MD224 Topic 2 Acetylcholine and Amino Acids PDF

Topic 2 Acetylcholine and amino acids 1 Acetylcholine (ACh) I I I I I I CH3 O I II + CH3 C O CH2 CH2 N CH3 I CH3 The first molecule to be implicated as a NT 2 Discovery of acetylcholine YEAR DISCOVERY 1900 Neurons transmit information via electrical signals along axons 1903 Britis...

Topic 2 Acetylcholine and amino acids 1 Acetylcholine (ACh) I I I I I I CH3 O I II + CH3 C O CH2 CH2 N CH3 I CH3 The first molecule to be implicated as a NT 2 Discovery of acetylcholine YEAR DISCOVERY 1900 Neurons transmit information via electrical signals along axons 1903 British neurologist, Thomas Elliott, suggests that the message crosses the synaptic cleft via chemical compound 1914 English physiologist, Henry Dale, working in fungus ergot, discovered it secreted a compound that produced effects on organs similar to those produced by nerves: Acetylcholine 1921 German-American pharmacologist, Otto Loewi, devised method to test Elliott’s hypothesis: showed that stimulation of nerves attached to frog’s heart caused release of chemical substances, one of which he named vagusstoffe Dale realised vagusstoffe might be the same compound as ACh 1936 Loewi and Dale share Nobel Prize 3 Synthesis of ACh 5 Storage of ACh into synaptic vesicles VAChT Vesicular ACh Transporter 6 ACh action Acetylcholine can bind to two receptor types: Ionotropic: Metabotropic: Nicotinic receptor Muscarinic receptor 7 The structure of the nicotinic ACh receptor channel Receptor is made of 5 subunits: 2α β γ δ 8 Nicotinic ACh receptor: ionotropic Each nAChR complex has 2 binding sites for Ach Mostly on α-subunits, though partial involvement of δ and γ subunits Ligand binding opens channel and allows influx of Na+ 9 Pharmacology terms Agonist Antagonist Compound which binds to the receptor and mimics the action of the physiological ligand Compound which binds to the receptor. Lacks intrinsic activity Instead it blocks the activity of the physiological ligand 10 Compounds that bind to the nicotinic AChR Agonist Acetylcholine is the endogenous agonist Nicotine is an agonist of ACh ionotropic receptor (a plant alkaloid identified in 1914) Antagonist -bungarotoxin is an antagonist of ACh ionotropic receptor (from snake venom) 11 Muscarinic AChR, metabotropic 5 different mAChRs that can activate different G proteins that link to different signalling systems • M1, M3, M5: • Activate phospholipase C (PLC) via Go or Gq • M2, M4: • inhibit adenylate cyclase via Gi • stimulate a K+ channel via Gs • mAChR present in striatum and in peripheral nervous system. They also mediate autonomic functions acting on heart, smooth muscle and exocrine glands 12 Compounds that bind to Muscarinic AChRs Agonist: Muscarine: • Fungal alkaloid that binds with high affinity Antagonists: Distinguish different subclasses of mAChR Amanita muscaria M1 (brain) M2 (heart) M3 (glandular tissue) Atropine + + + Pirenzepine + - - Himbacine - + 13 Summary of ligands for ACh Receptors Acetylcholine Nicotinic receptor Activates Muscarinic receptor Activates Nicotine Activates Does not bind Inhibits Does not bind Muscarine Does not bind Activates Atropine Does not bind Inhibits α-Bungarotoxin 14 Inactivation of ACh Enzymatic degradation • Degraded by acetylcholinesterase in synaptic cleft where ACh is at high concentration • Catalytic rate of enzyme is 104 – 105 mols per sec • One of the most rapid enzymes known • Choline is taken back up into the nerve terminal by high affinity Na+dependent uptake system • Acetylcholinesterase is inhibited by: • Sarin nerve gas – lethal dose is 0.5 mg • Organophosphates – used in insecticides • Neostigmine, donezepil (used in treatment of Alzheimer’s disease) 15 Localisation of Acetylcholinesterase ACh is degraded in the synaptic cleft 16 ACh neurotransmission and disease: Myasthenia gravis • Muscle weakness • Autoimmune disorder • Patients’ sera contain antibodies directed against their own nicotinic AChRs • a decreased number of functional AChRs on muscle cells • Defective neuromuscular transmission leading to muscle weakness 17 Myasthenia Gravis Muscle recordings 18 Excitatory amino acid neurotransmitters: Example: Glutamate 19 Excitatory amino acids (EAAs) • Glutamate, aspartate, homocysteic acid • Produce an excitatory response in neurons • i.e., the neuron is more likely to send an action potential • Glutamate: • Most important NT for normal brain function • Nearly all excitatory neurons in CNS are glutamatergic • BUT glutamate is present in all cells 20 Glutamate synthesis and cycling between neurons and glia Glutamate is synthesised from glutamine by glutaminase Glutamate that has been released is taken up by astrocytes Here it is converted back to glutamine by glutamine synthetase 21 Glu (EAA) receptor subtypes Ionotropic Metabotropic subunits 22 Ionotropic Glu (or EAA) Receptors Receptor Ions flux through open channel Opening of channel triggered by AMPA Influx of Na+ only Glu binding Kainate Influx of Na+ only Glu binding NMDA Na+ Glu binding plus membrane depolarization Influx of and Ca2+ The subunits are derived from a different ancestral gene than AChR 23 NMDA receptor D-AP5 MK801 NMDA-R antagonists: D-AP5 MK801 Mg 2+ 24 NMDA and AMPA receptors act together AMPA receptor Opens quickly upon Glu binding, <1 ms NMDA receptor Opens slowly, >2 ms Requires two signals to open: 1. Ligand-gated: Glu must bind 2. Mg2+ block must be removed from the channel: A change in the transmembrane potential due to Na+ entry removes the Mg2+ block from the channel Axon releases glutamate Axon releases glutamate Displaced Mg2+ ion Mg2+ AMPA receptor Na+ enters AMPA receptor Na+ enters NMDA receptor Na+ and Ca2+ enter 25 Metabotropic Glu receptors • 7 membrane spanning peptide • 3 classes: at least 8 subtypes, mGluR1-8, have been cloned • Group I mainly postsynaptic • Linked to activation of phospholipase C • Group II and III - mainly presynaptic • Linked to inhibition of adenylate cyclase • May also play a role in memory formation • mGluR inhibitors block memory formation at some synapses • NMDA receptor activation potentiates signalling via mGluRs 26 Stroke: role of excitatory amino acid receptors in neuronal damage Ischemia Ischemia • Block in blood supply • Excessive Glu release causes over-stimulation of NMDA receptors • Excess Ca2+ influx into postsynaptic neurons • Leading to excitotoxic cell death (excitotoxicity) Energy supply Cell depolarization Glutamate release NMDA, AMPA receptor activation Ca2+ channel opening Increase [Ca2+] Neuronal injury 27 Excitotoxicity and Ca2+ Large influx of Ca2+ • Can activate calpains, phospholipases, NOS, nucleases etc. • Leads to very rapid cell death by necrosis • Is sequestered into mitochondria resulting in swelling and eventual rupture of mitochondria • Leads to delayed cell death NMDA-R antagonists D-AP5 and MK801 provide protection in models of ischemia 28 Inhibitory amino acid neurotransmitters: Example: GABA 29 Excitatory and inhibitory NT receptors Excitatory: eg NMDA, AMPA, nicotinic AChR • Influx of positively charged ions such as Na+ • Inside of the cell becomes less negative • Depolarisation of post-synaptic cell making it more likely to initiate an action potential Inhibitory: • Influx of negatively charged ions such as Cl• Inside of cell becomes more negative • Hyperpolarisation of post-synaptic cell making it less likely to initiate an action potential 30 -aminobutyric acid (GABA) • Main inhibitory NT in the brain • Approx 33% of synapses in brain uses GABA • Synthesised from glutamate by glutamic acid decarboxylase • Presence of GAD indicates a GABAergic neuron VIAAT GAT: GABA transporter VIAAT: vesicular inhibitory amino acid transporter 31 GABA receptors • Ionotropic • GABAA and GABAC • Metabotropic • GABAB 32 GABAA Receptor • Structurally related to nAChR • Ion channel is selective for Clions • Inactivation: GABA is removed from synaptic cleft by transporters 33 Valium (diazepam) acts through modulating GABA-A receptor signalling • Chloride ions (negative charge) enter the cell  Hyperpolarization • Benzodiazepines (e.g., diazepam) and barbiturates enhance the hyperpolarizing effects of GABA 34 Presynaptic inhibition Inhibitory NT binds to receptors on the presynaptic cell ↓ Reduction in depolarisation of the presynaptic nerve terminal ↓ Less Ca2+ influx ↓ Less excitatory NT release Action potential Inhibitory NT Excitatory NT Excitatory NT Inhibitory NT 35 Presynaptic inhibition Inhibitory NT binds to receptors on the presynaptic cell ↓ Reduction in depolarisation of the presynaptic nerve terminal ↓ Less Ca2+ influx ↓ Less excitatory NT release Action potential Inhibitory NT Excitatory NT Excitatory NT Inhibitory NT 36 Summary: NT receptor classification Ionotropic Fast neurotransmission Metabotropic Slow neurotransmission Excitatory (G protein coupled receptor) • Influx of Na+ • Inside of the cell becomes less negative, ie depolarisation Inhibitory • Influx of Cl• Inside of cell becomes more negative, ie, hyperpolarisation • • • • Phospholipase C Adenylate cyclase Phospholipase A2 Ion channels 37