Lecture 10: Membrane potential and ion channels Fain ch 3 end 10/5/09.
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Transcript of Lecture 10: Membrane potential and ion channels Fain ch 3 end 10/5/09.
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Lecture 10: Membrane Lecture 10: Membrane potential and ion channelspotential and ion channels
Fain ch 3 end10/5/09
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Telomere - protects Telomere - protects chromosome endschromosome ends
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Chromosomes degrade w/o Chromosomes degrade w/o telomeretelomere
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Telomerase adds Telomerase adds telomerestelomeres
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QuestionsQuestions
1. How do you follow membrane potential?
2. What can you learn from evolutionary relationships of ion channels?
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Example - Membrane Example - Membrane potential in hair cellspotential in hair cells
Important systemsAuditory VestibularLateral line
Hair cell responds to mechanosensationBending causes electrical response
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Bullfrog inner earBullfrog inner ear
Very accessible Use the
sacculusLarge hair cellsResponds to head movement (slow frequency)May respond to sound
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Frog sacculusFrog sacculus
Maculus is sensory epithelium (location marked by | | |)
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Hudspeth and Corey 1977Hudspeth and Corey 1977
Hair cells of inner ear (bull frog)
BundlesKinociliumStereocilia -
microvilli
HC = hair cellSC = supporting cellArrows point to kinocillium
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Hudspeth and Corey 1977Hudspeth and Corey 1977
Remove otolithic membrane (OM) to reveal hair cells
Use stimulus probe (SP) to perturb hair cell
Record intracellular potential with microelectrode (ME)
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Hair cell motionHair cell motionTowards kinocilium Depolarize
Inside cell less negative
Away from kinocilium Hyperpolarize
Inside cell more -
Sideways motion had no effect
Depol >> Hyperpol
Fain fig 3.11
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How can we explain this How can we explain this result?result?
Are channels opening or closing?
What ions are moving?
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Cell membrane contains ion pumps Cell membrane contains ion pumps and channels - create concentration and channels - create concentration
gradientsgradients
Na/K ATPaseOutside cell Inside cell
Na+
K+
15 mM
120 mM
Na+
K+
141 mM
3.3 mM
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Pump sends NaPump sends Na++ out outChannel lets NaChannel lets Na+ in+ in
Na/K ATPaseOutside cell Inside cell
Na+
K+
15 mM
120 mM
Na+
K+
141 mM
3.3 mM
NaNa++ pumped out
NaNa++ flows in through open channel
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Pump sends KPump sends K++ in inChannel lets KChannel lets K+ + outout
Na/K ATPaseOutside cell Inside cell
Na+
K+
15 mM
120 mM
Na+
K+
141 mM
3.3 mM
KK++ flows out through open channel
KK++ pumped in
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Possible mechanismsPossible mechanisms
Motion rel kino
Away Toward
Cell Hyperpol Depol
Channel
Na+
Na+ channel
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Possible mechanismsPossible mechanisms
Motion rel kino
Away Toward
Cell Hyperpol Depol
Channel
Close Open
Na+ Pump out Flow in
Na+ channel
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Possible mechanismsPossible mechanisms
Motion rel kino
Away Toward
Cell Hyperpol Depol
Channel
Close Open
Na+ Pump out Flow in
Na+ channel
Motion rel kino
Away Towards
Cell Hyperpol Depol
Channel
K+
K+ channel
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Possible mechanismsPossible mechanisms
Motion rel kino
Away Toward
Cell Hyperpol Depol
Channel
Close Open
Na+ Pump out Flow in
Na+ channel
Motion rel kino
Away Toward
Cell Hyperpol Depol
Channel
Open Close
K+ Flow out Pump in
K+ channel
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Which is it?Which is it?K+
Na+
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Voltage clampingVoltage clamping
Hold cell at fixed voltage
Measure current flow across membraneDirectionSize
Fig 3.13
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Ohm’s lawOhm’s law V = I R
Voltage = current * resistance
Current = voltage / resistance
I = V / R
But conductance,g is 1/RI = V g
V
R
I
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Cell is a resistance / Cell is a resistance / conductanceconductance
Resistance and conductance depend on how many channels are open
Measure current to learn about conductance
Fig 3.13
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Voltage clampingVoltage clamping Current flow
Erev is potential at which no current flows Potential which balances ion concentration
gradient
Vm is membrane potential during stimulation
€
E rev =RT
FlnαNao +KoαNai +K i
€
i = g(Vm − E rev )
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Calculate ECalculate Erevrev for hair cells for hair cells equally permeable to Naequally permeable to Na++ and and
KK++
Na/K ATPase
Outside cell Inside cell
Na+
K+
15 mM
120 mM
Na+
K+
141 mM
3.3 mM
€
E rev = 59mV ln140 + 3.3mM
15 +120mM= −1mV=1
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For hair cells, because For hair cells, because EErevrev~0~0
€
i = gVm
Ion current is proportional to conductance
As stimulate hair cell, conductance changes
Voltage gated current is prop to conductance
€
Δg =ΔimVm
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Current flow directionCurrent flow direction
Fig 3.14
Displace toward kinocilliumDepolarization
Vm positive, current is positiveCurrent flows out
Vm negative, current is negativeCurrent flows in
Vm
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Hair cell stimulusHair cell stimulus
Conductance change
So movement towards kinocillium increases conductance €
Δg =ΔimVm
=−current
−60mV= positive
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Hair cell stimulusHair cell stimulus
Conductance change
So movement towards kinocillium increases conductance Channels openNa channels!
€
Δg =ΔimVm
=−current
−60mV= positive
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Evolution of ion channelsEvolution of ion channels
How are different ion channels related?
What are structural similarities?
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KK++ channel channel
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Simplified 2TM channelSimplified 2TM channel
Roderick Mackinnon used the Streptomyces lividans channel in his Xray crystallography studies
Found it was similar to vertebrate K+ channels because both are blocked by neurotoxins
Only need 2 transmembrane TM regions and the pore region
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KK++ ion pore formed from 4 ion pore formed from 4 subunitssubunits
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Ion selectivity determined Ion selectivity determined by S5, S6 and poreby S5, S6 and pore
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S1-S4 adds channel gatingS1-S4 adds channel gating
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How channels are gated by How channels are gated by voltagevoltage
Nature 423 (2003) 42-8
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Voltage sensitive paddles - Voltage sensitive paddles - move to open and close move to open and close
channelchannel
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Large motion of S4 helix in Large motion of S4 helix in response to charge : response to charge :
Arginines (+)Arginines (+)
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Family Family of ion of ion
channelschannels
Label Ion :K, Na, Ca How channel is gated: voltage Ca
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Root is Root is likely likely
the 2TM the 2TM channelschannels
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TM channelsTM channels
Bacteria group A
Bacteria group B
Bacteria group C
Verts+inverts
Verts+inverts
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Gain of S1-S4 enables Gain of S1-S4 enables voltage gatingvoltage gating
Bacteria group A
Bacteria group B
Bacteria group C
Verts+inverts
Verts+inverts
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Some Some species species
have have 4x6TM 4x6TM regions regions
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Voltage gated sodium channelVoltage gated sodium channelResult of gene duplication and Result of gene duplication and
fusionfusion
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Multimeric channelsMultimeric channels
Bacteria
Yeast
Jelly, cnidarians, inverts
Verts+inverts
Verts
Na+
Ca+
Na+
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Phylogenies of different Phylogenies of different channelschannels
What would be difficult about building a tree comprised of these kinds of genes?
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CNG CNG channels channels
are are importaimportant for nt for vision vision and and
smellsmell
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Ion channel summaryIon channel summary
Structure and function reasonably well understood
Domain and gene duplications followed by fusions played role
Diversity of ways to gate channels
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Crystal structure of KCrystal structure of Kvv channel in open statechannel in open state