Cell Systems Flashcards
What is the Hierarchy of Atoms to Organism?
Atoms –> Molecules –> Macromolecules –> Internal Cell Structures –> Cells –> Tissues –> Organs –> Organ Systems –> Organism
What is Histology?
The study of the microscopic structures of tissues.
What are the five stages of Histology?
1) Fixation
2) Embedding
3) Sectioning
4) Staining
5) Visualize
How do Light Microscopes work?
By passing light through a sample, or reflecting it off the sample (dissection microscope). Glass lenses magnify the image. Has a limited resolution - light scatters away from the focal plane.
What are the three different types of light microscopy?
- Brightfield
- Darkfield
- Phase contrast
What are Fluorescence Microscopes?
They are microscopes with a higher resolution than light microscopes. Using the energy of excitation light to get an electron to jump from its ground state to an excited state on a fluorescent tag. As the electron goes back to its ground state, it releases a photon. The photon is what is detected with the fluorescent atom. The excitation light needs a very specific wavelength which is achieved using a laser. They cost more and require more specific training.
What is the resolution of an Electron Microscope?
Vastly higher resolution compared to light microscopy (10,000,000 X). Using a beam of electrons rather than light. Instead of using a lens it uses electromagnetism to filter and direct the electrons into the required path (electrostatics control this). Heavy metals are used to stain the samples.
What does SDS_PAGE stain?
It linearizes the protein so that its in its primary structure. Done through heat but linear shape kept by SDS (gives protein a net negative charge). It is a protein specific stain within the gel, but stains ALL proteins. For specific staining you can do a process called Western blotting.
What are the four basic tissue types?
1) Epithelium
2) Connective tissue
3) Muscle
4) Neural
What are the characteristics of Simple squamous tissue?
- Epithelium tissue
- In a single layer (permeable e.g. gas diffusion, filtration, delicate, low friction)
- Built on a basal lamina
- Lines capillaries, alveoli and glomeruli
What are the characteristics of Stratified squamous tissue?
- Epithelium
- Cells are in layers
- Built on a basal membrane
- Non-keratinised (can’t dry out) found in mouth, eyes, internal membranes
- Keratinised (dry and impermeable) found in skin, especially palms and soles, gums, top of mouth, tongue
What are the characteristics of Simple cuboidal tissues?
- Epithelium
- Single layer on a basal lamina
- Cube shape means nucleus is not squished between two sides of the cell
- Common secretive tissues (passive or active release of materials)
- Found in kidneys, thyroid gland, eyes, salivary glands, ovaries, testes
What are the characteristics of Simple columnar tissue?
- Epithelium
- Line digestive tract (stomach, small and large intestine)
- More space for nucleus and for machinery for protein production or secretion
What are the characteristics of Stratified columnar tissue?
- Epithelium
- Layered
- Found in places with much higher levels of mechanical stress, but still need to be secreting things
- Found in salivary glands, conjunctiva (eyes), pharynx (back of throat), anus and male urethra
What are the characteristics of Pseudostratified and Ciliated tissues?
- Epithelium
- Ciliates columnar cells (similar to little hairs) move mucus and other liquids in respiratory tract, airways, Fallopian tubes, uterus and central spinal cord
- Pseudostratified = contain more than 1 cell type so looks like multiple layers, but is a single layer
General connections between structure and function of cells
Flat = Absorption Tall = Secretion Layers = Stress resistance
There will always be exceptions
What is the Glandular epithelium?
Epithelial tissue that secretes a substance. Can be a single cell or a complex organ. Can be arranged into complex structures (endocrine and exocrine glands).
What is the Endocrine Gland?
Secrete into extracellular space, circulated in the blood.
- e.g. pineal gland, pituitary gland, pancreas, thyroid
What is the Exocrine Gland?
Secrete into a duct, taken directly to another organ or the surface of an epithelium.
- e.g. sweat, saliva, ceruminous gland
What are simple exocrine gland types?
- Tubular (intestine)
- Branched Tubular (oesophagus)
- Coiled Tubular (sweat glands)
- Branched Alveolar (spacious glands e.g. skin)
What are Exocrine Gland compound types?
- Tubular
- Alveolar
- Tubuloalveolar
What are “proper” Connective tissues?
- Extracellular fibers
Can be dense (tendons) or loose (adipose). The type of proper connective tissue it is depends on the number of cell types, fiber density and the base solution. - mainly made from collagen fibres
What are Supporting Connective tissues?
- Densely packed fibers
- Uniform cell types
- Example of these is cartilage and bone
What are Fluid Connective tissues?
- Cells suspended in fluid
- Example is blood and lymph
What are Collagen fibers?
- “proper” connective tissue is usually made from these
- Very strong and flexible (strength varied by thickness of fibres)
- Key component in tendons and ligaments
What are Reticular fibers?
- also found in “proper” connective tissue
- Make up the other half of basal membrane (basal membrane is made up from the basal lamina plus reticular fibres)
- Important for keeping cells in the correct position in the tissue and allowing function
What are Elastic fibers?
- Can be found in “proper” connective tissue
- Allow connective tissue to stretch
Which cells create and are within “proper” connective tissue?
Create:
- Fibroblasts (Always found in connective tissue, secretes pro-collagen, cleaved to form collagen, also secrete hyaluronan which allows things to move around efficiently)
- Fibrocytes (Next level of differentiation from fibroblasts. Look after the connective tissue network, important for repair and regeneration )
- Adipocytes (also known as lipocytes or fat cells)
Within:
- Macrophages (Scavenger cells, remove dead cells and pathogens)
- Mast cells, lymphocytes and microphages (When tissue is injured or damaged)
- Mesenchymal cells (stem cell which can differentiate into bone cells, cartilage, muscle, adipocytes and more)
What is the structure and function of Connective tissue?
- Dense and aligned = strong in one direction, but can’t be twisted/sheared
- Mesh pattern =can counteract stress from multiple directions, making a protective shield
- Inclusion of higher levels of elastic fibers = connective tissue can return to original size after stretching
What are the structures and functions of Supporting connective tissue?
- Strength, structure, and protection
- Cartilage is thick, gel-like matrix made from proteoglycans (proteins that have sugar groups attached to them), collagen fibers and chondroitin sulphate (gives resistance to compression)
- Cartilage is avascular and aneural
- Cartilage matrix is composed of and maintained by chondrocytes
What are the general principles of cell signaling?
1) Synthesis of signaling molecule
2) Release by exocytosis
3) Transport to the target cell
4) Binding to and activation of specific receptor
5) Intracellular signal-transduction pathways
6) Change in cell a) short term changes in metabolism/movement b) long term changes in gene expression/cell development
7) Termination by inhibition of receptor OR Termination by removal of extracellular signaling ligand
What are different methods of cell communication?
- Autocrine
- Juxtacrine
- Paracrine
- Endocrine
What is Autocrine cell communication?
The cell targets itself. It releases a messenger which acts on a receptor on the membrane of the original cell. Cells use this to regulate their own function.
What is Juxtacrine cell communication?
Cell directly connected to one another. Important for growth factors.
What is Paracrine cell communication?
Cells targets a close, non-connected neighbour.
What is Endocrine cell communication?
Cell targets a distant cell via the bloodstream.
What are examples of Autocrine signaling?
- Interleukin 6
- Vascular endothelial growth factor
What are examples of Paracrine signaling?
- Transforming growth factor b (beta sign)
- Prostaglandins
What are examples of Juxtacrine signaling?
- Notch
What are examples of Endocrine signaling?
- Insulin
- Testosterone
Hormones are in which three classes according to biochemical structure?
- Peptide/protein hormones
- Steroids
- Amines
What is a Peptide/protein hormone?
Amino acid chains, most hormones are in this class
What are Steroids?
Steroid hormones are neutral lipids, based on cholesterol skeleton, not made from amino acids
What are Amines?
Amines (class of hormone) are derived from tyrosine. Amines from the adrenal medulla known as catecholamines.
What hormones are hydrophilic and lipophobic (storable)?
- Peptide/protein hormones
- Catecholamines
This means to cross the plasma membrane they need to be packaged in some sort of vesicle and moved by exocytosis or endocytosis.
What hormones are hydrophobic and lipophilic (non-storable)?
- Steroid hormones
Therefore cannot be stored they can only be produced on demand
How are hydrophilic signals recognized (peptide/protein hormones, catecholamines)?
By generating intracellular second messengers (cAMP, cGMP, Ca2+) once they have bound to a receptor on the target cell surface
How are hydrophobic signals recognized (steroid hormones)?
Carrier protein carries hormone in the blood. Hydrophobic signaling molecule released. Binds to intracellular receptors (nucleus or cytocol). Changes gene expression or increases in cGMP.
What is the process for peptide/protein hormone synthesis, storage, and secretion?
DNA template –> transcription –> mRNA translation on the rough endoplasmic reticulum –> pre-prohormone (used to store) –> prohormone –> hormone –> secretory vesicles –> stimuli (Ca2+) –> exocytosis –> extracellular fluid blood
Once used or if they arent needed, the liver will degrade the hormones (although sometimes kidneys can be used).
What can happen inside the cell due to a signal?
Short term changes are usually from activating or deactivating an enzyme.
Altering the cytoskeletal structure/proteins can have long or short term effects.
Long term changes can be from altering gene expression.
What makes a good signal?
- Specific (specific instruction to target cell)
- Small (diffuse rapidly, if a membrane needs to be crossed then lipid soluble)
- Speed (made, mobilized, or altered into active form very quickly)
- Amplification (one ligand binding to a receptor protein triggers many more downstream processes, maximizing efficiency)
How is a signal terminated?
- Receptor sequestration (receptor being taken out of the membrane and can be put into storage vesicle)
- Receptor down-regulation (receptor is taken out of the membrane and put into a vesicle which then fuses with a lysosome, receptor is then destroyed)
- Receptor inactive
- Inactivation of signaling protein (relay protein/signalling protein is inactivated)
- Production of inhibitory protein (inhibiting the pathway with a protein from a later stage in the pathway)
What are conserved components of signaling?
1) Changes in the activity/function of specific enzyme/protein already in the cell
2) Changes in the amounts of specific proteins produced by a cell (usually transcription factors to stimulate/repress gene expression)
What are the main plasma membrane receptors?
- G-protein coupled receptors (GPCR)
- Tyrosine Kinases (RTK)
- Ion Channel Receptors
What is the main intracellular receptor?
- Steroid Receptors
What are proteins in the cell membrane?
- Single or multiple alpha helices
- A rolled up beta sheet
- Linked to membrane via fatty acid chains or via a GPI anchor
What are examples of single membrane-spanning receptors?
- Protein tyrosine kinase-linked receptors (PTKRs)
- Serine/threonine kinase-linked receptors (S/TKRs)
- Particulate guanylyl cyclases (pGCs)
- Non-enzyme-containing receptors
What are examples of Multi-pass membrane-spanning receptors?
- Ion channel receptors
- G-protein-coupled receptors (GPCRs)
- Sigma receptors
What is Fixation in histology?
Fixing your cells in place so they don’t move during the procedure. Get a snapshot of what the system looked like at the time of dissection. Can be done using heat or perfusion to remove the water from the system.
What is Embedding in histology?
Embedding the system you are studying in wax, a light epoxy, acrylic or agar (agar most common). Used to have a solid structure of your sample to aid in slicing it. Can be embedded horizontally or coronally.
What is Sectioning in histology?
Sectioning is slicing the sample once it has been fixed and embedded. Usually slices are made using a microtome.
What is Staining in histology?
Staining is used to add colour to the structures so that the contrast between sections is more strong, making specific sections more visible.`
What is Visualisation in histology?
Visualisation is when you look at your samples usually using a light microscope. Requires a good quality microscope and good quality of samples (means the previous histology stages must be completed correctly).
What is immunochemistry?
Done using a fluorescence microscope. Use stained antibodies to look at your sample in more detail. Can be used to:
- see cells (resolution)
- the quantity of cells
- how cell systems change
- interactions between different systems
What is electrophysiology?
Allows you to look at living cells and see what is happening in real time. There are three categories for this:
- Extracellular (electrodes are close to the cell but not touching or inside, commonly neuronal preparations, good for picking up change in voltage in area around the cell during neuro transmission - used to examine exocytosis)
- Intracellular (electrodes are going inside the cell and can look at the changes in electrical potential within a cell)
- Patch-clamp (electrode gets pushed up against the cell and touches the membrane. The a very small amount of suction is applied to draw the membrane slightly up. Used to examine how ions move through a single channel. High resolution)
What is a general description of epithelial tissue?
Tissue that lines your organs thats on the outside edge of your organs. Also skin is epithelial
What are three main categories of supporting tissue?
Hyaline Cartilage - found between two surfaces that need to move past each other smoothly and easily
Fibrocartilage - a lot of collagen, found at important joints (knee), very smooth and slippery
Elastic Cartilage - can hold a fixed shape and if malformed they can spring back into place e.g. ear
What is fluid connective tissue?
Blood can be a fluid connective tissue as it connects organs in our body through the blood and lymph system.
How are steroid hormones synthesised, stored and secreted?
Steroid hormones are produced from cholesterol (cholesterol from low density lipids within the liver). Most cholesterol comes from the diet but some is produced de novo within the liver. Then the cholesterol is feeding through the synthesis pathway to produce various steroid hormones e.g. aldosterone, testosterone. These hormones are not stored.
What is signal to noise ratio in cell signalling?
Difference between an actual signal and the background noise.
What happens inside the cell when a signalling molecule has been made?
- Signalling molecule arrives at the target cell and receives the message.
- A transduction pathway (cell signalling pathway) is required for the signal molecule to be converted into an internal response.
- Receptor in the plasma membrane converts the extracellular signal into a language that the intracellular machinery can understand.
- Relay protein in then used to mass the message on from the receptor to other proteins/structures within the cell.
- Relay proteins pass to transducer proteins. The transducer proteins amplify the signal so that multiple other proteins can be activated (common the transducers are enzymes which synthesis second messengers).
- Integrator proteins are able to respond to multiple inputs. Bring together information from multiple branches of a signal cascade in order to give a more specific response.
- Distributor protein then binds with integrator protein. The distributor proteins then distribute the activity onto the final proteins which make changes to the cell. This can have a few different final cellular outcomes.
What are the two general key responses when receiving an external signal?
- Changes in the activity/function of specific enzyme/protein already in the cell.
- Changes in the amounts of specific proteins produced by a cell (usually transcription factors to stimulate/repress gene expression)
What are main plasma membrane receptors?
- G protein coupled receptors
- Tyrosine kinases (RTK)
- Ion channel receptors
The majority of receptor cell types are going to lead to a change in gene transcription (not exclusive)
What are main intracellular receptors?
Steroid receptors
Receptors can be separated into two groups:
- Single membrane-spanning receptors (most common type are Protein tyrosine kinase-linked receptors - PTKRs, there are also Serine/threonine kinases-linked receptors - S/TKRs)
- Multi membrane-spanning receptors (Ion channel receptors and G protein-coupled receptors - GPCRs which are the largest group)
What are tyrosine kinase receptors (RTKs)?
Classic example of single pass transmembrane receptors
Operate as dimers (two separate halves of the receptor, need bound to the ligand to be a functional receptor)
Have a transmembrane domain and an internal edge with kinase domains
Once dimerised, they can recruit a variety of different transducer proteins, which activate amplifier proteins and this often leads to the production of second messengers
What are Serine/threonine kinase-linked receptors (S/TKRs)?
Single pass membrane receptors
Can work as dimers or tetramers to create one receptor
Can also rely on the binding of >1 ligand
The internal portion of the receptor can act as both a transducer and an amplifier.
What are ion channels?
Can be ionotropic or voltage gated
Ionotropic are ligand gates. Binds on the extracellular side of the channel allowing ions to flow through the channel.
Good example is synapse.
Neurotransmitters are usually a stimulator but they can also be stimulated by sensory stimuli e.g. touch, temperature
They are the receptor, transducer and the amplifier.
What are G protein-coupled receptors?
Usually give short term effects by modifying existing proteins (enzymes, ion channels)
Effects seen in a short timeframe, though can have long-term effects by activating/repressing gene transcription
Over 900 types in the human genome
7 transmembrane domains, 4 extracellular domains, 4 intracellular (cytosolic) domains
Ligand usually binds to 3rd and 4th extracellular domain
This is a receptor that couples to a G-protein - g protein is completely separate. G protein is a molecular switch.
Its the GPCR and the G protein that work together to activate a membrane bound enzyme
They have a desensitising mechanism to stop signalling (short term event)
When its on its bound to GTP and when its off its bound to GDP
What is ATP and GTP?
Energy source, ATP is more common than GTP.
GTP is more used for switching things on and off.
What are some examples of Intracellular second messengers?
cAMP, cGMP, DAG, IP3
Ca2+ and several phosphatidylinositol derivatives also act as second messengers.
How can you downregulate a G protein with the protein beta arrestin?
Adds phosphate groups to one of the cyctosolic domains of the G protein receptor and that allows for the binding of beta arrestin. Beta arrestin brings multiple proteins together which trigger endocytosis.
How is a cAMP signal downregulated?
- Everything attached to the nuclear membrane (outside edge of the nucleus) by mAKAP. Two enzymes held in place by mAKAP (PKA and PDE)
- PKA adds phosphate groups to target proteins when activated
- PDE hydrolyses cAMP, keeping the levels low so PKA isnt activated
- The GPCR and membrane enzyme are active and levels of cAMP go up. The cAMP binds to PKA and a catalytic subunit is released.
- The catalytic subunit phosphorylates PDE
- PDE is super active and brings levels of cAMP down, PKA inactivates and returns to its regulatory domain
What is ubiquitination?
Proteins can be removed from a system using ubiquitination. This is when you add a ubiquitin tag to a lysine residues of a target protein. Used by RTKs and GPCRs. Can degrade signalling molecules and/or receptors.
What is the proteasome?
It is a protein complex which degrades proteins by proteolysis (proteases digest peptide bonds). Chops them up into single amino acids again.
What is a lysosome?
Membrane bound organelle filled with degradative enzymes at a low pH, can digest more than just proteins.
How does GPCR activate PKA
- 7 helix transmembrane receptor activates a G-protein once a signal is received.
- G-protein activates adenylyl-cyclase which takes ATP to make cAMP.
- cAMP second messenger releases catalytic subunit of PKA from regulatory subunit.
- Kinase activity on target proteins = short term changes in the cell
- Some isoforms of PKA can cross the nuclear membrane
- PKA (the kinase) activated CREB
- CREB interacts with DNA to activate gene transcription = long term changes in the cell
What are some examples of cellular responses to a rise in cAMP?
In the kidney, an increases in the vasopressin hormone will cause the kidney to respond by reabsorbing water.
In the ovarian follicle, an increase in FSH and LH hormones will cause the ovary respond by increasing the synthesis of oestrogens and progesterone
What are the characteristics of DAG and IP3?
- Both second messengers
- Both are derived from phosphatidylinositol (PI) - which is a membrane lipid
- Both have inositol heads which has various -OH groups which can be phosphorylated in various combinations which is controlled by specific kinases.
- PI -(PI kinase takes phosphate from ATP, then PIP kinase takes phosphate from ATP)-> PIP2 -(phospholipase C beta enzyme acts on lipids, cleaves the IP2 with hydrolysis)-> DAG or IP3
- DAG is lipophilic and remains embedded in the plasma membrane, doesnt diffuse into the cytosol
What are examples of a cellular response to a rise in IP3 and cytosolic (Ca2+)?
In the fibroblast, PDGF hormone will respond by DNA synthesis and cell division
In the blood platelets, Thrombin hormone will respond by aggregation, shape change and hormone secretion
What are scaffolding proteins?
Give you another way of controlling when it is that a protein is activated and where it is that the protein is activated.
What are Calcium ions?
- Ubiquitous second messenger
- A lot of functions
- Released from stores in the ER (sarcoplasmic reticulum in muscle cells)
- Come through voltage gated ion channels in the plasma membrane
- Calmodulin is a intermediary protein that communicates an increase in the calcium ion concentration to an actual effect on a target protein.
- Ca2+/calmodulin-dependant kinase 2 (CaMKII) = example
What are cellular targets of CaMKII?
Calcineurin - a phosphate involved in the regulation of many transcription factors
CREB - transcription factor involved in the expression of many genes including numerous neuropeptides
GSK3 - a major regulator of cell metabolism, cell proliferation and apoptosis
AMPA and NMDA receptors - regulators of synaptic plasticity (memory and learning)
What do tyrosine kinase receptors usually bind to?
Predominantly bind to growth factors and various other types of ligands.
What are the three structure aspects all tyrosine kinase receptors have?
- Extracellular ligand binding domain
- Single pass transmembrane domain formed from an alpha helix (this crosses the plasma membrane)
- Kinase domain which is on the inside edge of the plasma membrane (adds phosphate groups to proteins)
What are examples of RTK signal proteins?
When the signal protein EGF binds to EGF receptors this will stimulate cell survival, growth, proliferation, or differentiation of various cell types: acts as inductive signal in development.
How is RTK activated?
- Arrival of a extracellular ligand at the membrane on the outside edge of the cell and binding to a receptor
- Receptor wont function unless we have dimerization of the two halves of the receptor when the ligands are bound
- The sides of the receptor will begin to phosphorylate each other (cross phosphorylation) and will add multiple phosphate groups to the kinase domain on the inside edge of the membrane
- Final result is multiple phosphate groups as receptor is fully active. The phosphorylated tyrosine’s acts like docking sites that other proteins can bind to in order to interact with the receptor and continue the signalling pathway. For the proteins to interact they usually have a PTB domain or an SH2 domain.
What are the differences between the SH2 domains and PTB domains in RTK signal transduction?
Proteins with a SH2 domain can bind directly to a phosphate group on the receptor
The PTB domain is actually being shown on a scaffolding protein which together are forming a protein complex. Scaffolding protein binds to the receptor meaning it itself gets phosphorylated and then becomes a docking point for the other signalling proteins. Therefore a docking protein is recruited to the receptor via a PTB domain.
Any protein with a SH2 or PTB domain will have the ability to bind to phosphorylated tyrosine’s.
What is the PI 3-kinase and Akt pathway?
- Classic pathway for activated RTKs
- After receptor is activated, the PI3K is activated and directly interacts with the tyrosine
- PI3K has a regulatory domain and a catalytic domain
- PI3K is a lipid kinase
- PI3K is interacting with the inositol ring on PIP2 on the inside edge of the plasma membrane. It replaces the –OH group on position 3 of the inositol ring with a phosphate group. This is where PIP2 can diverge and can be used to produce PIP3 or second messengers.
- If PIP3 is produces then PIP3 allows proteins to bind the membrane
- Two key proteins bind PIP3 (PDK1 and Akt(PKB))
- Akt has a catalytic domain and an inhibitory domain
- Binding to the PIP3 in the membrane allows the catalytic domain to become physically free to react
- PDK1 also recruited to membrane, which phosphorylates Akt. Cytosolic kinase (PDK2) phosphorylates a second residue on Akt
What are examples of downstream effectors of Akt?
- Metabolism
- Translation
- Proliferation
- Survival
- Angiogenesis
These can either have metabolic effects (enzymes involved in the metabolism of carbohydrates or sugars, metabolism of lipids or proteins or amino acids) mitogenic effects (same route as mitogenesis, proliferation, differentiation, apoptosis and cell survival).
Can be used to look at metabolic diseases (diabetes) or mitogenic diseases (cancer).
Example of Akt and cell survival/ apoptosis when Akt is inactive.
- Akt is inactive - no signalling occurring
- When Akt is inactive a cytosolic protein called “BAD” is able to interact with two proteins called BCL-2 and BCL-XL.
- BAD is able to form a dimer with either of these proteins.
- BCL-2 and BCL-XL are pro-survival factors. If BCL-2 or BCL-XL arent bound to BAD they promote cell survival.
- If BCL-2 or BCL-XL are bound to BAD they cant act as pro-survival factors and the apoptosis process will start to happen.
Example of Akt and cell survival/ apoptosis when Akt is active.
- AKt is active (RTK activated, PI3K is making PIP3, PDK1 and PDK2 have activated Akt)
- Akt phosphorylates BAD
- Bad cant interact with BCL-2 or BCL-XL anymore
- BAD binds to 14-3-3 instead
- 14-3-3 forces BAD to stay in the cytosol so BAD cannot interact with BCL-2 or BCL-XL.
- Therefore the cell survives
What is the Ras, MAPK and the Akt pathway?
- Another classic pathway for activated RTK’s
- Note that Ras can activate PI3K and therefore Akt (e.g. cross talk)
What are Ras proteins?
- Ras is an integrator protein
- Ras is a small molecular GTPase (GTPase will hydrolyse GTP into GDP and inorganic phosphate)
- Most of the time Ras is bound to GDP = inactive
- To activate Ras = switch GDP to GTP
- Ras is a GTPase so can hydrolyse GTP to GDP, however is super slow
1) Switching GDP for GTP
2) Allowing GTP hydrolysis. Controlled by other enzymes. GEF - exchanges GDP for GTP. GAP makes Ras ~10,000X more effienct at hydrolysing GTP.
Ras activates quickly but can be quickly inactivated when needed.
How do MAPK’s work
- Can pass on messages with a series of phosphorylation’s
- Ras can activate MAPK through this cascade of phosphorylation events
- Tyrosine kinase receptor is activated which leads to Ras activation
- Series of phosphorylation = changes in gene transcription
Stimulus (Ras activation) –> MAP3K –> MAP2K –> MAPK –> Transcription factors
What are the three different types of muscle in our body?
- Skeletal (Voluntary)
- Cardiac
- Smooth
What is muscle specialised for?
Muscles are highly specialised for contraction. They are electrically excitable, contractile, extensible and elastic.
What are the general characteristics of Skeletal Muscle?
- Strong
- Quick
- Discontinuous
- Voluntary
- Striated
What are the general characteristics of Cardiac Muscle?
- Strong
- Quick
- Continuous
- Involuntary
- Striated
What are the general characteristics of Smooth Muscle?
- Weak
- Slow
- Involuntary
- Not Striated
What are the functions of skeletal muscle?
- 40-50% of total body mass
- Force production (for locomotion, postural support, for breathing)
- Support for soft tissue
- Control of entrances/exits (some are smooth muscle)
- Heat production (shivering)
- Nutrient store
What are opposing pairs in skeletal muscle?
Most skeletal muscles work in opposing pairs e.g. biceps contract and triceps relax for flexion, however triceps contract and biceps relax for extension.
Tendons attach muscles to bones which act as levers.
What are the three main layers of connective tissue in skeletal muscle?
- Epimysium: collagen for protection (which surrounds the entire muscle)
- Perimysium: collagen, elastic fibres, blood vessels, nerves (surrounds bundles of muscle fibres - fascicles)
- Endomysium: collagen, elastic fibres, blood vessels, nerves, capillaries and myosatellite cells (surrounds individual muscle fibre - cell)
How are tendons formed?
Muscle fuses together at either end to form tendons which attach to bone.
What are skeletal muscles made from?
Fibrous protein collagen
What are muscle fibres?
- Smallest unit of muscle that give a normal physiological response.
- Enormous (up to 300mm long)
- Multinucleate (multiple nuclei), but enclosed by single plasma membrane (formed by fusion of myoblasts)
- Composed of 100s of myofibrils each encased in intermediate filament network
- Composed of repeating longitudinal units (sarcomeres)
What are sarcomeres?
1000’s of sarcomeres are arranged in series. Each sarcomere consists of an orderly arrangement of thick and thin filaments.
Z line: Boundary between adjacent sarcomeres. Contains proteins that interconnect and anchor thin filaments
M line: Middle of sarcomere. Contains proteins that stabilise thick filaments
I band: Thin filaments only (bisected by Z line)
H zone: Thick filaments only (bisected by M line)
Zone of Overlap: Thick and thin filaments overlap (each myosin filament surrounded by 6 actin filaments)
A band: H zone and zone of overlap.
Sarcomere proteins: Stabilizing proteins, Elastic protein (Titin, largest known protein, restores sarcomere length after contraction), Proteins responsible for muscle contraction (Actin and Myosin).
What is Myosin?
Large family of motor proteins that move along actin filaments.
Isoform found in skeletal muscle: Myosin II (~500 kDa)
Head: Actin and ATP-binding sites - Generates force
Neck: Associated with MLCs - regulate head activity (4 light chains - MLCs)
Tail: MHCs form coiled coil (2 heavy chains - MHCs)
Bipolar organisation
What is Actin?
Family of globular multi-functional proteins
G-actin can polymerise to become F-actin. Actin filaments have polarity, +ve end attached to Z line.
Each G-actin molecule has an active site that can bind a myosin head.
What is the sliding filament model?
As the muscle contracts, actin filaments slide relative to myosin filaments
- -> Increasing amount of overlap
- -> Shorter distance between Z lines
Note compression of titin during contraction = compressed spring
1) Myosin head bound to ATP (low-energy state, 45° angle to filament
2) Myosin head mydrolyses ATP to ADP and Pi (high-energy state, 90° angle)
3) Myosin head binds to actin , forming a cross-bridge
4) Releasing ADP and Pi, myosin returns to low-energy state (45° angle), sliding the thin filament (power stroke)
5) Binding of new ATP releases myosin head
This cycle is repeated several times per second
Summary: Myosin is the motor molecule, coverts chemical energy (ATP) to mechanical energy (power stroke). Actin track along which myosin moves. ATP is the fuel.
Actin-myosin interactions are responsible also for other cell movements (e.g. cell division)
How is muscle contraction controlled?
A muscle would contract continuously until it ran out of ATP. This is NOT how a muscle is controlled. At a molecular level control is exerted by allowing or preventing myosin from binding to actin.
For muscle contraction to occur, myosin and actin need to bind. BUT in a relaxed muscle a Troponin-tropomyosin complex blocks myosin binding sites on actin filaments and prevents actin-myosin interaction.
Ca2+-binding by troponin C leads to conformational change
–> Troponin-tropomyosin complex slides away, exposing active sites on actin
What controls calcium?
In order to contract, a skeletal muscle must:
- Be stimulated by a motoneuron
- Propagate the electrical signal along its sarcolemma
Signal must be disturbed quickly throughout the large muscle cell, so all regions contract at the same time.
- Increase intracellular Ca2+ concentration, the final trigger for contraction
–> Whole process called excitation-contraction (EC) coupling - linking the electrical signal to the contraction
Nerve impulse –> Pre-synaptic Ca2+ channels open –> ACh release –> ACh binds to nAChRs –> EJP in muscle
Where does the motoneuron meet the muscle fibre?
Neuromuscular junction (NMJ)
What are muscle membrane action potentials?
The muscle membrane (sarcolemma) has voltage-dependant Na+ and K+ channels (similar to those in neurons)
EJPs are usually supra-threshold, so the muscle membrane spikes when its motorneuron spikes.
Muscle spikes enable signal to be conducted rapidly across large surface. Muscle spikes spread down T-tubules into interior of fibre (T = transverse)
What are muscle spikes?
Each muscle spike causes a twitch contraction
Individual twitch contractions can summate
At high frequency, twitches fuse into a tetanic contraction (this is the most force a muscle can develop)
How do we go from a muscle action potential to contraction?
2 membrane systems play a key role in EC coupling in skeletal muscle:
- External: T (transverse) tubules
- Internal: Sarcoplasmic reticulum (SR)
What is the function of T tubules?
Narrow tubes that are continuous with the sarcolemma. Conduct muscle spikes deep into the sarcoplasm.
What is the function of the Sarcoplasmic reticulum (SR)?
Elaborate smooth endoplasmic reticulum that surrounds each myofibril. SR tubules enlarge, fuse and form chambers (terminal cisternae) on either side of the T tubule - triad.
Muscle impulses trigger Ca2+ release from SR into sarcoplasm.
What are dihydropyridine and ryanodine receptors?
Ca2+ is stored in SR, pumped by ATP-dependant calcium pump. T tubules and SR are connected by 2 linked proteins:
- In T tubules: Dihydropyridine receptor (DHPR)
- In SR: Ryanodine receptor (RyR1: an intracellular calcium channel)
DHPR is voltage sensitive. DHPR is directly coupled to RhR. RhR is a calcium channel. Spike activates DPHR which opens RhR. Allows Ca2+ out of SR and into sarcoplasm. Contraction happens.
The duration of muscle contraction is dependant on what?
- Period of stimulation at neuromuscular junction
- Availability of free Ca2+ in sarcoplasm
- ATP availability
What happens as APs cease to arrive at neuromuscular junction?
ACh is broken down by AChE –> SR actively reabsorbs Ca2+ (calcium pump, also some Ca2+ transport into ECF, but less important) –> Ca2+ concentration declines –> Troponin-tropomyosin complex returns to normal position, blocking active sites –> Contraction ends, muscle returns passively to resting length
What are the stages of a skeletal muscle action potential twitch?
1) stable resting membrane potential
2) ligand-gated channels open, allowing Na+ or Ca2+ to depolarise cell to threshold
3) when the cell reaches threshold, voltage-gated Na+ channels open, causing an action potential
4) at this high membrane potential Na+ channels close, and K+ channels open, repolarising the cell
5) when the cell returns to the resting membrane potential, voltage-gated K+ channels close
What is the function of Cardiac muscle?
Very small proportion of body muscle mass. Only found in the heart wall and at the base of large veins.
Crucial function: Pump blood round the body (heart is our most heavily worked muscle contracting ~100,000 times)
Cardiac muscle must:
- Contract forcefully and rhythmically in a higher coordinated fashion - spiral arrangement
- Modify force according to circulatory needs
- Never tire or tetanise
What is the muscle structure of Cardiac muscle?
Cardiac muscle is quite similar to skeletal muscle. Cardiac muscle cells (cardiomyocytes) also consist of myofibrils and sarcomeres.
- Straited appearance
- Ensures good force of contraction. Mature cardiac muscle cells do not divide
- Non-fatal injuries repaired by fibrous connective tissue
- Loss of cardiac function at site of injury
Differences from skeletal muscle:
- Individual cells are much smaller
- Single nucleus
- Cells are joined together in branching linear array, so looks like long fibres that branch and converge
- Cells contain large, densely packed mitochondria (reliable energy supply)
- Have Intercalated disks (dense bands that cross muscle, typically in irregular lines, occur at junction between individual fibres, only found in heart muscle)
What are the characteristics of cardiac membrane systems?
Same basic components as skeletal muscle: T-tubules and SR. But some structural differences:
- Larger T tubules
- SR less well organised. Forms diad with T-tubules (rather than triad as in skeletal muscle)
- Also difference in mechanism of Ca2+ release from SR
What are intercalated discs?
Dense bands that cross muscle, typically in irregular lines. Occur at junction between individual fibres. Only found in heart muscle.
Specialised structures that join neighbouring cells end-to-end. Contain 2 types of membrane junction:
- Desmosomes (strengthen tissue mechanically hold cells together)
- Gap junctions (communicating junctions that connect cytoplasm of adjacent cells)
Essential for cardiac muscle function
- Connect cells mechanically, electrically, and chemically
- Allow for branching by joining together >2 cells
- -> strong and efficient force-conducting network
- -> synchronised contraction due to electrical synapses
What are Gap junctions?
Hydrophilic channels
- Electrical synapse –> electrical signal transmitted and synchronised throughout tissue
- Direct exchange of ions and small molecules (e.g. calcium) –> common response to local regulators
How does cardiac muscle contract?
Similarities with skeletal muscle:
- Begins with action potential across cell membrane
- This causes rise in sarcoplasmic Ca2+ level by release from SR
- This activates sliding filament mechanism
Key differences:
- Heart muscle contracts spontaneously - a myogenic rhythm. It is modulated by neural input, but it is NOT directly driven by it (skeletal muscle doesn’t contract unless activated by its moterneuron)
- Action potential involves calcium inflow as well as sodium
- Ca2+ release from SR through RyR is triggered by this inflow of Ca2+, rather than by direct coupling to DHPR.
How is pacemaker activity generated?
Two specialised types of cardiac muscle cells:
Contractile cells (~99%):
- These do the work
- They are NOT pacemakers
Pacemaker cells:
- Membrane potential shows regular spikes
- These generate the rhythm
Complex ionic mechanisms:
- Rising phase is driven by inflow of Ca2+ (skeletal is driven by Na+)
- Mediated through voltage-dependant Ca2+ channels which inactivate at +mV. These are similar to, but slower than, the standard Na+ channels of the nerve action potential.
- Falling phase is driven by outflow of K+ channels, very similar to those in neurons
- But the key to pacemaker activity is the slow depolarization before the spikes.
What is the “funny” current?
The depolarization is driven by the funny current (If). It is mediated by slow inflow of Na+.
Funny? –> the channel is activated by hyperpolarization and inactivated by depolarization - most unusual
Turned on by the extreme hyperpolarization after the spike. Turned off at about the level the Ca2+ channels open.
Its slope is key to controlling the timing of the heartbeat.
It belongs to HCN (hyperpolarization-activated cyclic-nucleotide gated channel) channel family
How is contraction spread in the Cardiac muscles?
Pacemaker cells lie in four sites: Atrioventricular node, Sinoatrial node, Bundle of His and Purkinje fibres
Generates APs at different rates:
- Fastest: SA (sino-atrial) node –> actual pacemaker, others supplementary
- AP spreads through gap junctions in atrium
- Delay in AV node so atria contract before ventricles
- AP spreads through ventricle
Organised into nodes and bundles to transmit contractile impulse to different parts of heart in precise sequence.
How do action potentials work in contractile cardiomyocytes?
Contractile (non-pacemaker) muscle fibres get driven by pacemaker fibres through gap junctions (electrical synapses) in intercalated disks.
They form an electrically-coupled network, and pass on the excitation to other contractile fibres
Key features: Contractile fibre APs have a very long duration (250ms) due to long-lasting Ca2+ current (skeletal muscle 1-2ms)
Prolonged refractory period
Mechanical muscle response occurs while membrane is still depolarised.
No tetanus - protective mechanism
How is calcium released in cardiac muscle?
- DHPR is a voltage-dependant Ca2+ channel (L-type for long-lasting)
- DHPR is not directly linked to RyR2
- Instead RyR2 is itself Ca2+ activated
- A small amount of Ca2+ enters through DHPR during AP
- Triggers a large amount of Ca2+ release from SR through RyR2
What are regulatory mechanisms of cardiac muscle?
Force and rate of heart muscle contraction have to be adjusted to circulatory needs e.g. heart beats faster and more strongly during exercise
Change in force: Inotropic effect
Change in rate: Chronotropic effect
Controlled by nervous and endocrine systems
Parasympathetic (“rest and digest”): Acetylcholine - slower, weaker
Sympathetic (“flight or fight”): Noradrenaline - faster, stronger
Neural control exerted from two paired control centres in medulla oblogata of brain stem
Cardioinhibitory centre drives parasympathetic activity through vagus nerve (cranial nerve X [tenth])
Normal resting heart rate ~65 bpm, Isolated heart beats at ~100 bpm - So at rest continual down-regulation via vagus activity
How is heart rate regulated?
Effect of sympathetic stimulation
Works through 2nd messenger cascade:
Increase in AC –> Increase in cAMP –> Increase in protein kinase –> phosphorylation
Enhances funny current, more Na+ enters (HCN = cyclic-nucleotide gated)
Speeds pacemaker depolarisation - Positive chronotropy
Enhances calcium current, more Ca2+ entres - strong contraction, positive inotropy
Beta-blockers counteract effects (prescription-only drugs: e.g. atenolol. Used to reduce blood pressure, anti-anxiety)
Acetylcholine activates m2AChR - works through 2nd messenger cascade
Inhibits AC abd reduces cAMP - counteracts sympathetic increase in cAMP
Down-regulates voltage-dependant Ca2+ channel (less triggering Ca2+ comes in through membrane, less Ca2+ induced Ca2+ release from SR through RyR, weaker contraction, Negative inotropy)
Up-regulates voltage-dependant K+ channel (more K+ leaves cell after spike, more negative pacemaker after-spike hyperpolarization, Negative chronotropy)
What properties does cardiac muscle need?
1) Has to contract forcefully
- Sarcomeres and myofibrils, plus desmosomes for strength
2) Has to contract rhythmically
- Pacemaker activity of some cardiac muscle cells
3) Has to contract in a highly coordinated fashion
- Contraction spreads through heart in precise sequence along nodes and bundles to facilitate blood expulsion, synchronised through gap junctions
4) Rate and force have to be modified according to circulatory needs
- Pacemaker activity and tension can be changed by parasympathetic and sympathetic regulatory mechanisms
5) Must NEVER tire or tetanise
- Reliable energy supply and long AP duration and refractory period
What are smooth muscle structures?
smooth muscle shares some properties with skeletal and cardiac muscle, but also has some unique features.
Tissue features:
- Single cells do not extend full muscle length
- Groups of cells are arranged in sheets
- Sheets can be circular or longitudinal and surround other tissues
- No tendons. Cells held together at dense bands so do not tear apart
Cellular features:
- Spindle-shaped
- Single Nucleus
- Doesn’t have T tubules - excitation-contraction coupling different from skeletal muscle
- No myofibrils and no sarcomeres, but thick and thin filaments scattered throughout cytoplasm (No Z lines, no striations, same contractile elements as skeletal muscle, but different arrangement)
How does smooth muscle sit while relaxed and then how does it contract?
RELAXED Contractile units (bundles of actin and myosin) arranged diagonally in diamond-shaped lattice
No Z lines, but dense bodies anchor actin filaments (act like Z line, chemically similar).
Actin filaments from either end overlap
Myosin heads along entire length.
CONTRACTED
Myosin heads pull in opposite directions on opposite sides.
Absence of Z lines allows more shortening than in straited muscle. Causes cell to bulge out.
How is contraction of smooth muscle regulated?
Contraction is triggered by increased cytosolic calcium (like in skeletal and cardiac muscle)
- Ca2+ mainly comes from extracellular fluid across plasma membrane
- Some from sarcoplasmic reticulum
- Balance between extracellular fluid and SR depends on muscle type
- However, no troponin in thin filaments
MLCs:
Lightweight protein chains attached to myosin heads (MLCs) - Crucial regulatory function in smooth muscle.
Myosin can only interact with actin when regulatory light chain is phosphorylated.
- -> Phosphorylation of MLCs allows contraction
- -> Dephosphorylation of MLCs prevents contraction
- -> Ca2+ regulates this phosphorylation
What are the steps of contraction in smooth muscle?
CONTRACTION
1. Intracellular Ca2+ concentration increase when Ca2+ enters cell and is released from sarcoplasmic reticulum
- Ca2+ binds to calmodulin (CaM)
- Ca2+-calmodulin activates myosin light chain kinase (MLCK)
- MLCK phosphorylates light chains in myosin heads and increases myosin ATPase activity
CHEMICAL CHANGE IN THICK FILAMENTS
5. Active myosin crossbridges slide along actin and create muscle tension
What are the differences in Skeletal and Smooth muscle?
Smooth: Muscle excitation
Skeletal: Muscle excitation
Smooth: Rise in cytosolic Ca2+ (mostly from extracellular fluid)
Skeletal: Rise in cytosolic Ca2+ (only from SR)
Smooth: Series of biochemical events
Skeletal: Physical repositioning of troponin and tropomyosin
Smooth: Phosphorylation of myosin in thick filament
Skeletal: Uncovering of actins myosin-binding sites in thin filament
Smooth: Binding of actin and myosin at cross bridges
Skeletal: Binding of actin and myosin at cross bridges
Smooth: Muscle contraction
Skeletal: Muscle contraction
What are the steps of relaxation in smooth muscle?
RELAXATION
1. Free Ca2+ in cytosol decreases when Ca2+ is pumped out of the cell or back into the sarcoplasmic reticulum
- Ca2+ unbinds from calmodulin (CaM)
MLCK NO LONGER ACTIVATED BY CALMODULIN
- Myosin phosphatase removes phosphate from myosin, which decreases myosin ATPase activity
- Less myosin ATPase results in decreased muscle tension
How is ATP used in smooth muscle?
ATP is used for cross-bridge cycling, just like straited muscle.
ATP is also used to phosphorylate MLC
Why is smooth muscle not energetically expensive?
Smooth muscle responds more slowly than skeletal muscle (skeletal = 100ms, smooth muscle = <5000ms)
Slower ATP splitting by myosin ATPase during cross-bridge cycle
–> Cross bridge activity and filament sliding ~10x slower
–> Uses less ATP per second
Slower Ca2+ removal - slower relaxation
What is the latch phenomenon?
De-phosphorylating MLC reduces affinity of myosin head for ATP
[remember ATP binding to myosin head necessary for detaching from actin]
So myosin head takes longer to detach after MLC dephosphorylation - force is maintained for a while after activation terminated
Helps smooth muscle produce sustained contractions - efficient tissue well adapted for its purpose
What is smooth muscles response to stretch?
Two Important features:
- Smooth muscle can develop near-maximum tension over greater range of lengths than skeletal muscle
- much higher actin:myosin ratio than skeletal muscle, If one actin filament doesn’t overlap with myosin, another will
- No sarcomeres, thin filament do not collide with Z line at full contraction (can slide past dense bodies) - Stress relaxation response
= sudden stretch of smooth muscle -> initial increase in tension -> Quick return to tension level prior to stretch
Reverse stress relaxation response
= decrease in tension -> tension will quickly return to initial value
More plastic than skeletal muscle, mechanisms not fully understood
-
Where do we find smooth muscle?
Smooth muscle occurs widely throughout the body. In the walls of hollow internal organs:
- Gastrointestinal tract
- Bladder
- Uterus
…and in tubes:
- Blood vessels
- Airways of respiratory system
- Tubules of urinary system
- Reproductive ducts
What is the function of smooth muscle?
Exert pressure and move content forward.
How does smooth muscle work in the eye?
Pupil dilation/constriction is controlled by radial and circular smooth muscle.
Lens shape (focus) is controlled by circular smooth muscle.
How does smooth muscle work in the skin?
Contraction of smooth muscle bundles that are attached to hair follicles (arrector pili muscles) makes hair stand up for thermoregulation.
How can smooth muscle serve different functions in different locations?
The arrangement of smooth muscle is important.
When arranged in a circle, smooth muscle can control tube diameter and regulate blood (or air) flow e.g. in capillaries.
Different types also with varying physiological properties. Each type adapted to particular function:
- Sphincter muscles controlling emptying of bladder and rectum must maintain long steady contraction and then suddenly relax
- In stomach and intestine, the muscles are constantly active to move and mix food
- In the uterus, muscle properties vary over time (quiescent during pregnancy and contract forcefully during labour)
What are classifications of different types of smooth muscle?
1) Differences in contraction
- Phasic smooth muscle (occurring in phases, not continuously)
- Tonic smooth muscle (continuous)
2) Differences in source of excitation
- Multi-unit smooth muscle (neurogenic, only contracts in response to nerve input)
- Single-unit smooth muscle (myogenic, muscle contracts spontaneously, nerves modulate)
Most smooth muscle is single-unit
What is phasic smooth muscle?
Contracts in bursts (pronounced increases in contractile activity)
Trigger: muscle APs that increase cytosolic Ca2+
Most abundant in walls of hollow organs that push content through them e.g. digestive organs, where phasic contractions mix food with digestive juices and move it forward
What is tonic smooth muscle?
Partially contracted at all times (tone).
Low resting potential (-55 to -40 mV)
- Some voltage-gated Ca2+ channels open
- Always some Ca2+ entry
Tone does not depend on APs
No bursts of contractile activity, but activity can be modulated above or below tonic level e.g. found in arteriole walls where ongoing tonic contraction squeezes blood and maintains blood pressure.
What are the differences between multi-unit and single-unit smooth muscles?
Multi-unit: Neurogenic, only contracts in responce to nerve input (cf skeletal)
Single-unit: myogenic, muscle contracts spontaneously, nerves modulate (cf cardiac)
Categorisation depends on:
- How muscle cells are organised
- How action potentials are initiated
What is multi-unit smooth muscle?
Cells not coupled with gap junctions. Each cell activated by autonomic nerve input.
- Neurogenic (involuntary)
- Muscle paralysed if innervation is cut
Single branching neuron can activate many muscle cells.
e.g. found in large blood vessels, eye (iris, ciliary body), hair follicles
What are single-unit smooth muscles?
Muscle cells contract as a single unit
- electrically coupled by gap junctions
Self-excitable (requires no nervous stimulation)
- Myogenic
- Muscle not paralysed if cut innervation
Tension modulated by autonomic nervous system - involuntary
Often found in walls of hollow organs (e.g. gu, uterus, urinary tract)
What is phasic single-unit smooth muscle?
Clusters of specialised non-contractile cells within syncytium display spontaneous rhythmic electrical activity
Two major types of spontaneous depolarisation:
- Pacemaker potential
- Slow-wave potential
Rhythm mechanism probably like cardiac funny current
Spreads throughout whole muscle via gap junctions
What influences slow-wave potential?
Influenced by:
- Autonomic nerve activity
- Hormones
- Local metabolites
- Mechanical stretch
How is contraction (Ca2+) regulated in phasic smooth muscle?
Most Ca2+ comes from extracellular fluid (ECF)
Enters through voltage-dependant dihydropyridine receptors (L-type Ca2+ channels)
Smooth muscle cells are much smaller in diameter than skeletal muscle cells -> Ca2+ entering from ECF can reach cell centre even without elaborate T tubule SR mechanism
How is contraction (Ca2+) regulated in tonic smooth muscle?
Tension modulated by extrinsic ligands increasing or decreasing release of Ca2+ from SR.
Ligand (e.g. ACh) binds to G-protein coupled receptor in membrane
Activates phospholipase C, causing increase in inositol triphosphate (IP3)
Activates IP3-receptor Ca2+ channel in SR, releasing Ca2+ into cytosol –> Contraction through standard Ca2+/CaM MLCK pathway
What is gradation of muscle force?
In skeletal muscle, contraction is graded entirely by neural control mechanisms.
More plasticity in smooth muscle:
- Autonomic nerve activity
- Hormones
- Local metabolites
- Mechanical stretch
More external influences on smooth muscle
How do neural inputs work in smooth muscle?
No standard motorneurons, no neuromuscular junctions
- Transmitter is released from varicosities along nerve branches
- Receptors are diffuse along muscle cell membrane
- Multiple muscle cells can be influenced by release from single neuron
What is the enteric nervous system?
The gut has its own brain
Has sensory, motor and interneurons. Has more neurons in it than are in spinal cord. But influenced by CNS via autonomic nervous system.
Can also feedback influence to the CNS.
Factoid: 90% of serotonin in body is in ENS
Whats an example of hormonal effects on a single-unit smooth muscle?
Uterus
–> Contraction affected by circulating and locally released chemical messengers that vary during menstrual cycle and pregnancy
During pregnancy: High levels of circulating progesterone decrease expression of proteins involved in gap junction formation –> reduced excitability and muscle activity
During labour: High oestriol levels increase gap junction formation –> increases excitability and muscle activity
Why do we study neuroscience?
Understand and treat medical conditions
Learn about natural phenomenon so we can respond appropriately to them
Develop appropriate protocols for managing human-wildlife conflicts
Understand and study physiology and behaviour
The nervous system (NS) is split into which two main divisions?
- Central nervous system (CNS) - brain and spinal cord
- Peripheral nervous system (PNS) - rest of the body
What are the two key types of cells in the nervous system?
- Neurons
- Glia cells
What are phospholipid bilayers?
Animal cells (including neurons) have phospholipid bilayers surrounding their contents
This forms a barrier to water soluble ions as it has a hydrophobic region
0 Hydrophilic region (phosphate)
{} Hydrophobic region
{} (made of hydrocarbon chains)
0 Hydrophilic region (phosphate)
How do water soluble ions pass through phospholipid bilayers?
Water soluable ions can pass the hydrophobic region via channel proteins (aka ion channels).
What are the 4 important ions in neuron cytosol and the surrounding extracellular fluid?
Potassium = K+ Sodium = Na+ Calcium = Ca2+ Chloride = Cl-
They all carry an associated charge
The concentrations of these ions either side of the phospholipid bilayer and their ability to move across it are what generate the membrane potential and the equilibrium potential of a neuron.
What is membrane potential?
The voltage (electrical potential), or the difference in charge between an anode (+) and a cathode (-), across a membrane.
What are microelectrodes used for?
Intracellular recording using a microelectrode which penetrates inside the cell and measures the voltage relative to the outside - difficult to do.
What range (in volts) could be a resting membrane potential for a nerve cell?
-40 to -80 mV
Usually -65 mV
Why is the inside of a nerve cell (and most other cells) negative relative to the outside?
One key is the ionic concentration gradients.
–> Particularly the K+ gradient
The inside world - needs metabolic energy to make it different from outside. The outside world - as produced by nature.
Another key is that the resting membrane is relatively permeable to K+ ions, and impermeable to other ions (like Na+)
So the two key factors:
- There is a higher concentration of K+ inside than outside the cell
- K+ can move across the membrane, but not much else can
What is the membrane selectively permeable to and how does the cell system reach equilibrium?
K+ which leads to leakage channels. Relatively impermeable to everything else.
K+ diffuses out of the cell, down concentration gradient. Takes positive charge with it, leaves inside negative.
Electrical gradient tends to pull K+ back into the cell. The more K+ leaves, the bigger the electrical gradient.
Not many ions have to move to set up the electrical gradient - the chemical gradient is effectively unchanged.
Eventually (v. quickly actually) electrical gradient = chemical gradient and the system is in equilibrium.
What equation is used to calculate the equilibrium potential?
Nernst Equation
Emv = 58 / z log10 xoutside / xinside at 20°C
The resting membrane is also permeable to what ion other than K+?
The resting membrane is mainly permeable to K+, but is also a bit permeable to Na+.
The actual membrane potential is a “compromise” between the K+ and Na+ equilibrium potentials, but weighted towards K+ because it has higher permeability.
Are the ions in equilibrium at resting membrane potential?
The ions are NOT in equilibrium, they are in a steady state.
How is the resting potential calculated?
The resting potential can be calculated with the Goldman equation
- the membrane potential is mostly set by K+ and Na+
Em = 61 log (Pk[K+] + P Na [Na+] outside / Pk[K+] + P Na [Na+] inside)
Pk and P Na is the permeability of the membrane to each ion. The terms in the [ ] are the concentrations inside and outside the membrane
What are the gradients like at the resting membrane potential: steady state not equilibrium?
Steady leakage of K+ out of cell Na into cell.
Would cause gradients to run down. Gradients replenished by Na/K ATPase pump.
Gradients run down in > 10 mins. Membrane potential gradually lost without these pumps.
Why do we have nerves?
The job of the nervous system is rapid sensing, decision and action.
Signals are carried by nerve cells, rapidly, over quite long distances.
How is information passed from one end of a nerve cell to the other?
A wave of change of charge in the membrane potential
What are voltage-dependant ion channels?
The resting membrane has a steady, relatively high-K low-Na permeability (leakage channels) which does not change. Overall leakage permeability is low.
The membrane ALSO has voltage-dependant Na+ and K+ channels.
These are SHUT at the resting potential, but OPEN if a stimulation makes the inside of the cell more +ve.
How do Na+ channels react when stimulated?
The Na+ channels open quickly, but then automatically shut shortly after, even if the inside stays +ve (inactivation).
How do K+ channels react when stimulated?
The K+ channels open more slowly and do not automatically shut, but shut if the inside goes back -ve.
How are action potentials generated in positive (Na) and negative feedbacks (K)?
Positive feedback (fast):
1. Depolarisation (makes the inside of the cell more positive)
2. Increase gNa past threshold and action potential occurs
3. Na inflow
After a certain period of time Na+ channel is shut and inactive because it automatically shuts
Negative feedback (slow)
1. Depolarisation (makes the inside of the cell more positive)
2. Increase gK
3. K outflow
4. Hyperpolarisation (makes the inside of the cell more negative)
Because the possasium conductance is a lot higher than normal because we have both the potassium channel and the leaky channels that let potassium through the membrane. This means we overshoot the resting potential and get after-hyperpolarisation (AHP).
g = conductance
What is an action potential threshold?
A weak stimulus does not cause a big enough increase in gNa to overcome the resting K+ conductance if the stimulus is lower than threshold.
How do biotoxins affect ion channels?
Ion channels are crucial to neuron function and blocking them disrupts neuron ability to generate action potentials. Biotoxins often target specific voltage gated ion channels.
e.g. Pufferfish - Tetrodotoxin + Na+, Scorpions - Numerous types + Na+, K+ and Cl-
What is the Absolute refractory period?
Early in the AHP (after-hyperpolarisation) after a spike (1-2 ms) it is impossible to get another spike, no matter how big the stimulus.
Due to voltage-dependant sodium channels still being shut from previous spike (inactivated).
What is the relative refractory period?
Later in the AHP (~ 3-10 ms) after a spike it is difficult to get another spike, needs a bigger than normal stimulus.
Due to some voltage-dependant potassium channels still being open from previous spike.
What are the consequences of refractory period?
- Limits maximum frequency of action potentials (once a neuron spikes, it can’t spike again for a while, maximum frequency is usually ~300 Hz)
- Keeps propagation going in only one direction
How do action potentials move along the axon?
Positive charge spreads out from the site of the spike
Stimulates the next region of axon, which generates a spike in turn.
Refractory period prevents spike going backwards (like a burning fuse).
What are myelinated axons?
Myelinated sheaths help axons transfer information rapidly. Some vertebrate axons are myelinated.
Non-myelinated: action potential regenerated at every point. Conduction velocity is 0.1-10ms-1
Myelinated: action potential jumps from node to node. Conduction velocity is ~120ms-1
Both depend on diameter of axon - fat axons are faster.
What are advantages and disadvantages of frequency code for stimulus strength?
The stronger the stimulus, the higher the frequency of the spiking rates although limited by the refractory period.
Advantages:
- Unambiguous (all or none)
- Can transmit long distances without loss
- Fast (but finite) conduction velocity
Disadvantages:
- Needs time to integrate (weak signals, low frequency)
- Low frequency ceiling (~300 Hz)
- So needs range fractionation to cover wide range of signal strengths (different neurons cover different ranges of signal strength)
How is frequency quantified?
Average: Measured in impulses per second (Hz). Needs horizontal scale bar to get time.
Average frequency: count number of events in set time and scale to 1 sec.
Instantaneous: Measure time interval between each spike in sec. Take reciprocal to get sec (=Hz) (i.e. 1/x)
What are the two main electrical recording methods?
- Intracellular (Place a microelectrode inside a neuron)
- Extracellular (Place an electrode outside but near the neuron)
What is intracellular electrical recording of neurons?
- Place a microelectrode inside a neuron
- Measure the voltage relative to the outside
- i.e. the trans-membrane voltage
Can record:
- resting potential
- synaptic potentials
- spikes
- all spikes same size and shape
However all only from this neuron. Doesn’t give a good representation of entire system. Difficult to do as hard to place electrode inside neuron.
What is extracellular electrical recording of neurons?
- Place an electrode outside but near to neurons
- Measure the field potential in extracellular space relative to a distant site
- Field generated by flow of current across membranes of nearby neurons as spike goes past
Cannot record:
- resting potential
- synaptic potentials (too small)
Can record:
- spikes from any active nearby neuron
- spike size depends on size of axon, distance to electrode
Relatively easy to do.
Extracellular spike waveforms can add together if 2 neurons spike at the same time. Can produce a distorted spike. No physiological significance. This can’t happen with intracellular recording.
How is information transmitted through neurons?
Transmission over long distances happens within neurons by action potentials travelling down the axons of neurons.
What is a synapse?
At any point where a neuron must connect to another cell, information is passed between cells via synapses.
What does Presynaptic mean?
This is the transmitting neurone that sends information to the other neurone.
What does Postsynaptic mean?
This is the receiving neurone that gets information from the other neurone.
What are the two types of synapse?
- Electrical synapse
- Chemical synapse
What are electrical synapses?
Allow direct transfer of ions from one cell to another. Gaps between cells are bridged at sites called gap junctions.
Distance between cells is extremely small and bridged by special proteins (connexins) that combine to form connexons.
Two connexons (one from each cell) make up a gap junction channel. These channels allow movement of ions between cells.
What are connexons?
Distance between cells is extremely small and bridged by special proteins (connexins) that combine to form connexons.
Two connexons (one from each cell) make up a gap junction channel. These channels allow movement of ions between cells.
Connexons are like tunnels linking inside of one cell to inside of the other.
What is transmission like for electrical synapses?
Any change in pre-synaptic membrane potenial, +ve or -ve, can get transmitted to post-synaptic cell as post-synaptic potential (PSP).
Can be bidirectional.
If there are lots of gap junctions there is little loss of signal across synapse. Spikes may transmit 1:1
If only a few gap junctions - strong attenuation (post-synaptic potential «_space;pre-synaptic potential)
FAST, little delay
When does electrical transmission occur?
Where high speed is required (e.g. escape circuits)
When groups of cells need to produce more-or-less synchronous activity
- motorneurons innervating the same muscle
- not exactly synchronised, but “share” membrane potential changes between group.
How do chemical synapses transmit information?
Presynaptic cell activity –> Neurotransmitter release into synaptic celft –> Postsynaptic cell activity
- Presynaptic terminals form swellings known as boutons
- Synaptic cleft widens to approx 20nm (but greater at nerve-muscle junctions)
- High density of mitochondria (to support high metabolic activity) in presynaptic terminal
- Vesicles present in presynaptic terminal (these contain molecules of neurotransmitter)
- Protein accumulations called membrane differentiations in pre (action zones) and post (postsynaptic density) synaptic membranes
What are the different ways to classify synapses in the CNS?
- By the type of postsynaptic membrane and axon connects to
- By the thickness of the membrane differentiations in the pre and post synaptic membranes
How do chemical synapses use neurotransmitters?
Chemical synapses use the movement of chemicals (neurotransmitters) to transfer information.
What cuases neurotransmitter release from a vesicle in the pre-synaptic cell?
- Spike comes down axon
- Invades pre-synaptic terminal and depolarises it
- depolarisation opens voltage-gated Ca2+ channels present in terminal
- Ca2+ flows in
- this triggers exocytosis of vesicles releasing neurotransmitter from vesicles into the cleft
Complex set of proteins occur on vesicle and pre-synaptic plasma membrane. Ca2+ is essentially link to lock them together and open fusion pore linking interior of vesicle to outside of cell.
In most synapses, the calcium inflow has little effect on the membrane potential (although Ca2+ channels are voltage dependant, Ca2+ does NOT generate a spike, too little calcium comes in and its very local)
The job of Ca2+ is to act as an intracellular message converting the electrical depolarisation caused by the spike into a chemical signal that triggers exocytosis.
What are non-spiking synapses in chemical synapses?
We know that pre-synaptic depolarisation opens voltage gated Ca2+ channels and Ca2+ inflow causes neurotransmitter release.
However, some cells do not necessarily need a pre-synaptic spike. At some synapses, sub-threshold depolarisation can open Ca2+ channels and release neurotransmitters.
At some synapse, neurotransmitter is released at normal resting potential.
- can be increased by depolarisation
- decreased by hyperpolarisation
- like a leaky tap
How do neurotransmitters interact with synaptic cleft via vesicles?
A single vesicle contains several thousand molecules of neurotransmitter
The amplitude of the postsynaptic response to the neurotransmitter release in the synaptic cleft is therefore a multiple of the response to 1 vesicle of neurotransmitter
How many vesicles?
- often 1 vesicle if success
- sometimes 2 or 3
Various ways of increasing or decreasing number of vesicles
- learning, forgetting, facilitation (increase with repetition), fatigue (running out of vesicles)
At neuromuscular junction (NMJ) normally always successful release (~50 vesicles)
What are the different types of neurotransmitter?
- Many and varied
- Broadly 3 types: amino acids, amines and peptides
Most important thing is the effect the transmitter has and that depends on the post-synaptic cell’s response which depends on the receptors that bind the transmitter
Same neurotransmitter can have different effects on different cells, or even different regions of same cell, depending on receptor population
What are receptors for neurotransmitters?
The membrane of the postsynaptic neurone is specialized where it faces a presynaptic terminal, contains receptors.
Receptors can specifically bind neurotransmitter molecules (and other molecules with a similar structure)
Causes ion channels to open in the membrane of the postsynaptic cell
Two main types of receptor:
- Transmitter gated ion channels (ionotropic)
- G-protein coupled receptors (metabotropic)
How are neurotransmitters removed?
Neurotransmitter effect is usually short-lived
Binding is reversible (if cleft concentration drops, neurotransmitter unbinds)
Neurotransmitter in cleft is either destroyed by enzymes or removed by uptake into pre-synaptic cell, post-synaptic cell, glia
Many drugs work by prolonging or reducing lifetime of a neurotransmitter (e.g. Prozac reduces uptake of serotonin)
PSPs (postsynaptic potential) can be what two things?
Excitatory = EPSPs
or
Inhibitory - IPSPs
What are EPSPs?
Excitatory postsynaptic potential
Ion channels opened during EPSP
- varied, depends on function e.g. ACh opens mixed Na+/K+ channel (non-specific cation)
- number of channels opening depends on amount of transmitter released
- If a membrane is permeable to Na+ then the net effect is typically depolarisation
in a post-synaptic neuron
- if not much transmitter arrives
- few post-synaptic channels open
- sub-threshold response
What are PSPs change with distance?
PSPs decrement with distance. If no post-synaptic spike, response dies away with distance.
However, spikes do NOT decrement with distance. If no post-synaptic spike occurs, it doesn’t die away with distance.
How do dendrites impact EPSPs?
Dendrite cable properties impact on EPSPs and can influence their ability to reach the axon hillock to generate an action potential.
What are the differences between synaptic potentials and action potentials?
- PSP does not actively spread along the axon (i.e. it is localized and sub-threshold)
PSP not caused by voltage-gated channels - instead channels are opened directly or indirectly by neurotransmitter
For ACh-receptor channel, Na+ and K+ ions flow through the same channel - increase in gNa and gk are simultaneous, not sequential
PSPs have no refractory period and so can summate
What are IPSPs?
in a post-synaptic neuron
- the more transmitter arrives
- more post-synaptic channels open
- the more the postsynaptic membrane is hyperpolarised
- makes it harder for an EPSP to depolarise the postsynaptic membrane beyond the threshold to make spikes
This type of inhibition is called a shunt
The key function of synapses is integration and decision making.
One post-synaptic neuron may have 1000s of separate pre-synaptic inputs.
The balance of excitation and inhibition is continuously shifting.
If excitation wins, then the post-synaptic neuron spikes and the message is passed on. If inhibition wins, then the message dies.
