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LECTURE 3: ION CHANNELS & THE RESTING MEMBRANE POTENTIAL. REQUIRED READING: Kandel text, Chapters 7, pgs 105-139. + + + + + + + + + + + + + + + + + + + + + + + + + + + + +. V m = V in - V out. - - - - - - - - - - - - - - - - - - - - - - - - - -. In resting neuron:. - PowerPoint PPT Presentation
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LECTURE 3: ION CHANNELS & THE RESTING MEMBRANE POTENTIAL
REQUIRED READING: Kandel text, Chapters 7, pgs 105-139
- - - - - - - - - - - - - - - - - - - - - - - - - -
+ + + + + + + + + + + + + + + + + + + + + + + + + + + + +
Vm = Vin - Vout
In resting neuron:
Vm ~ - 60 to - 75 mV
Membrane potential is a BATTERY providing power to drive currentswhen the cell is activated
This lecture discusses how membrane potential is establishedand maintained
- - - - - - - - - - - - - - - - - - - - - - - - - -
+ + + + + + + + + + + + + + + + + + + + + + + + + + + + +
MEASURING THE RESTING MEMBRANE POTENTIAL:MICROPIPET FILLED WITH HIGH SALT
For this method, recording pipet has a very fine tip and is filled with a high salt solution (e.g. 3M KCl), so that pipet has very low resistance.In this way, the voltage measured by amplifier accurately reflects the voltage across the cell membrane.
(method not useful in very small cells due to pipet salt poisoning of cells)
Vpipet Vm (actual)
Rm (actual)Rpipet
Vm (measured) = Vm (actual) + Vpipet
when Rpipet <<< Rm (actual)
VVmm (measured)(measured) = V = Vmm (actual)(actual)
MEASURING THE RESTING MEMBRANE POTENTIAL:PATCH PIPET IN “WHOLE-CELL” CONFIGURATION
Patch pipet filled with cytoplasm-like solution is touched to cell membrane; with negative pressure, the pipet makes a very tight
“cell-attached” or “on-cell” seal onto membrane (leak resistance > 10 G)
Applying gentle suction can break the membrane inside the pipet, making pipet fluidcontiguous with the cytoplasm. This is the “whole-cell” configuration.
When break is made into cell, the pipet can record the membrane potential
TWO TYPES OF PROTEIN COMPLEXES CONTRIBUTE TO ESTABLISHING THE RESTING MEMBRANE POTENTIAL
ION PUMP -- drives a specific ion or group of ions from one side of the plasma membrane to the other side
Pumps drive ions ONE-WAY and use energy from ATP hydrolysisto make the process energetically favorable
ION CHANNEL -- protein complex containing a small pore which allows a specific ion or group of ions to pass
Flow of ions through channels is PASSIVE and is driven by theprevailing chemical and electrical gradients
A channel is an ion-specific resistor with a certain conductance ( g )
For most channels, the conductance is the same for ions flowing IN or OUT
Other channels allow ions to pass with greater conductance in one direction;these are called RECTIFYING CHANNELS
e.g., a channel with greater conductance of inward current is calledan inwardly rectifying channel
Na/K ATPase PUMP
Na+
22
33
ATP ADP + Pi
insideinside
outsideoutside
K+Na+/K+ ATPase USES ENERGY FROM ATP HYDROLYSIS TO PUMP
SODIUM IONS OUT OF CELL &POTASSIUM IONS INTO CELLAT A 3 Na+ : 2 K+ RATIOAT A 3 Na+ : 2 K+ RATIO
CONSEQUENCES OF PUMP ACTIVITY
[ K+ ]in >> [ K+ ]out
[ Na+ ]in << [ Na+ ]out
Net positive charge pumped outof cell causes a matching amount
of permeable chloride anions to moveout passively through channels
[ Cl- ]in << [ Cl- ]out
IONS CHANNELS
insideinside
outsideoutside
K+
Na+POTASSIUMCHANNEL
(non-gated, “leak”)
Some types of ion channels are “gated”, meaning the ion-selective pore can be either open or shut (not in-between)Such channels can be gated by ligands, phosphorylation, or voltage
Other types of ion channels are open all the timeThese channels referred to as “leak” channels
POTASSIUM CHANNELS FAVOR A NEGATIVE MEMBRANE POTENTIAL
Potassium channels are the most abundant leak channels in neurons.
Because the Na/K pump makes [K+]in >> [K+]out , potassium ions move
outwards through channels due to the chemical driving potential, EK .
(EK can be thought of as a potassium “battery”)
Net outward ion flow continues until opposed by a membrane
potential, Vm , of equal force built up in the membrane capacitor.
When Vm = 0, large K+ effluxAT EQUILIBRIUM
When Vm = EK, zero net K+ flux
inin
outout
K+
K+ Na+
Na+
Cl-
Cl- A-
inin
outout
K+
K+ Na+
Na+
Cl-
Cl- A-
+ + + +
- - - -
+ - + -
+ - + -
CIRCUIT REPRESENTATION OF POTASSIUM CONDUCTANCE,POTASSIUM BATTERY, AND MEMBRANE CAPACITANCE
When Vm = 0, large K+ efflux When Vm = EK, zero net K+ flux
inin
outout
K+
K+Na+
Na+
Cl-
Cl- A-
inin
outout
K+
K+Na+
Na+
Cl-
Cl- A-
+ + + +
- - - -
+ - + -
+ - + -
+
-
IK
CM
EK
gK
+
-
IK = 0
CM
EK
gK + + + _ _ _VM = 0 VM = EK
What is the strength of the potassium battery EK ???
THE NERNST EQUATION
The cytoplasmic and extracellular concentrations of an iondetermine the chemical driving force for that ion and
the equilibrium membrane potential if this is the ONLY ionthat is permeable through the membrane
EK+ = 58 mV
1log 5
130
Nernst EquationNernst Equation
Where EX is the chemical potential and z is the charge of ion X
[K+]in = 130 mM [K+]out = 5 mM z = +1
EX = 58 mV
z log [X]out
[X]in
= - 82 mV
For potassium:
inin
outout
K+
K+Na+
Na+
Cl-
Cl- A-
+ + + +
- - - -inin
outout
K+
K+Na+
Na+
Cl-
Cl- A-
+ - + -
+ - + -
When Vm = 0, large K+ efflux
IK = 2 pA
When Vm = EK, zero net K+ flux
IK = 0 pA
Vm
IK
EK = - 82 mV
slope = K = 25 pS IK = 2 pA
VOLTAGE-CURRENT RELATION OF THE POTASSIUM BATTERY
inin
outout
+
-EK = - 82 mV
K = 25 pS
I K =
2
pA
inin
outout
+
-EK = - 82 mV
K = 25 pS
I K =
0
Conductivity of single K channelK = 25 pS
Total K conductivity (gK )
gK = K X NK
where NK is # K channels
IK = gK x ( Vm - EK )
Vm = EK + IK RK
gK and Cm DETERMINE HOW FAST Vm CHANGES TO EK
inin
outout
K+
K+Na+
Na+
Cl-
Cl- A-
+ - + -
+ - + -
Vm (mV)
t
- 82
0
~ Cm / gK
channels open
The greater the value of gK , the greater the potassium current ( IK ) and
the faster the transition to the potassium Nernst potential ( EK)
The greater the value of Cm , the longer the potassium current ( IK ) and
the slower the transition to the potassium Nernst potential ( EK)
RESTING POTENTIAL SET BY RELATIVE PERMEABILITIES
OF K+, Na+, & Cl- IONS
EK = - 82.1 mV 1.0
ENa = + 84.8 mV 0.05
ECl = - 63.6 mV 0.2
Nernst Potential Relative Permeability (P)
Resting membrane potential reflects the relative permeabilitiesof each ion and the Nernst potential of each ion
When the resting membrane potential is achieved, there isongoing influx of sodium and a matching efflux of potassium.
Na/K ATPase is continually needed to keep the ion gradientsfrom running down over time
gK EK + gNa ENa + gCl ECl gK + gNa + gCl
Vm =PK EK + PNa ENa + PCl ECl
PK + PNa + PCl
=~
THE GOLDMAN EQUATION
PK EK + PNa ENa + PCl ECl
PK + PNa + PClVm =
from before
Nernst equatiion EX = 58 mV
z log [X]out
[X]in
Goldman equation Vm = 58 mV log10
PK[K+]o + PNa[Na+]o + PCl[Cl-]i PK[K+]i + PNa[Na+]i + PCl[Cl-]o
( )The greater an ion’s concentration and permeability, the more
it contributes to the resting membrane potentialWhen one ion is by far the most permeable, Goldman eq. reduces to Nernst eq.
RELATIVE PERMEABILITY & THE RESTING POTENTIAL
PK EK + PNa ENa + PCl ECl
PK + PNa + PClVm =
[K+]o
[K+]i
PK
= 5 mM
= 130 mM
= 145 mM
= 5 mM = 8 mM
[Na+]o
[Na+]i
PNa
[Cl-]o = 100 mM
[Cl-]i
PCl= 0.2= 0.05= 1
EK= - 82.1 mV ENa
= 84.8 mV ECl= - 63.6 mV
VVmm = - 72.4 mV = - 72.4 mV
GRAPHIC AND CIRCUIT REPRESENTATIONS OF ION FLOWSACROSS THE MEMBRANE AT THE RESTING POTENTIAL
inin
outout
K+
K+
+ + +
- - -
Cl-
Cl-K+
K+
K+
K+ Na+
Na+
+ + ++ + ++ + +
- - - - - -- - - - - -- - -
+ + + + + +Vm = - 72.4 mV
EEKK = - 82.1 mV = - 82.1 mV EENaNa = + 84.8 mV = + 84.8 mV
IIK K + + IINaNa + + IIClCl = = 00
AT STEADY STATE:
inin
outout
+
-EK = - 82.1 mV
gK = 2 nS
I K =
1
9.4
p
A
RK = 0.5 G
inin
outout
-+ENa = + 84.8 mV
gNa = 0.1 nS
I Na =
-
15
.7
pA
RNa = 10 G
EEKK + I + IKKRRKK = Vm = EENaNa + I + INaNaRRNaNa = EEClCl + I + IClClRRClCl-82.1 mV + (19.4 pA)(0.5 G-82.1 mV + (19.4 pA)(0.5 G)) = -72.4 mV = +84.8 mV + (-15.7 pA)(10 G+84.8 mV + (-15.7 pA)(10 G)) = -63.6 mV + (-3.5 pA)(2.5 G-63.6 mV + (-3.5 pA)(2.5 G))
Vm =inin
outout
-72.4 mV
+++
-- -
outout
-+
ECl = - 63.6 mV
gCl = 0.4 nS
I Cl =
-3
.5 p
A
RCl = 2.5 G
INCREASING SODIUM PERMEABILITY UNDERLIES SODIUM INFLUXAND MEMBRANE DEPOLARIZATION DURING ACTION POTENTIAL
During action potential, the number of open sodium channels increases dramatically
EK = - 82 mV 1.0 1.0
ENa = + 85 mV 0.05 5.05.0
ECl = - 64 mV 0.2 0.2
Nernst Potential Prest Paction-
potential
GOLDMAN EQUATION-PREDICTED Vm
Rest During Action Potential
- 70 mV ++ 36 mV 36 mV
When sodium channels open, sodium ions flow in rapidly because of the negative membrane potential and the strong inward sodium battery
Inward sodium current depolarizes membrane and moves it towards the positive potential predicted by Goldman’s equation
(this positive potential is never fully achieved due to additional channel dynamics)
Next Lecture: MEASURING MEMBRANE CONDUCTANCE AND CAPACITANCE &VOLTAGE-CLAMP RECORDING
REQUIRED READING: Kandel text, Chapters 8, 9 (beginning), pgs 140-153