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PSL300 PSL300 - Lecture 01 Background – Cell Membrane – Cell Membrane permeability Simple Diffusion Facilitated Diffusion Active Transport Secondary Active Transport – Channels Voltage Gated Channels Ligand G...

PSL300 PSL300 - Lecture 01 Background – Cell Membrane – Cell Membrane permeability Simple Diffusion Facilitated Diffusion Active Transport Secondary Active Transport – Channels Voltage Gated Channels Ligand Gated Channels – Endo/Exocytosis Kiss and Run Full Fusion Membrane Potential – Na+/K+ pump – Resting Membrane Potential (RMP) Cell Membrane Cell membrane is not an inert bag holding the cell together Cell membrane = composed of Phospholipid bilayer Lipid-soluble molecules and gases diffuse through readily Water-soluble molecules cannot cross without help Impermeable to organic anions (proteins) Permeability depends on molecular size, lipid solubility, and charge Membrane Permeability If a substance can cross the membrane by any means, the membrane is permeable to that substance Gases can diffuse across the membrane Polar molecules and ions need the help of proteins (channels or carriers) to cross  lots- > little DOWN Simple Diffusion Simple Diffusion: Small, lipid-soluble molecules and gases (e.g. O2, CO2, ethanol, urea etc…) pass either directly through the phospholipid bilayer or through pores - Movement of substance is down its [ ] gradient The relative rate of diffusion is roughly proportional to the [ ] gradient across the membrane Passive: No energy input required from ATP Facilitated Diffusion Saturates when there are more subs than Facilitated Diffusion is a process of diffusion, where molecules diffuse transporter proteins across membrane, with the assistance of carrier protein Carrier protein aid the movement of polar molecules (e.g. sugars and amino acids) across cell membrane Movement of substance is down its [ ] gradient The energy comes from the [ ] gradient of the solute Passive: No energy input required from ATP  NatIp * Pump Active Transport little - lots What if we want to move molecules against its [ ] gradient? Active Transport is a mechanism to move selected molecules across cell membranes, against their [ ] gradient Substance binds to protein carrier that changes conformation to move substance across membrane Active Requires energy from ATP hydrolysis ATPases (Na+/K+ Pump)  Secondary Active Transport When a substance is carried up its concentration gradient without ATP catabolism, this is known as Secondary Active Transport Kinetic energy of movement of one substance down its concentration gradient powers the simultaneous transport of another up its concentration gradient Secondary transporters ride on the ‘coat-tails’ of primary active transport and do not themselves require ATP Sequential binding of a substance and ions to specific sites in the transporter protein induces a conformational change in the protein * Secondary Active Transport is powered by the chemical energy in the substance diffusing down its [ ] gradient and this energy is used to ‘push’ some other substance against its [ ] gradient 1st Down powers 2nd Up Against Channels Membrane spanning protein forms a ‘pore’ right through the membrane – 4-5 protein subunits fit together such that a central pore is created through membrane, through which specific ions can diffuse through These ‘Pore loops’ of the protein molecules dangle inside the channel Physical properties of the pore loops create a selectivity filter – Only specific molecules can diffuse through (by means of size and electric charge) These ‘pores’ are called Membrane Channels Gated Channels Membrane channels generally are not kept perpetually open Channels can be closed off by a branch of the protein structure which functions as ‘Gate’ Under certain condition, the gate is closed, and no diffusion takes place, under other conditions the gate is open and diffusion is allowed (remember that it is still selective) The protein components switch between 2 shapes; one creates an open pore, the other blocks the pore Factors determining channel protein shape: – Ligand gated channels: Binding of chemical agent – Voltage gated channels: Voltage across the membrane Ligand Gated Channels Cell membrane receptors are part of the body’s chemical signaling system The binding of a receptor with its ligand usually triggers events at the membrane, such as activation of an enzyme synaptic transmission  Voltage Gated Channels Some membrane channels are sensitive to the potential difference across the membrane (e.g. OUTSIDE CELL depolarization), changes the conformation of the channel subunits causing a diffusion pore to be created A The voltage sensing mechanism is in the 4th transmembrane domain of ↓XX the protein, the S4 segment ⑦ charged attracted downwards INSIDE CELL Voltage Gated Channels S4 sticks out to the side of the protein (like a wing) The natural position of the S4 ‘wing’ OUTSIDE CELL is up towards the outer surface of the cell membrane. But when the membrane is polarized, the positively charged wing is attracted downwards to the negatively charged inner surface of the membrane INSIDE CELL  Depolarized= lesso Repolarized : returns to MP Hyperpolarization more : Voltage Gated Channels less E Depolarization of the membrane to about -50 mV no longer provides sufficient electrical attraction to hold OUTSIDE CELL the S4 wing downwards, so it migrates back-up In the up position, S4 removes a structural occlusion from the pore such that ions can now diffuse through it OPEN depolarized INSIDE CELL  Endo/Exocytosis v Endocytosis: inward ‘pinching’ of membrane to create a vesicle; usually receptor- - J mediated to capture proteins, from outside to inside. bring in Exocytosis: partial or complete fusion of vesicles with cell membrane for bulk trans-membrane transport of specific molecules, from inside to outside. out going - Exocytosis Intracellular materials, packaged in vesicles, are either secreted or delivered to the plasma membrane by exocytosis There are 2 different types of exocytosis, with different docking mechanisms – Exocytosis 1: The more rapid mechanism has been dubbed the ‘Kiss and Run’ – Exocytosis 2: Full exocytosis Exocytosis 1 Kiss and Run: The secretory vesicles dock and fuse with the plasma membrane at specific locations OUTSIDE CELL called ‘fusion pores’ Vesicle can connect and disconnect several times before contents are emptied Since only part of the contents are emptied in one ‘Kiss’, the process can be repeated several times before the vesicle is depleted Generally only part of vesicle contents diffuse into the interstitial INSIDE CELL fluid, used for low rate of signaling Exocytosis 2 FULLY /ALLAT ONCE Full exocytosis: This involves complete fusion of the vesicle with OUTSIDE CELL the membrane, leading to total release of vesicle contents at once Necessary for delivery of membrane proteins and high levels of signaling Must be counterbalanced by - - endocytosis to stabilize membrane - surface area to avoid loose membrane INSIDE CELL  > - created by : * ion concentration gradients between ECF and ICF * selectively permeable cell membrane Membrane Potential All cells in the body generate Membrane Potential (MP) To generate MP we need 2 conditions: 1. Create a concentration gradient: an enzyme ion pump (functions as an ATPase) must actively transport certain ion species across the membrane to create a concentration gradient 2. Semi-permeable membrane: allows one ion species to diffuse across the membrane more than others Diffusion of that ion species down its conc. gradient creates an electrical gradient  3 Nat OUT 2k + EN Na+/K+ Pump Concentration Gradient All cell membrane is loaded with Na+/K+ pumps, this is the staple of all living cells. Na+/K+ dependent ATPase is enzyme that moves Na+ out of cell, and K+ into cell by breaking down ATP ATP- > ADD + P For each ATP molecule broken down, 3 Na+ ions are pumped out and 2 K+ pumped in (creates a concentration gradient) Consumes 1/3 of energy needs of body (in neurons it’s 2/3, i.e. huge consumer of energy) Na/K inequality > potential difference of -10 mV inside wh respect to outside inside is less , moreE excess o on the outside  does aga , ↳ Resting Membrane Potential So is our resting MP roughly -10 mV? No! the actual resting MP in neurons is not -10mV but it’s closer to -70 mV, why? Resting Membrane Potential Since our Resting MP is closer to -70 mV, something else is obviously happening > this is due to diffusion of K+ ions outwards The ‘resting’ membrane is most permeable to K+ ion K+ diffuses out of cell, down concentration gradient, via K+ channels K+ are Cations accumulate on the outside of the membrane, leaving a net negativityleaving inside membrane channels Leak  Resting Membrane Potential ECF = 0 This efflux will occur until there is such a build up of “+” charge on the outside of ICF = O the membrane that further diffusion of K+ is repelled by the electromagnetic force = i.e. we reach an equilibrium situation until they decide its enough I electromagnetic creates force of electrical REPULSION gradient  Equilibrium Potential At equilibrium, electrical work to repel outward cation diffusion equals chemical work of diffusion down conc. gradient A Membrane potential at equilibrium is determined by the concentration gradient Can be calculated using the Nernst Equation Nernst Equation In the ideal situation, the Nernst equation, describe the balance between the chemical work of diffusion with electrical work of repulsion The equation gives the potential difference across the membrane, inside with respect to outside, at equilibrium The result is valid if and only if one ion species (K+ in this case) is diffusing across the membrane K+ Equilibrium Potential If you calculate using the K+ ion you get: EK+ = (RT/F) ln([K+]o /[K+]i) = -90 mV (equilibrium potential for K+) This is what the MP would be if only K+ ions was involved and nothing else But in the typical neuron, the resting MP is NOT -90mV, it’s about -70 to -80 mV What’s happening? else something going on