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Recording Membrane Potentials: Extracellular and Intracellular Methods - Xia Qiang, MD & PhD

This article discusses the different methods of recording membrane potentials, including extracellular and intracellular recording. It also explains the concepts of resting membrane potential, depolarization, repolarization, and hyperpolarization.

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Recording Membrane Potentials: Extracellular and Intracellular Methods - Xia Qiang, MD & PhD

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  1. Membrane potentials膜电位 Xia Qiang, MD & PhD Department of Physiology Room C518, Block C, Research Building, ZJU School of Medicine Tel: 88208252 Email: xiaqiang@zju.edu.cn

  2. Electrocardiogram ECG（心电图）

  3. Electroencephalogram EEG（脑电图）

  4. Electromyogram EMG（肌电图）

  5. Extracellular Recording（细胞外记录）

  6. Intracellular Recording（细胞内记录）

  7. Recording of membrane potentials

  8. Video Extracellular and Intracellular Recording • Extracellular recording in earthworm giant axons • Intracellular recording in crayfish muscle cells

  9. Opposite charges attract each other and will move toward each other if not separated by some barrier.

  10. Only a very thin shell of charge difference is needed to establish a membrane potential.

  11. Resting membrane potential（静息电位） A potential difference across the membranes of inactive cells, with the inside of the cell negative relative to the outside of the cell Ranging from –10 to –100 mV

  12. （超射） Overshoot refers to the development of a charge reversal. A cell is “polarized” because its interior is more negative than its exterior. Repolarization is movement back toward the resting potential. （复极化） （极化） Depolarization occurs when ion movement reduces the charge imbalance. Hyperpolarization is the development of even more negative charge inside the cell. （超极化） （去极化）

  13. electrochemical balance - - - - - - - - - - - - - - - - - ++++++++++++++++ chemical driving force electrical driving force

  14. The Nernst Equation: K+ equilibrium potential (EK) (37oC) German physical chemist and physicist R=Gas constant T=Temperature Z=Valence F=Faraday’s constant （钾离子平衡电位）

  15. Begin: K+ in Compartment 2, Na+ in Compartment 1; BUT only K+ can move. Ion movement: K+ crosses into Compartment 1; Na+ stays in Compartment 1. At the potassium equilibrium potential: buildup of positive charge in Compartment 1 produces an electrical potential that exactly offsets the K+ chemical concentration gradient.

  16. Begin: K+ in Compartment 2, Na+ in Compartment 1; BUT only Na+ can move. Ion movement: Na+ crosses into Compartment 2; but K+ stays in Compartment 2. At the sodium equilibrium potential: buildup of positive charge in Compartment 2 produces an electrical potential that exactly offsets the Na+ chemical concentration gradient.

  17. Difference between EK and directly measured resting potential Mammalian skeletal muscle cell -95 mV -90 mV Frog skeletal muscle cell -105 mV -90 mV Squid giant axon -96 mV -70 mV Ek Observed RP

  18. Goldman-Hodgkin-Katz equation

  19. Role of Na+-K+ pump: • Electrogenic • Hyperpolarizing Establishment of resting membrane potential: Na+/K+ pump establishes concentration gradient generating a small negative potential; pump uses up to 40% of the ATP produced by that cell!

  20. Click here to play the Sodium-potassium Pump Flash Animation

  21. Origin of the normal resting membrane potential • K+ diffusion potential • Na+ diffusion • Na+-K+ pump

  22. Action potential（动作电位） Some of the cells (excitable cells) are capable to rapidly reverse their resting membrane potential from negative resting values to slightly positive values. This transient and rapid change in membrane potential is called an action potential

  23. A typical neuron action potential Positive after-potential Negative after-potential Spike potential After-potential

  24. Electrotonic Potential（电紧张电位）

  25. The size of a graded potential (here, graded depolarizations) is proportionate to the intensity of the stimulus.

  26. Graded potentials can be: EXCITATORY or INHIBITORY (action potential (action potential is more likely) is less likely) The size of a graded potential is proportional to the size of the stimulus. Graded potentials decay as they move over distance.

  27. Graded potentials decay as they move over distance.

  28. Local response（局部反应） • Not “all-or-none” （全或无） • Electrotonic propagation: spreading with decrement（电紧张性扩布） • Summation: spatial & temporal（时间与空间总和）

  29. Threshold Potential（阈电位）: level of depolarization needed to trigger an action potential (most neurons have a threshold at -50 mV)

  30. Ionic basis of action potential

  31. (1) Depolarization（去极化）: Activation of Na+ channel Blocker: Tetrodotoxin (TTX) （河豚毒素）

  32. (2) Repolarization（复极化）: Inactivation of Na+ channel Activation of K+ channel Blocker: Tetraethylammonium (TEA)（四乙胺）

  33. The rapid opening of voltage-gated Na+ channels explains the rapid-depolarization phase at the beginning of the action potential. The slower opening of voltage-gated K+ channels explains the repolarization and after hyperpolarization phases that complete the action potential.

  34. An action potential is an “all-or-none” sequence of changes in membrane potential. The rapid opening of voltage-gated Na+ channels allows rapid entry of Na+, moving membrane potential closer to the sodium equilibrium potential (+60 mv) Action potentials result from an all-or-none sequence of changes in ion permeability due to the operation of voltage-gated Na+ and K + channels. The slower opening of voltage-gated K+ channels allows K+ exit, moving membrane potential closer to the potassium equilibrium potential (-90 mv)

  35. Click here to play the Voltage Gated Channels and Action Potential Flash Animation

  36. Mechanism of the initiation and termination of AP

  37. Re-establishing Na+ and K+ gradients after AP • Na+-K+ pump • “Recharging” process

  38. Properties of action potential (AP) • Depolarization must exceed threshold value to trigger AP • AP is all-or-none • AP propagates without decrement

  39. Nobel Prize in Physiology or Medicine 1963 • "for their discoveries concerning the ionic mechanisms involved in excitation and inhibition in the peripheral and central portions of the nerve cell membrane" • Eccles Hodgkin Huxley How to study ? • Voltage Clamp

  40. Currents recorded under voltage clamp condition

  41. Nobel Prize in Physiology or Medicine 1991 • "for their discoveries concerning the function of single ion channels in cells" • Erwin Neher Bert Sakmann • Patch Clamp

  42. Video The Squid and its Giant Nerve Fiber "The Squid and its Giant Nerve Fiber" was filmed in the 1970s at Plymouth Marine Laboratory in England. This is the laboratory where Hodgkin and Huxley conducted experiments on the squid giant axon in the 1940s. Their experiments unraveled the mechanism of the action potential, and led to a Nobel Prize. Long out of print, the film is an historically important record of the voltage-clamp technique as developed by Hodgkin and Huxley, as well as an interesting glimpse at how the experiments were done.

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