Cholinergic Receptors Lecture 7 PDF

Cholinergic receptors Dr Chris Bax  Overview Cholinergic receptors Definition, types, structure and functions Pharmacology and disease Drugs and disease - examples Cholinergic receptors Signalling Centred on the properties of the neurotransmitter: acetylcholine (A...

Cholinergic receptors Dr Chris Bax  Overview Cholinergic receptors Definition, types, structure and functions Pharmacology and disease Drugs and disease - examples Cholinergic receptors Signalling Centred on the properties of the neurotransmitter: acetylcholine (ACh). ACh has actions in both the central and peripheral nervous systems (CNS and PNS), including the activation of skeletal muscle. The current therapeutic uses for cholinergic/anticholinergic drugs are limited (since effects are wide-reaching). Cholinergic receptors Neuromuscular junction (NMJ) Axon Synapse End plate potential Muscle contraction Cholinergic receptors Classes Muscarinic acetylcholine receptors (mAChR). Nicotinic acetylcholine receptors (nAChR). RECEPTOR RECEPTOR TYPE RECEPTOR TYPE Muscarinic G-protein coupled METABOTROPIC acetylcholine receptor Nicotinic Ligand gated ion IONOTROPIC acetylcholine channel Cholinergic receptors The nervous system VISCERAL: body cavity CNS SOMATIC: “bodily” Sensory Motor impulse impulse “ANS” EFFERENT Rest/digest and PNS Feed/breed PARA AFFERENT SYMPATHETIC Somatic Visceral Motor division Motor division SYMPATHETIC Fight or flight Sensory Smooth muscle Receptors Skeletal Cardiac muscle muscle Glands TISSUE VOLUNTARY INVOLUNTARY Neuronal layout in the PNS https://www.sciencedirect.com/topics/veterinary-science-and-veterinary-medicine/parasympathetic-nervous-system Cholinergic receptors The ANS (Structure) ANS Para- sympathetic Sympathetic Homeo- GANGLIA stasis Cholinergic receptors Neurotransmitters and the ANS Sympathetic Parasympathetic Somatic Spinal cord Preganglionic ACh neuron nAChR ACh nAChR Postganglionic ACh ACh neuron NA Tissue Adrenergic Muscarinic Nicotinic receptor Cholinergic receptors Biochemistry and physiology of cholinergic neurotransmission Occurs at the synapse of neurons (site of release). ACh is synthesised in one step from Acetyl Coenzyme A and Choline (in the cytoplasm). The rate limiting step is determined by the availability of choline, NOT choline acetyltransferase. Cholinergic receptors Synthesis Choline comes from three sources: 1. From the synaptic cleft (i.e. ACh which has been broken down by acetylcholinesterase) – 35-50%. 2. Plasma based stores. 3. Phospholipid (cell membranes) – phosphorylcholine. Cholinergic receptors Acetylcholine storage and release IMPULSE Negative membrane - - - -- - -- -- -- -- potential Ca2+ channel IMPULSE ++ + - - -- -- -- -- Na+ IMPULSE +++ - - - - -- -- -- Ca2+ IMPULSE +++ RELEASE OF - - - - -- -- - - - NEUROTRANSMITTER e.g. ACh Cholinergic receptors Acetylcholine storage and release Release of ACh into the synaptic cleft occurs through fusion of the synaptic vesicle with the plasma membrane. Nerve axon Process is dependent on axon terminal DEPOLARISATION  Influx of Ca2+ Botulinum toxin Synaptic cleft Cholinergic receptors Acetylcholine storage and release After synthesis, ACh is transported into synaptic vesicles for storage. Mediated by the vesicular ACh transporter (vAChT). ACh VESAMICOL ACh vAChT ACh ACh H+ ACh H+ H+ H+ H+ Synaptic vesicle Cholinergic receptors Degradation of ACh ACh is essential for rapid and repeated neurotransmission – hence there must be a mechanism to limit the duration of its action. CHOLINESTERASES are the enzymes responsible for the degradation of ACh: Acetylcholinesterase (AChE) Butyrylcholinesterase (BuChE) AChE hydrolyses about 4 x 105 molecules, per enzyme, per minute. Cholinergic receptors Acetylcholinesterase (AChE) AChE is concentrated on the post-synaptic membrane. Cholinergic receptors AChE inhibitors Inhibitors of AChE can cause: muscular paralysis, convulsions, bronchial constriction and death by asphyxiation. Two types: organophosphates (irreversible) and carbamates (reversible). Organophosphates: used in insecticides (malathion) and nerve gas (Sarin; novichok) – not used clinically (very long action). Carbamates: e.g. physostigmine, neostigmine, edrophonium. 1. Inhibit the catalytic domain of AChE (occupy active site for short periods of time - secs-mins). 2. Used to treat glaucoma and myasthenia gravis. Cholinergic receptors AChE inhibitors – clinical applications They have a number of uses: 1. Increase transmission at NMJ (diseases where there is insufficient ACh release or AChR number - e.g. myasthenia gravis). 2. Increase central cholinergic activity (e.g. donepezil for Alzheimer's disease – controls symptoms, doesn’t treat cause). Cholinergic receptors Types nAChR – ionotropic. mAChR – metabotropic. Structure is completely different, despite them sharing the same neurotransmitter ligand (ACh). mAChR (M2) nAChR Cholinergic receptors Muscarinic ACh receptors G-protein coupled receptors Latency of at least 100-250 ms associated with muscarinic responses to receptor activation (nAChR has 5 ms latency). WHERE DOES MUSCARINIC SIGNALLING OCCUR? Organs innervated by parasympathetic (ANS) nerves – hence the rest/digest and feed/breed functions CNS 5 types of cDNAs for human muscarinic receptors have been found: M1-M5 Cholinergic receptors Muscarinic ACh receptors (Function) M1, M3, M5 → Gq → ↑ PLC → excitatory. M2, M4 → Gi → ↓ AC, ↑ K+ channel → inhibitory. Second messenger molecules can modulate ion channel activity. βγ subunit of G-protein known to increase opening of various G protein-modified inwardly rectifying K+ channels (GIRKS). Cholinergic receptors Muscarinic ACh receptors (Function) TYPICAL RECEPTOR FUNCTION MECHANISM EXPRESSION CNS: Attention, Gq  PLC  M1 analgesia, arousal ↑ IP3 + ↑ DAG CNS Salivary gland/stomach secretions ↑ Ca2+ + ↑ PKC Cardiac tissue (e.g. Heart functions (mostly M2 βγ of G-protein  both nodes, ventricles inhibitory) – slows heart ↑ K+ (via GIRK) and atria) rate M3 Smooth muscle Contraction As M1 Glandular tissue Endo/Exocrine secretion M4 CNS Unknown As M2 M5 CNS Unknown As M1 Cholinergic receptors Nicotinic ACh receptors Ligand-gated ion channel. TWO ACh molecules bind to activate the channel. Conformational change induced by ACh-binding opens the receptor’s non-selective cationic channel. Opening causes influx of Na+ → depolarisation Depolarisation is brief (10 ms) due to action of AChE. Cholinergic receptors Nicotinic ACh receptors (Structure) Complex! Cholinergic receptors Nicotinic ACh receptors (Structure) Receptor is comprised of 5 subunits (pentamer). These are: α, β, γ, δ and ε (there are various subtypes for each subunit). The association of different subunits confers distinct structural and functional properties to the resultant nAChR. Can be divided into muscle type and neuronal type nAChRs. Cholinergic receptors Nicotinic ACh receptors (Structure) γ ε α subunit 1 2 3 4 α α β α β α γ ε TM2 region of each subunit is important for the cationic pore (structure  function) Cholinergic receptors Nicotinic ACh receptors (Structure) ε TM2 for each subunit contains negative amino acids which form a negatively charged ring which γ α draws positively charged ions through the channel 2 AC h 2 2 2 ACh binds to the α subunit 2 AC h N.B. 2 ACh molecules required for activation (which is why there is always, at least, 2 α subunits in α β each type of nAChR) Cholinergic receptors Nicotinic ACh receptors (Types) RECEPTOR TYPE LOCATION Muscle-type: (α1)2βδε or Neuromuscular junction (α1)2βδγ Ganglion-type: (α3)2(β4)3 Autonomic ganglia Heteromeric CNS-type: (α4)2(β2)3 Brain Further CNS-type: (α3)2(β4)3 Brain Homomeric CNS-type: (α7)5 Brain  Pharmacology and disease Pharmacology (m/n AChRs) Systemic application of cholinergic agonists/antagonists results in complex effects. Why? The presence of AChR at both sympathetic and parasympathetic ganglia Pharmacology and disease Pharmacology (m/n AChRs) Sympathetic Parasympathetic Somatic Spinal cord BLOCKADE OF AChR OCCURS TO BOTH SYMPATHETIC AND PARASYMPATHETIC NEURONS – HENCE EFFECTS ARE WIDE REACHING Preganglionic ACh neuron nAChR ACh nAChR Postganglionic ACh ACh ACh or neuron Nor/Adr Tissue Adrenergic Muscarinic Nicotinic receptor Muscarinic Pharmacology and disease Muscarinic AChR drugs: agonists AGONISTS (mostly non-selective). Acetylcholine, Carbachol, Pilocarpine, Methacholine, Bethanechol and Muscarine Inhibition of Urinary DRUG cardiac GI activity Activity contractility Acetylcholine ++ ++ ++ Methacholine +++ ++ ++ Carbachol + +++ +++ Bethanechol - +++ +++ Pharmacology and disease Muscarinic AChR drugs: agonists Topical administration can localise the effects, hence useful for: Glaucoma (optic nerve damaged – loss of retinal ganglion cells) Acetylcholine, Carbachol and Pilocarpine: eye drops Pharmacology and disease Muscarinic AChR drugs: antagonist ANTAGONIST CLINICAL APPLICATION Reduce saliva/mucus secretion during surgery Atropine Antidote to mushroom poisoning (muscarine) Scopolamine Motion sickness, nausea and vomiting Pirenzepine Peptic ulcer disease Ipratroprium COPD, Asthma (ACh causes bronchoconstriction) Oxybutynin Overactive bladder Mydriasis induced by atropine (mAChR antagonist) Pharmacology and disease Nicotinic AChR drugs: agonists Acetylcholine, Nicotine (!), Carbachol, Succinylcholine (AChE resistant), Epibatidine (frog poison). Little clinical use (again, too many adverse effects). Succinylcholine used in surgery: depolarising NM blockade. Resistant to AChE → persists in synaptic cleft → continuous nAChR activation → reduce chance of post-synaptic neuronal depolarisation. Pharmacology and disease Nicotinic AChR drugs: antagonist Not many clinical applications for nAChR antagonists. ANTAGONIST ACTION/CLINICAL APPLICATION α-bungarotoxin – snake venom = paralysis and death TOXINS  inhibit ACh binding α-conotoxin – marine snail venom TUBOCURARINE Induction of non-depolarising neuromuscular PANCURONIUM blockade in surgery Hypertension in patients with acute aortic dissection TRIMETHPHAN (via ganglionic blockade) Summary Cholinergic transmission is an integral part of both central and peripheral nervous system. Acetylcholine binds to and activates both muscarinic AChRs (GPCR) and nicotinic AChRs (ligand-gated ion channel). Cholinergic pharmacology is complex due to the physiology of the nervous system – the specificity and function of cholinergic drugs continues to be defined to this day.