1. Objectives Basics of electrophysiology 1. Know the meaning of Ohm’s Law 2. Know the meaning of ionic current 3. Know the basic electrophysiology terms 6. Understand the terms ‘activation’ and ‘inactivation’ 4. Know the effects of changing membrane potential in excitable cells 5. Know the effects of changing ionic conductances in excitable cells

2. antiarrhythmics What do the following categories of drugs have in common? anxiolytics anticonvulsants sedatives/hypnotics anesthetics antidiabetics antihypertensives They all include drugs that act on ion channels

3. Ion channels are interesting to pharmacists Therefore...

4. Channel selectivity Na + K+K+ Ca 2+ Cl - molecules

5. Voltage Extracellular ligand Intracellular ligand Channel gating

6. Ligand-gated ion channels (Dr. Ishmael) Voltage-gated ion channels

7. Voltage sensor Inactivation Voltage-dependent block

8. Voltage sensor

9. Inactivation + - intracellular extracellular

10. Voltage-dependent block + + - intracellular extracellular +

11. A guide to “Electrophysiologese” Membrane potential (E m ): The voltage difference across the cell membrane (inside vs outside) (millivolts) Resting potential: The membrane potential at which the membrane spends most of its time Action potential: The transient change in membrane potential due to active properties of the membrane Electrotonic potential: A change in membrane potential due to passive properties of the membrane

12. A guide to “Electrophysiologese” Depolarization: A change of membrane potential in the positive direction. Repolarization: Return of the membrane potential to the resting potential after a depolarization. Hyperpolarization: A change of membrane potential to a more negative value than the normal resting potential.

13. A guide to “Electrophysiologese” Inward current: Net movement of positive ions into the cell, or net movement of negative ions out of the cell. By convention, plotted as negative current. Outward current: Net movement of positive ions out of the cell, or net movement of negative ions into the cell. By convention, plotted as positive current. Inward current causes depolarization Outward current causes repolarization/hyperpolarization

14. A guide to “Electrophysiologese” Excitable cell: A cell that can fire action potentials Excitability: The ability to fire action potentials Threshold potential: The membrane potential at which an action potential fires

15. 2 msec mV Excitable cells fire action potentials

16. A nerve cell (neuron) Cell body axon

17. Hodgkin and Huxley Voltage clamp

18. Depolarization changes the conductance of the membrane

19. Inward current is carried by Na + ions Outward current is carried by K + ions

20. Hodgkin & Huxley reconstructed the action potential

21. Electrochemical gradients Which way will they go? At what rate will they go through? Ion channels allow ions to pass through Why would ions want to pass through?

22. Concentration gradient (chemical gradient) Net flow

23. Membrane potential (electrical gradient) + - + - Anion channel Cation channel

24. Membrane potential (electrical gradient) + - + - Anion channel Cation channel

25. Electrochemical gradient - - - - - - - - - - - - - - - - - - - -- - + + + + ++ + + + + ++ + + + + + + + + + + + -

26. The Nernst potential [X i ] = Ionic concentration inside the cell [X o ] = Ionic concentration outside the cell z X = ionic valence (number and sign (+ or -) of charges on ion) () E X = 60 zXzX. log [X o ] [X i ] (At physiological temperature) E X is in millivolts (mV)[X o ] and [X i ] are in millimolar (mM)

27. ion Extracellular concentration (mM) Intracellular concentration (mM) Nernst potential (mV) Na + K+K+ Ca 2+ Cl - 145 4 1.5 123 12 155 0.0001 4.2 67 -98 129 -90 If Cl - is passively distributed (not pumped), E Cl = resting potential

28. The different concentrations of physiological ions means that they have different Nernst potentials. Therefore, at any membrane potential, there is a driving force on at least some of the ions. (driving force = membrane potential – Nernst potential) At physiological membrane potentials, the driving force is inward for Na + and Ca 2+ ions and outward for K + ions. Therefore, at physiological membrane potentials, there are inward Na + and Ca 2+ currents and outward K + currents.

29. Ohm’s law: V=IR; I=GV V or E = potential (Volts); I = current (Amps); R = resistance (Ohms); G = 1/R = conductance (Siemens) The cell membrane is a resistor

30. I V High G Low G Slope = conductance (G) Ohm’s Law I=GV

31. I V Ohm’s Law I Na =G(E m -E Na ) I K =G(E m -E K ) E Na = 67 mV E K = -98 mV

32. +++++++++ ---------- I Na IKIK I Cl ATPase At rest, ionic gradients are maintained by the Na + -K + ATPase 2 K + 3 Na +

33. membrane potential = -90 mVG Na is low G K is high +++++++++ ---------- I Na IKIK -I Na = -((-90mV)-E Na ) x G Na = (I K ) = ((-90mV)-E K ) x G K I Cl E Cl = -90 mV outside inside (Ca 2+ channels not shown) If the membrane potential is not changing,

34. membrane is depolarizing G Na is very high G K is high +++++ ----- I Na > -(I K ) Na + channels just opened outside inside I Na IKIK I Cl (no significant effect on concentration)

35. G Na is very high G K is high +++++ ----- I Na = (30mV- E Na ) x G Na = -(I K + I Cl ) = -[(30mV- E K ) x G K + (30mV- E Cl ) x G Cl ] membrane potential = +30 mV I Na IKIK I Cl outside inside (outward current, inward Cl flow)

36. membrane potential = -90 mV +++++++++ ---------- I Na IKIK I Cl E Cl = -90 mV outside inside What will happen to the membrane potential if we open more Cl - channels? What will happen to excitability if we open more Cl - channels?

37. chord conductance equation

38. Electrical signaling changes intracellular Ca 2+ [Na + ] i, [K + ] i, [Cl - ] i don’t change significantly. Depolarization opens Ca 2+ channels. [Ca 2+ ] i increases. Ca 2+ Action potential axon Postsynaptic cell receptor Neurotransmitter

39. Here are the main points again: Nerves, muscles and other excitable cells use electrical signaling Physiologically, Na + channels always pass inward current; K + channels always pass outward current. In an excitable cell, depolarization causes activation of Na + channels, followed by inactivation of Na + channels and activation of K + channels. Inward current depolarizes the membrane. Outward current repolarizes/hyperpolarizes the membrane. These processes underlie the action potential of the nerve axon.

40. Ion selectivity varies among ion channels. Net movement of ions through channels is always down the electrochemical gradient. The membrane potential depends on the relative conductance of the membrane for K +, Na +, Cl - and Ca 2+ ions. In cells that don’t actively transport Cl -, opening Cl - channels decreases excitability by stabilizing the membrane potential. Concentration gradients are maintained by ATPases and ion exchangers The intracellular response to electrical signaling is a change in cytoplasmic Ca 2+.