Peripheral Neural Transmission Flashcards
What receptors does NA bind to
Binds to α»_space;> β
But still activates β eg in SNS to heart
How are small molecule NTs acquired?
synthesised locally within presynaptic terminals
Where are the enzymes needed to locally synthesise neurotransmitters produced?
in the soma and then transported to the nerve terminus by slow axonal transport
Where do the precursor molecules for synthesising new small molecule NTs come from
may pre-exist within the nerve terminal but more commonly they are brought inside by some transporters found in the plasma membrane of the nerve terminus.
True or false:
small molecule NTs can be synthesised inside the vesicles
True
For some small-molecule neurotransmitters, the final steps of synthesis occur inside the vesicles where the necessary enzymes are presen
Where are neuropeptides usually synthesised
What happens next
in the soma where they are packaged into vesicles where further processing takes place
peptide filled vesicles are transported along the axon to the nerve terminal via fast axonal transport
What do fast and slow axonal transport in nerves transport?
fast: peptide filled vesicles (neuropeptides)
slow: small molecule neurotransmitters
True or false
all neurotransmitters are loaded into vesicles soon after synthesis
False
Gaseous transmitters eg NO are not
all others are
Briefly describe the membrane and the interior of the vesicles for NT transport
phospholipid bilayer
aqueous interior
What is the size of vesicles of small molecule transmitters
40-60nm
Describe the appearance of small molecule NT vesicles
Hence what are they known as
appear clear in EM
small, clear core vesicles
What is another name for small, clear core vesicles
Why?
synaptic vesicles
they are more abundant and cluster at the active zone
Where are neuropeptides loaded into vesicles
What size are these vesicles
in the soma
90-250nm in diameter
What are neuropeptide vesicles known as?
why
large dense core vesicles
they appear electron dense in EM
How are neuropeptide vesicles different to those of small molecule transmitters
neuropeptide - larger,
more e- dense
do not cluster at the active zone
small molecule:
small, clear-core vesicles,
gather at the active zone
How are vesicles released (as discovered by Katz)
in a quantal manner where 1 quantum is the content of a single vesicle (as we saw in HOM last year)
Which ion channels in the presynaptic nerve terminal open in response to depolarisation ?
What types of channel specifically?
voltage gated Ca2+ channels
Cav2 series: mainly P/Q and N type
How long does it take for Ca to trigger exocytosis
required conc of Ca is very quickly reached
Why is the required conc of Ca for exocytosis very quickly reached in the presynaptic terminal
Cavs are clustered around the active zone and some vesicles are already docked to release sites so release of contents can be v rapid
What is the releasable pool?
the presynaptic vesicles that are already docked to release sites in the terminal
How much of the total amount of vesicles is represented by the releasable pool
Only a v small amount
Their numbers can be enhanced on demand when vesicles from the reserve pool join them
What cell structure is the reserve pool associated with
cytoskeleton
What are the 6 steps of vesicle release and recycle
1) Transmitter loading
2) mobilisation of vesicles toward active zone
3) Docking
4) Priming
5) Exocytosis
6) recycling of vesicles
Describe the transmitter loading stage of the presynaptic vesicle life cycle
What precedes and follows it
NT imported using ATP
Proton pump acidifies interior to create an electrochemical gradient across vesicle membrane
Gradient provides energy for NT uptake by specialized transporters
Preceded by recycling of vesicle
Followed by mobilisation of vesicles
What protein family is important for mobilization of vesicles towards the active zone
What modification is important in this process
synapsins
phosphorylation
Describe vesicle mobilisation toward the active zone in presynaptic terminals
non phosphorylated synapsins reversibly bind to vesicles and actin to cross link them to each other, keeping the vesicles tethered to the reserve pool
Ca2+ increase (following depolarisation) activates CaMKII, which phosphorylates synapsins, allowing dissociation from vesicles.
The vesicles are now free to move
What is CaMKII
Ca2+ /calmodulin dependant protein kinase (type II)
Describe the docking stage of the presynaptic vesicle life cycle
What step precedes and follows it
At the active zone vesicles dock to nerve membrane via SNARE proteins
preceded by vesicle mobilization
followed by vesicle priming
What occurs at the priming stage of the vesicle cycle
Docked vesicles undergo ATP dependent prefusion that primes them for Ca induced release
Which type of protein is vvv important in vesicle priming
Are these the only proteins involved
SNARE
no - many others involved
What is the main purpose of priming vesicles
to organize SNAREs into correct conformation for membrane fusion
Describe the exocytosis step of vesicle release
What follows and precedes it?
Ca2+ (sensed by synaptotagmins) triggers complete fusion of vesicle with membrane in a rapid reaction
Which protein sense Ca2+ to initiate vesicle-membrane fusion
synaptotagmin
What are the 3 different routes that can be taken for vesicle recycling after release of the contents
reversible fusion pore
endocytosis
bulk retrieval pathway
Describe the reversible fusion pore route of vesicle recycling
vesicle does not completely fuse with nerve membrane - contents are released via a fusion pore instead
Vesicle is retrieved by closing the pore
Spent vesicles can either stay fused to membrane (kiss and stay) or relocate to reserve pool ( kiss and run)
How quickly can reversible fusion pore occur
vvv quickly
in tens to hundreds of milliseconds
What does ‘kiss and stay’ and ‘kiss and run’ refer to
reversible fusion pore
stay: spent vesicle remains fused to nerve membrane
run: spent vesicle relocates to reserve pool of vesicles
Describe the endocytosis pathway for vesicle return
What happens after the vesicle is recycled
vesicle coated in clathrin
the coat is later shed and the vesicle returned to interior of synaptic nerve terminal
empty vesicles either refill immediately with NT or pass through endosomal sorting
Describe the bulk retrieval pathway for vesicle return
excess membrane re-enters the terminal by budding from uncoated pits - not very common .
Why do many congenital myasthenic syndromes arise
defects in ACh release due to inadequate number of synaptic vesicles available for release.
What happens in familial infantile myasthenia
synaptic vesicles are much smaller
NOT a shortage of vesicles
What happens in Lambert Eaton myasthenic syndrome (LEMS)
What other disease can this syndrome be a complication of
ACh release is reduced due to autoimmune destruction of neuronal Cav channels in active zone
cancer
Why is vesicle- membrane fusion difficult
Because a membrane bilayer is a stable structure, fusion of the synaptic vesicle and plasma membrane must overcome a large unfavourable activation energy
What does SNARE stand for
soluble N-ethylmaleimide-sensitive factor attachment receptors
How conserved are SNARE proteins
SNAREs are universally involved in membrane fusion, from yeast to humans
How are SNAREs categorised
v- SNARE (v for vesicle)
t-SNARE - (t for target membrane)
What part of the SNARE protein is highly conserved
cytosolic domain:
the SNARE motif, that is 60-70 residues long and form coiled-coil alpha-helical structures
What SNAREs does each synaptic vesicle have
a single type of v-SNARE:
synaptobrevin (AKA VAMP)
What is another name for synaptobrevin
vesicle associated membrane protein or VAMP
What SNARE proteins are present in the target membrane of the presynaptic active zone
2 types:
syntaxin
SNAP25
What happens to the SNARE proteins during exocytosis
form a trans-SNARE complex
What does the trans-SNARE complex consist of
4 alpha helices - one each from synaptobrevin and syntaxin and two from SNAP-25
Other proteins involved are:
Munc 13, Munc 18, complexins, Rab3A
What must happen to the trans-SNARE complex after fusion
dissemble via the action of NSF and SNAP (an adaptor protein)
What proteins mediate dissembly of the SNARE complex
What is the action
cytoplasmic ATPase called NSF (N-ethylmaleimide-sensitive fusion protein) binds to SNARE complexes via an adaptor protein called SNAP
NSF and SNAP use the energy of ATP hydrolysis to dissociate SNARE complexes, thereby regenerating free SNAREs.
What ion is involved in vesicle fusion?
What does this mean?
Ca2+
process should implicate a Ca2+ sensor - a role fulfilled by synaptotagmins
How are synaptotagmins associated to the vesicle
Give another fact about their structure
anchored to synaptic vesicles via their N termini while their C-termini contain two C2 domains - C2A and C2B, that bind Ca2+.
What do the C2 and 3 domains of synaptotagmins do respectively
C2: when Ca2+ is bound, C2 binds sytaxin and SNAP 25, and also binds to membranes containing acidic phospholipids
Describe the Ca2+ binding of the C2 domain of synaptotagmins
low binding affinity so large [Ca2+] required for activation
How do nerves ensure C2 domains of synaptotagmins get enough Ca2+
by co-localisation of synaptotagmin with Cav channels that cluster the active zone
Where do Ca2+ rich microdomains develop in the presynaptic terminal
in the vicinity of Cav pores due to the poor diffusibility of Ca2+
Why is the high [Ca2+] near Cav channels important
high local [Ca2+] can then be sensed by the synaptotagmins present in close proximity with these channels
Name 2 toxins that target SNARE proteins
How they both work very generally
botulinum toxins (BoTx)
Tetanus (TeNT)
Both selectively cleave SNARES
What are the 2 types of target proteins on the postsynaptic membrane for released transmitters
ionotropic (ligand gated ion channels) receptors
metabotropic receptors (GPCRs)
How do most small molecule NTs interact with postsynaptic receptors
bind to their cognate LGICs to mediate a fast post-synaptic response which could be the generation of either excitatory or inhibitory post-synaptic currents
or could bind to GPCR for a slower response (catecholamines and neuropeptides also do this second one)
Which postsynaptic receptor family is important in synaptic plasticity
GPCR - modulate strength of synaptic transmission by altering function/availability of LGICs in the postjunctional membrane
Can neurotransmitters change K+ position
binding to post synaptic GPCRs can open K+ channels leading to K efflux and reduced excitability of post synaptic membrane
True or false
Very little neurotransmitter is released upon stimulation
What does this mean for removal
False
Neurotransmitters are released typically in large amounts and timely removal of transmitters from the synaptic cleft is critical to synaptic transmission
How can NT be removed from the synaptic cleft
reuptake (either into presynaptic terminal or into surrounding, non neuronal cells eg glia)
Destruction of neurotransmitter (eg proteolysis of neuropeptides)
How is ACh removed from the synaptic cleft
hydrolyzed by AChE
Which are removed more quickly from the synaptic cleft:
neuropeptides or small molecule transmitters
small molecule transmitters are removed faster than neuropeptides
What is homotypic interaction at synapses
What is this also called
When released transmitters modulate further release of their own from the same nerve terminals
Homotropic interaction
What is heterotropic interaction at synapses
AKA?
when released neurotransmitters modulate release of other transmitters from different nerve terminals through activating cognate receptors (GPCRs) in the presynaptic terminal membrane
AKA heterotypic interaction
What receptors regulate homotropic and heterotropic interactions?
GPCRs called presynaptic autoreceptors (if homotropic interaction)
or
presynaptic heteroreceptors (if bind NTs from other synapses)
What are presynaptic hetero/autoreceptors usually coupled to
inhibitory G proteins (Gi or Go) leading to reduced [cAMP]] and enhanced K+ efflux - ultimately reduces NT release
Give an example of one of the few cases where released NT acting on a presynaptic GPCR leads to enhanced NT released
motor neurons connecting the skeletal muscle fibre
Is it only neurotransmitters that can act on presynaptic GPCRs?
No
sometimes, chemicals that are not canonical transmitters (e.g. prostaglandins, endocannabinoids, NO etc.) and produced by the post synaptic neurons/ effector tissues, can also modify neurotransmission through this pre-synaptic routes.
Give 3 chemicals that are not canonical transmitters but can modify NT release via presynaptic GPCRs
prostaglandins,
endocannabinoids,
NO
Other than action on presynaptic GPCRs what can affect release of NT
anything that interferes with the generation of Ca2+ signals in the presynaptic terminal
eg by inhibiting Cav channels or by chelating Ca2+
Give 2 agents which inhibit Cav channels in the presynaptic terminal
verapamil
aminoglycoside antibiotics
Which drugs chelate Ca2+ in the presynaptic terminal
tetracyclines
Where are LDCVs located in the presynaptic terminal
away from the active zone
What does exocytosis of synaptic/ clear core vesicles result from
What can trigger this
transient but large and local increases in intracellular Ca2+
a single depolarising event is enough to provide such Ca2+ signals.
What do LDCVs require to by exocytosed? (compared to clear core vesicles)
How is this requirement often met
smaller but sustained rise in global Ca2+ concentration
by a rapid train of action potentials (= repetitive firing of the nerve).
Why are the different requirements for exocytosis of LDCVs and clear core vesicles useful
specific patterns of electrical activity in a neuron may lead to the preferential release of a neuropeptide or a small-molecule NT
What are the 3 main broad sites of pharmacological intervention at the synapse
presynaptic nerve terminal,
sypatic/ junctional cleft
the post-synaptic/post-junctional target membrane
What are the 2 ways a pharmacological agent can modulate peripheral neurotransmission
direct (binds to postsynaptic target as an agonist, antagonist or pore blocker)
indirect (reduces/ enhances amount of NT released)
What was the first neurotransmitter to be discovered
ACh (1914)
Where is ACh found in the peripheral nervous system (5)
- motor nerves that innervate the skeletal (voluntary) muscles
- all autonomic pre-ganglionic nerves
- nerves terminating at the adrenal medulla
- postganglionic parasympathetic nerves at neuro-effector junctions
- postganglionic sympathetic nerves innervating the sweat glands
The availability of what chemical is vital for ACh synthesis?
How do we acquire it?
choline
primarily from diet as there is little de novo synthesis in neurons
can be recycled after ACh breakdown
What is ACh hydrolysed to
acetate and choline
What happens to the products of AChE
much of the choline is taken up by ChT1 for ACh synthesis
What is usually the rate limiting step in ACh synthesis
reuptake and availability of choline
What mediates choline reuptake into the presynaptic terminal
ChT1
Describe ChT1
What is it a family of
what can it transport
what inhibits it?
high affinity choline transporter
a member of the solute carrier family of proteins
transports Na+ and can be inhibited by hemicholinium
Is hemicholinium used clinically?
no
How is ACh formed from choline
Within the cytoplasm of the pre-synaptic nerves, choline is acetylated to form ACh by choline acetyltransferase (ChAT or CAT)
Where does the acetyl group for choline in ACh synthesis come from
acetyl CoA from the Mt
What is Triethylcholine
competes with choline as a substrate for CAT and is converted to acetyltriethylcholine which is in turn released instead of ACh
What does the release of acetyltriethylcholine mean
it is far less potent than ACh so acts as a false transmitter
How is ACh stored in the presynaptic terminal
in cholinergic vesicles
How does ACh enter the vesicles in the presynaptic terminal
by vesicular ACh transporters (VAChTs)
How do VAChTs work
high H+ gradient to allow exchange of H+ for ACh
What provides the H+ gradient across the ACh vesicle wall
V- type ATPase
What kind of vesicle is ACh stored in
clear core generally
contain ATP also at a ratio of 10:1
Do cholinergic presynaptic vesicles ever have dense core vesicles?
yes - some have vesicles containing VIP
Why do cholinergic vesicles sometimes contain ATP and VIP
ATP and VIP function as co transmitters at some synapses
What drug targets ACh storage
vesamicol
an experimental compound, is a non-competitive and reversible blocker of VAChT.
What is vesamicol
an experimental compound, is a non-competitive and reversible blocker of VAChT.
Which pharmacological agents target ACh release?
Botulinum (BoTXs/BoTNs)
β bungarotoxin (β-BuTx)
α- latrotoxin (α-LTX)
Tetanus neurotoxin (TeNT)
What produces botulinum
Clostridium botulinum
a Gram-positive bacterium responsible for botulism that may happen through food poisoning, colonisation of the gut in an infant or via exposed wounds
What do botulinum toxins target
cholinergic peripheral synapses (NMJ)
Describe the action of BoTX (think about their structure)
heavy chains bind to some gangliosides specific to the target nerves which allow them to be endocytosed specifically into these nerves.
C termini are proteases that are selective for SNARE
What are symptoms of botulism (2)
skeletal muscle weakness (which may lead to respiratory paralysis)
autonomic signs that would be associated with loss of cholinergic activity (e.g. constipation, blurred vision, dry mouth, difficulty in swallowing, urinary retention etc.).
Where is β - BuTx found
Venom of branded kraits
What is the action of β BuTx
Causes irreversible damage to pre-synaptic terminals which then lose synaptic vesicles
ACh release block is probs via its phospholipase A2 action - degrades phospholipids in active zone
Does β BuTx attack all nerves
Why
Only cholinergic motor nerves
May stem from its interaction with the substrate phospholipids that are unique to these nerves
How does β BuTx kill you
Respiratory failure
Where is α latrotoxin found
Black widow spider venom
How does α LTX cause paralysis
Causing a massive release and subsequent depletion of the ACh at the NMJ
ACh release in absence of depolarisation/ Ca entry
How do we think α LTX works
As a tetramer, It can form a calcium permeable ion channel
What produces TeNT
How does it enter the nervous system
Clostridium tetani (another Gram-positive bacterium)
enters the nervous system at the NMJs
How can TeNT enter the nervous system via the NMJ
the C-terminal heavy chain of TeNT binds to some gangliosides that are enriched on the peripheral terminals of motor neurons and this binding enables endocytosis.
What happens once TeNT enters through the NMJ
trapped in endosomes, TeNT progresses towards soma via retrograde axonal transport and is then discharged into the intersynaptic space within the spinal cord
What happens after TeNT is released into the inter synaptic space in the spinal cord
binds to presynaptic membrane of inhibitory interneurons to be endocytosed into the synaptic vesicles of these neurons
What happenns to TeNT after it is endocytosed into the synaptic vesicles of presynaptic inhibitory neurons of the spinal cord
acidic nature of vesicles causes insertion of N terminal light chain into cytoplasm and cleaves synaptobrevin
What is the result of TeNT’s N terminal cleaving synaptobrevin on the synaptic vesicles in the inhibitory presynaptic neurons in the spinal cord
the inhibitory interneurons are unable to release glycine, causing disinhibition of the motor neuron which then results in violent (tetanic) contractions of skeletal muscle.
True or false
ACh is a physiological inhibitor of itself
true
At postganglionic parasympathetic nerve endings, inhibitory muscarinic M2 receptors participate in auto inhibition of ACh release;
other mediators, such as noradrenaline (NA), also inhibit the release of ACh.
How does ACh modulate its own release at NMJ
What is the purpose
pre-synaptic nicotinic receptors (nAChRs) facilitate ACh release
allows synapse to function reliable during prolonged high frequency activity
What are cholinoceptors
cholinergic receptors that ACh binds to after its release
Where are cholinoceptors found and how can they be classified
post synaptic membrane
muscarinic and nicotinic
How can you experimentally determine a nicotinic and muscarinic AChR
on the basis of their different affinities for agents that mimic the action of ACh (cholinomimetic agents)
What are nAChRs
LGIC belonging to the ‘Cys Loop’ family
Where are nAChRs found
at the NMJs, autonomic ganglia, adrenal medulla, the CNS and some other non-neuronal (notably various immune and epithelial) cells
Describe the structure of a functional nAChR
a pentamer consisting of α,β, γ, δ and ε subunits with at least 2α subunits present. Each subunit consists of four transmembrane (M1-M4) helices and the M2 helix from all five subunits together form the pore. ACh binds to the interface between the α and neighbouring δ or γ subunits and minimally 2 molecules of ACh are required for the activation of the channel leading to the opening its pore.
What are most isoforms of the nAChR permeable to
Is this always the case?
Na and K
No- one of the CNS isoforms namely the (α7)5 channel, is highly Ca2+ permeable.
What does activation of nAChRs cause
depolarisation and therefore an EPP (muscle) or EPSP (peripheral ganglia)
What happens if an EPp is strong enough
An action potential is generated
True or false
nAChRs can be desensitised
True
Usually by prolonged or much stronger agonist stimulation
How can nicotinic AChRs be divided based on location
Nm (muscle) and Nn (neuronal)
What is the major isoform of the Nn nAChR
Where is it found
(α3)2(β2)3
Autonomic ganglia
What is the major variant of Nm nAChR?
Where is this found
(α1)2(β2)δε
Adult skeletal muscle membrane
What are the predominant Nm isoforms of the nAChR in the CNS
(α4)2(β2)3
(α7)5
True or false
All nAChRs are metabotropic
False
All mAChRs are metabotropic (GPCRs)
Why are they called muscarinic AChRs?
in addition to binding ACh, also recognize muscarine, an alkaloid that is present in certain poisonous mushrooms.
How do AChRs respond to nicotine?
nAChR: affinity
mAChR: low affinity
What do mAChRs mediate
postganglionic transmission in the peripheral nervous system, being present on the membranes of effector tissues including heart, tissue-specific smooth muscles (lung, some blood vessels, bladder, gut etc.) and glands (salivary, lacrimal etc). They also occur in the CNS
Where is the M2 mAChR found
What is its action here
pre-synaptically on the endings of noradrenergic nerves and cholinergic nerves
its activation causes a decrease in transmitter release
How many subtypes of mAChRs are there
Which are the most important in the periphery
5: M1-5
M1, 2, 3
Are M1, 2, 3 mAChRs found in autonomic ganglia
. Some of these isoforms may be present in autonomic ganglia but they are mostly present in the effector tissue membrane and they mediate the signalling leading to the biological functions of cholinergic stimulation.
Is AChE usually floating about?
no it is tethered to the post synaptic membrane
a soluble AChE is found in the cytoplasm of the presynaptic neuron (obvs doesnt affect life span of released ACh)
What does neurotransmission in autonomic ganglia involve
release of ACh by pre-ganglionic fibres and the rapid depolarization of post-ganglionic nerve membranes via the activation of neuronal nAChRs (Nn).
How is the structure of cholinergic autonomic ganglia different to the NMJ
Unlike the NMJs, ganglia do not have discrete end plates with focal localization of receptors; rather, the dendrites and nerve cell bodies contain the receptors
Which Nn is insensitive to alpha bungarotoxin
What can inhibit these
(α3)2(β2)3
trimetaphan (a competitive antagonists) and hexamethonium
What are the 4 different changes in membrane potential of postganglionic nerves from cholinergic stimulation
- an initial EPSP (via Nn receptors) that may result in an action potential
- an IPSP mediated by muscarinic M2 receptors
(Gi/o-coupled) - a secondary slow EPSP mediated by muscarinic M1 receptors (Gq/11-coupled)
- a late, slow EPSP mediated by various neuropeptides
What is the issue with agents acting at the autonomic ganglia
lack of selectivity between sympathetic and parasympathetic ganglia and few can also have effects on the CNS. Their peripheral action is largely governed by the extent of the sympathetic and parasympathetic control of the tone of the effector systems.
What are the 2 categories in which ganglionic stimulants can be grouped
Nn agonists
mAChR agonists
Name a member of the Nn receptor group of ganglionic stimulants
nicotine
What is nicotine
a tertiary amine found in tobacco leaves
What does nicotine do at low and high concentrations
stimulates Nn channels whilst at higher doses, it desensitizes the channel (typical of for LGICs)
Name some products that use nicotine as an aid for smoking cessation
gums, lozenge, e-cigarettes (‘vapes’) and transdermal patches as part of the nicotine replacement therapy (NRT)
What is varenicline used for
used as a smoking cessation ‘aid’ along with counselling and behavioural therapy to help people stop smoking
What drug is used to aid smoking cessation in conjunction with therapy
Describe its action
varenicline
high affinity partial agonist for the α4β2 receptors (mainly implicated in nicotine addiction) and also a full agonist on α7 receptors
both are in the CNS
What receptor is mainly implicated in nicotine addiction
α4β2 Nn receptor
Name 2 ganglionic stimulants that act as Nn agonists
Are these drugs
tetramethylammonium (TMA)
dimethylphenylpiperazinium (DMPP)
not drugs but useful for experiments
What are mAChR agonists blocked by
atropine
What do mAChR agonists mimic
the slow EPSP
How does nicotine act as an Nn receptor blocking agent
initially stimulates the ganglia by an ACh-like action and then blocks them by causing a persistent depolarization
Do all ganglionic blocking agents act in the same way as nicotine
no other ones
impair transmission either by competing with ACh for binding at ganglionic Nn receptors (e.g. trimethaphan) sites or by blocking the channel pores (hexamethonium).
equivalent to non depolarising blockers at NMJ
What does trimethaphan do
competes with ACh binding at ganglionic Nn receptors
Which drug blocks channel pores at ganglionic cholinergic synapses
What is important to remember when prescribing this drug
hexamethonium
Blockade of ganglionic Nn channels by hexamethonium manifests use dependence.
What were the earliest antihypertensive agents
Are they still used?
ganglionic blocking agents
no - clinically obsolete by now due to numerous side effects, largely coming from their inability to discriminate between sympathetic and parasympathetic ganglia.
How many nerves innervate one muscle fibre
only one nerve fibre supplies each muscle fibre - the end plate
What happens at the NMJ structurally
the pre-synaptic axon terminal boutons synapse with the muscle fibre membrane (sarcolemma) that is deeply-infolded as ‘junctional folds’
Where are Nm nAChRs present in the sarcolemma
What else is near them
on the crest of the junctional folds
volatage gated Na channels are deep in the folds and in the non junctional part of the sarcolemma
What happens when ACh binds to Nm receptors
the nAChRs open to allow Na influx and K efflux
generates the EPP and if threshold for Nav channels is reached an AP is elicited
Nm channels are equally permeable to Na and K - does this mean equal amounts of Na enter and K leave?
No
the prevailing electrochemical gradients for Na+ is much greater than that of K+, so more Na+ enters than K+ leaves the cytoplasm
How is ACh removed from the NMJ
hydrolysed by AChE present in the basal lamina within the junctional cleft.
where is ACh found in the NMJ
in the basal lamina within the junctional cleft.
hat do directly acting agents modulating NMJ transmission act on?
directly on the Nm receptors on the sarcolemma
How do the subunits of the Nm receptor at the NMJ vary between the adult and foetus
The adult Nm is represented by (α1)2β1δε
the foetal form has gamma in place of the ε subunit
What is the adult Nm receptor irreversibly antagonized by
α-bungarotoxin
What is the main clinical use of neuromuscular blocking agents
as an adjuvant in surgical anaesthesia to obtain relaxation of skeletal muscles.
How do muscle relaxants improve the safety of anaesthesia ?
less anaesthetic is required, minimizing respiratory and cardiovascular depression and allowing rapid post-anaesthetic recovery
What is used in conjuction with GA during laryngoscopy, bronchoscopy, and esophagoscopy
Why do we not have to worry about these drugs having a wide spread effect
Short-acting drugs are used to facilitate endotracheal intubation
charged so don’t cross BBB and action remains peripheral
What are the 2 types of NMJ blockers
Depolarising and • non-depolarizing neuromuscular blocking agents
How do depolarising NMJ blockers work
Nm agonists so open Nm channels but are not readily hydrolysed by AChE
depolarisation last longer leading to some fasciculations,
followed by blocking of neuromuscular transmission and flaccid paralysis (called phase I block).
The depolarisation from the Nm channels leaves Nav channels in a inactivated state
with increasing [depolarising blocker] over time the block converts from phase 1 to phase II (membrane gradually repolarizes but now the Nm channels become desensitized)
How can a depolarising blockade be potentiated
Anti-AChE agents
Name depolarising NMJ blocker
Suxamethonium (succinylcholine)
Why is succinylcholine considered unique
Suxamethonium (succinylcholine) represents the only depolarizing blocking agent being used in the clinics at present.
What is phase I and II of the depolarising block at the NMJ
Phase I: extended depolarisation leads to fasciculations before flaccid paralysis because the open Nm receptors maintain the depolarized state of the sarcolemma and thus leaving all the Nav channels in inactivated state
Phase II: agent increases and membrane gradually repolarizes but now the Nm channels become desensitized.
When is succinylcholine used despite its side effects
still in conditions where fast onset and brief duration of action are required, e.g. with tracheal intubation
Why does succinyl-Ch have a v brief duration of action
due to its rapid hydrolysis in the plasma by butyrylcholinesterase (BuChE, also known as pseudocholinesterase).
Where is BuChE synthesised
What does this mean when administering NMJ blockers?
liver
patients with liver disease or genetic deficiency of this enzyme therefore run the risk of prolonged action of suxamethonium (which is hydrolysed by BuChE)
Whyy must you check a patient doesnt have electrolyte imbalance before administering succinyl-Ch
suxamethonium is a depolarising NMJ blocker and causes
K+ loss from the muscles into blood, which may lead to hyperkalemia and can be life-threatening for patients with electrolyte imbalance
What are non depolarising NMJ blockers essentially
competitive antagonists of Nm channels
true or false
non-depolarising NMJ blockers lead to flaccid paralysis
true
they prevent depolarization of the sarcolemma and inhibit muscular contraction, leading to a flaccid paralysis
fasciculations are not seen with these
How can the competitive action of non depolarising NMJ blockers be overcome
Why is this useful clinically?
by administration of anti-cholinesterases (e.g. neostigmine), which increase the [ACh] in the NMJ.
gives anaesthesiologists an option to shorten the duration of the neuromuscular blockade.
What was the first known non depolarising block
curare - a cocktail of natural alkaloids used as arrow poison by South American Indians to paralyze prey
What is the major constituent of curare
Is it used clinically?
d-tubocurarine
no due to poor selectivity between the ganglionic and NMJ-specific nAChRs
What are synthetic non-depolarising blocking drugsdivided into
the aminosteroid group (the ones with uronium suffix) such as pancuronium, rocuronium etc.
and
the benzylisoquinolinium group (the ones with the urium suffix) such as atracurium
Describe the aminosteriod group of drugs
Give 2 examples
synthetic NMNJ non-depolarising blocking drugs
Ends in uronium
pancuronium, rocuronium
Describe the benzylisoquinolinium group of drugs
eg?
synthetic non-depolarising blocking drugs
(the ones with the urium suffix) such as atracurium, mivacurium
How does the onset of rocuronium compare to succinyl choline
succinyl Ch has a faster onset
non-depolarising neuromuscular blocking drugs have a slower onset of action than suxamethonium
How can non-depolarising neuromuscular blocking drugs be divided based on action?
by their duration of action as short-acting (15–30 minutes), intermediate-acting (30–40 minutes), and long-acting (60–120 minutes), although duration of action is dose-dependent
Which kinds of non-depolarising neuromuscular blocking drug is used more commonly in practice
Drugs with a shorter or intermediate duration of action, such as atracurium and rocuronium, are more widely used than those with a longer duration of action, such as pancuronium
Name 2 non depolarising NMJ drugs which have an intermediate duration of action
Which has a longer duration
atracurium and rocuronium
pancuronium
What is common about NMJ non depolarising blocker structure
quaternary ammonium (=charged) compounds therefore poorly absorbed (cannot be given orally) and rapidly excreted.
For the same reason, they do not cross the placenta (which allows them to be safe in obstetric anaesthesia) as well as the blood-brain barrier.
What do anesthetists use to monitor postoperative recovery of neuromuscular transmission?
tetanic fade
What is tetanic fade
non-depolarising blocking agents also block facilitatory pre-synaptic autoreceptors and thus inhibit the release of ACh during repetitive stimulation of the motor nerve
What are some of the clinical responses to NMJ blocking agents caused by
release of histamine
Name 4 NMJ blocking agents that cause histamine release
Suxamethonium, mivacurium,
atracurium
to a lesser extent than d-tubocurarine unless administered rapidly
Which NMJ blockers are less likely to cause histamine release
amino steroids such as pancuronium and rocuronium
Why is histamine released by muscle relaxants?
Histamine release typically is a direct action of the muscle relaxant on the mast cell rather than an outcome of individual’s allergic hypersensitivity
How does alpha- bungarotoxin work
What is it often used in
irreversibly binds to the ACh binding site on the adult Nm receptors at the NMJ and thus block the transmission.
a- BuTx, specially its various antibody/fluorophore conjugates are widely used in biochemical and structural studies of Nm receptors and NMJs.
How are distinct bungarotoxins different
α-Bungarotoxin inhibits the binding of acetylcholine (ACh) to nicotinic acetylcholine receptors; β- and γ-bungarotoxins act presynaptically causing excessive acetylcholine release and subsequent depletion
How is BoTX used clinically
BoTX-A
injected locally for cosmetics
can treat certain ophthalmic conditions associated with spasms of eye muscles (e.g., blepharospasm).
how do anti-AChEs at on NMJ transmission
act indirectly by prolonging the life time of release ACh within the junctional cleft.
Why must patients with myasthenia gravis be careful with certain antibiotics
members of the aminoglycoside antibiotics (e.g. streptomycin, neomycin etc.), in large doses inhibit Cav channels and thus can produce neuromuscular blockade, especially in patients with myasthenia gravis
Which antibiotics can block Cav channels at the NMJ
Give 2 examples
aminoglycoside antibiotics
e.g. streptomycin, neomycin
What is the effect of aminoglycoside antibiotics on patients treated with a NMJ blocker
What other drug class has a similar effect
in normal or low doses can unpredictably prolong muscle paralysis
tetracyclines
Where are mAChRs primarily found
What is a key exception
present in autonomic ganglia and vascular endothelial cells
also on autonomic effector cells innervated by post-ganglionic parasympathetic nerves
exception: postganglionic sympathetic nerves to sweat glands are cholinergic and the released ACh acts via mAChRs present in the gland
What is unusual about mAChRs being in vascular endothelial cells
they receive little or no cholinergic innervations
In the majority of cases how many types of mAChR are present in the effector tissue
at least two muscarinic receptor subtypes (typically M3 and M1/M2)
What is the ‘neural’ subtype of mAChR
M1
Where are M1 mAChRs mainly found
on CNS, autonomic ganglia and on gastric parietal cells
How to remember what the different classes of mAChR are coupled to
all odd ones (M1, 3, 5); coupled to Gq/11
what does agonist binding to mAChR M1 lead to
What other mAChR types would behave in the same way
activation of PLCβ -> hydrolyses PIP2 to IP3 and DAG
IP3 mediates intracellular Ca2+ release
DAG activates PKC
M3 and M5
Why can IP3 readily diffuse through the cytosol
What do the characteristics of DAG mean for it respectively
it is polar and activates the ER membrane-bound IP3 receptors which then mediate intracellular Ca2+ release
DAG being lipophilic, stays in the membrane and activates PKC
Does DAG only activate PKC ?
No
also activates several members of the TRP family of ion channels which are mainly involved in sensory perception and several of them are also Ca2+ permeable, mediating Ca2+ influx.
What else (other than PLC) mediates [DAG]
DAG lipase to produce arachidonic acid (AA).
What is the importance of the production of AA from the action of DAG lipase on DAG
may trigger a distinct Ca2+ influx pathway in some cells
most importantly, it is the precursor of biologically active lipid-derived molecules eg prostaglandins and leukotrienes .
What do M1 receptors in neurons inhibit
How does it do this (2)
What is the purpose?
K+ conductance (the m channel)
through the depletion of the PIP2 in the postganglionic nerve membrane. In certain case, the Ca2+ signals via calmodulin (CaM) and PKC, also contributes to the process
brings about a late depolarisation in the autonomic ganglion which occurs many seconds after the initial, fast EPSP induced by Nn receptors
What are M channels that conduct K+ known to be composed of
subunits of the Kv7/ KCNQ channel family.
Describe the M2 mAChR receptor
What are they coupled to
What does this mean they activate
cardiac subtype
Gi/o proteins
the inhibition of adenylyl cyclase (AC) which results in decreased cAMP production and thus decreased activation of protein kinase A (PKA)
What does M2 activation ultimately lead to
in decreased phosphorylation of a number of proteins that notably include the voltage-gated Ca2+ channels (CaVs), reducing probability of opening and Ca2+ influx
This decreases NT release from neurons
What is the predominant mAChR subtype in the heart
How are they distributed
M2
largely confined to the SAN and AVN to some extent in the atria but much less in the ventricles.
What does stimulation of the M2 receptors in the heart result in
negative chronotropic effect
How does activation of M2 channels in the heart
reduced Ca2+ current via the L and T-type Cavs due to reduced PKA-mediated phosphorylation of these channels
βγ opens g protein coupled inwardly rectifying K+ channels in the nodal pacemaker cells. This causes hyperpolarisation by K+ efflux
threshold for opening of the HCN channels which mediate the slowly-depolarizing inward Na+ current (the If current), is shifted towards a more negative value.
Reduces atrial contractility via triggering I(K-ACh)
What are the inwardly rectifying K+
GIRK1
Kir3.1
What is the K+ efflux current following M2 stimulations in the heart called
I (K-ACh)
What causes the threshold opening of the HCN channels which mediate the slowly-depolarizing inward Na+ current (the If current), to shift towards a more negative value.
reduced [cAMP]
What can the M3 mAChR subtype be classed as
glandular subtype
What are M3 receptors coupled to
What is usually the result of stimulation of these mAChRs
Gq and G11
stimulation of glandular secretions, e.g. saliva, sweat etc
What doe activation of M3 receptors in smooth muscle lead to
contraction because of the increase in cytosolic free [Ca2+]i
What causes [Ca2+]i to increase when M2 mAChRs are activated in smooth muscle
Why is the effect different in vascular smooth muscle
IP3 invoked Ca2+ release from ER
increased endothelial [Ca2+]i binds to CaM
Ca2+-CaM then activates eNOS to produce NO leading to smooth muscle relaxation
What are the following:
CaM
eNOS
EDRF
CaM = calmodulin
eNOS: endothelial nitric oxide synthase
EDRF: ‘endothelium-derived relaxation factor’ ie NO
What is the effect of CaM-Ca2+ activating eNOS in smooth muscle
NO activates soluble guanylyl cyclase, produces cGMP and ultimately relaxes the vascular smooth muscle
How can pharmacological agents modulate muscarinic cholinergic synapses (2)
the activity of mAChRs by physically binding to these receptors (directly-acting agents)
- the amount/life-time of ACh released from the post-gangilonic nerve terminal (indirectly-acting agents)
What happens to vascular smooth muscle if you rub away the endothelium and add ACh
no relaxation until the ACh is washed away
Which mAChR is mainly present in the eye and which parts of the eye
mainly M3 - constrictor pupillae; ciliary muscles; lacrimal glands
What does stimulation of M3 receptors in the constrictor pupillae lead to
contraction
shortens pupil diameter (miosis)
lowers intraocular pressure
What does stimulation of M3 receptors in the ciliary muscles lead to
contraction to reduce focal length of lenses, allowing accommodation
What does stimulation of M3 receptors in the lacrimal glands lead to
increased secretion
What is the distribution of mAChR in the detrusor muscle and mucosa/submucosa of the bladder
detrusor: M2=80%
M3=20%
(some beta 3 ARs present too)
mucosa and submucosa: M2 and M3
What is the PNS supply to the bladder
via pelvic nerve
What is the distribution of mAChRs throughout the lungs
pre-junctional terminal: M2 to reduce ACh release
postganglionic cell body: M1
airway smooth muscle cells: M2 and 3 for bronchoconstriction
at submucosal glands: M1, M3, to increase mucus secretion
How does the Vagus affect the lungs
tonically fires to contract (mainly via M3- PLC(beta)-IP3 mediated) airway smooth muscle during normal breathing - vagal tone
What effect does the vagus nerve have on the lungs when irritants are present
Who is this dangerous for
irritants stimulate sensory vagal afferents, activating a reflex PNS activity via the vagal efferents causing bronchospasm
it is major
bronchoconstricting factor for COPD patients
True or false:
It’s relatively easy to develop directly acting mAChR ligands that are subtype selective
False
No it is very challenging
the classical (orthosteric) binding site (= where ACh binds to) is highly conserved across mAChR subtypes
What have we used to discover that the ACH binding site is conserved across mAChRs
Analysis of amino acid sequence and recent X-ray crystallographic studies
What have we used to overcome the problems of the conserved binding site of mAChRs
these receptors possess distinct allosteric sites located often within the extracellular loops or the outer segments of different transmembrane (TM) helices.
Because these regions show a considerable degree of sequence variation across M1–M5 receptors, considerable progress has been made in developing subtype specific allosteric modulators.
How do allosteric modulators affect mAChRs
by altering the affinity or efficacy of orthosteric muscarinic ligands.
What are PAMs in relation to mAChRs
What is another related drug type
Positive allosteric modulators (PAMs)
enhance orthosteric activity,
negative allosteric modulators (NAMs) inhibit it
allosteric modulators have lead to development of new therapeutic agents with increased efficacy and reduced side effects. Name 2 diseases where this research is focussed
several severe disorders of the CNS, including Alzheimer disease and schizophrenia.
Other than allosteric modulators, what is another potential strategy for mAChR subtype selectivity
development of hybrid, bitopic ligands that interact with both the orthosteric binding cavity and an allosteric site
Why are hybrids that bind to both the orthosteric and allosteric sites of mAChRS being developed?
By targeting orthosteric and allosteric sites simultaneously, bitopic ligands can potentially achieve both high affinity and receptor subtype selectivity.
How can directly-acting parasympathomimetic or cholinomimetic agents (muscarinic agonists) be classified?
1) choline esters
2) naturally occurring cholinomimetic alkaloids
Name some choline esters
include ACh and synthetic esters of choline, such as carbachol (carbamylcholine) and bethanechol
Name 2 synthetic choline esters
carbachol (carbamylcholine) and bethanechol
name a cholinomimetic alkaloid
pilocarpine
True or false
. All of the direct-acting cholinergic drugs have a longer duration of action than ACh.
True
All of the direct-acting cholinergic drugs have a longer duration of action than ACh.
What is Bethanechol
What does it activate and where does it not have action?
chemically a hybrid of methacholine and carbachol
non-selectively activates mAChRs
lacks nicotinic action
What is the effect of bethanechol on the bladder
stimulates the detrusor muscle of the bladder (via M3 receptor), whereas the trigone and sphincter muscles are relaxed
ie produces urination
What is bethanechol licensed for
Is it very common?
for acute postoperative, postpartum and neurogenic urinary retention
its use has largely been superseded by catheterisation
What is pilocarpine used for
to treat glaucoma
What is the drug of choice for emergency lowering of intraocular pressure for glaucoma patients?
How is it administered?
pilocarpine
by instillation as eye drops and it is readily absorbed across the conjunctival membrane
How is pilocarpine absorbed
administered by instillation as eye drops and it is readily absorbed across the conjunctival membrane
What is the action of pilocarpine once it has been absorbed across the conjunctiva
contracts the ciliary muscle to produce traction on the trabecular meshwork around the Schlemm canal, causing an immediate drop in intraocular pressure as a result of the increased drainage of aqueous humour
How quickly does pilocarpine act
How long does it last
Is it repeatable?
action occurs within a few minutes
lasts 4 to 8 hours
can be repeated.
Why may patients who have taken pilocarpine struggle to focus their eyesight?
pilocarpine causes miosis, which is experienced as a spasm of accommodation and vision is fixed at a random distance
What is the effect of atropine on the eye
opposes the effects of pilocarpine as atropine is a muscarinic blocker
Name 2 diseases where pilocarpine is used to increase secretion
xerostomia
Sjögren’s syndrome
What is Sjögren’s syndrome
syndrome which is an autoimmune disease of glands that secrete fluid
What is xerostomia
chronic dry mouth problem, most common complications associated with radiotherapy of head and neck cancer)
How do pharmacological antagonists of mAChRs work
directly acting anti-cholinergic or cholinolytic agents
What site do competitive inhibitors of mAChRs block
Where are these receptors found?
the othosteric site
on effector cells at parasympathetic neuroeffector junctions in the peripheral ganglia (as well as the CNS).
True or false
mAChR competitive inhibitors can block SNS transmission
True - mostly acts on PNS but can act on cholinergic junctions eg sweat and salivary glands
Do mAChR competitive inhibitors affect AChRs
What does this mean
no
little or no action at NMJs or autonomic ganglia.
Describe the chemical structure of anti-muscarinic agents
contain an ester and basic groups like that of ACh but bulky aromatic group replacing the acetyl moiety
What kind of drug is scopolamine
Name another of this type
a naturally occurring alkaloid mAChR inhibitor
atropine (you need to know this one)
WHat are the 3 different types of anti-muscarinic agent
Name one of each
naturally occurring alkaloid (atropine)
natural alkaloid semisynthetic derivative (ipratropium)
synthetic agents (solifenacin)
Name 2 semi synthetic anti muscarinic agents
homatropine,
methscopolamine,
ipratropium
Name 5 synthetic muscarinic agents
tropicamide, cyclopentolate, pirenzepine, darifenacin, solifenacin
Give example of some relatively selective mAChR blockers (4)
What are most mAChR antagonists?
pirenzepine = M1
MT7= M1
darifenacin and solifenacin =M3
most are non selective
What is MT7
A toxin in the venom of the African green mamba that selectively targets M1.
What are mAChR blockers used for clinically
to inhibit effects of parasympathetic activity in the respiratory tract, eye, urinary tract, GI tract and heart
Which muscarinic blockers are used on the eye clinically
topically for mydriasis and cycloplegia in ophthalmic examinations
Would you want to use atropine for an ophthalmic examination?
Short-acting, relatively weak mydriatics, such as tropicamide (action lasts for 4–6 hours) and cyclopentolate (action up to 24 hours), are typically preferred over atropine (action may last up to 7 days).
What drugs would you use to make funduscopy easier (2)
What effects do they cause that make this beneficial
tropicamide
cyclopentolate
mydriasis
cycloplegia
What is used as the standard treatment of poisoning
atropine in large doses by irreversible, long lasting inhibition of AChE
What is the main effect of antimuscarinic agents on the heart
main effect is on rate - dominant effect= tachycardia
clinical doses may lead to transient bradycardia
How does the effect of atropine on the heart change as its dose changes
Larger doses of atropine cause progressive tachycardia by blocking M2 receptors on the SA nodal pacemaker cells, thereby antagonizing parasympathetic (vagal) tone to the heart
in which type of patient is the effect of atropine more profound
healthy young adults, in whom the vagal tone is considerable
What is atropine useful for clinically
to manage bradycardia associated with the excessive use of the ‘beta blockers’ and acute myocardial infarctions.
Which mAChR is involved in bronchoconstriction
What happens if these receptors are activated on lung epithelia
M3 on airway smooth muscle
mucous secretion
Describe a ‘vagally-mediated reflex bronchospasm’
lung M3 mAChR activation happens normally as a ‘vagal tone’ but many irritants (dust, allergens, histamine, some gases etc.) via local sensory nerves can make it more prominent in the form of a ‘vagally-mediated reflex bronchospas
What is the relevance of vagal tone in COPD
Is this the same in asthma
it is the major reversible brochoconstrictive component
no
What does M3 blockade in bronchial smooth muscle result in?
How does the effect of these blockers differ from a normal patient to a COPD patient
bronchorelaxation/ bronchodilation due to decreased muscle tone
minimal effect in normal airways but significant in COPD
What are the 2 categories of mAChR blockers used in COPD
Give an example of each
LAMA (long acting muscarinic antagonist) - tiotropium
SAMA (short acting muscarinic antagonist) - ipratropium
Name 2 LAMA drugs
tiotropium and glycopyrronium
True or false
All drugs in both SAMA and LAMA categories have higher selectivity for one type of mAChR
True
Higher selectivity for M3 than M2
LAMA also dissociate much faster from M2 than 3
Where does the longevity of LAMA action come from
their slow dissociation from M3 receptors
What kind of anti muscarinic agent can be used if asthma is not being adequately controlled by beta-AR stimulants
LAMA as additional bronchodilators
What do you use to treat urinary incontinence
Give 2 examples
How does this work
M3 selective antagonists
eg solifenacin and darifenacin
selective inhibition of M3 receptors that mediate contraction of bladder’s detrusor muscle
Which anti-muscarinic drug can be used to reduce excessive salivation
Which feature allows this
glycopyrronium
cannot cross BBB
Why might someone have the symptom of excessive drooling
drug induced or associated with heavy-metal poisoning and Parkinson’s disease
What are the 2 types of cholinesterase
Describe the specificity of each
BuChE (AKA pseudocholinesterase)
AChE
AChE = only ACh BuChE= much broader
Name 2 drugs that BuChE can break down
procaine (a local anaesthetic)
suxamethonium (depolarising NMJ blocker)
Where is BuChE made
Where can it be found in the body (4)
liver
mostly blood
traces in: skin, gut, and brain
Where is AChE found (2) and what form does it take in these locations
both in the presynaptic nerve ending (as a soluble form in the cytoplasm) and in the synaptic cleft (as tethered to the postsynaptic neuronal membrane/junctional fold).
What is the presynaptic purpose of AChE (2)
guards against the accumulation of cytoplasmic ACh outside storage vesicles
termination of ACh action
How is AChE attached to the post synaptic terminal
3 tetramers of AChE are attached by disulphide bonds to a long collagenous tail which tethers the enzyme to the postsynaptic/post-junctional target membrane.
Where in the synpase is the AChE primarily responsible for ending ACh action ?
membrane-bound oligomeric form of AChE
What is the turnover of AChE
> 10,000 mol per active site per second
What kind of enzyme are the cholinesterases?
serine hydrolases
What part of the Ser group in Ser hydrolases performs the nucleophilic attack
What is the purpose of this attack
hydroxyl (-OH) group
An intermediary adduct with the substrate is formed, which then undergoes hydrolysis
What is the 3D structure of AChE
Where is the catalytic site? Give measurements and a specific name
hourglass
at the base of a deep (~about 20Å from the surface) and narrow (~ 5Å wide) tunnel named the aromatic gorge.
What is the peripheral anionic site?
What is the purpose of this?
the entrance too the aromatic gorge of AChE
recognition of ACh
What is one of the key interaction s of the peripheral anionic site
a weak cation-pi interaction between the heteroaromatic ring of a particular tryptophan residue (W279 for mammalian AChE) and the charged quaternary nitrogen of ACh
What happens after the peripheral anionic site recognizes ACh
transferred down the aromatic gorge to the active site
How is the ACh helped down the aromatic gorge after being recognised by the peripheral anionic site?
14 conserved aromatic residues line the gorge and help in pushing the ACh downwards towards the catalytic centre through cation-pi interactions with ACh. An electrostatic gradient running down the gorge also favours such movement of ACh.
What happens to ACh when it reaches the end of the aromatic gorge
to the ‘catalytic anionic site’ (CAS) which helps in properly positioning ACh against the catalytic triad consisting of specific glutamate, histidine and serine
Describe the tetrahedral intermediate formed by nucleophilic attack of Ser on ACh in AChE (a Ser hydrolyase)
has a charged oxygen
intermediate is stabilised by backbone amide groups (oxyanion hole)
from this the ACh bond is broke, releasing choline, and leaving acetyl-serine behind
After choline has been released from AChE, an acetyl serine is left. What happens next
The serine¬-acetate bond is then spontaneously hydrolysed by water to release acetate and regenerate the enzyme
How fast is the action of AChE
one of the fastest known : >10^4 molecules of ACh are hydrolyzed per second by a single AChE molecule, requiring only about 100microsec.
What are anticholinesterases also referred to as
as indirectly acting cholinergic/cholinomimetic agents because they generally do not activate receptors directly
Why are anti-AChE agents often used as toxins
Give a famous example of how they were used recently
cholinergic synapses are widely distributed throughout the body
deployment of nerve gas (novichok) in an attempt to assassinate victims in Salisbury in 2018
What is a necessary feature of an anti-AChE agent used to treat Alzheimer’s
Give 2 examples
can cross BBB
rivastigmine
donepizil
How can you categorise anti-AChE agents by:
a) mechanism of inhibition
b) duration of action
Give examples
a) • Reversible: a) non-covalent (edrophonium) b) covalent inhibitors (physostigmine, neostigmine etc)
• Irreversible - organophosphates such as dyflos, parathion, ecothiophate
b) • Short acting (edrophonium)
• Intermediate acting (the ‘stigmines’)
• Long acting (the organophosphates
What is edrophonium
Non-covalent, reversible, short acting anticholinesterase
Describe the structure of edrophonium
Where can it act
a quaternary ammonium compound
restricted to the periphery
Name a non-covalent, reversible, short acting anticholinesterases and describe how it interacts with AChE
edrophonium
interacts only to the catalytic anionic site (CAS) of the enzyme, primarily through a cation- π interaction between its quaternary nitrogen and the aromatic (indole) ring of a tryptophan (W84) residue of AChE
the reversible nature of this interaction explains the short duration of edrophonium action
What is the duration of edrophonium action
2-10 min
What is edrophonium used for
…
the tensilon test -
in order to diagnose myasthenia gravis
HOWEVER:
Edrophonium is discontinued in the UK as its use as a diagnostic tool in myasthenia gravis has been largely superseded by antibody testing and the use of electrical conduction studies.
What does a positive result in the tensilon test look like?
Mention drug used and illness diagnosed
A myasthenia positive individual characteristically feels temporary improvement in the facial weakness and ptosis within 5-10min of edrophonium injection
Describe Covalent, reversible, medium acting anti-cholinesterases as a group
Give 4 examples
esters of carbamic acid (instead of acetic acid as with ACh) and have the suffix ‘stigmine’ and include physostigmine, neostigmine, pyridostigmine and rivastigmine.
All except rivastigmine are charged and therefore act only peripherally.
How does neostigmine interact with AChE?
What kind of drug is this
With their basic group, they interact with the anionic site of AChE and transfer carbamyl group to the hydroxy group of Ser 203. Carbamylated AChE is more stable than acetylated AChE and takes minutes to hydrolyze. Hence these are medium acting agents.
Covalent, reversible, medium acting anti-cholinesterases
How and when is neostigmine administered
intravenously to reverse neuromuscular blockade after surgery and orally to treat myasthenia gravis
How is pyridostigmine administered
When
Why this drug
orally
myasthenia gravis
longer action than neostigmine
How is ACh relevant to Alzheimer’s
there is significant loss of cholinergic neurons in the basal forebrain and the resultant reduced cholinergic neurotransmission is considered to underlie, at least in part, the dementia, intellectual deterioration, and personality changes associated with AD.
Describe irreversible, long acting anti-cholinesterases as a chemical group
organophosphorous compounds containing a pentavalent phosphorous atom connected to three oxygen atoms (sometimes two oxygen and one sulfur atoms) along with a labile group, such as fluoride (dyflos or DFP) or an organic group (ecothiophate, parathion)
How do irreversible, long acting anti-AChE work
labile group released and Ser’s -OH phosphorylated
this is v stable and lasts hundreds of hours
How is the AChE-organophosphate complex following administration of irreversible long acting anti-AChE drugs
subject to a process known as ‘aging’, in which oxygen–phosphorus bonds within the inhibitor undergo spontaneous rearrangement in favour of stronger bonding between the enzyme and the inhibitor. Once aging occurs, the duration of AChE inhibition is increased even further.
Thus, organophosphate inhibition is essentially irreversible, and the body must synthesize new AChE proteins to restore AChE activity.
How are irreversible, long acting anti-AChE used clinically
Use specific examples
What other uses do they have
limited clinical uses
Ecothiophate- used locally in glaucoma
insecticides
chemical warfare nerve gases eg sarin and novichok
Name 4 nerve gases
What kind of drug is this
sarin, tabun, soman, and novichok
irreversible, long-lasting anti-AChE
What does acute poisoning by irreversible anti-AChEs result in
When does it become fatal
excessive salivation, severe bradycardia, hypotension and difficulty in breathing
when combined with a depolarising neuromuscular block and central effects
How do you treat anti-AChE poisoning
large doses of atropine as it can reach brain and periphery
What is the effect of atropine of peripheral NMJ compromise
what other drug is used in a compromised NMJ
no effect
pralidoxime (2-PAM, a AChE reactivator) must be used instead
What is 2-PAM used for
How does it work
to reactivate AChE
cationic group of pralidoxime interacts with the anionic site of AChE, which brings the oxime group into close proximity with the phosphorylated serine. The oxime group is a very strong nucleophile and lures the phosphate group away from the Ser hydroxyl of the enzyme
How is the AChE reactivation by 2-PAM limited
Give the chemistry behind this
effectiveness is limited to within a few hours of exposure
phosphorylated AChE undergoes ageing, rendering phosphorylated groups no longer susceptible to nucleophilic attack
Name 3 catecholamines
Why are they known as catecholamines#?
noradrenaline
adrenaline
dopamine
they possess a catechol and amine connected by a 2 carbon linkage
How is the 2 carbon linkage in dopamine different to the other catecholamines
NA, Adr, and ISO all have an alcoholic -OH on the beta C (the C closest to the catechol)
DA does not
What is a catechol
What is an amine group
catechol: aromatic ring with two proximal hydroxyl groups
amine: -NHR, where R = H or alkyl group
What is the difference between noradrenaline and Adr
NA is effectively Adr lacking the methyl on the amine
What is the main NT in the SNS
NA
What is the precursor of NA?
Where does this precursor have important action
dopamine
mainly in the CNS and dopaminergic nerves
How does Adr mainly act
Where is it made
as a hormone
adrenal glands
True or false
NA is effectively the N-methylated version of Adr
False
Adr is effectively N-methylated NA
How could you convert NA to Adr
Where does this take place
methylation
place in the chromaffin cells of adrenal medulla in presence of the enzyme PNMT (phenylethanolamine N-methyltransferase
How much of the catecholamines in the adrenal medulla are Adr
80%=Adr
20%= NA
What is ISO
isoprenaline a synthetic (= N-isopropyl) derivative of NA, often used experimentally.
How does ISO differ from NA
amine group on NA is just -NH2
ISO: -NH-CH-(CH3)2
What is the starting material for biosynthesis of catecholamines? (Give the specific type)
How is this taken to to form catecholamines?
L-tyrosine
present in bodily fluid and taken up from the circulation by catecholaminergic nerves and converted into DOPA (dihydroxyphenylalanine) by tyrosine hydroxylase (TOH)
What is the rate limiting step in catecholamine biosynthesis
conversion of L-tyrosine to DOPA by tyrosine hydroxylase (TOH)
Where is TOH found
ONLY in catecholaminergic nerves
Give each step of Adr synthesis with the enzyme for each equation (5)
- L-tyrosine
enzyme: TOH - DOPA
Enzyme: DOPA decarboxylase - Dopamine
enzyme: Dopamine beta-hydroxylase - NA
enzyme: PNMT
5.Adr
what is DOPA decarboxylase also known as?
Why?
L-aromatic acid decarboxylase
it’s relatively non-selective and catalyzes decarboxylation of other L-aromatic acids such as L-histidine or L-tryptophan to produce histamine and serotonin (5-HT), respectively
What is the effect of L-aromatic acid decarboxylase on the following:
a) L-histidine
b) L- tryptophan
converts them to
a) histamine
b) 5-HT
What happens to dopamine after it is formed (not conversion to NA)
What drives this process
stored into vesicles via the vesicular monoamine transporter (VMAT)
transvesicular H+ gradient
What happens to dopamine inside the vesicles
hydroxylated at its β-carbon to form NA and this is catalyzed by dopamine β-hydroxylase (DBH).
Where can DBH be found
How do you stop DB being released with NA when NA vesicles fuse with presynaptic membrane
exclusively within noradrenergic nerves
you don’t - small amounts of DBH are co-released with NA
What happens to released DBH
What can we use this as an indicator for
not rapidly degraded or susceptible to any uptake mechanism, so its concentration in plasma and body fluids can be used as an index of overall sympathetic nerve activity.
Where is PNMT mainly expressed in the adrenal medulla
in the adrenaline-releasing (A) cells that are distinct from the smaller proportion of noradrenaline-releasing (N) cells
What controls PNMT expression
glucocorticoids eg cortisol from adrenal cortex
How does cortisol affect catecholamine production (2)
Reduction in cortisol levels causes a decrease in production of PNMT and, hence, in the Adr:NA ratio.
also increases activity of dopamine β-hydroxylase (DBH).
Which pharmalogical agent is used experimentally to block NA production
α-Methyltyrosine -
a competitive inhibitor of TOH
What is the importance of blocking TOH as a way of reducing amount of NT produced?
it is the rate limiting step using α-Methyltyrosine is the only effective way to reduce amount of catecholamine produced
Give an example of the clinical use of α-Methyltyrosine
in the past was used pre-op before treatment for pheochromocytoma (adrenal gland tumour, increasing Adr and NA secretion)
What is the most effective way to increase production of catecholamines pharmacologically
bypass the step catalysed by TOH through using substrate for one of the later enzymes
What happen in Parkinson’s disease
dopaminergic neurons in the substantia nigra pars compacta (SNc) are selectively degenerated
Why can we not just use exogenous DA to replace that which is lost by destruction of dopaminergic neurons in Parkinson’s?
What is used instead?
DA does not cross the BBB
L-DOPA, DA’s natural precursor, can cross
Therefore L-DOPA is used to boost DA synthesis in the Parkinsonian brain
What accounts for the unwanted peripheral effects of L DOPA therapy
A substantial fraction of the administered L-DOPA undergoes decarboxylation in the periphery by peripheral decarboxylase to produce DA and NA
How to overcome the peripheral side effects of L-DOPA therapy
use in combination with carbidopa
carbidopa cannot cross the BBB but can inhibit peripheral decarboxylases
this ensures L-DOPA conversion to DA only occurs in CNS
Which drug is used to inhibit DBH
How is it used clinically
Disulfiram
as an adjunct in the treatment of alcohol dependence
How does disulfiram work
What is the basis of its action in dealing with alcoholism?
act by chelating the Cu2+ ion which is an essential co-factor of DBH or it may attack the sulphur-handling system for the methyl donor, S-adenosyl methionine
inhibition of aldehyde dehydrogenase
How selective are the enzymes in the biosynthesis of catecholamine biosynthesis
What does this mean
TOH- selective
DDC and DBH - non selective
can result in the production and storage of a variety of other amines in the synaptic vesicle
What happens when α-Methyldopa is taken into noradrenergic nerve terminals
converted successively into α-methyldopamine (α-methyl DA), and α-methylnoradrenaline (α-methyl NA)
What happens to α-methyl NA in nerve terminals
stored within synaptic vesicles and released with NA as a false transmitter
Describe the action of α-methyl NA compared to NA
functional but less active than NA at alpha 1 ARs and more selective towards alpha 2
Name 2 drugs that target alpha 2 more selectively than NA does normally
What do they both result in? How?
clonidine
α-methyl NA
decrease BP
inhibition of NA release (via presynaptic alpha2 AR stimulation) and central action
What kind of vesicle is NA usually stored in
What is often stored with it
clear core
ATP
what transports NA and DA into vesicles
What family is it part of
What does it use to power the active transport?
VMAT2 (vesicular monoamine transporter)
SLC protein family
trans-vesicular H+ gradient
How many H+ are transported per amine by the VMAT2
2H+ extruded per amine imported into the vesicle
How is the H+ gradient in catecholamine vesicles set up
up by an ATP-dependent proton pump (a v-ATPase).
What is VMAT1 also known as
Where is it found
chromaffin granule amine transporter
in non-neural tissues including the chromaffin cells of the adrenal medulla and few other cells of the GI tract.
What is reserpine
What does it do
– a naturally occurring alkaloid (not used clinically),
binds to the amine site of the VMAT2 with high affinity and irreversibly blocks the uptake of monoamines (DA, NA and 5-HT) into vesicles
What does reserpine’s action eventually lead to
Why does this happen
long lasting depletion of stored NA (and 5-HT in the CNS)
vesicles do allow leakage of the stored catecholamines into the cytoplasm where it is metabolised
What was reserpine used for originally
Why was it discontinued?
as an anti-hypertensive
lead to profound psychological depression (due to decreased central 5-HT)
Have any VMAT2 inhibitors been approved by the FDA
What are they approved for
a few - namely tetrabenazine and valbenazine
to manage abnormal involuntary movements associated with Huntington’s disease, tardive dyskinesia etc
What do the 2 VMAT2 inhibitors have in common
both pro-drugs
upon in vivo metabolism, produce the active metabolite alpha-dihydrotetrabenazine (DTBZ)
How does DTBZ differ from reserpine, meaning it is safer
Which monoamine is most affected
DTBZ REVERSIBLY inhibits VMAT2 unlike reserpine
DA
How can you alter release of catecholamines
1) direct blockade of catecholaminergic neurons
2) triggering release in absence of any depolarisation (indirectly-acting and mixed-acting sympathomimetic drugs)
3) through stimulating pre-synaptic receptors.
4) alter storage
Give an agent that directly blocks adrenergic neurons by acting on vesicles in the presynaptic terminal
Guanethidine
How does guanethidine enter noradrenergic neurons
accumulates selectively into noradrenergic nerves by Uptake 1/NET
Is guanethidine selective?
yes - complex, selective action on noradrenergic nerves - not fully understood
How does guanethidine action vary based on dosage
At low doses, guanethidine blocks impulse conduction in these nerves, like the local anesthetics. It also accumulates into the synaptic vesicles via VMAT2, causing gradual, long-lasting depletion of NA (like reserpine).
At a very high dose, it irreversibly damages the nerve, effectively serving as a neurotoxin selective to noradrenergic neurons
What happens to guanethidine at low doses
guanethidine blocks impulse conduction in these nerves, like the local anesthetics. It also accumulates into the synaptic vesicles via VMAT2, causing gradual, long-lasting depletion of NA (like reserpine).
What happens to guanethidine at very high doses
irreversibly damages the nerve, effectively serving as a neurotoxin selective to noradrenergic neurons
What was guanethidine used treat
uncontrolled hypertension
no longer recommended in the UK
What is tyramine (2)
an indirectly acting sympathomimetic
a dietary amine
Name an Indirectly acting sympathomimetic (NOT tyramine)
How does it act generally
dexamphetamine
capable of releasing stored transmitter from noradrenergic nerve endings by a Ca2+-independent process.
Are tyramine and dexamphetamine AR agonists ?
NO - they are substrates for monoamide transporters
How are tyramine and dexamphetamine taken up into nerves
What happens to them inside the nerve
How is this helpful?
are avidly taken up into noradrenergic nerve endings by Uptake1/NET
loaded into vesicles by VMAT2, displacing NA
NA is instead expelled into synaptic cleft by reverse NET action
When can a tyramine rich diet become a serious problem
Tyramine-riched food intake in presence of MAO inhibition can lead to severe hypertensive crisis
What kind of drugs are MDMA and ecstasy
Indirectly acting sympathomimetics
they are CNS stimulants, often abused for recreational purposes
Name a mixed acting sympathomimetic amine
How do they work generally
ephedrine
indirectly release NE like tyramine and dexamphetamine but they also can directly activate adrenoreceptors (ARs)
How is ephedrine used usually
as a nasal decongestant as it results in NA-mediated vasoconstriction in some blood vessels to the nose
What does ephedrine act on
some direct action on bronchial β2-ARs and thus can help in relaxing the airways.
Also used to treat urinary incontinence in female dogs after the surgical removal of their reproductive organs.
What effect do tyramine, ephedrine, amphetamine, and related drugs cause?
How does this differ from reserpine action
a relatively rapid and brief liberation of the transmitter, producing a sympathomimetic effect indirectly
reserpine, by irreversibly blocking the loading of monoamines into synaptic vesicles, produces a slow, prolonged depletion of NA (and other monoamines) from the presynaptic nerves.
Through which presynaptic receptor is NA release most prominently regulated
α2-adrenoceptors (coupled to Gi/o) which, when stimulated, decrease the amount of NA released
Is it only presynaptic α2-adrenoceptors that modulate NA release
no
there can be pre-synaptically located β2-ARs (coupled to Gs) which upon stimulation, enhance NA release
How does cAMP affect NA release
increased cAMP production increases Ca2+ channel activating phosphorylation by PKA - more NA released
Other than decreasing AC activity via the Gi/o, how does activating presynaptic alpha2-ARs on noradrenergic nerves decrease NA release
beta-gamma subunit, also results in the opening of K+ channels (GIRK)s which causes hyperpolarisation of the nerve terminal membrane and reduces its excitability
Are the presynaptic the predominant α2-ARs and the occasional β2-ARs autoreceptors?
yes usually as released NA interacts with them
However, In some cases, there can be pre-synaptic heteroreceptors (e.g. muscarinic M2 receptors and δ-opioid receptors) which upon activation by cognate agonists, decrease NA release.
How are catecholamines inactivated?
What are the 2 processes involved?
Which process is more important pharmacologically
removal from cleft
one into the presynaptic neuron (Uptake 1) and the other into postjunctional effector cells (Uptake 2).
Uptake 1: inhibiting this process enhances and prolongs the actions of released NA whereas inhibiting Uptake 2 does not affect the response.
Describe Uptake 1 in noradrenergic synpases
This is a high affinity, low capacity system mediated by a transporter called NET (norepinephrine transport protein).
What is NET
a member of the solute carrier (SLC) family of transporters. It is Na+-dependent, and blocked by many drugs including cocaine
What kind of drugs block NET
cocaine
tricyclic antidepressants eg imipramine
Name a tricyclic antidepresant
imipramine
How much of the NA released from SNS neurons is recaptured by NET
> 75%
VMAT2 has a much higher affinity for NA than does MAO. What does this mean for NA storage
most of the recaptured NE is resequestered into storage vesicles.
What is in charge of transmission from noradrenergic neurons in the periphery
Uptake 1
What is 6-OHDA
6-Hydroxydopamine (AKA oxidopamine)
a synthetic neurotoxic organic compound that is experimentally used (injected into brain areas) to selectivity destroy dopaminergic (and noradrenergic) neurons in the brain.
How do you induce Parkinson’s disease in animals
inject oxidopamine which induces nigrostriatal damage
How selective is 6-OHDA
Where does this come from
selective for catecholaminergic
its selective uptake into these neurons via NET
What happens to oxidopamine once it has been taken up into the noradrenergic neuron via NET
it is readily oxidized into reactive metabolites including 6-OHDA quinone and hydrogen peroxide that damage to the neurones
Describe Uptake 2 at noradrenergic synapses
lower affinity, higher capacity system of uptake of NA that hasnt been collected by Uptake 1
What transporter mediates Uptake 2
ENT: extraneuronal amine transporter
What transporter family does ENT belong to
to a largely and widely distributed family of Organic Cation Transporters (OCTs).
Which of Uptake 1 and 2 is Na+ dependent
What does this mean
Uptake 1
ENT in Uptake 2 is not Na+ dependent (unlike NET)
they have different pharmacological profiles
What inhibits ENT
the irreversible α-AR blocking agent - phenoxybenzamine (which will also block NET at much higher concentrations).
normetanephrine, a metabolite of NA, corticosteroids (such as corticosterone and hydrocortisone)
Name a irreversible α-AR blocking agent of ENT
What is its effect on NET
phenoxybenzamine
will also block NET at much higher concentrations
Where does most metabolism of catecholamines take place?
Why?
in the same cells where the amines are synthesized and stored
There is ongoing passive leakage of catecholamines from vesicular storage granules of sympathetic neurons and adrenal medullary chromaffin cells
What happens to most of the catecholamines that leak into the neuron cytoplasm?
What happens to the rest
VMAT2 effectively sequesters 90% of the amines leaking back into storage vesicles
10% escapes sequestration and is metabolized.
Which 2 enzymes are responsible for catecholamine metabolism
monoamine oxidase (MAO) and catechol O-methyltransferase (COMT)
How should MAO and COMT work in theory vs practice
Although these enzymes in principle can act on the catecholamines in any order and the final metabolites should be the same but, in practice, two different metabolites (VMA and MOPEG) are found in the urine
VMA= from periphery MOPEG= from CNS
What do the following stand for:
VMA
COPEG
vanillylmandelic acid
3-methoxy-4-hydroxyphenylethyleneglycol
How does MAO work
removes the amine group from the catecholamines and converts them into corresponding aldehydes, which, in the periphery, are rapidly metabolised into the corresponding carboxylic acids.
Where does MAO action occur
in the outer membrane of mitochondria and it is abundant in noradrenergic nerve terminals but is also present in liver, intestinal epithelium and other tissue
What is the main role of MAO in
a) nerve terminals
b) liver
a) degrade NA and DA leaking from vesicles
b) inactivates circulating monoamines such as tyramine
What are the 2 isoforms of MAO
How do they differ
MAO-A and -B
MAO-A degrades 5-HT, NA and DA, while MAO-B degrades DA more rapidly than the other monoamines.
How does pharmacological treatment aimed at MAO differ between isoforms
Non-selective or selective MAO-A inhibitors are used in depression and anxiety disorders whilst selective MAO-B inhibitors is used in the treatment of Parkinson’s disease.
Why might there be an increase in blood pressure if you are eating lots of cheese and wine while on MAO inhibitors
What is this known as
Cheese and red wine contain tyramine, which is usually metabolised by MAO in liver/gut before reaching circulation
a large amount of tyramine-rich food is ingested, then significantly higher amount of tyramine reaches the blood and then can trigger release of NA. The latter can be sufficient to cause widespread vasoconstriction and a fatal hypertension
The cheese effect
What other foods can the ‘cheese effect’ result from
tyramine rich foods: red wines, pickled herring, yeast extracts, soya beans
Where is COMT found
in cytosol, primarily liver and chromaffin cells (adrenal medulla)
ABSENT FROM NORADRENERGIC NERVE CELLS
is COMT free in the cytosol of chromaffin cells
no - it is membrane
bound.
This is also true of COMT in the liver cells
what does COMT do chemically ?
methylates one of the aromatic hydroxyls (at position 3 of the aromatic ring) in the catecholamines (or their deaminated metabolites produced by MAO).
What is the main end product of catecholamine metabolism by MAO then COMT
How is it excreted
How is this fact used clinically
VMA
via urine
in phaeochromocytoma VMA excretion is markedly increased
What is used as an inhibitor of COMT
What is this used for
Entacapone
used in Parkinson’s disease in combination with L-DOPA and Carbidopa
Why is entacapone used with L-DOPA to treat Parkinson’s disease
. It reduces metabolism of L-DOPA in the periphery so that more L-DOPA could reach the CNS where it is needed to produce DA for the Parkinson’s patient and less side effects occur from activation of dopamine receptors present in various peripheral tissues.
How does ISO affect the different ARs differently?
it barely activates α adrenoceptor subtypes but it possesses significantly higher potency at β adrenoceptor subtypes compared to Adr and NA.
Give 9 major effects of stimulation of α1-ARs
- Dilatation of pupils (mydriasis)
- Constriction of blood vessels supplied to vicera, brain and skin
- Salivary secretion
- Decreased gastrointestinal motility and tone (due to relaxation of smooth muscles)
- Localized (e.g. in palms of hands) secretion of sweat
- Piloerection (goose bumps)
- Contraction of trigone and sphincter muscles of urinary bladder
- Ejaculation of sperms
- Increased glycogenolysis and gluconeogenesis
Give 4 effects of α2 AR stimulation
- Inhibition of neurotransmitter release (mentioned earlier)
- Platelet aggregation
- vasoconstriction
- Inhibition of insulin release from pancreatic beta cell
Give 2 results of β1 AR stimulation
- Increased force of myocardial contraction
* Increased heart rate
Give 4 results of β2 AR stimulation
- Relaxation of tracheal and bronchial smooth muscle
- Relaxation of smooth muscle of blood vessels supplied to heart and skeletal muscle
- Relaxation of uterine smooth muscle (for non-pregnants)
- Hepatic glycogenolysis
Give 2 actions of β3 AR stimulation
- lipolysis
- thermogenesis
This is the least studied thus far
What are the key effects of NA
Via which receptor?
Through α1 AR stimulation, Increases TPR due to vasoconstriction in most vascular beds (including kidney)
Both diastolic and systolic pressures increase enough to trigger reflex bradycardia which decreases pulse rate
Which receptors does adrenaline activate and what is this attributed to
α and β -ARs
The present of a missile group at the amino N atom
What are the 7 broad types of reaction caused by catecholamines or sympathomimetics
- A peripheral excitatory (= contractile) action on certain types of smooth muscle, such as those in blood vessels supplying skin, kidney, and mucous membranes; and on gland cells, such as those in salivary and sweat glands.
- A peripheral inhibitory (= relaxing) action on certain other types of smooth muscle, such as those in the wall of the gut, in the bronchial tree, and in blood vessels supplying skeletal muscle (we need to breathe more, our leg muscles need to have more blood supply to have energy for ‘fight or flight’).
- A cardiac excitatory action that increases heart rate and force of contraction. (remember what happens if we are exposed to stress/danger or we simply do some exercise)
- Metabolic actions that include enhanced glycogen breakdown in liver and muscle and liberation of free fatty acids from adipose tissue (remember the benefits of exercise which corresponds to increased sympathetic activity, we also need more energy for ‘fight or flight’).
- Endocrine actions, such as modulation (increasing or decreasing) of the secretion of insulin, renin, and pituitary hormones.
- Actions in the central nervous system (CNS), such as respiratory stimulation, an increase in wakefulness and psychomotor activity, and a reduction in appetite.
- Prejunctional actions that either inhibit or facilitate the release of neurotransmitters, the inhibitory action being physiologically more important
How does NA predominantly act (ie which receptors)
Why is this
NA predominantly acts via apha-ARs
and this is attributed to the presence of a methyl (-CH3) group at the amino N atom.
What is the effect of Adr when it activates β1-ARs
Positive inotropic and chronotropic affects (thus increase pulse rate)
What is the effect of Adr stimulating β2 ARs
Compensates for constriction of arterioles in skin, mucus membranes, and viscera by Faisel dilation of skeletal muscle vascular bed which also reduces TPR
What are the α1 mediated effects of Adr
Construction of arterioles in the skin, mucus membranes, and viscera
What is the net effect of adrenaline stimulating ARs
Increase in systolic pressure coupled with a slight decrease in diastolic pressure
True or false
Isoprenaline only manifests α1 AR mediated effects
False
Only manifests β-AR effects
This selectivity towards β adrenoreceptors stems from
A bulkier substituent (isopropyl) at the amino N atom
What are the effects of isoprenaline? Describe the effects through the receptors which are stimulated
What is the net effect
β1 AR: Intense stimulation of the heart, increasing rate, contractility, and cardiac output
β2: Delete arterioles of the skeletal muscle, resulting in marked decrease in TPR
Because of its cardiac stimulatory action, may increase systolic pressure slightly, but this is outweighed by reduction in TPR with a net reduction in mean arterial and diastolic blood pressures
What are adrenoceptor agonists also known ask
Directly acting sympathomimetic agents
What do non-selective AR agonists act on
Both α and β ARs
Give a non-selective adrenoreceptor agonist
When is this used
Adrenaline
Acute cardiac failure, acute severe asthma or anaphylactic shock. In all these cases it can be life-saving
Carried in an Epipen
Adr is also used by anaesthetists
What is status astmaticus
Acute severe asthma
How is adrenaline given in cases of anaphylactic shock
What does the Adr do (2)
Intramuscularly,
To counteract the systematic vasodilation and reduction in test you perfusion that is largely caused by massive histamine release
Also acts on bronchial smooth-muscle to relieve bronchospasm
How do you anaesthetists use adrenaline
Local anaesthetic preparations often contain low concentrations of adrenaline which greatly increases the duration of local anaesthesia by producing vasoconstriction at the sight of injection
Allows local anaesthetic to persist at the injection site before being absorbed into systemic circulation
Keeping anaesthesia local avoids systemic effects
What do selective α-AR agonists do
Selective for α ARs over β
But poorly discriminate between α-ARs
Name a selective α AR agonist
How is it administered
What is it used for
Xylometazoline
Topically
To relieve nasal congestion
How does xylometazoline relieve nasal congestion
Causes vasodilation in nasal mucosa – dilated blood vessels in nasal mucosa is an underlying feature of stuffy nose
Name a selective α1 agonist
What is it used for (3)
Phenylephrine (oxymetazoline is also one)
To treat nasal congestion, to produce mydriasis, and treat acute hypotension (eg in septic shock)
Name 2 α2 selective AR agonists
What are they used for
Clonidine and α-methylNA (from α-methyldopa)
Anti-hypertensives
How do clonidine and α-methylNA reduce BP
Partly by inhibiting NA release from peripheral nerve endings
Also on CNS neurons that reduces SNS discharge to periphery
What receptor, other than α2-AR, does clonidine bind to
What is the consequence
I1 receptors
Unknown
What is an example of an α2-AR agonist used in veterinary medicine
How is it used
Xylazine
As a sedative with analgesic and muscle relaxant properties
What is a non selective β AR agonist
What is it used for
What was the issue
What has replaced it
ISO
In asthma to relax bronchi through actions on β2- adrenoreceptors but
But increased heart rate from β1- AR stimulation was a major problem
β2-AR selective agonists
Name a β1 AR selective agonist
What is it used for
What is the issue
Dobutamine
In cases of cardiogenic shock to increase cardiac output
All β1-AR agonists can cause cardiac arrhythmias
Name 2 short acting β2 AR selective agonists (SABA)
What are they used for
Salbutamol (need to know)
Terbutaline
Mainly for bronchodilator effects in asthma
What is salbutamol
How is taken usually
How long does it take for maximum effect
How long does it last
What else can it be used for
The blue puff asthma inhaler
Inhaled as needed
Within 30 mins
3-5 hours
Prophylactically against allergen and exercise induced bronchospasm
Name 2 long acting β2 agonists
Which do you NEED to know
How long counts as long lasting
How are they used
Salmeterol (need to know)
Formoterol
8-12 hours
Used prophylactically in chronic asthma
What is clenbuterol
A horse LABA
What kind of drug that acts on ARs is used to treat COPD
Eg
LABAs
Especially very long lasting ones like indacaterol
What is mirabegron
A β3- AR agonist that relaxes detrusor smooth muscle to increase bladder capacity
Who would take mirabegron
Patients with an overactive bladder
Give 2 other names for AR antagonists
How are they classified
Adrenergic blockers or sympatholytics
Based on relative affinity to α or β ARs in the SNS
Name 2 α-AR antagonists that do not discriminate between α subtype (which do you need to know✅)
What did they treat but are now obsolete for? Why!
Phentolamine
Phenoxybenzamine ✅
Anti hypertensives (largely replaced by α1 antagonists) - as they may reduce blood pressure so much that it triggers reflex tachycardia
What does phentoxybenzamine do chemically
What is its specific clinical use therefore
Covalently binds to alpha-AR, causing long lasting inhibition
Preparing patients with phaeochromocytoma for surgery since surgical manipulation tends to cause massive catecholamine release
What is prazosin
What does it do
So what is it used for
Selective and competitive α1-AR antagonist
decreases TPR and BP by relaxing arterial and venous smooth muscle
as an anti-hypertensive without causing the same degree of reflex tachycardia observed with the non-selective α-AR antagonists
What are the negative side effects of α1-AR antagonists
Give an explanation
postural hypotension and incontinence, the latter occurring because α1-ARs mediate contraction of the smooth muscle of the bladder, which is now inhibited
Name 2 α1-AR antagonists (need to know both)
prazosin
tamsulosin
How are α1-AR antagonists used to treat some prostate pathologies
cause relaxation of the prostate capsule and inhibit prostate hypertrophy, which has lead to the development of drugs like tamsulosin. These are selective α1A-AR antagonists that allow better bladder emptying and thus reducing urinary retention associated with benign prostatic hypertrophy
What is tamsulosin
a selective α1A-AR antagonists
What is the benefit of selective α1A-AR antagonists
Since α1B-ARs largely mediate vasoconstriction, these α1A-AR antagonists produce much less postural hypotension which is a common problem with non-selective and α1-selective AR antagonists.
What is Yohimbine
What is it useful for
a naturally occurring alkaloid, is a selective α2-AR selective antagonist
no clinical use for the humans (but is in animals), being only a useful experimental tool to determine α-AR subtype response in a given tissue
Name a a naturally occurring alkaloid, is a selective α2-AR selective antagonist
yohimbine
True or false
All of the clinically available β-blockers are competitive antagonists for the beta-AR
true
What do cardioselective beta blockers primarily block
β1 -ARs.
What is the clinical use of β2 -AR blockers
there arent any
Do beta blockers cause postural hypotension?
What does this mean?
no
because the α ARs remain unaffected.
Therefore, normal sympathetic control of the vasculature is maintained
What do all beta blockers result in
What is the effect on blood pressure
negative inotropic and chronotropic effects (block β1-ARs)
decrease blood pressure in hypertensive patients, but have no effect in normotensive individuals
What contributes to the antihypertensive effects of beta blockers
decreases in peripheral vascular resistance and cardiac output both contribute to the antihypertensive effect of these drugs observed in patients with high blood pressure.
Name 7 general conditions that beta blockers can be used in
primarily indicated in hypertension and angina but can be useful in cardiac arrhythmias, myocardial infarction, heart failure, hyperthyroidism, and glaucoma
How to recognise a beta blocker name
exceptions?
ends in -olol
except for labetalol and carvedilol
What is the prototypical β blocker
Describe its selectivity
What is the effect
Propranolol
Non selective among β ARs.
Reduces hypertension
What are the mechanisms of propanolol that decrease hypertension (4)
Decreased cardiac output
Inhibition of renin release
Decrease TPR with long-term use
Decrease SNS outflow from CNS
What do the undesirable side-effects of propanolol and other non-selective β- blockers stem from
Give examples
Blockade of β2
β2 blockade in COPD and asthma patients can exacerbate their condition and lead to a serious crisis
β2 blockage also causes loss of vasodilation to cutaneous blood vessels leading to coldness in extremities
What are 2nd generation β blockers
Eg? (You need to know this drug)
β1 selective antagonists
AKA cardioselective β blockers
Atenolol
What is the benefit of 3rd generation β blockers
Eg
Polypharmacological benefits
Nebivolol - selectively blocks β1 But also offers additional benefit as vasodilator and cardioprotective agent
How is Nebivolol cardioprotective
Through promoting NO release from endothelium and in myocardium
What is carvedilol
A mixed α1/β-AR antagonist with anti-oxidant and anti - inflammatory properties
What are NANC transmitters
Eg
Non-adrenergic, non-cholinergic transmitters
Eg purines: ATP+ derivatives
Neuropeptides
Volatile substances: NO
What are small molecule NANC NTs stored in
What about neuropeptides
Clear core vesicles
LDCVs
Where are neuropeptides synthesised and stored
Synthesised, processed and packaged into large dense core vesicles in the cell body and then move down along the axon towards the nerve terminal via axonal transport
How does storage of ATP in adrenergic nerves compare to cholinergic nerves
Release?
ATP and ACh coexist in cholinergic vesicles whilst ATP, NPY, and catecholamines are found within storage granules in adrenergic nerves and the adrenal medulla
similarly, ATP is released along with NTs
Which 2 peptides are mostly implicated in ANS transmission?
Name 9 other notable neuropeptides or groups of peptides
What do all these work on
Vasoactive intestinal peptide (VIP) and neuropeptide Y (NPY)
the enkephalins, substance P and other tachykinins, somatostatin, gonadotropin-releasing hormone (GnRH), cholecystokinin (CCK), calcitonin gene–related peptide (CGRP), galanin chromogranins
their cognate GPCR
How are NANC transmitters often released in the periphery
eg?
concurrently with the classical neurotransmitters as the nerves are depolarised
eg: ATP is released with ACh and NA in the urinary bladder and vas deferens, respectively.
what is suramin
ATP antagonist (blocks P2X and some P2Y subtypes)
If you have a typical biphasic or twin-peaked contractions of the guinea pig vas deferens measured in response to electrical stimulation of sympathetic nerves innervating the tissue, how will the peaks be affected by suramin and prazosin
suramin: ATP antagonist
prazosin: α1-adrenoceptor antagonist
suramin alone abolishes early peak
prazosin alone abolishes late peak
response is completely eliminated with both S+P
How do neurons that release many NTs, do so?
How do they interact with the postsynaptic membrane
Many other neurons contain and release more than one transmitter, sometimes from the same vesicles and sometimes from different ones
each NT has its own receptor
Which NTs are commonly released alongside NA in the SNS
ATP and NPY
What does co-transmission allow
Elaborate
greater complexity of signalling
NT time course may vary
Often NTs are released independently of each other
What does the different time course of NTs in co-transmission allow
allow different types of responses to be elicited in the postsynaptic cell. Transmitters with a prolonged action, because of slow removal from the synaptic cleft, can also act on more than one target group of cells
Give example of NTs being released independently of each other from same presynaptic neuron
e.g. higher concentrations of Ca2+ are required to elicit release of peptide transmitters (discussed early in this handout), so they are preferentially released at high rates of neural stimulation (i.e depolarisation).
What does a substance have to meet to be considered a neurotransmitter
Dale’s criteria
How is the action of ATP and its derivatives mediated
through purinergic receptors or purinoceptors:
Adenosine receptors
Ionotropic P2X receptors
Metabotropic P2Y receptors
What are the 3 types of purinergic receptors?
Elaborate briefly on each
a) Adenosine receptors – A1, A2A, A2B and A3 (all are GPCRs)
b) Ionotropic P2X1-7 receptors – all are homo/heterotrimeric ATP-gated cation channels
c) Metabotropic P2Y1,2,4,6,11-14 receptors – all are GPCRs whilst few still remain as ‘orphan’ i.e. the natural agonists are yet to be known.
What is an orphan receptor
one where the natural agonists are yet to be known
How is ATP stored in nerve terminals (2) and how may it be released (3)
may be present in the cytosol or stored into vesicles by the vesicular nucleotide transporter (VNUT)
can be released by exocytosis in Ca2+-dependent way or through large membrane channels called the pannexins or via the nucleotide transporters (NtT).
How does ATP enter vesicles
VNUT: vesicular nucleotide transporter
When may cytoplasmic ATP cause neurotoxicity
released from dying or necrotic cells including degenerating neurons and excessive release in this way could lead to further cellular damage/neurotoxicity
What happens when ATP is released into the extracellular space
broken down into ADP, AMP and adenosine by action of ecto-nucleotidases
How is ATP converted to ADP etc in the extracellular space
What does this lead to
by action of ecto-nucleotidases
termination of transmission at the P2X and P2Y receptors
Conversion of ATP to its derivatives by ecto-nucleotidases ceases transmission at P2Y and P2X receptors. Does this mean transmission at the synapse has ceased
no adenosine can still act ?
- maybe ask Ritter about this not sure
Where is adenosine present
in the cytosol of all cells and is taken up and released via a specific membrane-bound nucleoside transporter(s) (NsT).
How is adenosine taken up and released from cells
via a specific membrane-bound nucleoside transporter(s) (NsT).
How can you get inosine?
hydrolyze adenosine by adenosine deaminase
What are P2X receptors
non- selective cation channels (i.e. equally permeable to Na+ and K+ and with significant Ca2+ permeability.
What is the permeability of P2X channels
equally permeable to Na+ and K+ and with significant Ca2+ permeability.
when do P2X channels open
open within milliseconds of the binding of extracellular ATP
How many P2X receptor isoforms are there in mammals
7
assemble as homo or heterotrimers to form a functional channel
Where are P2X receptors found
have a widespread tissue distribution, being expressed on both central and peripheral neurons, where they are involved in synaptic and/or neuromuscular transmission.
What are the dominant P2X receptor types in neurons
2, 4, 6
What is the dominant P2X subtypes in smooth muscle
P2X1
What is the importance of P2X receptors in pathology
Changes in the expression of P2X receptors have been characterized in many pathological conditions of the cardiovascular, gastrointestinal, respiratory, and urinogenital systems and in the brain and special senses
In the vas deferens and blood vessels, which NT produces the fast and slow response from SNS innervation
ATP produces fast responses mediated by P2X1 receptors, followed by a slower component mediated by G protein-coupled alpha-ARs.
Give an experiment demonstrating the importance of P2X receptors
Male mice lacking the P2X1 receptor gene have a drastically reduced fertility because of much reduced sperm count in the ejaculate
What receptor could be targeted for a male contraceptive
P2X1
How is adenosine an atypical NT
- it is neither stored in vesicles nor released by a Ca2+-dependent process
Where is adenosine found
found in the cytosol as well as in the extracellular fluid throughout the body, including the central and peripheral nervous system`
How is adenosine produced extracellularly
How is it then taken up
adenosine is produced following ectonucleotidase-catalyzed hydrolysis of ATP and then taken up by cells through a nucleoside transporter (NsT).
What does dipyridamole do
blocks NsT to increase [adenosine]e
What is methotrexate
What does it do and what is the mechanism
an anticancer and immunosuppressant drug, also appears to increase the extracellular concentration of adenosine, through mechanism(s) not very clear to date
What does adenosine boosting do
believed to contribute to its immunosuppressive action significantly.
What kind of range are [adenosine]i and e kept within
submicromolar
What are the 2 ways adenosine can be metabolised
to inosine (by adenosine deaminase) or to adenine nucleotides (by adenosine kinase)
Why can [adenosine]e increase under stressful conditions
large amount of ATP can be released from the injured cells (e.g. endothelial cells, neutrophils, cardiomyocytes, smooth muscle cells, glial cells etc.) via large pore-containing channels such as pannexins or connexins (or directly after necrotic cell death) and subsequently dephosphorylated to adenosine by ecto-nucleotidases
intracellular production may also increase
What is a autacoid
locally acting hormone
How are all biological actions of adenosine mediated
through adenosine receptors
through the activation (A2A and A2B) or inhibition (A1 and A3) of adenylyl cyclase and thus cAMP signalling
Which adenosine receptors are activatory and which are inhibitory
Do they have any other effects (3)
A2a and A2b activated AC and increase [cAMP]
A1 and A3 inhibit AC
Some, if not all, of these receptors have also been reported to activate phospholipase C (and thereby trigger intracellular Ca2+ release) and mitogen-activated protein kinase (MAPK).
Also modulate Cav and K+ channel function
How do adenosine receptors affect ion channels at the PM
modulate function of voltage-gated Ca2+ channels (via influencing their level of phosphorylation) and K+ channels (via βγ subunit and/or altering phosphorylation states).
What is the effect of adenosine on the heart
What the receptor mediating
negative inotropic and chronotropic effects (A1 receptor)
How can the effects of adenosine on the heart be used therapeutically
How is it administered
Is it safe? why?
to terminate supraventricular tachycardia
an intravenous bolus injection
due to its short duration of action (destroyed/taken up in seconds of intravenous administration) it is considered safer than other alternative drugs
What is the effect of adenosine on smooth muscle?
Which receptors mediate this effect and what are they coupled to
relaxation
A2A and A2B receptors that are Gs coupled
What is the effect to adenosine pre-synaptically
Via which receptor?
exerts pre-synaptic inhibitory effects on the release of excitatory transmitters in the CNS and periphery
presynaptic A1
How does caffeine work
blocks inhibitory action of adenosine on presynaptic A1 receptors
Name a methylxanthine agent
What kind of drug is it
caffeine
competitive antagonist of A1 receptors.
Name an adenosine derivative drug
What kind of drug is it
what effect is it used for
Regadenoson (Trade names: Lexiscan or Rapiscan)
selective A2A adenosine receptor agonist
coronary vasodilatory activity
What has the FDA approved regadenoson for?
for diagnostic purpose only in a type of heart scan called ‘radionuclide myocardial perfusion imaging’
What diagnostic procedure can regadenoson be used for?
What does this procedure assess?
Why is regadenoson useful?
‘radionuclide myocardial perfusion imaging’
to see the blood flow in the heart muscle
it acts as a stress agent that has a similar effecton the heart as exercise
How big is the neuropeptide family
> 100 members
How big are neuropeptides
3-36 amino acids
Where are neuropeptides made
soma
How are neuropeptides made
synthesis is a multistep process that takes places in the soma and begins with the transcription of the gene that encodes one or more prepropeptides in the nucleus, splicing of the resulting primary RNA transcript to produce a mRNA
Prepropeptide mRNA is exported to cytoplasm for translation, which makes the prepropeptide. This can become several different neuropeptides
Name a prepropeptide
pre-POMC
What do prepropeptides contain characteristically
contain an N-terminal signal sequence (or “pre” sequence) that directs the newly synthesized protein into the lumen of the rough ER and thus into the proper, regulated secretory pathway
What does remval of the pre sequence from a prepropeptide result in
What happens to it
in a propeptide (e.g. POMC)
released from the ribosome after translation is complete, transferred to the Golgi complex, and subsequently packaged within large dense core vesicles (LDCVs)
What happens to the propeptide within the golgi and LDCVs
undergoes additional posttranslational processing, which involves additional cleavages and covalent modifications.
What is the main neuropeptide at cholinergic synapses
VIP
Why do neuropeptides produce a longer response
What terminates their action
because they are not rapidly removed from the ECF
response is terminated either by diffusion or by extracellular peptidases
What receptors do neuropeptides activate
how can they thus be described
GPCRs
neuromodulators (more appropriate than neurotransmitter)
What do neuropeptides account for in autonomic ganglia
be specific
released neuropeptides (for e.g. GnRH) account for the late slow EPSP.
True or false
Receptors for neuropeptides bind their ligands with greater affinities than do the receptors for small-molecule transmitters
true
. This high affinity implies that neuropeptides can act at low concentrations, which is what would be expected if they often diffuse for great distances.
What is needed to make NO normally
How does this differ in pathology
nitric oxide synthase (NOS) which converts L-arginine to NO and L-citrulline
cells eg vascular smooth muscle can generate NO independent of Ca2+ after stress
What are the different isotypes of NOS
nNOS (NOS I)
iNOS (NOSII)
eNOS (NOS III)
Where is nNOS expressed
neuronal NOS (nNOS, or NOS I) expressed in the CNS and NANC nerves
Where is NOS II expressed
inducible NOS (iNOS, or NOS II) is usually absent but its expression is induced in macrophages and other cells by bacterial lipopolysaccharide and/or inflammatory cytokines, notably interferon-γ.
Which cells produce iNOS when induced to do so
fibroblasts, vascular smooth muscle cells, endothelial cells, Kupffer cells, neutrophils
Which inflammatory cytokine induces iNOS expression in some cells
interferon-γ.
Where is eNOS expressed
expressed in platelets as well as endothelial cells
Where does NO go after produced
diffuses all directions - across the presynaptic membrane and the membranes of non-neuronal cells producing it.
What does NO activate
sGC (soluble guanylyl cyclase)
How does NO activate sGC chemical
through interacting with the haem moiety of the enzyme
What does the cGMP produced by NO action do
can act directly on effector proteins eg ion channels but more commonly through the activation of (PKG). PKG phosphorylates various proteins and leads to relaxation of smooth muscle, particularly in the blood vessels and this accounts for the term endothelium-derived relaxation factor (EDRF) that is used to denote NO
what are nitrergic nerves
NO releasing
Where are innervations with nitrergic (i.e. NO releasing) postganglionic nerves present
in several peripheral tissues that notably include upper airways, gastrointestinal tract (the enteric neurons) and male sexual organs
What kind of NOS do enteric nitrergic neurons express
What role does the released NO play in the GI tract (6)
nNOS
NO regulates the muscle tone of the sphincter in the lower esophagus, pylorus, sphincter of Oddi, and anus.
regulates the accommodation reflex of the fundus and the peristaltic reflex of the intestine
What is reduced nNOS associated with in the GI tract
eg
GI tract motility disorders
nNOS-/- mice had ~50% loss of inhibitory NANC transmission in small intestine and showed distended stomach and pyloric hypertrophy
How is nNOS expression associated with diabetes (use a mouse experiment)
Diabetic mice often show delayed gastric emptying that has been attributed to negligible expression of nNOS protein and thus inadequacy in NO-mediated NANC transmission.
Insulin promotes nNOS protein expression and thereby restored NANC-mediated pyloric relaxation
How is insulin associated with nNOS
promotes nNOS protein expression
What happens to nitrergic neurons in achalasia
, there is selective loss of the nitrergic neurons supplied to oesophageal sphincter muscles, which incapacitates oesophagus to move food along.
What is the blood pressure like in mice who lack the gene coding NOS III?
Why?
they are hypertensive
The L-arginine/NO pathway is tonically active in resistance vessels, reducing peripheral vascular resistance and hence systemic blood pressure
How is Nebivolol associated with NO
the selective 1-AR blocker Nebivolol (more correctly speaking, its metabolite) also triggers NO production
What class of drug use NO to vasodilate
nitrovasodilators
donate NO to relax blood vessels
