Announcements Volunteer note taker Tutorial next Thursday.
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Transcript of Announcements Volunteer note taker Tutorial next Thursday.
Action Potentials
• What are they?– Rapid reversal of the resting membrane
potential
cell
0 mV
-80 mV
3 ms
Electrophysiology Techniques
Lolligo pealeii
Squid giant axon-1 mm in diameter-1000X larger than most
Action Potential
0 mV
-80 mV
Rising phase ordepolarization
Falling phase orRepolarization
Resting membranepotential
Threshold Potential
Undershoot or after-hyperpolarization
Overshoot
Reminders…
1. Action potential is a rapid reversal of Vm
2. Vm is dominated by the equilibrium potential of the most permeable ion
3. Permeability controlled by ion channels
K+
Na+ Voltage-gated channels
K+ leak channel
Na+
K+
Section of Squid Axon
Me
mbr
an
e P
ote
ntia
l
time
1. At rest only K+ leak channels open, PK>>PNa
2. With stimulus, voltage-gated Na channels open, PNa>>PK
Na+ flows into the cell carrying positive charge
3. Delayed opening of voltage-gated K channels, PK>>PNa
K+ flows out of cell removing positive charge
How do we know Na+ important for depolarization?
0 mV
-80 mV
Replace Na+ in extracellular bath with impermeable
cation - cholineNormal
Low Sodium
Ion currents underlying the AP
• Use voltage-clamp technique to measure currents
• Measure currents in the presence and absence of Na+
• What are ionic currents?– So far, voltage (V)– When ions move current (I)– Movement through channel is resistance (R)
• Reciprocal is conductance (g)
Ohm’s law
I=V/R or I=gV
Where g = 1 / R
• More properly
Iion = gion X emfion
• Iion is ionic current
• gion is ionic conductance
• emfion is the electromotive force acting on an ion
• emfion = Vm - Eion
Total membrane potential Nernst potential for the ion
Therefore,
• gion is controlled by ion channels
• If all channels closed, g = 0 and no ions flow
• if Vm = Eion then emf = 0, and no ions flow
( )ion ion ionI g Vm E
Electrophysiology TechniquesVoltage clamp
Squid axon
Recording electrode
Reference electrode
Command Signal
ImCurrent output
Control amplifier
Voltage output
+ -
Currentelectrodes
Membrane currents
• Measure ionic currents from squid axon– To determine contribution of Na+ and K+
Measure in normal saline with Na+ and Na+-free saline
Ion currents underlying the AP
Membrane
Potential
Ionic
Currents
Total current normal saline
K+ current Na+ free saline
outward
inward
Ion currents underlying the AP
Ionic
Currents
Na+ current
outward
inward
Ionic
Currents
Total current normal saline
K+ current Na+ free saline
outward
inward
Subtract K+ current from total
Ion currents underlying the AP
1. The Na+ current activates quickly and then inactivates quickly
2. The K+ current activates more slowly and persists longer
Ion currents underlying the AP
Membrane
Potential
Ionic
CurrentsK+ current
After-hyperpolarization
Pk(leak) + Pk(volt)Pk(leak)
Stimulus & Threshold
• The stimulus depolarizes the membrane– Experimentally applied current– Synaptic potential– Receptor potential
Threshold
• The membrane potential at which
Na flowing into the cell exactly equals the
K flowing out of the cell
• A fraction more stimulus depolarization is required to ‘fire’ an action potential
Membrane
depolarization
Increased Na
permeabilityNa+ entry
Positive Feedback
The AP is regenerative and
displays all-or-none behaviour
Why does the AP stop rising?
1. As VmENa, Na+ inflow stops
2. Na+ channels
inactivate
3. K+ channels open, K+ outflow starts
ENa
Refractory Period
1. A second stimulus very soon after the first will not fire an AP (Absolute)
2. With a delay, a second stronger stimulus will cause a small AP (Relative)
3. With longer delay a second AP can be fired
Why is there Refractory Period?
• The Na channel stays inactivated for a short period of time after it closes
InactivatedOpenClosed
Active
Closed
Active