Signalling and Transport PDF

- Additional processes influencing homeostasis: a. Circadian rhythm: - Some physiological variables changes according to circadian rhythm, including but not limited to: - Body temperature - Concentrations of growth hormone and cortisol in blood...

- Additional processes influencing homeostasis: a. Circadian rhythm: - Some physiological variables changes according to circadian rhythm, including but not limited to: - Body temperature - Concentrations of growth hormone and cortisol in blood - Urinary excretion of ions - Circadian rhythm follows a 24h light-dark cycle: - Central nervous system receives input from eyes and in response, influences various organ systems b. Feedforward control (anticipatory response): - While negative feedback loops can stabilise the internal environment, they are unable to prevent changes from happening in the first place - The human body also employs feedforward control mechanisms to make adjustments before the changes happen - Example: - When a person is about to begin exercising, their heart rate will increase before moving commences 64 c. Positive feedback: - Not all physiological functions are regulated by negative feedback loop - Some non-homeostatic physiological functions are driven by positive feedback loops - Unlike negative feedback (which eliminates the input signal), positive feedback loops amplify the input signal - Positive feedback loops can bring about an effect very quickly (amplification of the input signal accelerates the process) - Additional mechanisms in place to turn off the positive feedback loops - Examples: - Coagulation cascade - Childbirth - Surge of luteinising hormone after follicular phase of menstrual cycle 5.7.5. SUMMARY OF HOMEOSTASIS 1. State the hierarchical and interactive organisation of body functions: - Various organ systems: cardiovascular, digestive, endocrine, immune, integumentary, lymphatic, musculoskeletal, nervous, reproductive, respiratory and urinary - Body fluid compartments: intracellular fluid and extracellular fluid 2. Describe the concept of homeostasis: - Homeostasis is about maintaining stable physical/ chemical properties of internal environment (extracellular fluid) - Homeostatic control systems (sensor, control centre and effector) maintain regulated variable within predictable range - Mechanism of control: neural, endocrine and neuroendocrine 3. Explain the importance of homeostasis in the maintenance of physical health - Essential for survival and good health - Some abnormalities in physiological variables are tolerated short-term, but are harmful/ lethal in long run 4. Describe the principles of negative and positive feedback controls of physiological functions - Most homeostatic control systems are negative feedback loops, but some non-homeostatic functions are driven by positive feedback loops - Negative feedback loops are self-limited: they eliminate the input signal (deviation of regulated variables from setpoints) - Positive feedback loops amplify the input signala quick effect - Circadian rhythm and feedforward control also influence homeostasis 65 6. SIGNALLING AND TRANSPORT 6.1. CELL MEMBRANE TRANSPORT 6.1.1. OVERVIEW - Structure of a typical somatic cell: - 2 types of organelles: - Blue – membranous organelles - Orange – non-membranous organelles - Most membranous organelles have a phospholipid bilayer like the plasma membrane - Structure of plasma membrane: 66  - Very thin, ranging from 10-12 nm - 3 main components: 1) Lipids: - Majority of the surface of the plasma membrane (phospholipid bilayer) - Each phospholipid molecule have one hydrophilic head near the membrane surface and two hydrophobic tails inside - Other lipids like cholesterols are also present 2) Proteins - Account for around 55% of the total weight of the plasma membrane - 2 types of membrane proteins: - Integral proteins: part of membrane structure, cannot be removed without damaging plasma membrane - Peripheral proteins: bind to either the inner of outer side of membrane, can easily be separated - There are more integral proteins that peripheral proteins 3) Carbohydrates: - Account for around 3% of the total weight of the membrane - Components of complex glycoproteins - Membrane Transport: - The plasma (cell) membrane is a barrier, but: - Nutrients must get in - Products and wastes must get out - Permeability determines what moves in and out of a cell - Plasma membrane is selectively permeable: - Allows some materials to move freely - Restricts other materials - Membrane proteins (integral or peripheral) allow the passage of substances across the membrane 6.1.2. TYPES OF MEMBRANE TRANSPORT - Mechanisms of membrane transport: - Transport through a plasma membrane can be: - Active (requiring energy and ATP) - Passive (no energy required) - Three different mechanisms: A. Diffusion (passive) B. Carrier-mediated transport (passive or active) C. Vesicular transport (active) – non-selective 67 A. Passive Transport 1) Diffusion: - Molecules in solution move randomly causing mixing - Steeper concentration gradient in one part of a solvent than another - Factor influencing diffusion: - Distance (shorter distance) - Molecular size (small molecules) - Temperature (higher) - Concentration gradient (higher) - Electrical forces (oppositely charged) - Diffusion tends to spread materials from a region of high concentration to another of low concentration – “down a concentration gradient” - Diffusion across plasma membrane: - Simple diffusion: - Lipid-soluble compounds (alcohols, fatty acids, and steroids) - Dissolved gases (oxygen and carbon dioxide) - Channel-mediated diffusion - Involves transport proteins (~1 nm in diameter) - Water-soluble ions - Factors in channel-mediated diffusion: - Size - Charge - Interaction with the channel – leak channels 2) Osmosis: - Osmosis is the diffusion of water across the cell membrane - More solute molecules, lower concentration of water molecules - Membrane must be freely permeable to water, selectively permeable to solutes - Water molecules diffuse across membrane toward solution with more solutes - Volume increases on the side with more solutes 68  - Osmolarity: - Refers to the solute concentration of the solution - Describes solution itself - Tonicity: - A description of how the solution affects a cell - Isotonic (tonos = tension): - No osmotic flow occurs - Hypotonic: - Results in the osmotic flow of water into the cells; cell swells - Hypertonic: - Results in the movement of water out of the cell, cell shrinks 3) Facilitated diffusion: - Passive (no ATP required) - Channel proteins transport small organic ions - Leak channels (always open) vs gated channels (open upon stimulation) - Carrier proteins transport molecules too large to fit through channel proteins (glucose, amino acids) - Conformational changes to transport molecules across plasma membrane - Once the carrier proteins are saturated, no change in the rate of transport by increasing the concentration gradient - At this point, concentration gradient is no longer a factor affecting membrane transport 69 - Summary: - Diffusion is a passive transport mechanism, which occurs until the concentration gradient is eliminated - Lipid-soluble molecules and dissolved gases pass through the plasma membrane by simple diffusion - Channel and carrier proteins are required to passively transport small water-soluble molecules/ions and large molecules down their concentration gradients, respectively B. Active Transport 1) Primary active transport: - Move substrates against concentration gradient - Require ATP energy supply - Ion pumps for specific ions in one direction only (E.g. Na+, K+, Ca2+) - Exchange pumps counter-transport two ions at the same time E.g. (Na-K ATPase) 2) Secondary active transport: - The transport mechanism itself does not require energy, but the cell often needs to use ATP at a later time to preserve homeostasis (e.g. sodium-glucose linked transporter for glucose absorption along the intestinal tract) - E.g. Na-glucose transporter: - Move glucose and Na+ at the same time - Na+ is a free rider in this process - Na+ is needed to provide a driving force for the use of ATP via the Na+-K+ pump 3) Vesicular transport (bulk transport): - Materials move into or out of cells in vesicles - Active transport using ATP - Endocytosis: - Receptor-mediated; Pinocytosis; Phagocytosis - Exocytosis: - Granules or droplets are released from cells 70 - Receptor-mediated endocytosis: - Produce vesicle that contains a specific target molecule in high concentration - Begins when materials in the extracellular fluid binds to a receptor in the plasma membrane - Transport protein or hormone binds to surface receptor - Area coated with ligands from deep pockets in the plasma membrane - Pockets pinch off, forming vesicle structures called endosomes - Endosome fuses with lysosome, where degradative enzymes are enriched - Lysosomal enzyme causes removal of ligands and absorption into the cytosol - Lysosomal and endosomal compartments will be separated - Endosome fuses with the plasma membrane for exocytosis - Pinocytosis: - “Cell-drinking” - Formation of endosomes that are filled with extracellular fluid - Termed pinosomes - Process is not as selective as receptor-mediated endocytosis (no receptors involved) - Phagocytosis: - “Cell-eating” - Macrophages protect tissue by engulfing bacteria - Pseudopodia form around object and fuse to form a phagosome - Fuse with lysosome, contents digested, debris released into the cytosol 71 - Summary: - Carrier-mediated transport involves the binding and transporting of specific molecules by integral proteins - Active transport consumes ATP and is not dependent on concentration gradients - Vesicular transport moves materials (ligands, fluids, or solids) into or out of the cell in membrane vesicles 6.1.4. SUMMARY - Clinical relevance of membrane transport: - Many clinically important drugs affect the plasma membrane: - For some anaesthetics (e.g. chloroform, ether, halothane, nitrous oxide), potency is directly correlated with its lipid solubility - High lipid solubility accelerates the drug’s entry into cells and enhances its ability to block ion channels or change other properties of plasma membranes (and thereby reduce the sensitivity of neurons and muscle cells - Some common anaesthetics have relatively low lipid solubility, e.g. procaine and lidocaine, used as local anaesthetics - Both affect nerve cells by blocking sodium channels in their plasma membranes, which reduces or eliminates the responsiveness of cells to painful stimuli 72 6.2. CELL MEMBRANE SIGNALLING 6.2.1. OVERVIEW - General features of cell signalling: - 5 steps common to all signalling pathways: 1. In response to a stimulus, the chemical messenger is secreted from a specific cell 2. The messenger diffuses or is transported through blood or other extracellular fluid to the target cell 3. A molecule in the target cell, termed a receptor (a plasma membrane receptor or intracellular receptor), specifically binds the messenger 4. Binding of the messenger to the receptor elicits a response 5. The signal ceases and is terminated - Chemical messengers: - Also called signalling molecules - Transmit messages between cells - Nervous system – neurotransmitters - Endocrine system – hormones - Immune system – cytokines - Others – retinoids, eicosanoids, growth factors - Receptors: - Proteins that contain a binding site specific for a single chemical messenger and another binding site involved in transmitting the message - Either plasma membrane receptors or intracellular receptors - When a chemical messenger binds to a receptor, the signal it is carrying must be converted into an intracellular response - This conversion is called signal transduction 73  6.2.2. TYPES OF CELL-CELL COMMUNICATION 1. Direct contact (juxtacrine): - Direct contacts of proteins on plasma membrane for communication - The molecule stays attached to the signalling cell and binds to a receptor on an adjacent target cell, establishing physical contact between the two cells - E.g. Antigen presentation by macrophages: 2. Paracrine - Over short distances - The molecule released by the signalling cells diffuses to neighbouring target cells of a different cell type - E.g. Synaptic transmission using neurotransmitters: - E.g. Allergic reaction by histamine: 74 3. Autocrine - The secreting cells themselves express cell surface receptors for the signalling molecule - Cells release chemical messengers and can respond to itself - May lead to rapid mitotic feedback and tumour formation - E.g. T-cell self-proliferation 4. Endocrine - Long-distance communication - The molecule secreted by the signalling cell is transported to the target cell via the bloodstream - Binding to cells with suitable receptors to produce a lasting effect - E.g. Growth hormone - Types of signalling molecules and receptors: 1) Lipophilic signalling molecules and receptors: - Lipid-soluble (hydrophobic) molecules that freely diffuse through the lipid bilayer of plasma membranes to interact with specific receptors inside the target cell - Tend to have long half-lives (hours to days) – taken daily - E.g. Oral contraceptives contain lipophilic signalling molecules (e.g. ethinyl estradiol – a derivative of estradiol) - Binding of the signalling molecule to its receptor leads to alterations in gene transcription - Cytoplasmic receptors: The signalling molecule–receptor complex then translocates to the nucleus, where it binds to a specific DNA sequence - Nuclear receptors: These receptors alter the transcription of specific genes only when the appropriate signalling molecule diffuses to the nucleus and binds to them 2) Hydrophilic signalling molecules and receptors: - Water-soluble molecules that cannot diffuse through the hydrophobic core of cell membranes - Instead, these molecules bind to specific receptors at the cell surface, which triggers the activation (or inhibition) of signalling events downstream from the signalling molecule–receptor complex - Tend to have short half-lives (seconds to minutes) - E.g. Epinephrine is administered to treat severe acute allergic reactions that may lead to anaphylactic shock - Binding of the signalling molecule to its receptor will cause conformational change to the receptors > trigger signalling cascade - Two important classes of receptors for hydrophilic signalling molecules: - G-protein coupled receptors (GPCRs) - Receptor tyrosine kinases (RTKs) 75 - Signal termination: - Some signals need to turn off rapidly, some signals can turn off more slowly, while other signals may persist throughout our lifetime - Chronic diseases are caused by failure to terminate signal response at the appropriate time - Means of termination: - Chemical messenger (no longer secreted, catabolised) - Receptor desensitised to the messenger by phosphorylation, internalisation, degradation - GTPase activity (intrinsic and/or accelerated by GAPs) - Degradation of second messenger - Removal of phosphate groups from proteins by phosphatases 6.2.3. G-COUPLED PROTEIN RECEPTORS - General features of signalling via G-protein coupled receptors (GPCR): 1) Trimeric G-proteins: - Act as “molecular switches” - Most physiological processes based on GPCR - Composed of 3 subunits: 𝛼, 𝛽, 𝛾 - Inactive G-protein has GDP bound to 𝛼-subunit which is in turn attached to the 𝛽- and 𝛾- subunits - Activation involves exchange of GDP to GTP via the action of GEF - Activated G-protein has GTP bound to 𝛼-subunit and separated from the 𝛽- and 𝛾-subunits - Inactivation involves intrinsic GTPase activity of G-protein, which hydrolyses the bound GTP into GDP (accelerated by GAP) 76 2) Typical signalling of GPCR: - Involves extracellular domain which binds to signalling molecule - Involves intracellular domain which interacts with trimeric G-protein - Steps: 1. A signalling molecule (first messenger) binds to the extracellular domain of the receptor and causes a conformational change in the protein - α- and 𝛾-subunits have covalently bonded lipid tails that are anchored to the membrane - In the inactive state, the α-subunit of the G‐protein binds GDP - GPCR bound to signalling molecule is now the GEF for the trimeric G-protein 2. The intracellular domain (acting as a GEF) activates its associated trimeric G-protein by triggering the exchange of GDP for GTP 77  3. The activated GTP-bound G-protein interacts with a membrane-bound effector protein, which is usually an enzyme that produces (or hydrolyses) a second messenger - Second messengers, e.g. cAMP, cGMP, DAG, IP3, Ca2+ 4. Signalling is terminated by various mechanisms: dissociation of signalling molecule, inactivation of G-protein, and reduction of cellular concentration of second messengers - Common GPCR signalling pathways: - Different GPCRs can interact with different G-proteins: Gs, Gi, Gq, Gt A. Signalling via Gs: - GPCR-mediated activation of the α-subunit of Gs activates the effector protein adenylate cyclase, an enzyme that converts ATP to cAMP - The second messenger, cAMP, stimulates the activity of protein kinase A (PKA), an enzyme that alters the activity of various proteins via phosphorylation (attachment of phosphate group to activate/ inactive enzyme) 78 B. Signalling via Gi: - GPCR-mediated activation of the α subunit of Gi inhibits the effector protein adenylate cyclase and prevents the production of cAMP and other downstream events - cAMP is not produced. PKA is not activated. C. Signalling via Gq: - GPCR-mediated activation of the α subunit of Gq activates the effector protein phospholipase C (PLC), an enzyme that cleaves the membrane lipid phosphatidylinositol 4,5-bisphosphate (PIP2) into DAG and inositol 1,4,5-triphosphate (IP3) - DAG activates protein kinase C (PKC) - PKC phosphorylates the serine/threonine residues of protein targets, which has several consequences on cellular metabolism, growth, and differentiation - IP3 causes release of calcium from the endoplasmic reticulum - Ca2+ binds to other proteins to activate them 79 D. Signalling via Gt: - GPCR linked to Gt is activated by light - GPCR-mediated activation of the α subunit of Gt activates the effector protein cGMP phosphodiesterase, an enzyme that hydrolyses cGMP into the noncyclic 5′-GMP. - The reduced levels of cGMP cause hyperpolarisation of visual cells, which is important for vision - Diversity of GPCR signalling: - Same hormone may induce different physiological responses in different cells - Binding of epinephrine to 𝛽-adrenergic receptor (type of GPCR): - cAMP causes relaxation of bronchial and intestinal smooth muscle - Relieve bronchospasm (asthma attacks) - cAMP causes contraction of heart muscle - Restore cardiac rhythms (cardiac arrest) 80 - Clinical implications of GPCR: - E.g. Cholera: - An acute diarrhoeal illness caused by infection of the intestine with Vibrio cholerae bacteria - After swallowing food or water contaminated with the cholera toxin - Due to signalling via Gs: - Involves conversion of ATP into cAMP - cAMP needs to be terminated after some time by inactivating the 𝛼-subunit of Gs - Cholera toxin (from Vibrio cholerae) prevents inactivation of Gs 𝛼-subunit - Covalently modify the alpha subunit and decrease its intrinsic GTPase activity → Remains in the active GTP-bound form → Signal cannot terminate - Continued stimulation of adenylate cyclase eventually leads to overproduction of cAMP - In intestinal cells, the elevated cAMP in cells lead to opening of chloride channels and a loss of electrolytes and water - Manifests as diarrhoea [Similar mechanism for the action of heat-labile enterotoxin secreted by E. coli (traveller’s diarrhoea)] 6.2.4. RECEPTOR TYROSINE KINASE - General features of signalling via receptor tyrosine kinase (RTK): - Play important role in cellular processes, including growth, motility, differentiation and metabolism 1) Monomeric G-protein: - E.g. RAS - Inactive when bound to GDP, active when bound to GTP - RAS possesses intrinsic GTPase activity, and cycles between the inactive and active forms - Similar to trimeric G-protein, activation involves GEF and inactivation involves GAP 81 2) Typical signalling of RTK: - Involves extracellular domain which binds to signalling molecule - Involves intracellular domain which possesses tyrosine kinase activity, interacts with adaptor and docking proteins (when receptors dimerised) - Steps: 1. A signalling molecule binds to the extracellular domain of the RTK (existing as 2 monomers) – this induces a conformational change that causes receptor dimerisation 2. The dimerised receptor phosphorylates specific tyrosine residues on itself (auto- phosphorylation/ cross-phosphorylation) 3. These phosphotyrosine residues are recognised and bound by adaptor and docking proteins, which activate downstream signalling pathways that are either dependent or independent of the small, monomeric G-protein RAS - RAS is activated after binding to GEF 82  4. RAS-dependent signalling is facilitated by members of the mitogen-activated protein kinase (MAPK) family, whereas RAS-independent signalling is facilitated by other types of kinases. Both signalling pathways trigger the phosphorylation of protein targets in the nucleus, plasma membrane, and cytoplasm, leading to alterations in gene transcription and protein activity 5. Signalling via RTKs is terminated via multiple mechanisms that include degradation of the signalling molecules by extracellular proteases, ligand-induced endocytosis of the receptors followed by lysosomal degradation, accelerated RAS inactivation, and dephosphorylation of target proteins by phosphoprotein phosphatases - Example of RTK pathway signalling – RAS and MAP kinase pathway: 1. Part of the intracellular domain of the receptor forms a binding site 2. The adaptor protein Grb2 binds to it 3. Protein SOS binds to Grb2: a. SOS is a guanine nucleotide exchange factor (GEF) for RAS b. SOS catalyses the exchange of GDP for GTP on RAS, activating RAS 4. RAS binds to the protein Raf: a. Raf is a protein kinase which phosphorylates and activates another protein kinase MEK, which subsequently phosphorylates and activates another protein kinase ERK b. The MAP kinase cascade ultimately leads to an alteration of gene transcription affecting cell survival and proliferation 83 - Clinical relevance of RTK: 1. E.g. RAS and cancer: - Mutant forms of RAS - 30-50% of lung and colon cancer, 90% of pancreatic cancers are related to point mutations in RAS - Mutation causes decrease the intrinsic GTPase of RAS - Lock in the active (GTP-bound) state for a much longer period - Excessive signalling from mutated or overexpressed RTKs → human cancer - May cause more proliferation of cells/ overexpression of genes - RTKs are targets for drugs that inhibit such excessive signalling (e.g. breast cancer drug herceptin targets HER2, an RTK) 2. RTK and achondroplasia - Fibroblast growth factor receptor 3 (FGFR3) is an RTK - Important role in regulation of bone growth - In chondrocytes (cells that synthesise cartilage in long bones): - FGF bind to two FGFR3 → dimerisation → autophosphorylation → activation of signalling pathways → inhibits chondrocyte differentiation and proliferation - Reduced number of chondrocytes → decreased cartilage deposition (template for bone formation) - Most people with achondroplasia have mutated FGFR3 → increases FGFR3 activity (without FGF) → impaired chondrocyte proliferation and cartilage deposition → reduced length of long bones 84 85 6.3. MEMBRANE EXCITABILITY 6.3.1. OVERVIEW - In our body, some cells carry a transmembrane potential by which signals can be transmitted from one site to another in the same cell, or transmitted from one cell to another - Types of transmembrane potentials: 1) Resting potential: - The transmembrane potential of resting cells 2) Graded potential - Temporary, localised change in resting potential 3) Action potential - Is an electric impulse produced by graded potential - Propagates along surface of axon to synapse 6.3.2. RESTING MEMBRANE POTENTIAL - Measuring the resting membrane potential: - Can be measured with glass pipette with fine tip - Uneven electrical charge distribution - Connected to a voltmeter which can measure the potential difference across the cell membrane - Inside of membrane is negative - Movement of ions influenced by diffusion: - Influx of Na+, Cl-; efflux of K+ - Both chemical and electrical in nature: - The concentration difference in extracellular fluid and cytosol will generate passive force to move the ions across the cell membrane - As they are charged, this will also generate a potential generation through the movement of these ions 86 - Electrochemical gradient: - The sum of chemical and electrical forces - Chemical gradients: [Na+], [K+] - Electrical gradients: separate charges of positive and negative ions - Membrane is selectively permeable, so in (a), since there are no membrane proteins present, there is no movement of ions - In (b), a leak channel for K+ ions is inserted into the membrane, so there is movement of K+ ions from left to right compartment down concentration gradient (channel-mediated diffusion) - In (c) a state of equilibrium is reached, where diffusional force is equal to the electrical force - Reality is more complicated than the model: - The chemical gradients involve the concentration different across the cell membrane of different ions, while the electrical gradient is contributed by the uneven distribution of charge ions across the cell membrane - There are leak channels for both the positive and negative ions - Resting potential: - ~ –70 mV, measured by a voltmeter - The resting potential varies widely with the type of cell - A typical neuron has a resting potential of approximately – 65 to – 70 mV - Resting membrane with closed chemically gated sodium ion channels as the ligands are absent 87 - Active forces across the membrane – the sodium-potassium exchange pump: - Powered by ATP - Carries 3 Na+ out and 2 K+ in - Balances passive forces of diffusion to maintain the resting potential - Utilises as much as 70% of ATP in the brain - The sodium-potassium exchange pump stabilises the resting potential, which involves steady- state movement of ions through leak channels - Classes and states of gated-ion channels: 1) Chemically gated channels: - Open or close when bind to specific molecule called ligand - E.g. Receptor of ACh at neuromuscular junctions (ACh receptor) - When ACh binds to extracellular domain of ACh receptor, the channel opens, allowing positively charged ions to go into the cell 2) Voltage gated channels: - Open or close in response to change in the transmembrane potential - Major features of excitable membrane (where membrane is capable of conducting and generating action potential) 3) Mechanically gated channels: - Open or close in response to distortions in the cell membrane surface - When pressure is applied to the cell, the channel opens - Mechanosensitive receptors: important in touch, pleasure and vibration 88  6.3.3. GRADED POTENTIAL - Neuron is a highly polarised cell - Contains short dendrites, which receive information and transmits signal to the cell body - Contains long axon, which transmits signal from the cell body to the axonal terminal - Each neuron receives information in the form of graded potential in the dendrite and at the cell body - Stimulus produces graded potential - Graded potential at the synaptic terminal triggers the release of chemicals (neurotransmitters) - Graded potential: - Also called local potential - Cannot spread far from the site of stimulation - Includes EPSP and IPSP discussed below - Generation of graded potentials: 89 - Excitatory neurotransmitters: - Activated receptor is to open non-selective cation channels - Membrane depolarisation brings the membrane potential closer to the threshold [increase in membrane potential] - The potential change is called an excitatory postsynaptic potential (EPSP) - Measured by glass electrode connected to a voltmeter - Inhibitory neurotransmitters: - The postsynaptic response to the neurotransmitter is a hyperpolarisation, which reduces the likelihood to generate action potential [decreased transmembrane potential] - The potential change is called an inhibitory postsynaptic potential (IPSP) - Interactions between EPSPs and IPSPs: - EPSPs and IPSPs reflect the activation of different types of chemically gated channels - Interactions between EPSPs and IPSPs determine the transmembrane potential - If EPSP and IPSP are equal in size but have opposite effects, the membrane potential remains at the resting level 90  - Graded potential vs action potential: 6.3.4. ACTION POTENTIAL - Measuring action potential in a neuron: - Electrophysiological setup - A way to study the properties of action potential is to artificially inject current into a neuron by an (stimulating) electrode, and another electrode records the change in the membrane potential in response to the injected current - When current is injected, this will generate many action potentials in succession - The rate of action potential generation depends on the magnitude of the continuous depolarising current - Every spike represents on action potential, and the spikes have a distinctive pattern of the change in the transmembrane potential versus time 91 - Depolarisation and hyperpolarisation: - The properties of the channels determine the nature of the change: - The opening of Na+ channels causes depolarization - The opening of K+ or Cl- channels causes hyperpolarization - The stronger the stimulus, the greater the change in the transmembrane potential - Effect of membrane depolarisation or hyperpolarisation persist only when there is a chemical stimulus - When the stimulus is removed, transmembrane potential returns to normal (repolarisation) - Change in transmembrane potential (depolarisation or hyperpolarisation) is below the threshold value (about 10 mV above the resting potential) - When the chemical stimulus is strong enough to depolarise the membrane potential, reaching the threshold potential, it causes action potential - Generation of an action potential: - Propagate changes in the transmembrane potential - Affect an entire excitable membrane - Link graded potentials at cell body under stimulations - At the resting membrane potential, the axolemma contains both voltage-gated sodium channels and voltage-gated potassium channels that are closed [Voltage-gated sodium channels have two gates: the activation gate and the inactivation gate; Voltage gated potassium channels only have one gate] 92 - Steps: 1) Depolarisation to threshold - Stimulus is large enough to open the voltage-gated sodium channel - Occurs at threshold value (-60 mV to -55 mV) [Purple: sodium ions, orange: potassium ions] 2) Activation of sodium channels and rapid depolarisation - Rapid depolarisation - Na+ ions rush into cytoplasm (membrane is more permeable to Na+ ions) - Inner membrane changes from negative to positive 3) Inactivation of sodium channels and activation of potassium channels - At +30 mV, inactivation gates close (Na+ channel inactivation) - K+ channels open - Repolarisation begins 4) Closing of potassium channels - K+ channels begins to close at -70 mV - K+ channels finish closing at -90 mV - Transmembrane potential return to resting level, action potential is over 93 - Summary: 6.3.5. SIGNAL PROPAGATION ALONG AN AXON - More on signal propagation along an axon: 94  - Previous segments of axon still in refractory period (incapable of producing new action potential), so action potential always proceeds away from a site of generation - Action potential generated at the synaptic terminal is identical to the one generated at the initial segment - Rate: ~ 1 m/s - Voltage-gated sodium channels: - Voltage clamp: “clamp” the membrane potential at any chosen value - To measure the ion currents through the membrane of excitable cells - Properties of voltage-gated Na channel: - Open with little delay - Stay open for about 1 ms - Channels become inactivated, and cannot be open again by depolarisation (refractory period) - The membrane cannot respond to further stimulation until sodium channel inactivation ends → the refractory period ensures the unidirectional propagation of signals 6.3.6. FACTORS AFFECTING SIGNAL PROPAGATION A. Myelination - Neuroglial cells: - Diverse, many types - Account for around 50% of the volume of the nervous system - Weighs around 1.5 pounds in brain (3 pounds) - 2 types of neuroglial cells are involved in myelination: 1) Oligodendrocytes (CNS) 2) Schwann cells (PNS) 95 1) Oligodendrocytes: - In the CNS - Form cytoplasmic extension - The processes of oligodendrocytes are generally in contact with the exposed surface of the axon - Many axon in the CNS are completely covered by oligodendrocytes, which insulate them from contact with the extracellular fluid - The myelin serves as electrical insulation, increases the speed at which an action potential travels along the axon - Each oligodendrocyte myelinates several axons - Internode: the large area of the axon that is wrapped in myelin, 1-2 mm - Node: small gap that separates adjacent internodes [Astrocytes form the blood brain barrier] 2) Schwann cells: - In the PNS - Schwann cells form myelinated sheath - Each Schwann cell myelinates one segment of a single axon - A series of Schwann cells required to myelinate the whole length of an axon - Growth: membrane continuously wraps around the axon 96 - Saltatory conduction: - Action potentials “jump” from node to node - Depolarisation only occurs at nodes - Myelin allows current to spread farther and faster between nodes - Uses less energy - Myelinated axons cannot have continuous propagation - Ions rapidly cross cell membrane only at the node region (node of Ranvier) - Saltatory propagation carries nerve impulses many times more rapidly than continuous propagation - Propagation along myelinated axons: 97 B. Axon diameter - Axon diameter affects action potential speed - Affects myelinated axons more than unmyelinated ones - The larger the diameter, the lower the resistance, the faster the speed - The most important information (vision, balance, motor commands) is carried by large- diameter, myelinated axons - Classification of axons: 98 6.4. AUTONOMIC NERVOUS SYSTEM 6.4.1. OVERVIEW OF THE NERVOUS SYSTEM - Functions of the nervous system: - Receive, process, integrate, and respond to information - Enable perception and cognition - Control the activities of the somatic and visceral systems 99 - Organisation of the SoNS and ANS: - Division of the ANS: 1. Sympathetic division - “Kicks in” only during exertion, stress or emergency - Prepares the body for heightened activity (“fight or flight” response) - Stimulates tissue metabolism and increase alertness 2. Parasympathetic division - Predominates under resting conditions - Conserves energy and promotes sedentary activities, e.g. digestion - Two divisions may work independently, and most often have opposing effects - Two divisions may work together, with each controlling one stage of a complex process 100  6.4.2. THE SYMPATHETIC NERVOUS SYSTEM - Overall organisation of the sympathetic division: - Consists of shorter preganglionic neurons and longer ganglionic neurons - Preganglionic neurons originate from the spinal cord (T1-L2 segments) - Preganglionic neurons make contract with ganglionic neuron at autonomic ganglions Distribution of the sympathetic innervation: [Red: preganglionic; Black: ganglionic fibres] 101 - 3 types of sympathetic ganglia: 1. Sympathetic chain ganglia - Innervates eyes, salivary glands, thoracic organs; skin, fat and sweat glands 2. Collateral ganglia - Celiac ganglion - Superior mesenteric ganglion - Inferior mesenteric ganglion 3. Adrenal medulla - Innervates adrenal endocrine cells (release of epinephrine and norepinephrine) - Sites of ganglia in sympathetic pathways: - Grey matter: inside, butterfly-shaped - White matter: outside, due to myelination - Dorsal horn – sensory nuclei - Ventral horn – motor nuclei - Neurons/ Nuclei more laterally placed in spinal cord section - The cell bodies of the preganglionic neurons are in the lateral grey horns - Their axons enter the ventral roots of these segments - Preganglionic fibres running between the sympathetic chain ganglia interconnect them 102 - Ventral root of spinal cord gives rise to preganglionic neurons - Connection with ganglion neuron → ganglionic neuron comes out through the spinal nerve, innervates skin/ sweat glands - If there is no contact with the sympathetic ganglia, preganglionic neurons continue to grow to reach the collateral ganglia → innervates visceral organs in the abdominopelvic cavity 103 - If there is no contact with the collateral ganglia, preganglionic neurons continue to grow to reach the adrenal medulla and endocrine cells → controls the secretion of neurotransmitters into circulation 104 - Sympathetic varicosities – for smooth muscles: - Sympathetic preganglionic neurons release ACh at synapses with ganglionic neurons - Cholinergic synapses are always excitatory - Postganglionic fibres form extensive connection with the target cell (branching on smooth muscle cell surface) - These ganglionic neurons then release NE (adrenergic synapse) from vesicles at specific target organs - The synaptic terminals form a branching network, resembling a string of pearls - Each “pearl” is packed with neurotransmitter vesicles, called a varicosity - Synthesis and release of norepinephrine (NE) and epinephrine: - NE and epinephrine activate adrenergic receptors (α and β receptors, both are G protein‐coupled receptors (GPCRs)) - 2 Main Classes of Adrenergic Receptors: 105  1) Alpha receptors: - α1 (most common): activates Gq release of intracellular calcium ions; excitatory effect on the target cell - α2 : Binding of norepinephrine to α2 receptor activates Gi, which in turn inhibits adenylyl cyclase 2) Beta receptors: - β1‐3: Binding of norepinephrine to the β receptor activates Gs, which in turn activates adenylyl cyclase to generate cAMP for PKA activation [β-receptor and α2-receptors have opposite effects – β has excitatory effects while α2 has inhibitory effects] 6.4.3. THE PARASYMPATHETIC NERVOUS SYSTEM - Organisation of the Parasympathetic Division of ANS: 106  - Consists of longer preganglionic neurons and shorter ganglionic neurons - Ganglionic neurons are closer to the target organ/tissue (intramural) - Preganglionic neurons originate from the brainstem and sacral segment - Distribution of parasympathetic innervation: - Parasympathetic stimulation and ACh release: - All parasympathetic neurons release ACh as a neurotransmitter - Effects vary widely, depending on the types of receptors (nicotinic or muscarinic receptors) and the nature of the second messenger - Nicotinic receptors: - It works by opening chemically (ligand) gated ion channels in the membrane - Pentameric; conformational change allows Ca2+ and Na+ to pass through - Occur on ganglionic cells of both parasympathetic and sympathetic divisions - ACh always cause short‐lived excitation of the ganglionic neuron or target (immediate depolarisation of axonal membrane) 107 - Muscarinic receptors: - Muscarinic receptors are GPCRs in the parasympathetic division - Activated by muscarine from toxic mushrooms - Stimulation produces longer‐lasting effects than nicotinic receptors - Can regulate multiple downstream proteins → eventually moderate Ca2+ channels - The response can be excitatory or inhibitory depending on downstream effectors 6.4.4. SUMMARY - Anatomical differences between two ANS divisions: 108  Sympathetic Parasympathetic Consists of short preganglionic and long ganglionic Consists of long preganglionic and short neurons ganglionic neurons Location of ganglionic neurons further from target Location of ganglionic neurons closer to target region region Makes use of ACh as a neurotransmitter from Makes use of ACh as a neurotransmitter from preganglionic to ganglionic neurons (nicotinic) preganglionic to ganglionic neurons (nicotinic) NE/E activates α and β adrenoreceptors (GCPR) No NE/E activity No activation of muscarinic receptors ACh activates nicotinic AND muscarinic receptors Has widespread impact (involves circulatory Innervates visceral structure only (does not system) involve circulatory system) - Summary: - SNS includes voluntary movement of the muscles and organs, while ANS makes routine and unconscious homeostatic adjustment in physiological systems - The sympathetic division consists of short preganglionic neurons (ACh) and long ganglionic neurons (NE/E) involved in using energy and increasing metabolic rates - The parasympathetic division consists of long preganglionic neurons (ACh) and short ganglionic neurons (ACh) involved in conserving energy and lowering metabolic rate - Some organs are innervated by just one division, but most receive dual innervation 109 6.5. NEURAL ARCHITECTURE 6.5.1. OVERVIEW - Classification: - General architecture: - CNS: - Consists of the brain and the spinal cord - Exception: CN II (optic nerve) is part of CNS rather than PNS - PNS: - Consists of nerve fibres and nerve ganglia outside the brain and spinal cord, and special nerve ending - Divisions of the nerve system: 110  - Somatic nervous system (SoNS): - Voluntary, conscious control of activities of the body - Motor and sensory innervations to all parts of the body - Does not include the internal organs, cardiac muscles, smooth muscles, and glands - Autonomic nervous system (ANS): - Involuntary control of the body - Provide motor and sensory functions - Includes the internal organs, cardiac muscles, smooth muscles, and glands - Sympathetic and parasympathetic divisions - Basic components of the nervous system: - Neurons/ Nerve cells: - Process and integrate information, enable all sorts of functions - 10-200 billion neurons in the brain - Glia/ Neuroglial cells: - Supporting neurons - 90% of cells in the nervous system are glial cells 6.5.2. DEVELOPMENT OF NEURAL CELLS 111 - CNS: - Grey matter: - Consists of neuronal cell bodies, neuropil, glial cells and capillaries - For synaptic integration, information process, motor control and sensory perception - Grey: because of colour of neuronal cell bodies and capillaries - [Neuropil: unmyelinated axons, dendrites and processes of glial cells] - White matter: - Consists mostly of glial cells and myelinated axons - White: because major component of myelin sheaths is lipids, which is white-ish - For forming tracts (“information highway”) and transmitting signals - PNS: - Ganglia: - Collections of nerve cell bodies outside the CNS together with their supporting cells - Includes sensory ganglia such as cranial nerve ganglia and dorsal root ganglia; and motor ganglia for the sympathetic and parasympathetic system - Nerves: - Almost all nerves are in the PNS - Bundles of myelinated and/or unmyelinated axons with their supporting cells; for transmitting signals 112  6.5.3. CLASSIFICATION OF NEURONS - Structure of nerves: - 2 types of cells in the nervous system: 1) Neurons: - Structural and functional unit, can not divide, 10-200 billion 2) Glia: - Supporting cells, forming myelin sheath, can divide throughout the adult life - Neurons: - Structural and functional unit - 3 physiological properties: - Excitability, conductivity, secretion - Neurons gather and transmit electrochemical signals (through action potentials) - Structure: - Soma: - Containing nucleus and organelles - Dendrite - Propagate the electrochemical stimulation to soma - Axon - Conducts electrical impulses away from the soma - One or more, myelinated or unmyelinated - Synapse - Specialised contacts between neurons for transmission of information 113 - Classification of neurons: a. Based on morphology: - Pseudounipolar neurons - Looks unipolar but is not - One axon, one dendrite; fuse together to form one stem - Peripheral axon and central axon – looks like one process but is actually formed from two processes - Usually sensory neurons - Neuronal cell bodies are located in the dorsal root ganglia or in the cranial nerve ganglia - Peripheral process links to the special sensory receptors → collects signals and conducts them back to the nerve cell body → signal sent to CNS - Bipolar neurons - Axons closely related to special senses, e.g. inner ear, retina, olfactory cells - One end is dendrite (collecting stimuli), other end is axon (sending information towards the CNS) - Multipolar neurons - Lots of processes - Usually one long big axon, with lots of dendrites - Most of the neurons in the nervous system are multipolar neurons b. Based on function: - Sensory neurons: - Detecting stimuli - Pseudounipolar or bipolar neurons 114 - Motor neurons - Responding to stimuli - E.g. multipolar neurons in the ventral horn of the spinal cord: - Axon projecting into effectors such as skeletal muscles - In the sympathetic/ parasympathetic system, there is a 2 step motor neuron system (presynaptic and postsynaptic) → send motor signal towards smooth muscle - Upper motor neurons: - Big projecting neurons like pyramidal cells - Pyramidal cells found in primary motor cortex of the brain - Project signals into the motor neurons in the spinal cord - Can also be classified as interneurons - Lower motor neurons: - Spinal motor neurons - Interneurons - Receive signals from other neurons - Locally projecting small neurons forming networks with unmyelinated neurons - Integration of information - Process, store, retrieve - Make decisions 115 - General structure: - Cell body (Soma): - Big - Dilated region of the neuron - Protein (and neurotransmitter) producing cell - Lots of specialised organelles - Prominent nucleus and nucleolus - Nissl bodies: blue staining, rough ER - sER: produces hormones and lipids, regulates and releases calcium ions - Vesicles: containing neurotransmitter and other secretory substances - Neurofilaments: shape and structural support for the axon and to regulate axon diameter - Microtubules: for intracellular transport between cell body and axon terminals - Dendrites: - Located in vicinity of the soma - Multiple branching short extensions of cytoplasm of soma - Have greater diameter than axons - Normally unmyelinated - Form dendritic tree (field) for detection of stimuli - Dendritic trees significantly increase the receptor surface area of a neuron - Receive stimuli from other neurons via synapses or from the external environment - Contain all the organelles found in soma (occasionally Golgi apparatus) - Axons: - A single (or more), long extension from the soma, may branch at distal end (synaptic terminal) - Convey information away from soma - Myelinated or unmyelinated - Substructures of the axon: - Axoplasm: - Cytoplasm within the axons - Lacks Golgi apparatus, rough endoplasmic reticulum, free ribosomes and mRNA - Axon hillock: - Transition zone between soma and axon, conically shaped region - Initial segment: - Region of the axon between the apex of the axon hillock - The beginning of the myelin sheath and site of action potential generation - Telodendria end with synaptic terminal - Synaptic terminal: - Ramified end branches to synapse with other neurons or effector cells via synapses 116  - Axonal transport within neurons: - Newly synthesised protein molecules are transported to distant locations within a neuron in a process - Anterograde transport: from soma to axon terminal - Retrograde transport: from axon terminal to soma - Synapse: - Small junction across which a nerve impulse passes from an axon terminal to a neuron, a muscle cell or a gland cell - Classification: a. Based on function: - Chemical synapse - Electrical signals (ions) is transferred to chemical signals and then transferred back to electrical signals - Conduction of impulses is achieved by the release of chemical substances (neurotransmitters) from the presynaptic neuron - Neurotransmitters diffuse across the narrow intercellular space (synaptic cleft) - Electrical synapses - Electrical signals (ions) is directly moved from one synapse to another synapse - Mainly present in smooth muscle cells, cardiac muscle cells, bone cells and retinal cells b. Based on morphology - Axosomatic synapses – binding to axon body - Axodendritic synapses – binding to dendritic spine - Axoaxonic synapses – binding to another axon to form feedback loops (enhancing/ inhibitory effect) 117 - Structure of synapses: - Presynaptic terminal and presynaptic membrane - Synaptic vesicles containing neurotransmitter - Postsynaptic terminal and postsynaptic membrane - Contain membrane receptors and iron gates (protein coupled or inotropic) to receive chemical signals - Synaptic cleft - A narrow space (20-30 nm) between pre and postsynaptic membrane - Direction of neurotransmission: - ‘One-way’ transmission - From presynaptic terminal to postsynaptic terminal - Initiated by depolarization of presynaptic membrane - Modulated by positive or negative feedback of presynaptic auto-receptors, adjacent neurons or environment 118  6.5.4. CLASSIFICATION OF GLIAL CELLS - Overview: - A large population (neuron:glia = 1:10) of supportive non-excitable cells in the nervous system - Glial cells are able to divide and multiply throughout life - Central neuroglia (CNS): 1. Astrocytes 2. Oligodendrocytes 3. Microglia 4. Ependymal cells - Peripheral neuroglia (PNS): 1. Schwann cells - Form myelin sheath 2. Perineuronal satellite cells - Locate within the ganglia, surrounding the nerve cell bodies, controlling micro-environment - Provide electrical insulation and pathway for metabolic exchanges - Analogous to Schwann cells - Do not form myelin 119 - General features of glial cells: 1. Astrocytes: - Largest neuroglia - Do not form myelin - Irregular shaped with many processes - Two types of astrocytes: - Fibrous astrocytes: - Mainly in white matter - Long, usually unbranched processes - Astrocytoma is one of the most common adult primary brain tumour - Protoplasmic astrocytes: - Mainly in grey matter - Shorter, thicker, highly branched processes - Endfeet surround capillary - Functions: - Physical and metabolic support for neurons - Providing nutrition and trophic factors - Buffering K+ and neurotransmitters - Keep extracellular K+ low; keep electrolyte balance - Provide guidance for migrating neurons - Some astrocytes span the entire thickness of the brain forming a scaffold structure for neural cell migration for adult neurogenesis - Maintain and facilitate the blood-brain-barrier (BBB) - Processes stretch from blood vessels to neurons - The ends of the processes expand, forming end feet that cover large areas of the outer surface of the vessel or axolemma - Participating in the injury responses (astrogliosis) - Phagocytise neuronal debris and fill in space to form glial scar after injury - Reduce the spreading and persistence of inflammatory cells - Maintain and repair of the BBB [BBB: - Active metabolic filter - Restriction for selective communication between blood-borne substances and neural tissue - Provide protective environment for the CNS] 120 2. Oligodendrocytes and Schwann cells: - Both myelin-forming cells - Schwann cells in PNS, oligodendrocytes in CNS - Myelin sheath: - White lipids electrically insulating layered structure formed by specific types of glial cells - Surrounds and protects the axon - Greatly accelerates transmission of action potential along axon (from 0.5-2 m/s to 80-120 m/s) - Track for axon regeneration in PNS - Interrupted at regular intervals by nodes of Ranvier - One Schwann cell forms myelin sheath wrapping around only one internodal segment of one axon in PNS - One oligodendrocyte sends out multiple processes to form myelin sheath, wrapping around one or several internodal segment of up to 50 axons in CNS - Nerve impulse jumping (saltatory) from node to node along the myelinated → acceleration of conducting velocity 121 - Myelination: - Initiated when the Schwann cell mesaxon or processes of oligodendrocyte surround the axon - The mesaxon wraps around the axon in a spiralling motion, forming concentric layers - Cytoplasm is squeezed out from between the membrane of the concentric layers - Decreases capacitance across the cell membrane, and increases electrical resistance - Oligodendrocytes (CNS): - Bare, not embedded in glial cell processes - Lack of basal lamina or connective tissue - Schwann cells (PNS): - A series of Schwann cells covers the length axons abutting tightly - Nodes of Ranvier - Axons fit into grooves in the surface of the Schwann cells - A single axon or a group of axons may be enclosed in a single groove of the Schwann cell surface - Large Schwann cells may have 20 or more grooves 122 3. Microglia: - Smallest neuroglia - Immune surveillance cells - Antigen presenting cells - Production of inflammatory factors (e.g. NO, cytokines) - Phagocytic cells - Activated after injury - Become enlarged and ameboid shaped - From circulation 4. Ependymal cells: - Epithelial-like cells lining of the ventricle - Ventricles – internal fluid-filled cavities - Lack an external lamina - Forming choroid plexus for CSF production 6.5.5. RENEWAL AND REPAIR OF THE NERVOUS SYSTEM - Neural stem cells: - Multipotent - Self-renewing, multipotent adult stem cells that generate the main phenotypes of the nervous system - Generated throughout an adult's life via neurogenesis - Differentiate to replace lost or injured neurons or glial cells/ - Neurogenesis occurs in subventricular zone (SVZ) of lateral ventricles and the dentate gyrus of hippocampus - Population of of neural stem cells is very small so will not replenish lost neurons 123 - Injury response of the nervous system: A. PNS: - Release of debris which are toxic and secrete factors that inhibit regeneration - Vasculation provides inflammatory cells and macrophages; remove debris quickly - Schwann cells form myelination – myelin sheath reaches original target where axon will grow back to function normally - Sometimes myelinations will not grow back to reach original target → functional deficit B. CNS: - Release of debris which are toxic and secrete factors that inhibit regeneration - Due to presence of BBB, microglial cells cannot effectively remove debris - Astrocytes arrive but form glial scar and cut off the regeneration route - Debris still present to inhibit neuroregeneration - CNS can technically not regenerate - Demyelinating diseases: - Characterised by preferential damage to the myelin sheath - Lost ability to transmit electrical impulses along nerve fibres, causing physical, mental and psychiatric problems - Several immune-mediated diseases affect the myelin sheath, e.g. multiple sclerosis 124 6.6. MUSCLE TISSUE 6.6.1. OVERVIEW OF THE MUSCULOSKELETAL SYSTEM - Muscle tissue: - Tissue aggregates of specialised and elongated cells - Primary role of contraction - Can convert chemical energy into mechanical/ kinetic energy - 3 types of muscle: 1) Skeletal muscle (voluntary): - Striated - Skeleton attached or visceral striated 2) Cardiac muscle (involuntary): - Striated - Heart, heart walls, S/I vena cava, pulmonary veins 3) Smooth muscle (involuntary): - Non-striated - Walls of blood vessels, internal organs, near hairs, pupil, viscera - Myofilaments: - Tissue or molecule responsible for muscle contraction - Thin filament, F-actin - Thick filament, myosin II - Organisation of skeletal muscles: - Myofilaments form myofibrils - Myofibrils are responsible for contraction - Skeletal muscle cell is a multinucleated synthesium – myogenic cells fuse together to form a chain that becomes a single cell - Skeletal muscle cells can be as long as 1m 125  - Endomysium: delicate layer of loose connective tissue that supports the muscle cells - Muscle fascicle: muscular cell bundle that perform tasks together - Perimysium: wrapped around muscle fascicle, thicker layer of connective tissue; thickened blood vessels and nerve endings - Epimysium: very thick, dense irregular connective tissue; observable with bare eyes; larger blood vessels and nervous supply - Histological representation of skeletal muscle cells: - Nucleus located peripherally - Regular, parallel arrays of myofibrils 126 - Distribution of striations: - Due to difference in density of tissues - H band: myosin only, shortens during contraction - A band: myosin and actin, always equals to the length of myosin - I band: actin only, shortens during contraction 6.6.2. CLASSIFICATION OF SKELETAL FIBRES - Classification is based on: a. Velocity – how fast the muscle fibre can contract and relax b. Enzyme velocity – how fast the enzyme can break down ATP to provide energy c. Metabolic profile – how to produce ATP (oxidative phosphorylation or glycolysis) 1) Type I (slow oxidative): - Slow-twitch fatigue-resistant - Generate less tension - Slower ATP-enzyme velocity - Utilise oxidative phosphorylation to produce ATP - Need large oxygen supply - Large amount of myoglobin (red meat), mitochondria and blood capillaries - Long and slow contraction, sustain aerobic activity (e.g. postural muscle) - Marathon runners - Stains darker in histological slides 127 2) Type IIb (fast glycolytic): - Fast-twitch - Fatigue-prone - Generate more power - Faster ATP-enzyme velocity - Utilise glycolysis to produce ATP - Lower amount of myoglobin (white meat) and mitochondria - High anaerobic enzyme activity - Larger in fibre size - Present in muscles such as biceps and triceps - Fine movements (e.g. movement of digits, eye movement) - Short-distance sprinters - Stains lightest in histological slides (lowest oxidative enzyme activity) 3) Type IIa (fast oxidative glycolytic): - Intermediate in properties and functions between type I and type IIb fibres - Fast-twitch - Intermediate ATP-enzyme velocity - Utilise both glycolysis and oxidative phosphorylation - 400-800 m athletes 128  6.6.3. ULTRASTRUCTURE OF SKELETAL MUSCLES - Myofibrils: have mitochondria, glycogen, peripheral nucleus - Delivery and removal of calcium - Sarcoplasmic reticulum: smooth ER, forms extensive network, wrap around each myofibril, separate myofibrils completely - SR serves as large Ca2+ reservoir - Separates from A-I junction to A-I junction - Perpendicularly placed in between two sections of the networks of SR, cell membrane penetrates into the A-I junction to give a special tubule structure – T-tubules - T-tubules: invagination/ insertion into the cell membrane - Adjacent to T-tubules and the A-I junction is a terminal cisterna - Each T-tubule is accompanied by 2 terminal cisternae (at either end of the A-I junction) - Depolarisation from cell membrane goes into T-tubules → Conducted into A-I junction → release of Ca2+ from internal store → initiate muscle contraction - Muscle contraction – the sliding filament theory: 129 - Stage 1 – attachment stage/ rigour formation - Stage 2 – detachment/ release stage - Stage 3 – bending by ~ 5 nm - Stage 4 – returning of myosin head to original (unbent) position by losing ADP and phosphate → power stroke 6.6.4. DEVELOPMENT, REPAIR, AND RENEWAL OF MUSCLES - Skeletal muscle proliferation stops at the 24th week of embryonic development - Muscle grows in size but number remains constant or even decrease - Muscle satellite cells: - Myoprogenic cells - Located in between the cell membrane of skeletal muscle fibre and the external lamina that wraps around the muscle cells - Can differentiate for repair and renewal - Pool of cells is quite small and repairing function is limited - Hypertrophy: increase in size of muscles [stamina, blood supply, number of myoglobin in blood cell] - Atrophy: muscles are wasted away [associated with anorexia/ starvation] - Dystrophy: affecting production of proteins within the muscle cells → cause muscle cell death 130 - Motor and sensory innervation of muscles: - Muscle spindles and tendon organs: - Capsulated structures with special structures - Can find capsular fluid within - Stretch receptors – detect stretch of muscles and conduct signal back to dorsal ganglion for spinal nerves or towards the cranial ganglions for cranial nerves - Neuromuscular junction: - Also called the motor end plate - Specialised chemical synapses - Release acetylcholine - Structural integrity - Junctional folds increase surface area for signal conduction/ transmission - Each muscle fibre has one NMJ - One axon and terminal ends branched out can control multiple muscle fibres - Every muscle fibre controlled by one neuron is called one motor unit - Muscles in charge of very precise movement: for the same amount of muscle cells there are more neurons controlling them – each neuron control less muscle cells 131  6.6.5. ULTRASTRUCTURE OF CARDIAC MUSCLES - Overview: - Cross-striation - Mostly one nucleus (sometimes two), located in the centre of the cell - Same sarcomere and myofibril structure as skeletal muscles - Junction between each cardiac muscle cell is called an intercalated disc - Cell-to-cell attachment via intercalated discs, numerous cardiac muscle cells forming a cardiac muscle fibre - May form branched fibres - Involuntary spontaneous rhythmic contraction - Ultrastructure: - Intercalated disc has a transverse and lateral component - Three major structures in the intercalated disc: - Fascia adherens/ Zonula adherens – similar to in epithelial cells; for attachment or anchoring of thin filaments to the cell membrane → so that CM cell can effectively contract - Macula adherens/ Desmosome – found in both transverse and lateral compartments; tightly bind two cells together to protect the gap junction - Gap junction – type of (electrical) synapse; direct cell-to-cell communication, allow ions and molecules to pass through easily – CM fibre can work in synchronised way - T-tubule penetrate at the Z-line instead of A-I Junction (one T-tubule at each Z-line) - Adjacent to the T-tubules, the SR can form a big cisternae and travels parallel to the T-tubule - Diad instead of triad (1 T-tubule, 1 cisternae) - A lot of mitochondria, centrally located nucleus - Calcium-induced calcium release mechanism: - Depolarisation → Voltage gated Ca2+ channels open → Ca2+ flow in from extracellular fluid → triggers secondary release of internal Ca2+ within the SR storage 132  6.6.6. CONDUCTION, DEVELOPMENT, AND REPAIR OF CARDIAC MUSCLES - Cardiac conduction system: - Purkinje fibre: - Special type of cardiac muscle cell - In charge of conducting impulses - Looks thicker than normal cardiac fibre - Stained lighter (foamy colour) – due to large glycogen store - Myofibrils in lower amount and located more peripherally 133 - Development, repair and renewal: - Cardiac muscle cells normally do not proliferate nor regenerate after foetal development - Localised injury replaced by fibrous/ connective tissue (as seen in non- fatal myocardial infarction) - Ischemic heart attack causes death of a certain region of cardiac tissue/ cells - Adult cardiac stem cells do not exist 6.6.7. ULTRASTRUCTURE AND CONTRACTION OF SMOOTH MUSCLES - Smooth muscle: - Bundles or sheets of elongated fusiform cells - A single nucleus located in the centre of each cell - Thin and thick filaments do not arrange in an orderly formation → no striated appearance - Interconnected by gap junction to regulate contraction - Contractile apparatus arrangement does not give striated appearance - Involuntary contraction innervated by autonomic nervous system, initiated by a variety of impulses - Can respond to mechanical, electrical or chemical stimulation - Some smooth muscle cells can contract almost permanently and relax upon stimulation, some are relaxed and contract upon stimulation, some are semi-contracted, some contract in wavy fashion, etc. 134 - Structure and contraction: - Caveolae: small depressions on membrane - sER: small amount of calcium stored (peripherally) – endoplasmic store of calcium very important from smooth muscle - Thin and thick filaments constitute contractile apparatus - Intermediate filaments have dense bodies for attachment of thin filament - Myosin heads on either side of thick filament faces in opposite direction (side-polar arrangement) – more thorough interaction between the thin and thick filaments - Forms web rather than myofibrils in smooth muscles → contracting SM cell is squeezed - Lack of T system - Specialised for slow, prolonged contraction [Contraction of skeletal muscles is voluntary and directly under neural control] - Renewal and repair: - Smooth muscle cells are capable of dividing to maintain or increase their number (proliferation) - Undergoing mitosis to aid physiological functions or in response to injury - Containing regularly replicating populations of cells (uterus, blood vessels, stomach and colon) 135 6.6.8. SUMMARY OF DIFFERENT MUSCLE FEATURES 136 6.7. MUSCLE CONTRACTION 6.7.1. CHARACTERISTICS OF SKELETAL MUSCLE FIBRES - Formation of multinucleated skeletal muscle fibre: - Myoblasts fuse with each other to form muscle fibre - Some myoblasts are not fused and are present as myosatellite cells (muscle stem cells - They will divide and fuse with a damaged muscle fibre - They are used to assist in the repair of muscle tissue - Skeletal muscle fibres are very long (up to 30 cm) 137 - Myofibril: - Each muscle fibre contains hundreds to thousands of myofibrils - Made up of 2 filaments: - Actin – thin filament - Myosin – thick filament - Sarcolemma: - Acts as a plasma membrane - Has characteristic transmembrane potential due to the unequal distribution of positive and negative charges across membrane - Change in transmembrane potential is the first step of muscle contraction - Transverse tubules (T-tubules): - The signal to contract must be distributed quickly throughout the interior of the cell to ensure uniform contraction of muscles - This is carried out by the T-tubules - Narrow tubes that are continuous with the sarcolemma and extend deep into the sarcoplasm - Same general properties as the sarcolemma; action potential travels along T-tubules to the interior of the cell - Terminal cisternae: - T-tubules are associated with the swelling region of the neighbouring sarcoplasmic reticulum - Swelling region is the terminal cisternae - 2 terminal cisternae of the SR and 1 T-tubule together is known as a triad - Triad is responsible for controlling the calcium concentration inside a cell - Mitochondria: - Abundant in skeletal muscles - Longitudinal view of sarcomere structure: - Sarcomere is the smallest (repeating) functional unit of the muscle fibre - Dark (A) band; Light (I) band - Z line marks boundaries of sarcomere, M line is middle line of the sarcomere 138 - Actin (thin filaments): - Involves 4 proteins: F-actin, nebulin, troponin, and tropomyosin - Accessory proteins are nebulin, troponin and tropomyosin - F-actin: a twisted strand of two polymers of about 300-400 individual actin (G-actin) molecules - Long strand of nebulin extends along F-actin strand in the space between the two F-actin polymers to hold the F-actin together - Each G-actin molecule contains an active site that can bind to the myosin heads in the thick filament - At the resting state, the active sites are covered by the tropomyosin-troponin complex - Tropomyosin: double strand protein that covers the active site of actin - Troponin: receptor that binds to calcium ions - Myosin (thick filaments): - A thick filament contains about 300 myosin molecules, each made up of a pair of myosin subunits twisted around one another - The free head, which projects outwards towards the thin filament has 2 globular protein subunits - Head and tail functions as a hinge that allows the head to flip in different orientations - When the head pivots, it makes and generates a force that can move the thin filament towards the M line - All the myosin heads are arranged with tails pointing towards the M line - Each myosin has a core of titin - The exposed titin strands are elastic, so they will recoil after stretching - In resting sarcomere, titin strand is completely relaxed 139  6.7.2. MUSCLE EXCITATION - Sliding filament theory: - A band stays the same width - Z line moves closer (larger overlap between the thin and thick filaments) - I band gets shorter - Control of skeletal muscle activity: - Skeletal muscle activity is different from cardiac or smooth muscles since it is under neural control - When the stimulus in the form of an action potential travels along the axon of the motor neuron towards the synaptic tunnel, the motor neuron communicates with a skeletal muscle as a specialised connection structure - Communication between the nervous system and a skeletal muscle fibre occurs at a specialised intercellular connection known as a neuromuscular junction (NMJ) - Neural stimulation of a muscle fibre is coupled to the contraction of the fibre (excitation- contraction coupling) - The excitation-contraction coupling is also the linkage between the generation of an action potential into the sarcolemma and the start of a muscular contraction - Peripheral chemical synapse – NMJ: - Found in the PNS; presynaptic and postsynaptic cells are communicating with each other by the use of a chemical - Presynaptic nerve terminal contains the clusters of synaptic vesicles at the active zone - Postsynaptic muscle membrane contains the clusters of acetylcholine (ACh) receptors, facing the active zone of the nerve terminal - The clusters in the presynaptic and postsynaptic cells are perfectly aligned so as to increase the efficacy and efficiency of neurotransmission 140 - Acetylcholine receptors: - Ligand-gated cation channel (mainly Na+ and K+) - Allows passage of cations into the plasma membrane in the presence of a ligand - Pentameric structure with 5 subunits: - 2 alpha, beta, gamma and delta - The ligand binding sites are located on the alpha subunits - Once the ligand ACh binds to both the alpha subunits, there is a conformational change in the structure of the protein, which translates into the opening of the ion channel - Results in an influx of Na+ and a small efflux of K+ - Induces membrane depolarisation and muscle contraction - If depolarisation exceeds the threshold potential, there will be an action potential 141