Lecture 2 Physiology of the Nervous System

Excitability and ionic transport

Dr. Ana-Maria Zagrean

Discipline of Physiology and Fundamental Neurosciences

Excitability and Ionic transport

1) input
Excitability

2) integration
3) output

The capacity / condition for a

live system to recognize
and respond to specific
signals, as a form of
updated information,
necessary for its adaptive
and continuous organization.

physical

stimulus

chemical
?

Receptor

Interaction stimulus - receptor

response

The excitability of neurons is based on ion gradients

across the cell membrane, and on transport properties
of the cell membrane.

Premises
Interrelation between cell membrane properties and the
complex characteristics of biological systems.

acquiring
excitability

identity

Selective
membrane
permeability

adaptability

coordinating

disseminating
evolution

Selective membrane permeability:

The lipid barrier of the cell membrane and cell membrane transport proteins

Chemical compositions of
extracellular and intracellular fluids.

Diffusion Versus Active Transport

Diffusion - random molecular movement of substances molecule by molecule, either
through intermolecular spaces in the membrane or in combination with a carrier
protein.
The energy that causes diffusion is the energy of the normal kinetic motion of matter.
Active transport - movement of ions or other substances across the membrane in
combination with a carrier protein in such a way that the carrier protein causes the
substance to move against an energy gradient, such as from a low-concentration state
to a high-concentration state; requires an additional source of energy (ATP) besides
kinetic energy.

Cell membrane and its selective permeability

TRANSPORT OF SUBSTANCES THROUGH THE CELL MEMBRANE

1.Diffusion
-Simple diffusion:
- lipid-soluble subst. (O2, CO2, alcohols) through intermolecular
spaces of the lipid barrier
- through a membrane opening - protein channels (e.g., water,
lipid-insoluble molecules that are water-soluble and small
enough):
selective permeable channels
non-gated OR gated (open/closed by gates)
voltage-gated
ligand-gated (chemical-gated)
-Facilitated diffusion = carrier mediated diffusion
e.g. transport of most of aminoacids and glucose
Driving force of diffusion and net diffusion depends on:
-Substance availability, kinetic energy, membrane permeability
-Concentration difference/gradient
-Membrane electrical potential effect on diffusion of ions
(see Nernst potential)
2.Active transport
- Primary active (pumps)
- Secondary active (co- and counter-transport)

Simple diffusion through protein channels

Pores/channels integral cell membrane proteins
Always open
Pore diameter, its shape and its internal electrical charge/chemical bonds
provide selectivity
Aquaporins/water channels (13 different types) - protein pores which
permit rapid passage of water through cell membranes but exclude other
molecules (a narrow pore permits water molecules to diffuse through the
membrane in single file). The pore is too narrow to permit passage of any
hydrated ions. Density of aquaporins (e.g., aquaporin-2) in cell
membranes is not static but is altered in different physiological conditions.

Transport of Na+ and K+ through protein channels

Na +

K+
Conformational changes in the protein molecules
open or close "gates" guarding the channels:
voltage or ligand gated

Na+ channel is only 0.3 by 0.5 nm in diameter,

the inner surfaces of this channel are strongly
negatively charged and pull small dehydrated
sodium ions into these channels selective
permeability for Na+.
K+ channels are up to 0.3 nm in diameter,
but their inner surfaces are not negatively
charged K+ are not pulled away from the
water molecules that hydrate them the
smaller hydrated potassium ions can pass
easily through the channel, whereas the
larger hydrated sodium* ions are rejected,
thus providing selective permeability for K+.

K+ are slightly larger than Na+

but Na+ attracts far more water molecules...

The structure of a potassium channel.

- four subunits, each with two transmembrane helices.

- a narrow selectivity filter formed from the pore loops
carbonyl oxygens line the walls of the selectivity filter, forming sites for
transiently binding dehydrated K+. The interaction of the K+ with carbonyl
oxygens causes them to shed their bound water molecules, permitting the
dehydrated K+ to pass through the pore.

Gating of protein channels to control channel permeability

VOLTAGE GATED:
-for Na+ channel:
strong negative charge on the inside of the cell
membrane outside Na+ gates remain tightly closed
when the inside of the membrane loses its negative charge, these gates would
open suddenly and allow tremendous quantities of Na+ to pass inward through the
sodium pores.
-for K+ channel:
gates are on the intracellular ends of the K+ channels, and they open when the
inside of the cell membrane becomes positively charged.
CHEMICAL (LIGAND) GATING:
- protein channel gates are opened by the binding of a chemical substance (a
ligand) with the protein; this causes a conformational or chemical bonding change
in the protein molecule that opens or closes the gate.

Diffusion: simple and facilitated

Effect of concentration of a substance on rate of

diffusion through a membrane by simple
diffusion & facilitated diffusion.
Note that facilitated diffusion approaches a
maximum rate Vmax. what limit it?

Postulated mechanism for

facilitated /carrier mediated diffusion
(e.g. glucose or amino acids transport)
- glucose transport by GLUT4 is increased
10-20x by insulin (but not in the brain).

Diffusion and the cell membrane potential

A. Net Diffusion Rate Is Proportional to the

Concentration Difference Across a
Membrane.
B. Effect of Membrane Electrical Potential on
Diffusion of IonsThe Nernst Potential.

C. Effect of a Pressure Difference Across the

Membrane.

Diffusion and the cell membrane potential

Net Diffusion Rate Is Proportional to the

Concentration Difference Across a Membrane.

Diffusion and the cell membrane potential

Effect of membrane electrical potential on diffusion of ions: the Nernst Potential
The concentration of (-) ions is initially the same on both sides of the membrane, but a (+) charge
applied to the right side of the membrane and a (-) charge to the left, creates an electrical gradient
across the membrane, moving the ions... When the concentration difference rises high enough, the
two effects balance each other.

EMF (mV) = -RT/zF log C inside/C outside

EMF = +/-61 log C inside/C outside at 37C for any univalent ion (z=1), as Na+ or K+
R = gas constant, F = Faraday constant, z = valence, T = temp; C = ion conc.
(+) for negative ions / (-) for positive ions, diffusing from inside to outside.

Diffusion and the cell membrane potential

Effect of a Pressure Difference Across the Membrane.

Diffusion and the cell membrane potential

A, Establishment of a "diffusion potential" across a nerve fiber membrane, caused by

diffusion of K+ from inside the cell to outside through a membrane that is selectively
permeable only to K+:
within ~1 msec. diffusion potential becomes great enough to block further net K+
diffusion to the exterior, despite the high K+ concentration gradient. For a normal nerve
fiber, the potential difference is ~ 94 mV, with negativity inside the fiber membrane.

Diffusion and the cell membrane potential

B, Establishment of a "diffusion potential" when the nerve fiber membrane is

permeable only to Na+: the membrane potential rises high enough within msec. to
block further net diffusion of Na+ to the inside; however, this time, in the mammalian
nerve fiber, the potential is ~ 61 mV positive inside the fiber.
Internal membrane potential is negative when K+ diffuse and positive when Na+
diffuse because of opposite concentration gradients of these two ions.

Calculation of the diffusion potential when the membrane is

permeable to several different ions Goldman equation

(Actual potential)

Calculation of the diffusion potential when the membrane

is permeable to several different ions Goldman equation
The membrane is permeable to several different ions and the diffusion
potential that develops depends on three factors:
(1) the polarity of the electrical charge of each ion,
(2) the permeability of the membrane (P) to each ion,
(3) the concentrations (C) of the respective ions on the inside (i) and outside
(o) of the membrane.
Goldman /Goldman-Hodgkin-Katz (GHK) equation gives the
calculated membrane potential (Vm) on the inside of the membrane
when two univalent positive ions, sodium (Na+) and potassium (K+), and
one univalent negative ion, chloride (Cl-), are involved.

The passive properties of a plasma membrane

A plasma membrane has resistance, capacitance and conductance

Electro-neutrality principle

The membrane maintains a separation of charges, as an electrical dipole layer.

Electrostatic forces between charges keeps them in close proximity to the membrane.
To establish the normal "resting potential" of -90 mV inside the nerve fiber, only about
1/3,000,000 to 1/100,000,000 of the total (+) charges inside the fiber needs to be
transferred. Also, an equally small number of (+) ions moving from outside to inside
the fiber can reverse the potential from -90mV to as much as +35 mV within 0.1 msec !
Rapid shifting of ions in this manner causes the nerve signals.

Cell Membrane Potential

Proteins & phosphates are negatively charged at normal cellular pH.
These anions attract positively charged cations that can diffuse through the membrane
pores.
Membrane more permeable to K+ than Na+.

Relation between cell membrane potential - membrane ionic transport system

Membrane ionic transport system (MITS)

1 - Ion channels
2 - Ion pumps
3 - Ion exchangers, carriers, co/counter transporters

Components of membrane ionic transport systems

H+

H+

K+

3Na+

Ca2+

CARRIERS

PUMPS

K+
Cl-

CHANNELS

K+

Cl-

Ca2+

AA
Na+

Cl-

1. Ion channels
Gated (active) Ion Channels
- Voltage gated
- Ligand gated
- Mechanic gated
Non-gated (passive) Ion Channels
The diversity of ion channels is significant, especially in excitable cells
of nerves and muscles. Of the more than 400 ion channel genes currently
identified in the human genome, about 170 encode potassium channels,
38 encode calcium channels, 29 encode sodium channels, 58 encode
chloride channels, and 15 encode glutamate receptors. The remaining are
genes encoding other channels such as inositol triphosphate (IP3) receptors,
transient receptor potential (TRP) channels and others.

Gated (active) Ion Channels

Gated (active) Ion Channels

VOLTAGE-GATED ION CHANNELS:
ion selective pore, voltage sensor, activation/inactivation gate

1
2

Voltage-gated K+ channel

At rest, negative cellular potential keep voltage-gated K+ channels closed.

Depolarization cell potential becomes positive K+ channel is activated and can
conduct K+ ions (yellow arrow, right).

The conformation change from the closed to the open state is driven by the
movement of the positively-charged amino acids (orange "+" symbols) located in a
region of the protein called the voltage sensor surrounding the central ionic pore.

The voltage-gated Na+ channel

- used in the rapid electrical signaling
- components:
- ion selectivity filter for Na+:
Na+ discard the water molecules associated with them in order to pass in single
file through the narrowest portion of the channel
- activation gate that can open and close, as controlled by voltage sensors,
which respond to the level of the membrane potential
- inactivation gate limits the period of time the channel remains open, despite
steady stimulation.
a subunit: polypeptide chain of >1800 am.ac. embedded in cell membrane.
* Nonpolar side chains coil into transmembrane alpha-helices and face
outward where they readily interact with the lipids of the membrane.
* By contrast, the polar peptide bonds face inward, separated from the lipid
environment of the membrane.
b subunit: anchor the channel to the plasma membrane
- activation:
- at resting membrane potential (-90-70 mV) the channel is closed;
- the voltage sensor moves outward and the gate opens if any factor depolarize
the membrane potential sufficiently (threshold ~ -50 mV).

Voltage gated Na+ channel

From Basic Neurochemistry, 7th Edition

An experimental strategy to study the ionic currents passing through

the membrane, is one using agents that specifically block either the
voltage-gated Na+ channels or the voltage-gated K+ channels.
Tetrodotoxin (TTX) is a highly potent toxin that inhibits voltage-gated
Na+ channels. The source of TTX is the puffer fish (fugu), that is a
delicacy in some countries. Even minute quantities of ingested TTX
are fatal!

Local anesthetics / nerve blocking agents such as lidocaine

(Xylocaine) and procaine (Novacaine) prevent the generation of
APs by inhibiting voltage-gated Na+ channels of sensory neurons.
Thus, depolarization elicited by sensory stimulation does not lead to
the generation of action potentials that can travel to the CNS.
Tetraethyl ammonium (TEA) is a chemical agent that inhibits the
voltage-gated K+ channels.
Blockers such as TTX and TEA have been instrumental in revealing
the workings of ion channels and their roles in neuronal function.

Voltage-gated Na+ channel

Gated (active) Ion Channels

Ligand-gated ion channels : ionotropic vs metabotropic
Ionotropic directly gate ion channels
Metabotropic indirectly gate channels via 2nd messengers

Ligand-gated ion channels - glutamate receptors:

- NMDA & AMPA ionotropic receptors
- metabotropic group I & II receptors (G-prot. coupled)

PCP- phenylciclidine

Gated (active) Ion Channels

Mechanic gating ion channel
extracellular

Anchoring
situs

Cell
membrane

intracellular
Fibrillary
protein

gate

Non-gated (passive) Ion Channels

K+ leak channels

2. Ion Pumps
Functional particularities:
-active transport of ions and organic molecules
against concentration gradient

- involve enzymatic reactions, ATP consume

-decreased transport rate

Ex: Na+/K+ pump, H+ pump, Ca2+ pump...

Ion Pumps

Na+-K+ ATPase pump can run in reverse: If the electrochemical gradients for
Na+ and K+ are experimentally increased enough so that the energy stored in their
gradients is greater than the chemical energy of ATP hydrolysis, these ions will move
down their concentration gradients and the Na+-K+ pump will synthesize ATP from
ADP and phosphate.
The phosphorylated form of the Na+-K+pump can either donate its phosphate to ADP
to produce ATP or use the energy to change its conformation and pump Na+ out of
the cell and K+ into the cell. The relative concentrations of ATP, ADP, and phosphate,
as well as the electrochemical gradients for Na+ and K+, determine the direction of the
enzyme reaction.
For nerve cells, 60 to 70 % of the cells energy requirement may be devoted to
pumping Na+ out of the cell and K+ into the cell.

The Na+-K+ Pump

- The Na+-K+ Pump Is Important For Controlling Cell Volume.

Inside the cell are large numbers of proteins and other organic molecules that
cannot escape from the cell. Most of these are negatively charged and
therefore attract large numbers of potassium, sodium, and other positive ions
as well. All these molecules and ions then cause osmosis of water to the interior
of the cell. Unless this is checked, the cell will swell indefinitely until it bursts.
The normal mechanism for preventing this is the Na+-K+ pump.
Na+-K+ ATPase pump initiates osmosis of water out of the cell
- Electrogenic Nature of the Na+-K+ Pump

Calcium Pump

Ca2+

Ca2+

3. Ion Exchangers/
Carriers/Cotransporters
- Na/Ca
- Na/H
- Na/HCO3
- Na/ aa, Na/G

- Cl/HCO3 - Na/K/2Cl

- K/Cl, etc

Ion gradients, channels, and transporters in a typical cell (Boron, 2009)

Cell membrane potential

Resting Membrane Potential
The basis for the resting membrane potential:
Na+-K+ pump

K+-Na+ leak channels

Humans
[Na+]

[K+]

Out

142

In

14

140

Ratios
Na+ In:Out = 0.1
K+ In:Out = 35.0

Membranes are 100X more

permeable to K+, as there
are more leakage channels
for K+ (see no. of genes)

Slow rate of Na+ influx is accompanied by slow rate of K+ efflux.

Na+/K+ pump maintains the concentration gradients.
Electrochemical gradient membrane potential of ~ - 65 mV

diffusion
2 K+

3 Na+
pump

Factors that influence the resting membrane potential

The Na+ /K+ pump contributes to

resting membrane potential in 2 ways:
Pumping Na+ & K+ ions in a 3:2 ratio
Maintaining a high K+ concentration
in the cells interior
The membrane conductance to K+
far exceeds that to Na+ :
K+ leakage results in internal
electronegativity

How is membrane potential measured?

When the neuron is inactive, the

membrane is said to be at rest and
has a resting membrane potential
When the neuron is active, the flow
of information is from soma to axon
terminal action potentials (AP).

A Motor Neuron

Membrane responses to stimulus current

Hyperpolarizing currents produce
responses 1 and 2.
A small depolarizing current
produces response 3.
These are all graded local
responses which dissipate locally.
A sufficiently large current
(threshold) produces an action
potential (4), which can be
propagated along the axon.
Animation at http://www.sumanasinc.com/webcontent/animations/neurobiology.html

-A stimulus initiates a membrane

electrical change that depend on the
passive properties of the neuronal
membrane

-Electrical signal /potentials are

initiated by local current flow
-Local potentials then spread
electrotonically over short distances,
and decay with distance from their
site of initiation (as some of the ions
leak back out across the cell
membrane and less charge reaches
more distant sites);
Considering the Ohms law and a
stable membrane resistance, the
diminished current with distance
away from the source results in a
diminished voltage change.

- When the potential is equal/over

threshold, it propagates over a long
distance
- at the axon hillock level, the
potential initiates an action potential
(AP) that propagates without changing
its amplitude

- APs depend on a regenerative wave

of channel openings and closings in
the membrane

Action Potential (AP)

nerve impulse = action potential:

cycle of depolarization & repolarization
needs no direct energy
all-or-none principle
The action potential is essential to our understanding of
nervous system function. Its shape, velocity of conduction,
and propagation fidelity are essential to the timing,
synchrony, and efficacy of neuronal communication.
G. J. Kress and S. Mennerick / Neuroscience 158 (2009) 211222

Action Potential
-The necessary actor in causing both depolarization
and repolarization of the nerve membrane during the
action potential is the voltage-gated Na+ channel
-A voltage-gated K+ channel also plays an important
role in increasing the rapidity of repolarization of the
membrane.
-These two voltage-gated channels are in addition to
the Na+-K+ pump and the K+-Na+ leak channels.

Na+ permeability
increases 500-5000 x

The nerve action potential

Profile of a Nerve Action Potential

Threshold
-Occurs when Na+ entering exceeds K+
leaving
-A rise in potential of 15-30 mV is required
The All-or-None principle
An action potential will not occur until the
initial rise in membrane potential reaches
threshold. However any larger stimulus
produces no greater response than that
produced by the threshold stimulus, i.e.,
the threshold stimulus produces the
maximal effect the action potential.

The nerve action potential

Resting Stage
Depolarization Stage
Result of
Voltage-gated
Na+ channels

Membrane is polarized
i.e., a 90 mV membrane resting potential present
Membrane becomes very permeable to Na+ ions
Influx of Na+ ions
Polarized state is neutralized
Potential merely approaches in smaller CNS fibres
Membrane potentials overshoots beyond zero in
large fibres

Repolarization

Na+ channels get inactivated

Permeability to K+ increases

After-Hyperpolarization

K+ channels remain open after repolarization

Cation conductances during an action potential

action potential

Ion conductance
Na+ conductance increases faster
and lasts for a shorter duration.
K+ conductance is delayed,
increases slowly and lasts longer

The Action Potential and the

positive feedback of the
Na+ channels activation START

+ feed-back

Roles of other ions than Na+ and K+ during the AP

Impermeant anions inside the nerve axon
Calcium Ions:
- calcium pump pumps calcium ions from the interior to the exterior of the cell
membrane (or into the endoplasmic reticulum of the cell), creating a calcium ion gradient
of about 10,000-fold (internal cell conc ~10-7 molar).
- voltage-gated calcium channels slightly permeable to sodium ions as well as to
calcium ions; when they open, both calcium and sodium ions flow to the interior of the
fiber = Ca++-Na+ channels. The calcium channels are slow to become activated (slow
channels), requiring 10 -20 x as long for activation as the sodium channels
Increased permeability of the Na channels when there is a deficit of Ca2+
- extracellular Ca concentration effect on the voltage level at which the Na channels
become activated: a deficit of Ca2+ of ~50% determine Na channels to become activated
by very little increase of the membrane potential from its normal, very negative level
nerve fiber becomes highly excitable, sometimes discharging repetitively without
provocation rather than remaining in the resting state spontaneous discharge in
peripheral nerves, often causing muscle "tetany" (lethal when triggering tetanic
contraction of the respiratory muscles).
-mechanism: Ca2+ appear to bind to the exterior surfaces of the Na channel protein
molecule. The positive charges of Ca in turn alter the electrical state of the channel
protein itself, in this way altering the voltage level required to open the sodium gate.

Membrane Refractoriness

Refractoriness = non-responsive state

Involves Na channel inactivation
Absolute refractory period (ARP)membrane is not responsive to any
stimulation
Relative refractory period (RRP) membrane is responsive to
supra-threshold stimuli

Distribution - function relation for different types of

channels on nerve cell membrane
-Nongated ion channels - throughout the neuron
-Ligand-gated channels - more at sites of synaptic contact
(dendritic spines, dendrites, somata); also, at non-synaptic sites.
-Voltage-gated channels - predominantly on axons and axon
terminals

A spinal motor neuron.

Sodium channels (red);
Microtubule-associated
protein 2 - MAP2 (green)
(Shrager Lab)

Na channels distribution and

generation of AP in axon hillock
The soma membrane has few Na+
channels it is harder to have sufficient
Na+ influx to change membrane potential
to the threshold potential (-45 mV).
A voltage change up to +30 mV is required
Axon hillock membrane has 7x more Na+
channels than the soma membrane and
the threshold potential is lower (a voltage
change of only +10+20 mV is required to
bring the membrane potential to threshold)
= trigger zone for AP
Action potentials in postsynaptic
neurons are initiated at the axon hillock.

Simultaneous recording of
action potentials from different
parts of a neuron.
A, an excitatory synapse on a
dendrite is stimulated, and the
response near that dendrite is
recorded in the soma and at the
initial segment. The excitatory
postsynaptic potential (EPSP)
attenuates in the soma and the
initial segment, but the EPSP is
large enough to trigger an action
potential at the initial segment.
B, The threshold is high (-35 mV) in
regions of the neuron that have few
Na+ channels but starts to fall
rather steeply in the hillock and
initial segment. Typically, a stimulus
of sufficient strength triggers an
action potential at the initial
segment.
C, The density of Na+ channels is
high only at the initial segment and
at each node of Ranvier.

AP generation and conductance along the axon

- initial depolarization at the axon hillock +f.b. for Na+ channels critical
membrane potential = threshold (all-or-none response)

-AP: depolarization and repolarization, followed by afterhyperpolarization, as

K+ channels remain open and membrane permeability for K+ is higher
- propagation of AP to the axon terminals synapses
- also backpropagation in the soma & dendrites, without regenerating in the somal
membrane, as somal membrane has too few Na+ channels to regenerate APs; also,
inactivation of Na channels at axom hillock (here, refractory period).

-Speed of propagation depend on axon diameter & presence of myelin sheath

-in unmyelinated axons, Na & K voltage-gated channels are

uniformly distributed AP as a traveling wave
-large diameter axons allow a grater flow of ions grated length of
the axon to be depolarized increase of the conduction velocity
-in myelinated axons, myelin sheath insulate the axon membrane
generation of AP between the myelinated segments, at the nodes of
Ranvier saltatory conduction

Propagation of impulses from the axon hillock

Once the action potential begins, the potential travels forward
along the axon and usually also backward toward the soma.
However it does not regenerate in the soma membrane.

Why is regeneration impossible in the soma membrane?

EPSPs arrive and an AP is generated at the axon hillock.
The AP is regenerated forward to the axon, depolarization
spreads backwards to soma and dendrites, but impulse
potential decays dies because the somal membrane has
too few Na+ channels to regenerate APs.

Saltatory Conduction

current flows electronically to the next node

action potentials are regenerated only at nodes
action potential jumps from node to node

Propagation of an Action Potential

Action Potential travels along the membrane as a wave

of depolarization. Directional propagation of an AP

Speed of propagation depend on

the presence of myelin sheath

AP generation and conductance in the sensory neurons:

trigger zone is near the peripheral target.

Condition associated with channelopathies

(congenital or acquired (often resulting from autoimmune
attack on an ion channel).
Alternating hemiplegia of childhood
Congenital hyperinsulinism
Cystic fibrosis
Episodic Ataxia
Erythromelalgia
Generalized epilepsy with febrile seizures plus
Familial hemiplegic migraine
Hyperkalemic periodic paralysis

Channel type
Na/K-ATPase
Inward-rectifier potassium ion channel
Chloride channel
Voltage-gated potassium channel
Voltage-gated sodium channel
Voltage-gated sodium channel
various
Voltage-gated sodium channel

Hypokalemic periodic paralysis

Voltage-gated sodium channel

or voltage-dependent calcium channel (calciumopathy)

Long QT syndrome
Malignant hyperthermia
Mucolipidosis type IV
Myasthenia Gravis
Myotonia congenita

various
Ligand-gated calcium channel
Non-selective cation channel
Ligand-gated sodium channel
Voltage-dependent chloride channel

Neuromyotonia
Nonsyndromic deafness
Paramyotonia congenita (a periodic paralysis)

Voltage-gated potassium channel

various
Voltage-gated sodium channel

Retinitis pigmentosa(some forms)

Ligand-gated non-specific ion channels

Short QT syndrome

various potassium channels suspected

Timothy syndrome

Voltage-dependent calcium channel

Seizure

Voltage-dependent potassium channel