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Bi / CNS 150 Lecture 2
Friday, October 4, 2013
Voltage-gated channels (no action potentials today)
Henry Lester
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http://www.cns.caltech.edu/bi150/
The Bi / CNS 150 2013Home Page
Please note:
Henry Lester’s office hours
Read the book
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If you drop the course,
or if you register late,
please email Teagan Wall
(in addition to the Registrar’s cards).
Also, if you want to change sections,
please email Teagan
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In the “selectivity filter” of most K+ channels,
K+ ions lose their waters of hydration and are co-ordinated by backbone carbonyl groups
(Like Kandel Figure 5-15)
From Lecture 1
Gate
5
[neurotransmitter]
openclosed
chemical transmission atsynapses:
electric field
openclosed
electrical transmission inaxons:
actually, E
Major Roles for Ion Channels
Future lectures:
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The electric field across a biological membrane, compared with other electric fields in the modern world
1. A “high-voltage” transmission line1 megavolt = 106 V.The ceramic insulators have a length of ~ 1 m.The field is ~ 106 V/m.
2. A biological membraneThe “resting potential” ~ the Nernst potential for K+, -60 mV.The membrane thickness is ~ 3 nm = 30 Å.The field is (6 x 10-2 V) / (3 x 10-9 m) = 2 x 107 V/m !!!
Dielectric breakdown fields (V/m)
Ceramic 8 x 107
Silicone Rubber 3 x 107
Polyvinyl chloride 7 x 106
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open channel = conductor
Na+ channel
=
From Lecture 1
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1973
Max Delbruck
Richard Feynman
H. A. L
Carver Mead
http://en.wikipedia.org/wiki/Carver_Mead
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Intracellular recording with sharp glass electrodes
V
= RC = 10 ms; too large!
C = 1 F/cm2
E
R = 104 -cm2
intracellular
extracellular
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A better way: record the current from channels directly?
Feynman’s idea
A
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5 pA = 104 ions/ms
20 ms
A single voltage-gated Na+ channel
-80 mV
-20 mVA
Dynamic range
10 s to 20 min : 108
2 pA to 100 nA
50,000 chans/cell
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http://www.nobel.se/medicine/laureates/1991/press.html
Press release for 1991 Nobel Prize in Physiology or Medicine:
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Simulation of Shaker gating
http://nerve.bsd.uchicago.edu/model/rotmodel.html
Francisco Bezanilla's simulation program at the Univ. of Chicago.
“Shaker”, a Drosophila mutant first studied in (the late) Seymour Benzer’s lab
by graduate students Lily & Yuh-Nung Jan (now at UCSF);
Gene isolated simultaneously by L & Y-N Jan lab
& by Mark Tanouye (Benzer postdoc, then Caltech prof, now at UC Berkeley).
“Shaker”, a well-studied voltage-gated K+ channel
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Today we emphasize H & H’s description of channel gating
(although they never mentioned channels, or measured a single channel)
Channel opening and closing rate constants are functions of voltage--not of time:
The conformational changes are “Markov processes”.
The rate constants depend instantaneously on the voltage--not on the
history of the voltage.
These same rate constants govern both the macroscopic (summed) behavior and
the single-molecule behavior.
The Hodgkin-Huxley formulation of a neuron membrane
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This channel is actually Shaker with inactivation removed (Shaker-IR).
Based on biochemistry, electrophys, site-directed mutagenesis, X-ray crystallography,
fluorescence.
Two of 4 subunits. Outside is always above (show membrane). Green arrows = K+.
C1 and C2 are closed states, A is “active” = open.
6 helices (S1-S6) + P region, total / subunit.
Structure corresponds roughly to slide 7,
The two green helices (S5, S6 + P) correspond to the entire Xtal structure on slide 4.
First use manual opening. Channel opens when all 4 subunits are “A”.
Note the charges in S4 (5/subunit, but measurements give ~ 13 total). Alpha-helix
with Lys, Arg every 3 rd residue.
Countercharges are in other helices.
Note the S4 charge movement, “shots”. Where is the field, precisely? Near the top.
Note the “hinge” in S6, usually a glycine.
Demonstrating the Bezanilla model, #1
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Read the explanation on the simulation.
Show plot. Manual. Then Voltage (start at default, 0 mV ““delayed rectifier”.
Although we simulate sequentially, the cell adds many channels in parallel.
Not an action potential; this is a “voltage jump” or “voltage clamp” experiment.
Describe shots (measure with fluorescence, very approximately).
I = current. Note three types of I.
Describe gating current (average = I(gate); its waveform does not equal the
I(average).
Show -30 mV (delayed openings,) -50 mV (no openings), 0 (default).
Note tail current.
Note I(gate).
There are many V-gated K channels, each with its own V-sens and kinetics.
Demonstrating the Bezanilla model, #2
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Inactivation: a property of all voltage-gated Na+ channelsand of
Some voltage-gated K+ channels
http://nerve.bsd.uchicago.edu/
http://nerve.bsd.uchicago.edu/Na_chan.htm
Site home:
This model is ~ 10 years older than the K+ channel simulation.
Na+ channel has only one subunit, but it has 4 internal repeats(it’s a “pseudo-tetramer”).
The internal repeats resemble an individual K+ subunit. The “P” region differs, as in Lecture 1, Slide 22.
Orange balls are Na+.Note that the single-channel current (balls inside cell) requires two events: a) All 3 S4 must move up, in response to V;b) Open flap. When the flap closes, the channel “inactivates”.The flap may be linked to the 4th S4 domain.The synthesized macroscopic current shows a negative peak, then decays.
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http://www.krl.caltech.edu/Projects/physicscourses/index.htm
Monday’s lecture employs electrical circuits
See also Appendix A in Kandel
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End of Lecture 3