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.

4. 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. Polar head non-polar tail

5. 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

6. Integral-Membrane Proteins Can Serve as Receptors

7. Integral-Membrane Proteins Can Serve as Adhesion Molecules, ex-integrins, Cadherins. Can Form a Submembranous Cytoskeleton, Ex- spectrin,ankyrin

8. Hereditary Spherocytosis • 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.

9. Membrane Transport • Cell membrane is permeable to: • Non-polar molecules (02& C02). • Lipid soluble molecules (steroids). • H20 (small size, lack charge). • Cell membrane impermeable to: • Large polar molecules (glucose). • Charged inorganic ions (Na+ , K+ etc.,).

10. Types of Membrane Transport • 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 substances move across the membrane down their electrochemical gradient • -- and active movement, which uses metabolic energy to drive solute transport against this gradient. Passive transport Processes Includes: A) Diffusion & B) Osmosis

11. BulkTransport (Endocytosis and Excytosis) • Movement of many large molecules, that cannot be transported by carriers. • Exocytosis: • A process in which some large particles move from inside to outside of the cell by a specialized function of the cell membrane • Endocytosis: • Exocytosis in reverse. • Specific molecules can be taken into the cell because of the interaction of the molecule and protein receptor.

12. Exocytosis • Vesicle containing the secretory protein fuses with plasma membrane, to remove contents from cell.

13. Endocytosis • Material enters the cell through the plasma membrane within vesicles.

14. Types of Endocytosis • Phagocytosis - (“cellular eating”) cell engulfs a particle and packages it with a food vacuole. • Pinocytosis – (“cellular drinking”) cell gulps droplets of fluid by forming tiny vesicles. (unspecific) • Receptor-Mediated – binding of external molecules to specific receptor proteins in the plasma membrane. (specific)

15. Example of Receptor-Mediated Endocytosis in human cells

16. 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. • Facilitated: requires a carrier protein which aids in passage of molecules through the membrane by binding chemically and shuttling them through the membrane.

17. 1. Simple Diffusion • Molecules/ions are in constant state of random motion due to their thermal energy. • Simple diffusion occurs • whenever there is a concentration difference across the membrane • the membrane is permeableto the diffusing substance.

18. Membrane Transport C1 C2 ● • C1 ● ● ● ● ● ● particles ● ● ● ● ● ● ● ● C2 ● ● ● time

19. factors that influence net flux are: (Fick`s law of diffusion) 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

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

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

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

24. 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,channels can be opened or closed by gates • 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+. • Regulation of ion channels • Ion channels gates may be open or closed and the time and frequency of opening may be regulated. There are three major factors that are involved in the regulation of the frequency and duration of channel opening. 1.Ligand-gated channels 3.Mechanosensitive channels 2.Voltage-gated channels

25. 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)

26. Ligand-Operated ACh Channels • Ion channel runs through receptor. • Receptor has 5 polypeptide subunits that enclose ion channel. • 2 subunits contain ACh binding sites.

27. Voltage Gated Ion Channel • change in membrane potential(Vm)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 -

29. Organ of Corti • When sound waves move the basilar membrane it moves the hair cells that are connected to it, • but the tips of the hair cells are connected to the tectorial membrane • the hair cell get bent . • There are little mechanical gates on each hair cell that open when they are bent. • K+ goes into the cell and Depolarizes the hair cell. (concentration of K+ in the endolymph is very high)

30. Facilitated Diffusion via carrier • Ex- glucose, amino acid transport. • Down concentration Gradient • Chemical Specificity: Carrier interact with specific molecule only, cysteinurea. • Competitive inhibition: • Molecules with similar chemical structures compete for carrier site. • Saturation: • Vmax(transport maximum): • Carrier sites have become saturated. glucose transporter 4 (GLUT4) activated by insulin

31. 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.

32. Active transport • When the cell membrane moves molecules or ions uphill against a concentration gradient • (or uphill against an electrical gradient), • the process is called active transport • Primary active transport • Secondary active transport:

33. Active transport • 1 Primary active transport: • the energy used to cause the transport is derived directly from the breakdown of ATP or some other high-energy phosphate compound • 2 Secondary active transport: • The energy is derived secondarily from energy • That has been stored in the form of ionic concentration differences between the two sides of the membrane • created by primarily active transport

34. Intracellular vs extracellular ion concentrations Ion Intracellular Extracellular Na+ 5-15 mM 145 mM K+ 140 mM 5 mM Mg2+ 0.5 mM 1-2 mM Ca2+ 10-7mM 1-2 mM H+ 10-7.2 M (pH 7.2) 10-7.4 M (pH 7.4) Cl- 5-15 mM 110 mM

35. 3.1 Primary Active Transport • Hydrolysis of ATP directly required for the function of the carriers. • Molecule or ion binds to “recognition site” on one side of carrier protein.

36. 3.1 Primary Active Transport • Binding stimulates phosphorylation (breakdown of ATP) of carrier protein. • Carrier protein undergoes conformational change. • Hinge-like motion releases transported molecules to opposite side of membrane.

37. Mechanism of Acid Secretion • The key player in acid secretion is a H+/K+ ATPaseor "proton pump" located in the parietal cell membrane. • Hydrogen ion is pumped out of the cell, into the lumen, in exchange for potassium through the action of the proton pump.

38. Na+/K+ Pump

39. A Model of the Pumping Cycle of the Na+/K+ ATPase

40. Importance of the Na+-K+ Pump • Control cell volume • Develop and Maintain Na+ and K+ concentration gradients across the membrane • Electrogenic action influences membrane potential • Provides energy for secondary active transport

41. 2 Secondary Active Transport • Energy needed for “uphill” movement obtained from “downhill” transport of Na+. • Hydrolysis of ATP by Na+/K+ pump required indirectly to maintain [Na+] gradient.

42. Secondary active transport co-transport counter-transport (symport) (antiport) out in out in Na+ Na+ glucose ca2+ Co-transporters will move one moiety, e.g. glucose, in the same direction as the Na+. Counter-transporters will move one moiety, e.g.ca2+, in the opposite direction to the Na+.

44. (b) Beta cell secreting insulin. Closure of the KATP channel depolarizes the cell, triggering exocytosis of insulin. 1 2 3 4 5 KATP channels close. Metabolism increases. ATP increases. High glucose levels in blood Cell depolarizes and calcium channels open. 6 Ca2+ entry acts as an intracellular signal. Ca2+ Glucose Glycolysis and citric acid cycle ATP Ca2+ 7 GLUT transporter Ca2+ signal triggers exocytosis, and insulin is secreted. Insulin Secretion and Membrane Transport Processes

45. Epithelial Transport

47. A 7-year-old boy is brought to the pediatrician because of a chronic cough, fatty diarrhea, and failure to thrive. Pseudomonas aeruginosa is cultured from his respiratory tract. The physician informs the patient’s parents that their son has a disease that is caused by a mutation in a specific ion transporter. This patient has a mutation in the ion transporter of which of the following electrolytes? (A) Bicarbonate (B) Calcium (C) Chloride (D) Potassium (E) Sodium

50. factors that influence net flux are: (Fick`s law of diffusion) 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