Lec4

• Cellular Pharmacology


  Many short-term physiological effects of chemical mediators involve changes in excitation, contraction or secretion
• Excitation


  Excitable cells generate an action potential in response to membrane depolarisation

  
• Excitable cells


  • Neurons, muscle cells (skeletal, smooth, cardiac), endocrine gland cells

  
• Patterns of Excitation


  Neurons and muscle cells - AP can propagate to all parts of cell membrane and often to neighbouring cells<|>Neurons & striated muscle - AP propagation over long distances<|>Cardiac & smooth muscle, some central neurons - spontaneous rhythmic activity<|>Gland cells - AP amplifies secretory signals

  
• Resting cell


  Membrane relatively impermeable to Na+<|>Na+ actively transported out in exchange for K+ (Na+-K+-ATPase)<|>[K+]i higher, [Na+]i lower than extracellular<|>Other ions e.g. Cl- Ca2+ also actively transported and unequally distributed

  
• Resting membrane potential


  Cells have -ve internal potential -30 to -80 mV<|>Membrane permeable to K+<|>Resting membrane potential of neurones (-60—80mV) close to equilibrium potential of K+ (-90mV)<|>In smooth muscle membrane potential less dependent on K+ and is lower (-30 to -50 mV)
• Action potentials


  Rapid, transient increase in Na+ permeability due to opening of Na+ channels - depolarisation<|>Slower, sustained increase in K+ permeability due to opening of K+ channels - repolarisation<|>Later discovery that voltage-gated Ca2+ channels important e.g. in smooth and cardiac muscle cells

  
• Ventricular myocyte action potential

  Phase 0: due to activation of voltage-gated Na+ channels. There is an inward current and the cell moves towards ENa.<|>Phase 1: Early repolarisation due largely to inactivation of sodium channels.<|>Phase 2: The 'plateau' phase and is due to inward current through voltage-gated calcium channels. These are slow to activate and to inactivate.<|>Phase 3: The repolarisation phase is brought about by inactivation of calcium channels and an increase in permeability to potassium.<|>Phase 4: Corresponds to the resting membrane potential and is largely determined by permeability to K+.

  
• Discharge patterns of excitable cells

  Skeletal muscle cells quiescent until activated by neurotransmitter<|>Cardiac muscle cells discharge spontaneously at a regular rate<|>Neurones and smooth muscle cells may be silent, or may discharge spontaneously, either regularly or in bursts, at varying frequencies

  
• Patterns due to


  Characteristics of ion channels expressed

  
• Effects of mediators and drugs on channel function

  Increased opening of Na+ or Ca2+ channels - increased excitability<|>Increased opening of K+ channels - decreased excitability<|>Increased opening of Cl- channels - decreased excitability<|>Channels blockers - opposite effects
• Channel openers and blockers

  • Na+ channel: veratridine (opener), tetrodotoxin (blocker)
  • K+ channel: cromokalim (KATP opener), tetraethylammonium (TEA, blocker)
  • Ca2+ channel: BayK 8644 (opener), nifedipine (blocker)
  • Cl- channel: glycine (opener), strychnine (blocker)
• Sodium channels


  S4 helix forms voltage sensor (basic amino acids) - moves outwards on depolarisation - pore opens<|>Movement of S4 also causes movement of inactivating particle to block the pore
• Cycle of activation\inactivation

  1. Resting

  2. Open

  3. Inactivated
• Refractory period


  Duration determines the maximum frequency at which action potentials can occur
• Use dependence


  Drugs can show selective affinity for different states<|>Drugs which bind most strongly to inactivated state show use-dependence - most effective when rate of action potential discharge is greatest
• Lidocaine (lignocaine)


  Blocks the sodium channel in its inactive state<|>The drug then dissociates from the sodium channel before arrival of next AP<|>Normal heart beat not affected<|>During frequent APs drug not dissociated by time next AP arrives<|>These drugs are effective in blocking high frequency pacemaker activity without affecting the normal sinus rhythm
• Pharmacology (Chemistry) of local Anaesthetics

  Lidocaine takes~2-4 min to act. Why?<|>Anaesthetic e.g. Lidocaine is neutrally charged.<|>Site of action on cytosolic side.<|>TTX acts on external side - instant action.<|>Diffuses through membrane.<|>Once in cytosol, becomes ionised (+ve).<|>Binds to channel, in a use dependent manner.
• Voltage-gated Potassium Channels


  Action potential repolarisation<|>hERG - K+ channel proteins important in heart. Disturbance by genetic mutation or drug side effects can result in LQT and dysrhythmias<|>Blocked by 4-aminopyridine, cisapride, dofetilide, quinidine
• Inwardly-rectifying Potassium Channels


  Inwardly-rectifying K+ channel (KATP)<|>Sulfonylureas stimulate insulin secretion by blocking KATP channels in pancreatic b-cells<|>Potassium channel openers e.g. Cromokalim relax smooth muscle (relaxation of arteriolar SM lowers BP)
• Potassium Channels


  Potassium channel (KATP) openers e.g. Cromokalim hyperpolarise and therefore relax vascular smooth muscle<|>Used to treat hypertension<|>Minoxidil - Management of hypertension/hair loss
• Functional subtypes and drug effects on potassium channels
• gNa


  Sodium ion channel
• gCa


  Calcium ion channel
• gK


  Potassium ion channel
• hERG


  Human Ether-à-go-go-Related Gene
• Blocked by 4-aminopyridine, cisapride, dofetilide, quinidine
• Lidocaine concentration is ~24 μM
• Amino acids, glucose, K+, Ca2+, ATP, mitochondria are involved
• Insulin secretion


  1. Mitochondria

  2. ATP:ADP ratio closes KATP

  3. Depolarisation

  4. Voltage-gated Ca2+ channel

  5. Ca2+ entry

  6. Insulin secretion
• KATP


  Inwardly-rectifying Potassium Channel
• Sulfonylureas


  Stimulate insulin secretion by blocking KATP channels in pancreatic β-cells
• Potassium channel openers


  Relax smooth muscle (e.g. Cromokalim relaxes arteriolar smooth muscle, lowers BP)
• Potassium channel structural classes

  • Voltage-gated
  • Inward-rectifying
  • Two-pore

  
• Potassium channel functional subtypes

  • Voltage-gated potassium channels (hERG/IKr)
  • Ca2+-activated potassium channels (IKs)
  • G-protein-activated (GIRK)
  • ATP-sensitive (IKATP)
  • TWIK, TRAAK, TREK, TASK
• Voltage-gated potassium channels


  • Involved in action potential repolarisation, limit maximum firing frequency
  • Inhibited by tetraethyl-ammonium, 4-aminopyridine
• Ca2+-activated potassium channels


  • Inhibited by apamin, charybdotoxin
• 1. protein-activated potassium channels


  • Mediate effects of G-protein coupled receptors
• ATP-sensitive potassium channels


  • Open when [ATP] is low, involved in insulin secretion
  • Blocked by glibenclamide, opened by diazoxide, pinacidil
• Two-pore domain potassium channels


  • Modulated by G-protein coupled receptors, activated by volatile anaesthetics