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PHARIntroduction  Pharmacology= study of the effects of drugs on the function of living systemsDefinition of a Drug  A chemical substance of known structure, other than a nutrient or essential dietary ingredient, which, when administered to a living organism produces a biological effect  May be s...

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PHAR002

Introduction  Pharmacology= study of the effects of drugs on the function of living systems Definition of a Drug  A chemical substance of known structure, other than a nutrient or essential dietary ingredient, which, when administered to a living organism produces a biological effect  May be synthetic, obtained from animals or plants, or a product of genetic engineering  To count as a drug, must be administered rather than released by physiological mechanisms. Eg. Insulin or thyroxine which are endogenous hormones but are drugs when administered intentionally  Must also have an effect – eg. H20 is not a drug Origins  Before the 19th century: herbal remedies were widely used, but weren’t based on scientific principles  At the time therapeutics were materia medica-- doctors were skilled at clinical observation and diagnosis but didn't have many effective treatment methods  Death was regarded as semisacred and treatment was a discussion for authorities  Initially was largely focused on understanding natural substances eg. Plant extracts and some toxic chemicals  Once developments in chemistry allowed purification of active compounds from plants, many substances followed, proving that chemicals were responsible for the effects they were seeing.  Growth of synthetic chemistry and physiology made rapid progress in relation to chemical mediators and chemical communication.  Concept of receptors for chemical mediators was first proposed by Langley in 1905 and was quickly taken up by other pharmacologists. Drug targets  Paul Ehrlick insisted that drug action must be explicable in terms of conventional chemical interactions between drugs and tissues  One of the main beliefs in pharmacology is that drug molecules must interact with cell constituents to alter the function of the cell molecules  Cell molecules vastly outnumber the drug molecules, a non-uniform distribution of the molecule within the body or tissue is required for a likely chance of the drug interacting with a specific type of cell or tissue.  Drug targets: macromolecules with which drugs interact to produce their effects – mostly proteins except antimicrobial and antitumor that can interact with DNA directly.  Can be receptors, enzymes, ion channels, carrier molecules etc.  Receptor: protein molecules whose functions to recognise and respond to endogenous chemical signals. Drug Specificity  Specificity is reciprocal: individual classes of drug only bind to certain targets, and individual targets only recognise certain classes of drug  No drugs are completely specific - increasing drug dose can cause it to affect targets other than the principal one resulting in side effects

Drug Receptor interactions  Binding and activation (generation of an alternate physiological behaviour) are two separate steps  The tendency of a drug to bind to the receptors is governed by its affinity  The ability of the drug to activate the receptor is its efficacy  Drugs of high potency generally have a high affinity --> occupy a significant proportion of the receptors even at low concentration  Agonist - chemicals that activate receptors  Antagonists - chemicals that block the effect of agonists on that receptor  Agonists also possess significant efficacy, antagonists have zero efficacy  Drugs with intermediate levels of efficacy--> even 100% receptor occupation will result in a submaximal tissue response - partial agonists  Full agonists - are able to stimulate a maximal tissue response Origin of Drugs  Plants (eg. Poppy- opiates eg. Morphine, analgesics Aspririn/ Quinine from tree bark Cannabis, Cocaine, Caffeine)  Animals (eg. Insulin- cow/pig antibodies- rats/mice/sharks/llamas Human Growth Hormone- pituitary gland Maggots- anticoagulants, snake viper venom, antihypertensive drugs)  Micro-organisms (eg. Penicillin, mold, antibacterial eg. Streptomyscin, kanamycin, neomycin- transplantation)

Important properties of drugs  Potency- how much drug needed (log drug conc) vs effect  Toxicity  Selective- how specific of an effect  Duration of action (half-life- how easily metabolised)/ bioavailability)  Side effects eg. Anxiety but causes sleep  Efficacy- therapeutic value The Nervous System Consists of 

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CNS (brain and spinal cord) and PNS (all nerves outside of the CNS)

The autonomic nervous system conveys all the outputs from CNS to the rest of the body, except for motor innervation of skeletal muscle The enteric nervous system (consisting of intrinsic nerve plexuses in the GI tract) can function without the CNS, the sympathetic and parasympathetic can't The Autonomic NS is largely under voluntary control The autonomic efferent pathway consists of 2 neurons in series,- preganglionic and postganglionic In the sympathetic nervous system, synapses lie in autonomic ganglia outside of the CNS

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In parasympathetic, the postganglionic cells are mainly found in the target organs.

Transmitters in the Autonomic Nervous System  The two main neurotransmitters that operate in the Autonomic system are Acetylcholine and Noradrenaline  All autonomic nerve fibres leaving the CNS release Acetylcholine which acts on nicotinic receptors (although in autonomic ganglia there is some excitation due to muscarinic receptors)  All post ganglionic parasympathetic fibres release Acetylcholine which acts on muscarinic receptors  All postganglionic sympathetic fibres release noradrenaline which may act on alpha and beta adrenoreceptors Membrane Potential  

Resting membrane potential depends on permeability and electrochemical gradients of ions in the cell The Na+/K+-ATPase is a pump that actively transports 3Na+ outside the cell and 2K+ inside the cell at the same time, using energy released from ATP hydrolysis.

K+:  

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This creates a K+ concentration gradient: It is higher inside the cell than outside the cell (120mM vs 5mM) The K+ ions therefore would like to leave the cell. The cell is actually permeable to K+ ions, so some of these will flow outside the cell, leaving behind negatively charged ions/proteins they were associated with. This creates a negative charge inside the cell. This negative charge would in theory attract the K+ ions, but the concentration gradient is stronger. Therefore, the K+ ions will be allowed to flow down their electrochemical gradient until they reach the point where they two gradients balance out  they reach equilibrium at the equilibrium potential of potassium (roughly -90mV)

Na+ 

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There is a concentration gradient: it is higher outside of the cell than inside the cell (120mM to 10mM) so they would like to flow inside of the cell. They are also attracted to the negative potential inside the cell, However, the membranes are relatively impermeable to Na+ ions, so they aren’t allowed to reach their equilibrium potential. (roughly +55mV)

The Nernst Equation 

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This equation calculations the equilibrium potential for individual cells depending on the concentration of potassium inside and outside that cell

The Goldman Equation  

This equation calculates the resting potential of any cell, including the effect of other ions on the membrane Chloride ions for the most part, have little appreciable effect on the potential

 Where Pions= permeability of the membrane for that ion Cell Excitability    

Tendency to initiate an Action Potential Depends on the state of the Na and/or Ca channels, and the K channels Anything that increases number of Na channels or reduces sodium inactivation increases excitability Anything that increases K conductance or blocks sodium channels will decrease excitability incl. mutations of channel proteins

Generating an Action Potential

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An increase in Na+ permeability causes an inward depolarising current of Na+ ions, whereas an increase in K+ permeability causes an outward repolarising current

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A small depolarisation of the membrane is produced by either transmitter action or by the approach of an action potential passing along the axon. If the small depolarisation is above threshold (-40mV) an all-or-nothing Action potential of one standard size will be generated. The outer gate of the Na+ channel opens rapidly, while the inner gate (which is normally open) begins to close. Na+ permeability increases, allowing an influx of Na+ ions. The depolarisation opens voltage-gated sodium channels, depolarising the membrane further, bringing the membrane potential close to ENa. The increased Na+ conductance is transient, because the channels inactivate rapidly  sodium inactivation

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At resting state membrane potential is about -70mV. The nerve cell is not excited. Stimulation causes membrane potential to begin to change becomes positive to roughly +50mV- depolarisation The potential comes back down, repolarisation The signal goes too far, becomes more negativehyperpolarisation Returns to the resting state of -70mV

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Repolarisation is assisted by the opening of voltage-gated K+ channels (not the ones that are already open) which open later than sodium channels K+ ions are allowed to flow outside of the cell to balance the depolarization The channels take a long time to close, resulting in hyper-polarization before the normal permeability of K+ ions are resumed.

Travel of an Action potential    

An action potential can travel down both sides of the axon However, there are no synapses at the cell end, so the single will only be transmitted at the dendritic end. Action potentials cannot travel back on themselves, they are oscillatory responses This is due to the inner gate closing, causing Na inactivation.

Action Potential propagation: Unmyelinated nerve fiber -

The depolarized membrane will begin to depolarize the next region of membrane, due to the flow of Na+ ions along the membrane

Myelinated nerve fibers -

Action potentials can only be created at the nodes of Ranvier so the action potential jumps from node to node This is much faster  saltatory conduction

Structure of Nerve Fibres  UNMYELINATED  Associated with schwann cell (glial) in the PNS  Associated with oligodendroctye (glial) in the CNS  Grows around axons, supporting axon but not a part of  MYELINATED  Schwann cell wraps a myelin sheath around the axon except at the node of ranvier  Increase speed of induction of impulse from about 0.5-2.5 m/s to 120m/s  One myelinated axon per schwann cell, many schwann cells per myelinated axon Diseases to do with Action potential Transmission 

Multiple Sclerosis (MS) – a neurodegenerative disease in which the patient’s axons will have plaques (areas without myelin) - Wherever the plaques occur there is a failure in transmission causing pain, fatigue, difficult in speech, muscle stiffness and spasms - Treatments include fampridine, tetraethylammonium, amifampridine, 4-aminopyridine, which blocks the activity of K+ channels delaying repolarization, allowing increased Na influx

Na channel blockers   

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Many drugs at high concentrations can block Na channels, but the only drugs used clinically are local anesthetics, antiepileptic and analgesic drugs. These drugs act exclusively outside of the membrane and have very low lipid solubility Tetrodotoxin (TTX) – from the puffer fish, is toxic in nM quantities - Chefs who serve have to be trained to make the flesh safe to eat

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Binds inside Na channels preventing influx of Na Fish are unaffected because the aa combination of the puffer fish Na channel is different – arginine is replaced by a threonine residue. Therefore, TTX does not bind to the fish Na channels Saxitoxin (STX) – produced by a marine micro-organism, and it accumulates in marine shellfish Batrachotoxin – from frog skin. DDT and oyrethrins – insecticides Veratridine – plant alkaloids Scorpion and sea anemone venoms - Facilitates Na channel activation so that Na channels open are more negative potentials close to normal resting potential - They also inhibit inactivation - The membrane becomes hyperexcitable and the AP is prolonged - The cells eventually become permanently depolarized and in excitable

Local Anaesthetics (LA)  

Local anaesthetics reduce pain, reduce sensation and prevent arrythmias They consist of an aromatic part linked by an ester/amide bond to a basic side chain

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they are weak bases and are mainly ionized at physiological pH (pka in the 8-9 range) LA + H+ LA+

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Its amide link allows hydrolysis  the anaesthesia is temporary The ability to exist unionised  allows easy travel into nerve cells Ester containing compounds are less stable than amide containing ones pH can be important clinically, because extracellular fluid of inflamed tissues is relatively acidic and therefore LA will be mostly present as ionised molecules, so somewhat resistant to LA at lower concentrations they decrease rate of rise of AP, increase the duration and increase the refractory period – reducing the firing rate at higher concentrations they prevent AP firing

Mechanism of action:    

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LA+ can’t travel through the cell membrane, so is transported as LA Therefore, activity of the LA is increased at alkaline pHs, because of low penetration Once the LA penetrates the nerve sheath, primarily the ionised form binds to the channel and blocks it Many local anaesthetics have been shown to be use-dependent  the more Na channels open, the greater the block becomes (blocking molecule enters the channel more readily when it’s open) For local anaesthetics that rapidly dissociate from the channel, block only occurs when the refractory period is too short for it to do so

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Many local anaesthetics bind most strongly to the inactivated state of the channel, shifting the eqm of channels towards the inactivated state, both increasing refractory period and reducing number of channels available for opening Blocks conduction more readily in small nerve fibres than large ones Hydrophilic Pathway - B diffuses through the membrane - Picks up H+ in the cytosol - Blocks channel

Hydrophobic Pathway - Dissolves into the plane of the membranes - Pokes through the side of the ion channel - Picks up H+ and blocks channel - Even if the channel is closed. The LA can act in both mechanisms at once 

Recovery    

LA only injected near the nerve Dissociates from the channel Diffuses out through the membrane into blood vessels Reducing blood flow slows recovery – eg. Adrenaline

Uses and Administration of Local Anaesthetics  

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Types of administration include Topical /Surface anaesthesia - Nose, mouth, bronchial tree, cornea, urinary tract, uterus - Eg. Lidocaine, tetrocaine, dibucaine, benzocaine - Risk of system toxicity over high concs/ large SAs Infiltration anaesthesia - Direct injection into tissues to reach nerve branches and terminals - Most drugs can be used - Adrenaline often added as a vasoconstrictor to prevent tissue damage Peripheral nerve blocks - Injection close to major peripheral sensory nerves for a certain area – nerve trunks - Most drugs can be used - Less LA needed than for infiltration anaesthesia - Accurate placement of the needle is important Central nerve block - Spinal Anaesthesia – LA injected into the subarachnoid space containing cerebrospinal fluid producing waist down anaesthesia - Mainly lidocaine - Have to control cranial spread to prevent respiratory depression - Epidural Anaesthesia – LA injected into epidural space, blocking spinal roots - Mainly lidocaine and bupivacaine - Less chance of cranial spread

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Most LA have a direct vasodilator action, increasing the rate they are absorbed into the systemic system increasing their potential toxicity and reducing their potency as local anaesthetics

Cholinergic transmission Acetylcholine receptors 

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Nicotinic receptors nAChR ganglionic and CNS types). Ligand-gated ion channels - its antagonist is Tubocurarine Muscarinic receptors mAChR -typically G-protein coupled receptors - its antagonist is Atropine

(muscle

The neuromuscular junction 1.Acetylcholine Release -

ACh is synthesised within the nerve terminal from choline and packaged into synaptic vesicles (100mM conc) Arrival of an AP at the pre-synaptic bulb stimulates voltage-gated Ca2+ ion channels to open, allowing Ca2+ ions to flood into the nerve terminal The Ca2+ influx causes movement of ACh vesicles to fuse with the membrane – Ca2+ mediated exocytosis

2. Propagation of the AP there is a short synaptic delay while the Ach diffuses across the small snyaptic cleft (1 micrometre) – The Ach binds with the nicotinic ligand gated Ach receptor on the post-synaptic membrane. Ach molecules bind for roughly 2ms, then dissociate and are quickly hydrolysed to prevent multiple bindings. = fast and brief transmitter action – This stimulates an end-plate potential E.P.P, Typically 300 synaptic vesicles are released (roughly 2 mill Ach molecules released) which is usually enough to reach threshold and propagate the AP in the motor neurone. This causes muscle contraction. –

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– E.P.PS are local potentials- don't propagate and is a graded potential - dependent on Ach conc 3. Recycling of choline – The Acetylcholine is hydrolysed by acetyl cholinesterase (AChE) to form choline and acetate, of which the choline is recycle back to the axon terminal via a choline uptake carrier that cotransports Na+ and choline into the cell – More than 50% of the choline is normally recaptured by the nerve terminals – The mitochondria produce Acetylcholine coenzyme A (AcCoA) from acetate. – Free choline is acetylated by choline acetyltransferase (CAT) forming Ach – Rate is determined by choline transport (happens in the background at the most part, but faster after Ach release)

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At fast cholinergic synapses eg. Neuromuscular and ganglionic synapses (but not at slow ones e.g. Smooth muscle, gland cells, heart etc) the released Ach is hydrolysed quickly, so only acts briefly Transmissions at the muscle cells are on a much larger scale than at neurons and require much more synaptic current to generate an AP in the post synaptic membrane. At the neuromusclular junction, only one nerve fibre innervates supplies each muscle fibre, and the amplitude of the pp is normally more than enough (can be reduced by 80% and still initiate an AP), so fluctuations in transmitter release do not affect transmission

Diseases to do with Cholinergic transmission 

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Myasthenia Gravis – autoantibodies are produced against nAChR at neuromuscular junctions, affecting muscles such as eye, face, limbs, pharynx, respiratory being the last affected and first to recover - Receptor numbers are reduced, reducing the EPP below threshold  muscle weakness - Muscles are unable to produce sustained contractions - Treatment: immunosuppression, and inactivation of the AChE allowing the ACh to float for longer, stimulating more than one Ach receptor. - Acetylcholinesterase inhibitors eg. Pyridostigmine, edrophonium, physostigmine, neostigmine - Immunosupressive drugs eg. Prednisolonek azatthioprine

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Lambert Eaton Myasthenic Syndrome- patient makes Antibodies against pre-synaptic voltage-gated calcium channels preventing acetylcholine release - Treatment: also inactivating ACHe to increase Ach activation - Can also use potassium channel blockers to extend the length of an AP and increase ACh rel...