

LTP/LTD

Whole-cell Recordings

Patch-Recording Methods

Whole-cell recordings Current-clamp vs Voltage-clamp “EPSP” “EPSC”

Whole-cell recordings Current-clamp vs Voltage-clamp

Voltage-clamp

mEPSCs (minis) mEPSC amplitude reflects synaptic AMPAR content Beique et al. (PNAS, 2006)

mEPSCs (minis) mEPSC frequency can reflect: - total number of synapses - presynaptic function

Release 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)

What about Molecular interventions?

Biolistic transfection of cortical neurons in organotypic slices Beique & Andrade (J. Physiol, 2002; cover photo)

Biolistic transfection of cortical neurons in organotypic slices

Barria & Malinow (Neuron, 2002)Cull-Candy et al. (Science STKE, 2004)

Drawbacks to electrical stimulation: - Difficult to isolate individual synaptic events - Spatial Parameters left unknown

2P-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

Single Synapse Ca 2+ Imaging Time  Uncage Simultaneous Glutamate Uncaging and Calcium Imaging

Fundamental dendritic properties probed by glutamate uncaging and calcium imaging Losonczy & Magee (Neuron, 2006)

Fundamental dendritic properties probed by glutamate uncaging and calcium imaging Losonczy & Magee (Neuron, 2006)

Experimental Wizardry to Envy Kravitz et al. (Nature, 2009)

Experimental Wizardry to Envy Kravitz et al. (Nature, 2009)

Multiple Simultaneous Recordings in Slices Olah et al. (Nature, 2009)

The End