Synapses and NTs

Lectures 8 and 9

Describe a synapse

junction between two neurons allowing signals to pass from one to the other

Compare and contrast the key features, functions, and capabilities of chemical vs. electrical synapses

Both:

• are ‘plastic’ (i.e., can be modified), but chemical synapses probably more so

• allow summing up inputs by the post-synaptic neuron

Most synapses are chemical synapses

Describe the main steps of chemical neurotransmission and how these steps are regulated

1. Package NTs into vesicles at the axon terminal

2. Action potential arrives 🡪 voltage-gated Ca²⁺ influx

3. Ca²⁺ influx 🡪 vesicles fuse via SNAREs

4. NTs bind to receptors on postsynaptic side

5. NTs removed from synaptic cleft

What is synaptic transmission?

process of signalng via synapses

Where does the word synapse come from?

from Greek, ‘syn’ = together, ‘haptein’ = to fasten, join (coined by Sherrington in 1897)

How many synapses does the brain have? How does this compare to the nb of neurons?

~100 trillion (10¹⁴) synapses

~100 billion (10¹¹) neurons so 1000 synapses per neuron

How was the disagreement over distinct neurons or a continuous net of fibres resolved?

• Golgi stain (Ramón y Cajal, late 1800s)

• Physiological evidence from study of reflexes (Sherrington, around 1900)

• Final evidence from electron microscopy (1950s)

What’s the neuron doctrine vs the reticular theory?

existence of distinct neurons vs continuous net of fibres

How do neurons allow flexibility in complex organisms vs in very simple animals?

thanks to integration of inputs from interneurons, sensory neurons etc = output through motor neurons = behaviour // sensory input = stereotypic behaviour

What structures allow electrical synapses to function correctly and how? What are their subtypes?

gap junctions to let current pass between neurons

connexons made up of connexins

What’s the diameter of a gap junction? What flows through them?

1 - 2 nm

ions = current passing through

How are electrical changes in one neuron passed directly to the other? What types of currents are transmitted?

gap junctions !

hyperpolarisation and depolarisation

How can we block hyper.depolarisation from being passed from one neuron to the next?

by deleting a connexin gene (shakB2 mutant)

How was communication between neurons shown visually?

through a red dye, turning the green neurons yellow - this made adjacent neurons yellow too

What are 2 main advantages of synapses?

• fast communication

• synchronising neurons

What was the first evidence of chemical synapses? Who did this experiment?

using two isolated frog hearts that nerves release a chemical which slows the heartbeat - ACh!

Otto Loewi, 1921

What are types of post-synaptic cells?

• neuron in a neuron-neuron synapse

• skeletal muscle in a motor neuron

• hormonal gland, smooth muscle, heart in an autonomic neuron

What are secretory granules?

dense and large vesicles carrying peptide NTs and used once

What are the 5 stages of chemical synaptic transmission?

• NT put in vesicles at pre-synaptic terminal

• AP arrives, voltage-gated Ca²⁺ channels open

• Ca²⁺ influx 🡪 vesicles fuse to membrane, NTs released

• NTs diffuse across the synaptic cleft, activate Rs on the postsynaptic cell

• NTs removed from cleft

What are the main differences between synaptic vesicles and dense-core secretory granules?

• size and density: ‘clear’ small (40 - 50 nm) // dense and large (100nm)

• content: small molecule NTs // peptide NTs

• Filling: by transporter proteins at presynaptic `terminal // ER/Golgi secretory apparatus

• Termination: recycled // one and done

What are 2 ways that NTs can be packaged to be brought to the presynaptic terminal?

in synaptic vesicles or in dense-core secretory granules

When using a calcium-sensitive fluorescent protein, what does more fluorescence translate?

more calcium

How does AP affect calcium channels in presynaptic terminals?

opens the channels = influx of calcium

Is the synaptic cleft of electrical or chemical synapses larger?

chemical

How does a sudden influx of calcium in presynaptic terminals affect vesicles and their content?

vesicles fuse to membrane, NTs released

After a chemical synapse, the vesicle was recycled through endocytosis, this means the vesicle contained…?

small molecules, NOT peptides

What structures allow vesicle fusion? What ion is involved in this interaction and what role does it play?

SNAREs

calcium binds to synaptotagmin = conformational change so SNAREs ‘zipper’ together, forcing the vesicle to fuse to the plasma membrane

What are SNAREs targeted by?

 toxins (botulinum toxin, tetanus toxin)

What types of postsynaptic receptors can NTs bind to after their release into the synapse?

ligand gated ion channels and GPCRs eg

What type of receptor leads to direct de.hyper.polarisation of the postsynaptic cell?

ligand-gated ion channels

How does a NT affect the postsynaptic neuron? (does it enter the cell?)

by binding to receptors which lead to ion influxes or downstream effects like phosphorylation of target effectors

DOES NOT ENTER CELL

What types of channels do NTs go through?

ones allowing their uptake, NEVER pores in postsynaptic neurons

What are 3 ways in which NTs can be removed from the synaptic cleft, terminating their communication?

• diffuse away

• actively taken up by transporters for recycling (into presynaptic neuron or glia)

• destroyed in the cleft by enzymes

What is the issue with NTs always only diffusing away as a method of termination?

passive and slow, unreliable bc would mean prolonged communication when no longer necessary

What are common points between electrical and chemical synapses?

• both are plastic but chemical synapses a bit more

• allow summing up inputs by the post-synaptic neuron

Are most synapses chemical or electrical?

chemical

What distinguishes electrical vs chemical synapses in terms of their direction, their transformation and their speed?

• both directions // only in one

• passed directly, can only be attenuated // can be radically transformed (inverted, amplified, modulated)

• fast (under 0.3 ms) // slow (0.3 - 5 ms)

What types of action potentials do motor neuron APs cause?

always muscle cell APs

What is the speed of transmission in the neuromuscular junction (NMJ) and the NT used? Does failure/attenuation of AP occur?

fast and reliable

ACh

no!

What is a neuromuscular junction?

junction between motor neuron and muscle cell

How do presynaptic neurons contribute to the efficiency of the NMJ?

large nb of active zones

How do postsynaptic (motor end-plate) contribute to the efficiency of the NMJ?

Contains junctional folds, densely filled with NT receptors

How are active zones and junctional folds relatively located in the NMJ?

precisely aligned

What is the qualitative size of the NMJ?

one of largest synapses in the body

How was it understood that NTs are released from vesicles?

• release always produced the same amplitude so must be a repeatable way of releasing the same amount every time

• no halves, not irregular or continuous, always multiples = NTs come in quantal packets

What are different varieties of CNS synapses and what informs on this?

• dendrosomatic

• dendrodendritic

• axosomatic

• axodendritic

• axoaxonic

the morphology of the synapse

Why does botulinum toxin cause paralysis?

• targets SNAREs

• = no fusion of vesicles with neuron membrane = no NT release

• = no synapse so no motor mvt/no contraction

If you block vesicle endocytosis, how would this affect release of small molecule vs. peptide neurotransmitters?

small molecule vesicles would be affected bc they are recycled so we would run out // peptide would be fine bc they are a one use anyway

What is the Nernst potential of Ca²⁺ at 37 ºC, if the concentration outside is 1 mM and the concentration inside is 0.0001 mM? Why does this explain the movement of calcium ions when calcium channels open?

+122 mV

calcium moves out into IC down concentration/chemical gradient + towards the negative membrane potential bc is a positive ion (-65mV inside so defo mvt in considering nernst equation)

What is a quantum?

one vesicle full of neurotransmitter

What makes a molecule a NT?

• present in a presynaptic terminal

• released in response to stimulation

• acts on the postsynaptic neuron

• its inhibition should prevent synaptic transmission

What are 4 ways of determining if a molecule acts as a NT experimentally?

• immunostaining = presence of NT

• in situ hybridisation = enzyme/transporter expression?

• collect fluid around neuron = release?

• drugs, delete genes encoding enzymes/transporters/receptors = block it to study it

What are 3 origins from which NTs can come from?

• amino acids

• amines

• peptides

What are 3 characteristics of NTs that come from amino acids and amines?

• small (100-200 Da)

• stored in synaptic vesicles

• bind to ligand gated ion channels or GPCRs

What are 3 characteristics of NTs that come from peptides?

• large (1K - 3K Da)

• stored in secretory granules

• bind to GPCRs

What will peptide-releasing neurons also release alongside the NT?

small molecule transmitter, called a ‘co-transmitter’

What type of receptor will lead to direct depolarisation/hyperpolarisation of the postsynaptic cell?

ligand-gated ion channels (ionotropic receptors)

How many transmitters, receptors and effectors are involved in divergence?

1 // multiple receptors and effectors

How many transmitters and effectors are involved in convergence?

multiple transmitters and receptors // 1

What’s the major excitatory NT, its role, mode of action and how is it synthesised then degraded/removed?

• amino acid glutamate

• EPSP, coincidence detector

• binds to 3 different types of ionotropic Rs + mGLuRs

• found everywhere

• selective uptake into presynaptic terminals and glia

What types of receptors does glutamate bind to?

ionotropic receptors:

• AMPA

• NMDA

• Kainate

metabotropic receptors:

mGluR1, mGluR2, etc

What’s the function of glutamate AMPA receptors?

fast excitatory transmission

sodium and potassium currents = EPSP

Can AMPA and NMDA glutamate receptors co-exist? When do NMDA receptors open?

yes!

voltage-dependent magnesium block, magnesium blocks the gate at resting potential, evacuates and opens up the gate at -30mV eg, so only open when the neuron is already depolarised

What’s the function of NMDA receptors?

let calcium in, leading to downstream signalling

coincidence detector: when a neuron is activated right after it was already activated 🡪 important for learning

In what case can glutamate be inhibitory?

when binding to some metabotropic receptors like in the retina

What’s the major inhibitory NT, its role, mode of action and how is it synthesised then degraded/removed?

• GABA, amino acid but never used to make proteins

• IPSPs

• binds to ionotropic and metabotropic receptors

• synthesised from glutamate

• selective uptake into presynaptic terminals and glia

Which enzyme is responsible for synthesising GABA from glutamate?

glutamic acid decarboxylase

How does GABA produce IPSPs?

GABA-gated chloride channels

In what condition do GABA channels open?

when GABA binds

What’s the major modulatory NT, its role, mode of action?

• glycine

• inhibits neurons via glycine receptors + binds to glutamate receptors

Name disorders where NT systems are impaired. How are these systems linked to the identified disorders?

• too much GABA = coma/loss of consciousness

• too little = seizures

Why does it matter how excitatory and inhibitory synapses are arranged spatially?

if an inhibitory synapse is closer to the soma, it can inhibit the excitatory synapse/EPSP

When could a GABA receptor not produce an IPSP? Why?

e.g. if V_m is near chloride’s Nernst potential (-65mV)

instead of chloride exiting, it would flow out of the cell to stay closer to -65mV, instead of coming in and causing hyperpolarisation

When does inhibition often occur? What are mechanisms for this?

presynaptically

inactivating calcium channels = reduced calcium = fewer NTs = reduced effect/inhibition

What is runaway excitation? What avoids this and what type of neuron can this role be compared to?

too much excitation!

like interneurons - they balance excitation and inhibition here

What does shunting inhibition mean?

reduction in membrane resistance caused by the opening of membrane pores = decrease in neuronal activity acting as CNS-wide inhibition

How do GABA receptors leak current out of the membrane? What does this cause?

Opening chloride conductance decreases the membrane resistance

When other chemicals bind to GABA-Rs, what is their effect and a condition to this?

modulate the response to GABA binding ie no effect if GABA is not already bound - termed an allosteric drug

What are examples of chemicals that can bind to GABA receptors?

ethanol (alcohol), benzodiazepines (diazepam for anxiety), barbiturates (sedatives/anti-convulsant), neurostreroids (progesterone)

What’s an example of inhibition caused by alcohol binding to GABA-Rs? What does continuous then absent stimulation of these receptors cause (like sobering)?

not being able to walk in a straight line

withdrawal - will need more stimulation to work bc of this continuous stimulation = big adjustment needed

What are examples of ways in which metabotropic GABA Rs work in different cells?

• open K⁺ channels

• close Ca²⁺ channels

• trigger other second messengers like cAMP

Are metabotropic GABA Rs pre or postsynaptic? Are they inhibitory or excitatory?

presynaptic and/or autoinhibitory

Give examples of pharmacological agents that modify neurotransmission in a variety of ways and explain how they work

modulate GABA effect so can be positive or negative modulators, those leading to relaxation increase will increase GABA affinity

alcohol (+ GABA agonist), diazepam (+ GABA affinity), sedatives, anti-convulsants, progesterone (natural regulators?)

What is an allosteric drug?

bind to specific sites on proteins distinct from the active site modulating protein activity and offering advantages like enhanced selectivity