Electrophysiology

the science and branch of physiology that pertains to the flow of ions in biological tissues and, in particular, to the electrical recording techniques that enable the measurement of this flow and the potential changes related to them

Intracellular Recording

Intracellular Recording: A recording electrode is inserted into a cell, so that the intracellular potential can be measured against the extracellular potential

cell

Sir John Carew Eccles

Alan Lloyd Hodgkin

Andrew Fielding Huxley

The Nobel Prize in Physiology or Medicine 1963

"for their discoveries concerning the ionic mechanisms involved in excitation and inhibition in the peripheral and central portions of the nerve cell membrane"

http://nobelprize.org/medicine/laureates/1963/

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Membrane potentials in squid axons:

The first experiments examining howchanges in ion concentrations affect themembrane potential were done on squid.

Why squid axons?

Voltage Clamp

Allows you to ‘clamp’ the membrane potential and record the ionic current Normally, the voltage is ‘stepped; and the resulting current is measured Difficulties with Voltage Clamp and sharp intracellular recordings?

E. Neher B. Sakmann

1991 Nobel Prize in MedicineFor the development of the Patch clampSmall patch of membrane is sealed to the tip of a micropipetteThe high resistance seal (called a gigaohm seal)ensures that currents flow through the amplifier rather than escaping through the rim of the patch

Recording Configurations

• Why select cell-attached vs. whole-cell recording?

Cell-attached recording

• Allows the recording from a single ion channel located in the area of the patch under the pipette

• As the ion channel opens or closes there will be an abrupt increase or decrease in the conductance of the patch of membrane

• Ion channels can be characterized by their conductance, their open time, and probability of channel opening (in addition to pharmacological inhibition and ion substitution)

Figure 4.6 Effect of Potential on Currents

Permeability = open channelConductance = permeability + ions

Single spontaneously active K+ channel; 150 mM KCl on both sides

Channel current as a function of voltage; the slope of the line indicates the channel conductance; the ability of a channel to pass current

A linear IV curve indicates no voltage-dependent gating of the channel

Equilibirum Potential for K+ Currents

[K+]i = 90mM [K+]o = 3mM; (B) with no electrical gradient K+ flows out(C) with 20mV applied to the cell the flow of K+ out of the cell increases(D) -50mV inside the cell reduces the current amplitudeNon-linear curve indicates the channel is voltage-dependent

What is the difference between Patch Clamp and Voltage Clamp?

Can they be used simultaneously?

Recording Configurations

• Why select cell-attached vs. whole-cell recording?

• How do you decide what solution to include in your whole cell solution?

• Electrical access vs. dialysis?

Figure 7.1 Membrane Currents Produced by Depolarization

• Ion substitution experiments

Figure 7.3 Dependence of Early and Late Currents on Potential

How can we find synaptically connected neurons?

EPSPs

IPSPs

Figure 11.8 Reversal Potential for Synaptic Currents Measured by Voltage Clamp Recording

STDP

Fiumelli & Woodin 2007 Current Opinion in Neurobiology

Perforated Patch Clamp Recording