WINDSOR UNIVERSITY SCHOOL OF MEDICINE

1. WINDSOR UNIVERSITYSCHOOL OF MEDICINE Movement of Substances Across Membranes Dr.Vishal Surender.MD.

2. Objectives Understand how proteins and lipids are assembled to form a selectively permeable barrier known as the plasma membrane. Explain how the plasma membrane maintains an internal environment that differs significantly from the extracellular fluid. Explain the importance and characteristics of carrier-mediated transport systems. Understand how voltage-gated channels and ligand-gated channels are opened. Explain, using specific examples, the difference between primary and secondary active transport. Explain how epithelial cells are organized to produce directional movement of solutes and water. Explain how many cells can regulate their volume when exposed to osmotic stress. Understand why the Goldman equation gives the value of the membrane potential. Understand why the resting membrane potential in most cells is close to the Nernst potential for K+.

3. The Structure of the Plasma Membrane • Lipid bilayer– two sheets of lipids (phospholipids)-Polar (water-soluble) heads face out and the non-polar fatty acids hang inside. • Found around the cell, the nucleus, vacuoles, mitochondria, and chloroplasts. • Embedded with proteins and strengthened with cholesterol molecules. • Integral proteins (or intrinsic proteins) are embedded in the lipid bilayer, Include channels, pumps, carriers and receptors. • Peripheral proteins (or extrinsic proteins) do not penetrate the lipid bilayer. They are in contact with the outer side of only one of the lipid layers either the layer facing the cytoplasm or the layer facing the extracellular fluid

4. Hereditary Spherocytosis • A normal erythrocyte has a marked biconcave structure, which permits the cell to deform for easier passage through narrow capillaries, especially in the spleen. • The unusual shape is maintained by the cytoskeleton, formed by both integral and peripheral plasma membrane proteins. • A key player is the long filamentous protein known as spectrin, a peripheral protein that forms a meshwork on the cytoplasmic surface. • Hereditary spherocytosis (HS) is a genetic disease that affects proteins in the erythrocyte membrane, and the result is a defective cytoskeleton. • The most common defect is deficiency of spectrin, and the result is that regions of the membrane break off because they are no longer anchored to the cytoskeleton. • cell eventually becomes small and spherical. • Hemolysis (cell bursting) is present because the spherocytes are fragile to osmotic stress.

5. Transport across Membranes • The movement of large molecules is carried out by endocytosis and exocytosis, the transfer of substances into or out of the cell, respectively, by vesicle formation and vesicle fusion with the plasma membrane. • Cells also have mechanisms for the rapid movement of ions and solute molecules across the plasma membrane. • These mechanisms are of two general types: • -- passive movement, which requires no direct expenditure of metabolic energy, • -- and active movement, which uses metabolic energy to drive solute transport. Passive transport Processes Includes: A) Diffusion & B) Osmosis

6. Diffusion • By which molecules move from areas of high concentration to areas of low concentration and cations move to anions. • Its of 2 types. 1. Simple 2. Facilitated Diffusion • Simple Diffusion: means molecules move through a membrane without binding with carrier proteins. • SD occur via: 1. through the interstices of the lipid bilayer and 2. thru watery channels in transport proteins. • Facilitated: requires a carrier protein which aids in passage of molecules through the membrane by binding chemically and shuttling them through the membrane.

7. Simple Diffusion C1 C2 v ● ● ● ● ● ● ● particles ● ● ● ● ● ● ● ● ● ● ● time

8. factors that influence net flux are: 1. Electrical gradient. If the molecule is charged then its net flux across a membrane will be increased if the charge on the other side is opposite. 2. Temperature. Higher temperature ⇒ greater net flux 3. Surface area of membrane. Greater the surface area greater the net flux. 4. Molecule mass. Higher mass molecules move slower so the net flux would be less 5. Membrane permeability. The greater the membrane permeability for the molecule the greater the net flux

9. Diffusion Rate ↑Px Jx Flux ↓Px [X] Concentration

10. Simple Diffusion • Most molecules partition poorly – i.e. soluble in water but not lipid – therefore cannot cross lipid bilayer • Need – pores, channels and transporters

11. Pores, Channels and Transporters • Pore – transmembrane protein that is open • Channel – transmembrane protein with a pore that can open and close • Transporter – transmembrane protein that undergoes a conformational change and facilitates the transport of a ‘packet’ of substrate across the membrane

12. Ion Channels. • The cell membrane has ion channels that increase the permeability of the membrane for that ion species and allows the movement of those ions down their electrochemical gradient. • These channels can show a high degree of specificity for a particular ion species, e.g. the epithelial sodium channel is 30 times more permeable to Na+ than K+. • The inside of a cell has a net negative charge, therefore, this favors the flow of cations into the cell and anions out of the cell. • Regulation of ion channels • Ion channels may be open or closed and the time and frequency of opening may be regulated. This is important to realize as the state of the ion channels of the membrane have a major effect on the permeability of the membrane. There are three major factors that are involved in the regulation of the frequency and duration of channel opening. 1.Ligand-gated channels 2.Voltage-gated channels 3.Mechanosensitive channels

13. Ligand gated Ion Channel • The channel is a channel/receptor complex • Upon ligand binding there is a conformational change that opens the channel • Selectivity is conferred by charged amino acids and size (selects for cations or anions and then selects for size e.g. K+ ion much larger than Na+ ion – hydrated form)

14. ATP Ligand Gated Ion Channel e.g. P2X receptor

15. Voltage Gated Ion Channel • change in membrane potential moves charged molecules within the channel changing channel conformation either opening or closing the channel. • Charged amino acids inside the channel pore detect the electric field across the membrane – and conformational change can occur in response to a change in electric field -

16. Voltage Gated Ion Channel Vm e.g. voltage gated sodium channel

18. Facilitated Diffusion

19. Graph showing the relationship between net flux and concentration gradient of a substance moved across the membrane via facilitated diffusion. If the concentration gradient (and hence concentration) increases enough the transporters will become saturated and the net flux cannot be increased, this net flux value is called the transport maximum.

20. Primary Active Transport • Requires the direct expenditure of energy, in the form of ATP, in order to move ions against an electrochemical gradient – and in turn can generate a voltage across the membrane • e.g. Na+-K+-ATPase (stoichiometry 3:2 electrogenic) • V-H+-ATPase (hydrogen ion pump – generates voltage as it transports positive ions in one direction)

21. Primary Active Transport ATP → ADP + Pi 3+ ions out only 2+ in Vm -ve

22. Primary Active Transport • The ionic gradients are maintained by a combination of energy expenditure and Vm • Since the inwardly directed Na+ gradient is actively set up by the Na+-K+-ATPase – this acts as a membrane energy source, i.e. there is a large electrochemical gradient for Na+ to flow into the cell down an energy gradient – some transporters can use this Na+ gradient to transport other substrates (secondary active transport)

23. Secondary Active Transporters • Do not rely on the hydrolysis of ATP directly • Co-transport – both substrates transported in same direction (generally the substrate is driven into the cell up a chemical gradient – driven by the energy of Na+ flow into the cells down its electrochemical gradient)

24. Secondary Active Exchangers e.g. Na+/Ca2+ exchanger

25. Exchangers • e.g. Na+/Ca2+ exchanger - the energy of the inwardly directed Na+ gradient drives Ca2+ out of the cell up its electrochemical gradient • In general exchangers utilize the electrochemical gradient of one substrate to drive another substrate in the opposite direction and generally up its electrochemical gradient

26. Receptor Mediated Endocytosis • Receptors on the plasma membrane detect and attach to the macromolecule • The receptors are concentrated in a region of the plasma membrane – clathrin coated pits • The pits bud from the membrane and eventually form a vesicle that is internalized • e.g. uptake of cholesterol by cells (low density lipoproteins bind to the LDL receptor and are taken into the cell in this way).

27. Exocytosis/Endocytosis Exocytosis Endocytosis

28. Epithelial Transport