1. ION CHANNEL : A SITE OF DRUG ACTIONS
Heny Ekowati
Pharmacy Department
Medicine and Health Sciences Faculty
Unsoed, 2013

2. Ion Channels
The plasma membrane is 6-8nm thick, and consists of a mosaic of lipids and proteins. The lipid is hydrophobic, and will not allow ions through.
Ions are surrounded by waters of hydration. To move through the hydrophobic lipid bilayer, water molecules would need to be stripped off the ion. This takes too much energy.

3. Ion Channels
The solution is to provide ions with specialized pathways such as ion channels that will permit ions to cross with most or all of their water molecules.
Ion channel pores therefore provide ions with a polar environment.

4. Structure of Ion Channels
Ion channels are large assemblies of proteins, which make up subunits, which combine to form functional channels.

5. Important features: Gating (opening and closing)
What are the features that cause the channel to open and close?
Ion selectivity
Why does a K+ channel not allow Na+ through?
Molecular features related to their function.
What are the structural features that determine the function of the channel?

6. Gating
What are the features that cause the channel to open and close?

7. Controlled by different types of stimuli:
Gating:
Ligands
Phosphorylation
Controlled by different types of stimuli:
Voltage
Stretch

8. Gating 3 modes of gating:
Involves conformational changes in the ion channel protein.
Each channel protein has two or more conformational states (e.g., open & closed) that are relatively stable.
Each stable conformation represents a different functional state.

9. Ion Selectivity: Why does a K+ channel not allow Na+ through? 0.095 nm

10. Ion Selectivity: Why does a K+ channel not allow Na+ through?
Concept of waters of hydration: Increase the effective diameter of the Na+ ion.
Thus: Pore Size is 1 mechanism for selectivity.

11. Ion Selectivity: So why does an Na channel exclude K?
Channels have a specialized region that acts as a molecular sieve  The SELECTIVITY FILTER.
This is where an ion sheds its waters of hydration & forms a weak chemical bond with charged or polar amino acid residues that line the walls of the channel.

12. Klasifikasi
Kanal ion terdapat pada hampir di setiap sel, fungsinya untuk transport ion, pengaturan potensial listrik melintasi membran sel, dan signaling sel.
Klasifikasi :
- Kanal ion teraktivasi voltage (voltage-gated channel)
- Kanal ion teraktivasi ligan (ligand-gated channel)
- Kanal ion teraktivasi nukleotida siklik atau kalsium
(cyclic nucleotide-gated channel atau calcium-activated
channel)
- Kanal ion teraktivasi oleh kekuatan mekanik (stretch
activated channel)
- Kanal ion terhubung protein G (G protein-gated channel)

13. Voltage-Gated Ion Channels
CLOSED
OPEN
Depolarisasi
Repolarisasi

14. Voltage-Gated Ion Channels
A class of ion channels gated (opened and closed) by the trans-membrane potential difference (voltage).
There are many, many, types. Among these are:
Na+ Channels
K+ Channels
Ca2+ Channels
Cl- Channels.
There are actually many types of Na, K, Cl, and Ca Channels, classified according to pharmacology, physiology, and more recently- molecular structure.

15. Voltage-gated ion channels
Involved in:
Initiation and propagation of action potentials
Control of synaptic transmission
Intracellular ion homeostasis
Other aspects of intracellular function
Acting as activators of intracellular enzymes
Coordinating signals between cell membrane and internal organelles (e.g., mitochondria).

16. Ligand-Gated Ion Channels
Typically, these are ion channels located on the postsynaptic (receiving) side of the neuron
Some act in response to a secreted (external) ligand- typically a neurotransmitter such as
Acetylcholine (Ach)
GABA
Glycine
Glutamate
Some act in response to internal ligands such as cGMP and cAMP, and are also regulated by internal metabolites such as phosphoinositides, arachidonic acid, calcium.

17. Ligand-Gated Ion Channels
Among the first ligand-gated channels to be thoroughly characterized and cloned is the Ach channel.
5 subunits, each made of 4 membrane-spanning components (M1-M4)
2 Ach molecules need to bind in order to open the channel pore.
Fluxes Na and K.

18. Potensial sel Depolarisasi Repolarisasi Hiperpolarisasi
Resting potensial

21. Na+ Channel

22. Sodium Channels - Structure
Composed of α, β-1 and β-2 subunits, but the large α-subunits carries most of the functional properties
4 repeated motifs, each with 6 transmembrane domains
All linked together
Contain a voltage “sensor”/ligand binding domain (method of activation)
The hydrophobic S4 segment (voltage “sensor”) is found in all voltage gated Na+ channels and is absent in ligand gated Na+ channels
Selectivity filter (shell of hydration)
Inactivation gate

23. Cartoon representation of the “typical” voltage-activated
sodium channel

24. Types Of Na+ Channels
Voltage gated – Changes in membrane polarity open the channel
Ligand gated (nicotinic acetylcholine receptor) – Ligand binding alters channel/receptor conformation and opens the pore
Mechanically gated (stretch receptor) – Physical torsion or deformation opens the channel pore

25. Sodium Channels - Function
Play a central role in the transmission of action potentials along a nerve
Can be in different functional states (3)
-A resting state when it can respond to a depolarizing voltage changes
-Activated, when it allows flow of Na+ ions through the channel
-Inactivated, when subjected to a “suprathreshold” potential, the channel will not open

26. The theory is
that the inactivation gate
“swings” shut, turning off
the channel

27. Na+ Channel Modulation
Phosphorylation
sodium channel function is modulated by serine/threonine and tyrosine kinases as well as tyrosine phosphatases (Yu et al, Science 1997)
Mutation – altered amino acid sequence/structure can change the biophysical properties of the Na+ channel
Pharmacology – block Na+ channel to reduce the conductance
Proteolysis- (cleavage) Proteases may cleave specific residues or sequences that inactivate a channel, or significantly alter the biophysical properties

28. Why Na+ Channels/Modulation Are Important
Neuronal depolarization, Action Potential
Neuronal Excitability
Cardiac Excitability
Muscle Excitability
The basis of neuronal/cardiac/muscular function relies on the propagation of action potentials, down axons, sarcolemma, myocardium, as well as requiring synaptic transmission.
Differential excitability alters the electrical conduction/transmission properties of the “circuit”

30. Na + Channel Blockers/Pharmacological Agents
Tetrodotoxin (TTX)
Amiodarone
Lidocaine
Procainamide
Mexilitine
Ketamine
Many, many others

31. Some Na+ Channels Outside The Nervous System
Naf – “Funny Current” in pacemaker cells of the heart (SA node/ectopic pacemakers)
Nav in the myocardium, sarcolemma, and T-tubules and motor endplate

33. K+ Channels

34. K+ Channels:

35. Shaker K+ Channel: Each Channel is made of 4 Subunits.
Each Subunit is made up of a large protein having 6 trans-membrane segments (S1-S6).
Between S5 and S6 there is a loop (red) that, along with the S6 segment, lines the conduction pore.

36. Shaker K+ Channel:

37. What they look like:

38. What they look like:

39. Voltage-gated K+ channels
mediate outward K+ currents during nerve action potentials.
Important advances in understanding have come from:
physiological studies, including the use of patch clamping
mutational studies of the Drosophila voltage-gated K+ channel protein, product of the Shaker gene
crystallographic analysis of the structure of bacterial K+ channels.
molecular dynamics modeling of permeation dynamics.
4 identical copies of the K+ channel protein, arranged as a ring, form the channel walls.

40. Hydropathy analysis & topology studies predicted the presence of 6 transmembrane a-helices in the voltage-gated K+ channel protein.
The core of the channel consists of helices 5 & 6 & the intervening H5 segment of each of the 4 copies of the protein.

41. Helices 1-4 function as a voltage-sensing domain, with helix #4 having a special role in voltage sensing.
This domain is absent in K+ channels that are not voltage-sensitive.

42. The N-terminus of the Shaker channel (or part of a separate subunit in some voltage-activated channels) is essential for inactivation.
Mutants that lack this domain do not inactivate.
Adding back a peptide equivalent to this domain restores the ability to inactivate.

43. A "ball & chain" mechanism of inactivation has been postulated, in which the N-terminus of one of the 4 copies of the channel protein enters the channel from the cytosolic side of the membrane to inhibit ion flow.
In some voltage-gated K+ channels, entrance of the N-terminus into the channel is followed by a conformational change in the selectivity filter that contributes to the process of inactivation.

46. Ca2+ Channel

54. Cl- Channel

59. Obat pada Kanal Ion Cl-
Lubiproston (Amitiza ®), mengaktifkan kanal ClC-2 sehingga meningkatkan sekresi cairan ke lumen usus.
Obat ini diindikasikan untuk mengatasi obstipasi kronis idiopatik

60. See You Next week
Thank you