Cell Ultrastructure Flashcards
What can be caused by abnormal signalling/membrane transport?
include:
53. Oedema
54. Palpitations/abnormal heart rhythm
55. Raised blood pressure
65. Diarrhoea
91. Abnormal serum sodium
92. Raised serum calcium
What is a cell?
Fundamental functional unit of a tissue
What makes up a cell?
- Lysosomes:
•Cell’s dustbin - Mitochondria:
•TCA cycle
•Oxidative phosphorylation
➢Maternal inheritance only - Smooth Endoplasmic Reticulum:
•No ribosomes
•Site of lipid synthesis
•Some drug metabolism - Microtubules:
•Give structure to cell - Rough Endoplasmic Reticulum:
•Studded with ribosomes
•Site of protein synthesis - Golgi body:
•Mediates protein sorting to specific sites - Ribosomes:
•Translate mRNA into protein - Plasma membrane
•Keeps stuff in/out
•Selectively permeable - Nucleus
Genome:
•Instructions…
•Inherited disease
•Cancer
Cytosol?
??
Anti-bodies hormones?
Receptor ion channels?
Membrane vesicles:
●Intra-cellular tran
- endo/exocytosis
Why are membranes and their proteins needed?
- Cell signalling
➢Source of lipid precursors - Cell polarisation
➢Epithelia – this lecture - Compartmentalisation
➢Ionic gradients – next lecture
●Diffusion (Nernst potential)
●Membrane potential - Tightly regulated
- Disease disrupts this
➢Heart disease
➢Kidney failure
➢Cancer
➢Inflammation
Glycerophospholipid
Fatty acid tails:
•Non-polar
•Hydrophobic
•Saturated C-C bonds
•Unsaturated cis C=C bonds
➢Kinks hydrophobic tail
•Significant variation in length
Glycerol backbone (3C)
Phosphate group
-ve charge at physiological ph
One of either…
➢Serine (PS - phosphatidyl-serine)
➢Choline (PC - phosphatidyl-choline)
➢Inositol (PI - phosphatidyl-inositol)
➢Ethanolamine - PE (phosphatidyl-ethanolamine)
➢Sphingomyelin - SM
Glycerophospholipids
•Hydrophobic and hydrophilic
•Amphipathic molecules
Phospholipid head
- polar charged
- hydrophilic
Glycerophospholipid
Fatty acid tails:
•Non-polar
•Hydrophobic
•Saturated C-C bonds
•Unsaturated cis C=C bonds
➢Kinks hydrophobic tail
•Significant variation in length
Glycerol backbone (3C)
Phosphate group
-ve charge at physiological ph
One of either…
➢Serine (PS - phosphatidyl-serine)
➢Choline (PC - phosphatidyl-choline)
➢Inositol (PI - phosphatidyl-inositol)
➢Ethanolamine - PE (phosphatidyl-ethanolamine)
➢Sphingomyelin - SM
Glycerophospholipids
•Hydrophobic and hydrophilic
•Amphipathic molecules
Phospholipid head
- polar charged
- hydrophilic
What is the structure of the lipid bilayer?
Leaflet specific orientation
•Leaflet “flip”
➢Thermodynamically unfavourable
Outer:
•Sphingomyelin
•Enriched for PC
Inner:
•Enriched for PE
•PS and PI majority in inner leaflet
•PS – presence in outer leaflet is “eat me” signal for apoptosis
What is the fluidity of the bilayer?
Membrane fluidity is important…
➢Movement of phospholipids and proteins
➢Facilitates inter- and intra-cellular signalling (phosphatidylinositol)
➢Temperature- and structure-dependent phase transitions.
Liquid-ordered (raft)
•Highly ordered
•Relatively fluid
Solid gel
➢Highly ordered
➢Limited lateral mobility
Liquid-disordered
➢Low density packing
➢High lateral mobility
What are the integral membrane proteins?
Many sorts
➢Channels, pores, structural
• Defined secondary structure
➢a-helix in transmembrane region
➢Hydrophobic amino acids interface with hydrophobic phospholipid tail
➢Membrane helps maintain 3D structure
K2P§ K+ channel
What are anti-bodies to phospholipids?
Anti-phospholipid syndrome
•Systemic auto-immune disease.
•Anti-bodies bind to b2-Glycoprotein-1 on cell membranes
➢Initiate inappropriate blood clotting (thrombosis)
•Significant cause of pregnancy morbidity and early miscarriage
What is epithelia like?
Epithelia…
•Require polarisation of plasma membrane – apical vs basolateral surfaces
•Permits cell-specific function – secretion/absorption
•Strongly adhere to neighbours – tight junctions
What is the structure and layers of epithelia?
What are epithelia like?
Epithelia…
•Require polarisation of plasma membrane – apical vs basolateral surfaces
•Permits cell-specific function – secretion/absorption
•Strongly adhere to neighbours – tight junctions
•Three examples:
➢parietal cell (gastric pits)
➢intestinal epithelium
➢nephron
What occurs in parietal acid secretion?
On base lateral membrane
Osmosis
ECF
Duct lumen
Parietal cell
Omeprazol
How is substrate movement affected by water?
Substrate movement across membranes occurs with movement of water…
Sodium:glucose co-transport (symport) - basis of oral rehydration therapy
What is the nephron like?
The Nephron…
•Morphology and permeability of tubular epithelial cells changes along the tubule
•Reflects specific function of each aspect of tubule:
How is membrane function affected by environmental changes?
pH
➢Both extremes damage protein
➢Inhibits cell function
Critical role for acid:base homeostasis
Plasma Ca2+
Cell membrane
Excitability/permeability
How is membrane function affected by environmental changes?
pH
➢Both extremes damage protein
➢Inhibits cell function
Critical role for acid:base homeostasis
Plasma Ca2+
Cell membrane
Excitability/permeability
How are calcium concentrations tightly regulated?
Total serum calcium measured clinically (ionised [Ca2+] + unionised [Ca])
Serum Calcium:
•45% Free ionised Ca2+
➢Biologically active
➢Change Ca2+ (active): Ca (inactive) ratio with no change in total calcium
Acidosis
●fewer Ca2+ ions bound to plasma proteins
●H+ ions buffered by albumin instead
Alkalosis
●more Ca2+ bound to plasma proteins
●fewer H+ ions on albumin
➢Alkalotic patients more susceptible to hypocalcaemic tetany
➢Due to increased neuronal membrane Na+ permeability
How does temperature affect membrane function?
Temperature
➢Too cold – proteins slow down; membrane less fluid
➢Too hot – proteins denature; increased membrane
Effects of temperature on membrane function?
Heat exhaustion core temp >37°C but ≤40°C
Heat stroke core temp ≥ 40°C
Dehydration…
Everything slows down ‘umbles’
Lowest survivable core temp 13.7°C
What are the signs of hypothermia?
Signs of hypothermia: progressively
- Shivering - increased pulse, BP and respiratory rate with shivering/pulse, BP and respiratory rate slowly reduce until undetectable
- cardiac symptoms - atrial fibrillation, ventricular fibrillation inductively by rough handling, asystole
- neurological symptoms - listlessness and confusion, reduced consciousness, unconsciousness
How is SAN AP regulated?
Hypothermia
• depolarization rate of cardiac pacemaker cells
➢Bradycardia (not vagally mediated)
•Abnormal heart rhythms
➢Fibrillation (atrial and ventricle)
What is Hypovolaemia is and the lethal triad?
- Hypothermia
Decreased plasma - Acidosis
Plasma PH - Coagulopathy
Core temperature
What are membranes like?
Phospholipid bilayer, proteins, cholesterol,
•Temperature sensitive
•Leaflet-specific use of phospholipids
•Main proteins in correct 3D orientation
➢Facilitates rapid interaction (signal transduction)
•Proteins
➢Integral/peripheral
➢Glycosylated
➢Many functions
What is epithelia like?
Epithelia
•Need polarisation of apical/basolateral surfaces
➢Gastro-intestinal epithelium
➢Differential solute permeability based on function (nephron)
•Compartmentalises cell functions
What is membrane permeability like?
Phospholipid bilayer…
•Fluidity modified by
➢C=C bonds
➢Cholesterol
➢temperature
What is the cell membrane permeable to?
Freely Permeable…
•Water (aquaporins)
•Gases
➢(CO2, N2, O2)
•Small uncharged polar molecules
➢(Ammonia, urea, ethanol)
What is the cell membrane impermeable to?
Impermeable to…
•Ions
➢(Na+, K+, Cl-, Ca2+ etc.)
•Charged Polar molecules
➢(ATP, Glucose-6-phosphate)
•Large uncharged polar molecules
➢(Glucose)
What are the three main types of carriers in membrane proteins?
- Uniport
➢Single substance
●Glucose via Glut-1
●Passive - Antiport
➢Two substances in opposite directions
●3Na+/2K+ ATPase
●Energy from ATP hydrolysis
3.Symport
➢Two (or more) substance in the same direction
●Na+/Glc nutrient transport
●Indirectly from ATP hydrolysis
●Ion gradient used
What are some examples of membrane transport?
Simple diffusion
➢Blood gases, water
➢NH3, Urea, free fatty acids
➢Ketone bodies
●Facilitated diffusion
➢Glucose (glc; hexose sugars)
➢GLUT family
●Primary Active transport
➢Ions (Na+, K+, Ca2+, H+, HCO3-)
➢Water-soluble vitamins
➢Energy direct from ATP
●Ion Channels
➢Many sorts…
➢Voltage-gated
➢“Leak” channels
●Secondary active transport
➢Ions (renal tubule; fluid balance)
➢Symporters (Na+ + X)
➢Indirect energy from ion gradient
●Pino- / phago-cytosis
➢Vesicles
What are ion channels like?
Many sorts (Na+, K+, Cl-…)
•Integral membrane protein
•Defined secondary structure
➢a-helix
➢Hydrophobic amino acids interface with hydrophobic phospholipid tail
➢Hydrophilic pore for ion passage
•Selective
➢Charge
➢Ion size
•Gated (usually)
What are the cell membrane receptors?
Signal Transduction
•Internalise extra-cellular signal…
➢First message into second message
What are cell surface and nuclear receptors?
Signal Transduction
•Pharmaceutical targets…
Nuclear steroid receptors (NSRs)
•Direct effect on gene expression
•ER – tamoxifen
•PR – mifepristone
•AR – testosterone
•GR – cortisol; dexamethasone
•MR – aldosterone; spironolactone
Neurotransmission (NTRs)
•AchR – Muscarinic cholinergic blockade – atropine
•GABA – benzodiazipines
•Seretonin (5-HT3) - ondansatron
Ion channels
•Ca 2+ - nifedipine
•Na+ - Amiloride
•K+ - Amiodarone
GPCRs - Many sorts, many effects
•Membrane-bound steroid receptors
•In-direct effect on gene expression
•E – cardiovascular effects?
•P – uterine function/sperm function
Growth factors (GFRs)
•Tyrosine kinases mechanism
•EGFR – pertuzumab (Perjeta) – breast cancer
•VEGF - bevacizumab (Avastin) – ovarian cancer
•Growth Hormone – (Genetropin)
•Insulin; IGFs
What are GPCRs like?
GPCR –
•Ubiquitous (>800 sequences)
•>50% of all drugs mimic or inhibit various GPCR
•Significant drug target
•How do they work?
GPCR – six parts:
•Receptor – gives primary specificity
•Three G-proteins – a, b, g
➢Ga further specificity
•Enzyme to modulate second messenger
➢(e.g. cAMP)
•Enzyme to terminate signal
➢Phosphodiesterase
What is Ga subunit and fluid secretion?
+ve; makes cAMP:
•b2 agonists, PGE2 via EP2 receptor (uterine relaxation)
•Many intra-cellular actions
Lungs - Activates CFTR
•Hydrates mucus
•Mucociliary staircase
Cystic fibrosis
•DF508 – inactivation of CFTR
• Cl- secretion
•Airway/intestinal mucus underhydrated
•Reduced mucociliary clearance
Intestinal epithelium
•Cholera toxin – permanent activation of Gas and AC
• Cl- secretion (CFTR) – osmotic drag
•Significant fluid
What is Ga subunit and fluid secretion?
+ve; makes cAMP:
•b2 agonists, PGE2 via EP2 receptor (uterine relaxation)
•Many intra-cellular actions
Lungs - Activates CFTR
•Hydrates mucus
•Mucociliary staircase
Cystic fibrosis
•DF508 – inactivation of CFTR
• Cl- secretion
•Airway/intestinal mucus underhydrated
•Reduced mucociliary clearance
Intestinal epithelium
•Cholera toxin – permanent activation of Gas and AC
• Cl- secretion (CFTR) – osmotic drag
•Significant fluid loss
How does Ga subunit determine second messenger?
-ve; prevents cAMP:
•a2 adrenergic agonists – Ergometrine
●(uterine contraction)
•a, m, d, k, opioid receptors
•PGE2 via EP1 and EP3 receptors
●Misoprostol (uterine contraction)
•M2 ACh receptors
+ve; makes IP3 and DAG (membrane phosphatidyl inositol):
•Oxytocin receptor, PGF2 via FP2a receptor
●(uterine contraction; severe PPH; Carboprost)
•M3 ACh receptors
What is ion transport and RMP like?
Important driving force for trans-membrane ion transport
•Three components
➢Diffusional force (hight to low)
➢Electrical force (+ve to –ve)
➢Electrochemical (combination of both)
•Each ion carries a small charge when it diffuses across a membrane
➢ion diffusion potential
➢“Electrical force required across a membrane to counterbalance chemical diffusion forces of a given ion through that membrane”
➢NO net diffusion of that ion
•Collective ion diffusion potentials contribute to membrane potential
What is the diffusion potential in trans-membrane ion gradients?
What is the diffusion potentials driving force?
Nernst equation ?
Describes ion diffusion work done (ratio) – chemical driving force
Equilibrium: diffusional (chemical) and electrical forces balance
At
Ion gradients?
Convention dictates:
•Extracellular fluid potential = 0 mV (Reference)
•Diffusion potential is that on intra-cellular membrane
Hyperpolarised - (less +ve)
Polarised (more +ve)
Hyperpolarised - more -ve/Cl- can cause depolarisation too
What is the RMP like (Em)?
Each ion carries a small charge when it diffuses across a membrane
➢ion diffusion potential
•Collective ion diffusion potentials contribute to membrane potential
•Must consider ion diffusion potential collectively…
Goldman- Hodgkin-Katz equation
Don’t need to learn
What is the MP (Em) like?
•Ion conductance (permeability) is key determinant of Em
•Stable in most cells (but sensitive to ionic imbalance)
•Transient variability in excitable tissue
➢Ventricular myocytes Em ~ -90mV
•Permeability dependant on
➢Channel numbers
➢Channel gating
•Change ion permeability…
➢Change Em
What is the RMP like?
High K+ permeability (10; leak channels, -94.5 mV)
•Low Na+ permeability (0.05;+61.8 mV)
•Low Cl- permeability (0.45; -69.27 mV)
➢Na+/K+ together -90.6 mV
•Cl-
➢negligible contribution to resting Em (Cl- ~1.8 mV)
•Na+/K+ ATPase contributes small voltage to Em (~10 mV)
•K+ primary driving force in setting resting Em
➢Muscle damage, Type-II diabetes, renal patients - [K+]E can be out of range
What is K+ membrane potential like?
Serum [K+] range: 3.5 - 5.3 mmol/l (increased K+ - hyperkalaemia)
Em less -ve (tending to depolarisation)
Reaches threshold more easily
Cell depolarisation more likely (weaker)
‘Car misfiring’
Decreased SAN firing/ bradycardia
Decreased [K+]E (Clinically – hypokalaemia):
•Em more –ve (tending to hyperpolarisation)
•Disrupts various K+ channels
•Abnormal heart rhythms (arrhythmias)
Kidneys and adrenal steroids
•Major role in K+ homeostasis
•Renal failure – don’t excrete excess K+
•Adrenal gland problems (Conn’s – too much aldosterone)
Hyperkalemia and therapeutic membrane transport?
Insulin + dextrose
•Insulin activates Na+/H+ antiport
• [Na+]I drives Na+/K+ ATPase
•Cells take up excess K+ ions; [K+]E
•What about the dextrose?
What occurs in regular ventricular AP?
Phase 0:
•Depolarisation
➢Fast Na+ channels
Phase 1: K+ leak – partial repolarisation
Phase 3 – K+ efflux - repolarisation
Phase 4:
•Diastolic resting potential ~ -90 mV
Phase 4: Return to resting potential
•Ca2+/Na+ channels close
• K+ permeability
Phase 2 – Plateau /contraction
•Slow Ca2+ channels (L-type) open
• K+ permeability – no repolarisation
What occurs in regular ventricular AP?
Phase 0:
•Depolarisation
➢Fast Na+ channels
Phase 1: K+ leak – partial repolarisation
Phase 2 – Plateau /contraction
•Slow Ca2+ channels (L-type) open
• K+ permeability – no repolarisation
Phase 3 – K+ efflux - repolarisation
Phase 4:
•Diastolic resting potential ~ -90 mV
Phase 4: Return to resting potential
•Ca2+/Na+ channels close
• K+ permeability
Role of digoxin in ventricular AP?
Role of digoxin in ventricular AP?
How does ischameia affect ventricular AP?
Ischaemia:
Hypoxia - [ATP]I
Opens KATP channel
•EM less –ve (~-55mV) (4)
•Depolarises easily
•Fast Na+ channels inhibited ~ -55mV
Slow Ca2+-mediated depolarisation (0)
•Early repolarisation (1); Plateau (2);
• Action potential duration (3,4; KATP?)
What occurs at the neuromuscular junction at AP transmission?
Motor nerve impulse open synaptic voltage-gated Ca2+ channels
• Increased [Ca2+]i promotes synaptic vesicles migration to pre-synaptic membrane
•ACh released into synaptic cleft
➢Two ACh bind ACh-gated Na+ channel
➢Nicotinic cholinoreceptor (NAChR)
• increased [Na+]I depolarises muscle fibre
•T-tubule system propagates action potential
•Release of sarcoplasmic reticulum (SR) Ca2+
•Signal terminated by AChE
➢EPP terminated
➢Ca2+ returns to SR
➢Myasthenia Gravis – destroys NAChRs
What occurs in the depolarising blockade of neuromuscular junction?
Prolonged activation of NAChR
➢Abolishes effector response
•Presence of nicotinic agonist inhibits post-synaptic membrane recovery
➢Can’t initiate muscle fibre action potential
•Skeletal muscle relaxes
•“depolarising blockade”
➢Suxamethonium (Sux; succinylcholine)
➢Anaesthetics Rapid Sequence Induction (RSI)
➢Sux not metabolised by AChE
What is the membrane like?
Semi-permeable barrier
•Proteins facilitate transport and signalling functions
➢Carriers
➢Ion Channels
➢Receptors
•Maintains differential ion concentrations between intra-cellular and extra-cellular environments
➢Basis of membrane potential
●Influenced by acid:base balance and extra-cellular ion concentration
