Ligand-gated ion channels 1 Flashcards
What are ligand-gated ion channels
- multiple subunit proteins which form cation or anion channels
- Incorporate a receptor and open only when the receptor is occupied by an agonist
What are channel properties determined by
- The subunit composition
What channel properties are changed
- agonist activation
- ion permeation
- conductance properties
- deactivation / desensitisation kinetics
- receptor localization
What are examples of different sizes of ligand gated ion channels
- Puinergic- 3 subunits
- Glutameric- 4 subunits
- GABA- pentameric
What are the different subunits of GABA
- Alpha
- Beta
- Gamma
- Delta
- Epsolon
- Rho
What are the different types of GABA receptors
- GABAA - Cl- ion channel
- GABAB - GPCR
- GABAC - Cl- ion channel
Describe structure of GABAARs
- Heteropentameric ion channels
- 2 alpha subunits, 2 beta subunits and gamma subunit
- Biding site for GABA at interfaces of alpha and beta subunits
- Aqueous pore in centre
- 4 transmembrane spanning domains
- Cis loop on extracellular surface on N-terminal domain
What is the most common subunit combination for GABAAR
- 2X Alpha 1
- 2x beta 2
- 1x gamma 2
What does different combinations of GABAA subunits mean
- One transmitter can do lots of different jobs that is determined by the function of the GABAA receptor- governed by subunits
What do all alpha subunits have
- Characteristic cys-cys pair in the N-terminal extracellular domain, essential for agonist binding
Where is there homology between the receptor subunits
- Transmembrane domain M1, M2, M3, M4
- Ligand binding region are variable
- Intracellular loop interacts with proteins inside of cell- variable
Describe agonist activation o
- Binding of GABA to N-terminal domain at interface of a/b subunits
- Opening of ion-selective pore- anion
- Transmembrane 2 domains- Conserved leucine residue is implicated in the gating of the ion channel
Describe channel opening of ACh (analogous to GABAA)
- Agonist binding induces change to cause channel to open
- Closed state - two alpha subunits and transmembrane 2 domain facing into pore
- Weak interactions between inner helices causes hydrophobic constriction of ion channel
- In presence of ACh, the ion channel opens
- Binding induces rotation in extracellular beta sheets- conformational change which opens channel
Describe a model for agonist activation of LGIC with two agonist binding sites
- Agonist binding increases the probability of ion channel opening
- Measurement of functional binding-gating can only be done using single-channel analysis
- K = binding constant
- *= channel opening
- Efficacy = Beta/alpha
- Beta= opening rate constant
- Alpha= shutting rate constant
- If both binding sites are occupied get bigger opening
- Model of alpha and beta- if ligand only binds to one part it doesn’t open fully- otherwise just partial opening
- Need both to be ligated to open fully
Describe example of how subunit combination of GABAA can change pharmacology
- Functional GABAA receptor is a pentamer of at least a/b or a/b/g
- Agonist affinity and efficacy at the GABA binding site is altered by a subunit
- THIP and P4S are partial agonists at the GABAAR have the highest affinity and efficacy on a6b1g2 and the lowest affinity and efficacy on a4b1g2
- Depending on the composition of channel agonists can bind and open channel different amounts
- Compared effects of agonist on electrophysiological recordings
- GABA only – Alpha 4- low efficacy
- But alpha 6 much higher efficacy
What is ion selectivity based on
- usually refers to relative permeability
- pore size
- presence of charged groups lining the pore
Describe what effect GABAA when agonist bound
- GABAA agonist induces conformational change which increases permeability of central pore to Cl-
- Generally leads to hyperpolarization and inhibition of network activity
What determines the specificity to ions
- Permeable to specific ions – size of pore and charge of groups determines which ions
- Depends on amino acids - Basic, polar, acidic etc
What is an inhibitory synapse and give example
- e.g. g-aminobutyric acid
- Generates inhibitory postsynaptic potential (IPSP)
- When ion channel opens cl- diffuse into cell taking negative charge into cell- hyperpolarises the membrane away from resting membrane making it more difficult to cause action potential
What is the Nernst Equation measuring
- Ion flow through a channel
- The equilibrium potential for any ion (Eion) can be calculated using the Nernst equation
What is the Nernst Equation
- Eion = 2.303 RT/zF log [ion] outside/[ion] inside
- Constants
- R = gas constant
- T = absolute temperature
- F = Faraday’s constant
- z = charge of the ion
- So really based on ions outside to inside – everything else are constants
What type of ions do nAChR/5HT3 channels allow
- nAChR/5HT3 are cation channels
What are the two models of ligand binding and channel opening
- Induced fit model
- Allosteric model
What is the induced fit model of ligand binding
- binding of agonist to the receptor induces a conformational change in the region of ligand-binding site which propagates to the pore region and causes opening of the ion-conducting pore
What is the the allosteric model of ligand binding
- receptor constantly undergoing spontaneous transitions between different conformations.
- Different conformations confer different affinity for ligand.
- Binding of an agonist typically stabilizes the channel in the open state.
What are the determinants of IPSC kinetics
- Lifetime of GABA at the synapse- diffusion/reuptake mechanisms
- Postsynaptic GABAA receptor composition- IPSC decay rates vary from 10-100s of ms
- Single channel recording of receptors- some open for short some for longer- just based on different of one subunit
Do ligand-gated ion channels show desensitisation
- Also exhibit desensitisation
- Mechanism not as clearly worked out as for gpcrs
- Same gabaa receptor just different GABA application times – longer time= desensitisation
What is desensitisation of GABAARs altered by
- Desensitization of GABAARs is altered by subunit composition
- Desensitization is altered by endogenous factors (Zn, phosphorylation) and by pharmacological agents (e.g. BZ)
- Effects of subunit composition not well established – big difference between delta and gamma-2 containing proteins
Describe GABAA receptor localization
- in situ hybridization study for subunit mRNA in brains
- some individual alpha subunits have discrete distribution
- others have overlapping distribution
- g subunit important for trafficking Rs to cell surface
Where can GABAAR be found
- In synpase
- Extra synaptically
Describe activity of synaptic GABAAR
- Phasic activation – when nerve fires
- Transient inhibition
- Hi [GABA]
- a1,2,3 (5), g2 containing Rs
- BZ-sensitive
- Anxiolytic
- anticonvulsant
Describe activity of extrasynaptic GABAA receptors
- Persistent activation
- Tonic inhibition at low levels
- Low [GABA] but are active more or less all the time
- a4,6 (5), delta containing Rs
- BZ in-sensitive (not a5)
- Alcohol
- Anaesthetics
- Neurosteroids
