The sodium-potassium
pump and the membrane
potential at rest
LESSON NR. 14 - PSYCHOBIOLOGY
The sodium-potassium pump
Ion pumps, in contrast to ion channels, mobilizing the ions across the plasma membrane against their electrochemical
gradient, and then with an active transport performed by continuous energy expenditure.
Ion pumps are varied, but for the neuron the most important of these is undoubtedly the pump Na + / K +, also called ATPase
Na + / K + -dependent.
The sodium-potassium pump
The great importance of this pump is made evident by the fact that its operation requires about 25% of the total
requirement of cellular ATP of the entire organism at rest, and in neurons that number rises to around 70%.
The main function of such a pump is the maintenance of the cellular homeostasis through the regulation of osmotic balance
and the construction of the transmembrane gradient of Na + ions.
The sodium-potassium pump
From the structural point of view the Na + / K + pump is composed of 2 sub-units alpha and beta.
The alpha subunit binds ATP, the Na + and K + ions and also contains the phosphorylation sites that regulate the functioning
of the pump itself.
The beta subunit is responsible for localization of the pump in the plasma membrane and activation of the alpha unit.
The sodium-potassium pump
The operation of the pump can be schematised according to a model based on the acquisition of sequential conformational
changes, each of which is a consequence of the previous and because of the later.
These changes, induced by a series of phosphorylations and de-phosphorylations lead the pump to open alternately
inwards and outwards and then return to the initial configuration, ready to perform a new cycle of operation.
During each cycle, the pump extrudes from the inside of the cell 3 Na + ions and move in 2 K + ions, obtaining the energy for
both counter-gradient transport, through the degradation of a molecule of ATP to ADP.
The sodium-potassium pump
1)
In the initial configuration pump is open to
the inside, and link 3 Na + ion
2)
The bond with the 3 ions, activates the
enzyme ability to bind ATP and to selfphosphorilates releasing ADP.
3)
The condition of phosphorylation, induces a
conformational change that consists in
closing the pump on the intra-cellular side
and the simultaneous opening on the
opposite side. At the same time the links
and the affinity of the pump with the Na +
ions is reduced and these are released
outside of the cytoplasm.
4)
The release of 3 Na + ions, allows 2 K + ions
to bind to the external side of the pump
causing the phosphate on the inner side
from coming off in the form of inorganic
phosphate (de-phosphorylation). This event
induces a second conformational change
that closes the pump outside the reopening it in and releasing the two K + ions.
The sodium-potassium pump
In summary: at each pump cycle,
consuming 1 ATP, it causes the leakage of 3
Na + ions, and lets in 2 K + ions.
So if we consider only the electric charges,
the overall result can be summed up in the
net loss of a positive electrical charge into
the intra-cellular OR in increasing in the
cytoplasm of a negative electric charge.
For this reason, the Na + / K + pump is
defined electrogenic, that is, generating a
potential difference between electrical
charges intra- and extra-cellular. However,
the contribution of the pump to the
potential difference between the two
compartments is equal to only 10%
Protonic pump
The proton pumps perform the movement of H + ions
against their concentration gradient, using the energy
provided by the hydrolysis of ATP.
Important proton pumps are placed in the thickness
of the mitochondrial membrane and allow the H +
accumulation in the mitochondrial outer chamber
Relative to SN, the proton pumps of interest are those
of synaptic vesicles.
Such pumps H + accumulate inside of the vesicles by
creating a concentration gradient that will then be
used in a antiporter to insert the neurotransmitter in
the vesicles.
The neurons are able to take advantage of the concentration gradients and therefore the electrostatic charges of the
ions downstream and upstream of their plasma membrane for generating and transmitting along the same membrane
and between different cells, the nervous stimuli
++++++++ ++++++++ ++++++++ ++++++++
---------- ---------- ---------- ----------
The neurons are able to take advantage of the concentration gradients and therefore the electrostatic charges of the
ions downstream and upstream of their plasma membrane for generating and transmitting along the same membrane
and between different cells, the nervous stimuli
----------
++++++++ ++++++++ ++++++++
++++++++ ---------- ---------- ----------
Two forces act on a charged particle:
The DIFFUSION FORCE,
generated by the
concentration gradient
The POWER SUPPLY,
generated by the electric
gradient
The combination of these two forces pushes the ion flow inside
or outside the cell.
Diffusion
Sodium – potassium pump activity
The different concentrations of free ions within the cell
INTRA (mM)
EXTRA (mM)
Na+
15
150
K+
140
5
Ca2+
10-7 (M)
3
Cl -
4
118
A-
146
1
CATION:
ANION:
The intracellular nature of the negative charges corresponds for the most part to the electronegative phosphate
groups of the main macromolecules that make up the cell:
Nucleic acids
phosphorylated proteins
ATP
Nucleotides mono-, di- and tri-phosphates
All these structures confer largely stable electronegativity to the cell, as by nature they can not abandon the
cytoplasm, being associated with structural and obliged constituents.
For this reason they are defined as fixed anions (A-). In contrast to the free ions (Na +, K +, Cl-, Mg2 +, Ca2 +, etc
...), which instead can move quickly through the plasma membrane whenever they have provided a passage (ion
channel).
The electronegativity caused by the fixed anions within the cell, makes it highly attractive for free cations and
strongly repulsive for free anions
Potassium+
Potassium ions are a good example of balance with respect to internal and external concentrations of the cell (ions
entering = ions that come out).
This is possible thanks to the action of both the Na + / K + pump, which puts the potassium in, and to the presence
of multiple leaks or resting channels for potassium, perpetually open. which allow the exit thanks to facilitated
diffusion according to the concentration gradient.
In other words you can define the plasma membrane as a highly permeable to K + ions.
Despite this, the concentration of potassium is higher inside the cell because most of the potassium ions remains
attracted to the strong negative charge of the intracellular environment. In other words, only the K + ions in excess
on which the force of electrostatic attraction operated from the cytoplasmic environment has no effect, manage to
get out of the cell according to the gradient.
Sodium+
In the case of Na + ions, their exclusion from the intra-cellular environment by the Na + / K + pump is not
counterbalanced by the presence of constantly open channels, and the only sodium that is able to enter it does
thanks to some membrane transporters, using the Na + gradient as a form of energy for secondary active transport.
In other words, the plasma membrane is highly impermeable to Na + ions.
The result of this impermeability is the generation of two different concentrations of Na + highly unbalanced
between interior and exterior (about 10 times).
ChlorineAs regards the Cl- ions, the concentrations, and then the relative balance between inside and outside it is
comparable to the situation described for the K +, although with inverted values.
In fact, even the Cl- can easily pass through the membrane thanks to the presence of always open resting channels.
However compared to K +, the entrance according to the gradient is hindered by the general intracellular negative
charge.
Under normal conditions this ion is then distributed around the membrane.
Calcium+
As already seen, calcium is an important second messenger within
the cells (metabotropic channels) for these reasons its
concentration inside the cell are kept very low thanks to the action
of pumps for the Ca2 + that eliminate these ions as soon as their
concentration rises above a certain level, so as to be able to use the
increase of calcium as a signal for a series of processes:
These include muscle contraction, neuronal transmission as in
an excitatory synapse, cellular motility (including the movement
of flagella and cilia), fertilisation, cell growth or
proliferation, neurogenesis, learning and memory as with synaptic
plasticity, and secretion of saliva. High levels of cytoplasmic calcium
can also cause the cell to undergo apoptosis. Other biochemical
roles of calcium include regulating enzyme activity, permeability
of ion channels, activity of ion pumps, and components of
the cytoskeleton.
Magnesium+
The Mg 2+ ions, are essential for many phenomena associated with enzyme activity and metabolism of nucleic
acids. For example, the presence of Mg2 + is required to assemble the minor and major sub-unit of the ribosome, as
well as for the activity of various enzymes and as already seen, they are involved in the activity of NMDA receptors.
In summary:
• The interaction of the most abundant free ions (Na +, K + and Cl-)
with fixed anions, contributes in constituting an intracellular
environment more electronegative as compared to the
extracellular environment
• While fixed anions are non-diffusible, the membrane is highly
permeable to K + and Cl- which are found in almost equilibrium
conditions, and poorly permeable to Na + that this is in a strong
state of electrochemical imbalance, actively maintained thanks to
Na + / K +.
• A sudden opening of ion channels for Na + can cause an immediate
and impressive alteration of trans-membrane distributions of ions
The intra-cellular electrophysiology
A field of study that comes from the research of Young
Hodgkin & Huxley.
It is based on the idea that the external and internal
environment of the cell are two distinct compartments
characterized by a high electrical conductivity, separated by a
membrane with a high electrical resistance.
Is therefore possible to measure the difference in electrical
potential (voltage difference) just as you do in electric circuits,
thanks to voltmeter.
Volt = potential difference
Ampere = intensity of power
The measurement of the membrane potential
Membrane potential at rest
The measurement of the membrane potential
Thanks to the patch (cell region contained in the diameter of the micropipette) clamp technique is possible to
study and analyze the flow of ions across a few or even a single ion channel.
The measurement of the membrane potential
All cells (not only the excitable cells) have a resting potential: an electrical charge across the plasma membrane,
with the inside of the cell more negative than the outside. The value of the resting potential varies, but in excitable
cells is between -90 and -70 mV.
The measurement of the membrane potential
Goldman equation
It allows you to calculate the value of the resting membrane potential as a function of the variables that determine the value,
such as the concentration and the resistance of the membrane.
R = Gas constant
T = Temperature in Kelvin degrees
F = Faraday constant (96500 coulomb / gram equivalent charge)
Ln = Natural logarithm
Pna, PK, PCl = membrane permeability to ions in question
[...] = Internal and external concentrations
The measurement of the membrane potential
Nernst equation
The relationship that exists between the balance between chemical gradient and electrical gradient was established by the
German chemical-physicist Walter Nernst's, thanks to its formula that allows to calculate the equilibrium potential of the single
ion, i.e. the value of membrane potential in condition of maximum permeability of the membrane for this ion
RT [X]E
E x 
ln
zF [ X] I
R = gas constant
T = absolute temperature (Kelvin)
z = valence of the ion
F = Faraday constant (96500 coulomb / charging
gram equivalent)
[X] i = intracellular concentration of the ion X
[X] E = extracellular concentration of the ion X
Potential at rest
In conclusion, the condition of the greater intra-cellular electronegativity as compared to the outside has important
consequences with regard to the distribution of electrical charges in the immediate vicinity of the two sides of the plasma
membrane.
This condition causes a thickening of positive electrical charges on the exterior of the membrane and this thickening attracts
negative electric charges on the inside.
Therefore the two sides of the membrane are covered with opposite charges that attract but which can not be canceled
because of the separation of the membrane.
If we analyze the nature of these charges, it is clear that the positive charges on the outer face can not be other than Na + ions,
which of the three major ions (Na, K and Cl) are the only with strong electrochemical imbalance and a strong concentration in
the extra-cellular environment.