Exam 2: Topic 4 Flashcards
reticular theory
all of the units of the nervous system are interconnected in a continuous way
neuron doctrine
individual units that are connected in a contiguous matter
contiguous
things that are individual but next to one another
Golgi stain
based on the precipitation of silver ions
- Randomly stains a small subset of all cells
- Reveals detailed morphology of single cells
how do we know neuron doctrine is correct?
if you do multiple Golgi stain section on the same tissues, different neurons will take it up different times
- it was the reticular theory everything would be stained
what are the two classes of synapses?
- electrical synapses (gap junctions)
- chemical synapses
electrical synapses
cells cytoplasm is continuous because of the gap junction channels but their membranes are not continuous
gap junction
channels that connect the cells via cytoplasm
chemical synapses
a presynaptic neuron and a postsynaptic neuron where information goes across the synaptic cleft
connexon
6 subunits that bind together made up of alpha helices that make up the gap junction channels ⇒ the pore in the middle allow ions to flow through
- You need connexons in the pre and postsynaptic membrane to line up and bind to one another
hemichannel
each half of the connexion
- forms the functional channel
- Some are rectified and some are bidirectional depending on the subunits
T/F electrical signals have vesicles?
False
- no synaptic vesicles are released
what is the distance and pore size of an electrical synapse?
- Distance between cells = < 5 nm
- Pore size: > 1 nm ⇒ relatively large
what things diffuse through gap junctions?
ions, ATP, amino acids
what is the directionality of gap junctions?
bidirectional ⇒ some are rectified depending on subunit
how fast are gap junctions?
very fast ⇒ short synaptic delay
- delay of microsections between the cells
what is the modulation of gap junctions?
low
- post synaptic cell likely does not have voltage gated channels which causes passive decay ⇒ follows the current coming in from presynaptic cell
- Fidelity is high and most of the time when the first cell fires an action potential the second one will as well ⇒ sometimes the second one doesn’t fire not due to the gap channel but instead because of electrical pulses from somewhere else in the cell
what do the action potentials of the second electrical synapsing cell look like?
typically truncated so amplitude is lower and width is longer
what are the structure of chemical synapses?
the chemical transmitters in vesicles fuse with the plasma membrane and are released
- In the postsynaptic cell there is a dense region where receptors are located
active zones
where the vesicles are attached/locked on to the interior membrane of the presynaptic cell and when it gets a signal these vesicles are released to the postsynaptic cell
- The ligand will bind to the channel and often causes it to open (sometimes close)
- They can also activate biochemical cascades through G protein coupled receptors
what is the distance and pore size of a chemical synapse?
- Gap size between cells: 10-100 nm (broader than gap junctions)
- Pore size: < 1 nm
what diffuses through chemical synapses?
ions only
what’s the speed of chemical synapses?
longer synaptic delay (milliseconds instead of micro)
what it the directionality of chemical synapses?
unidirectional
what is the modulation of chemical synapses?
high ⇒ functionality can be changed
- Such as having another transmitter which can affect how the channel binds to the main ion ⇒ increase/decrease current flow
what is the general purpose of chemical synapses?
neurotransmission
what is the general purpose of electrical synapses?
ensures synchronization ⇒ escape behaviors, breathing
cleft
where the neurotransmitters diffuse across to bind to channels
what are properties of inhibitory neurotransmitters? (2)
- Binds to a receptor that opens and allows only chloride ions to come in which hyperpolarizes the cell
- Closes the channel when bound so nothing can excite the cell ⇒ such as sodium being unable to come in
what are CNS tripartite synapses?
astrocytes surround the synapse and play a role in synaptic function
what functions do astrocytes have in CNS synapses?
they are important for clearing neurotransmitters from the synaptic cleft
- There are multiple ways of getting rid of transmitters but one way is that it diffuses away from the synapse and is taken up by astrocytes
- Ensures there isn’t too much neurotransmitter in the cleft at a time
what kinds of events can cause the astroglia to dump neurotransmitters?
injury or trauma to the brain
- glutamate excitotoxicity
excitotoxicity
when there is too much glutamate being taken up and cells become overactive ⇒ it can lead to cell death
- glial cells can communicate with pre and postsynaptic cells which helps modulate their neurotransmitter release
how do we know neurons release something?
there was an experiment where saline between one chamber and another chamber with a heart it in was done
- when the vagus nerve was stimulated in chamber 1 and then this saline flowed to chamber two it caused a secondary activity of the heart in chamber 2
- same response with a slight delay
- Showed there is a chemical messenger responsible for nerve cells to pass signals on
what happens when you stimulate the vagus nerve? (2)
- Reduction in frequency of contraction
- Reduction in amplitude (strength) of contraction
what did early electron microscopy tell us?
it hinted neurotransmitters could be inside membranous vesicles
- the presynaptic cell has many vesicles
how do you create a good study for neuromuscular junctions? (2)
- Readily accessible animal system ⇒ abundant in nature or breedable in a lab setting
- You want the synapses to be readily accessible ⇒ motor neurons in periphery are right under the skin
what type of neuromuscular junction endplate studied were done on which animal?
frogs ⇒ restrictions were less than when working with mammals
- if you leave the motor neuron in tact and you can stimulate the muscle to record postsynaptic membrane potential
endplates
Saucer like structures in the muscles are called end plates and within this the axon terminal will heavily branch
synaptic boutons
innervate the muscle fiber in a specific way ⇒ high density of synaptic vesicles and release acetylcholine
end plate potential
the release of acetylcholine at the end plate in muscle cells ⇒ acetylcholine binds on the receptor cell (evoked response)
- These changes can be recorded ⇒ the cell gets highly depolarized since many receptors are receiving which causes the muscle contraction
spontaneous events
when we simply record but do not evoke action potentials ⇒ such as in the end plates of muscles
Miniature end plate potentials (MEPP)
small spontaneous events of depolarization but are not enough to cause an action potential
- When we stimulate the motor axon with reduced calcium we get a subthreshold endplate potential
- Sometimes there are no post synaptic events/end plate potentials
- They have different amplitudes ⇒ sometimes there is a spontaneous MEPP
what steps happen during MINIs? (4)
NT release spontaneously, NT crosses synaptic cleft, NT binds ligand gated ion channel receptors, Receptor channel opens
excitatory receptors (EPSPs)
NA+ and K+ current flow according to DF ⇒ small postsynaptic depolarization (mEPSP)
inhibitory receptors (IPSPs)
Cl- flows according to DF ⇒ small post synaptic hyperpolarization (mIPSP)
what are minis not caused by?
synaptic vessel release in response to an AP
- they play an important biological role in letting neurons know about their synapse ⇒ I’m here signal
- Neurons may change expression of neurotransmitter receptors if the frequency of minis changes
what happens if we do not have MINIs during development?
it is likely the synapse would no longer exist during development
- Cells that fire together wire together
what is the least common multiple for mEPP?
we get a bell shape that peaks around 0.4 mV in amplitude
- For spontaneous MEPPs they have unitary structure
- It’s possible the fusion and binding of a single vesicle is possible for the end plate potentials
quantal content
amount of postsynaptic depolarization caused by release of neurotransmitter from a single synaptic vesicle
- The least common multiple reflects the quantal content => 0.4, 0.8, 1.2, etc.
how did we “prove” a single/multiplet vesicle leads to quantal content?
we used 4-AP and then stimulated neuromuscular junctions while recording mEPPs
- the tissue was immediately frozen right after a stimulation happened and then analyzed under electron microscopy
- the fused vesicles were counted
4-Aminopyridine (4-AP)
blocks VG K+ channels and potentiates VG Ca2+ channels
- Increasing 4-AP prolongs depolarization and increases transmitter release
- intracellular calcium levels must increase in the presynaptic cell for NT to be released
MINIs
Measurement of change in postsynaptic Vm caused by spontaneous release of synaptic vesicles from the ready releasable (docked and primed) pool of vesicles on the presynaptic side
- the quantum (singular of quanta) is the amount of postsynaptic depolarization caused by the release of NT from 1 synaptic vesicle ⇒ more quanta are released as more vesicles fuse to the membrane
what are MINIs called in neurons?
mEPSPs or mIPSP for mini excitatory or inhibitory postsynaptic potential
what are MINIs called in muscles?
mEPP for mini end plate potential
how does quantal content affect amplitude?
The amplitude of the depolarization curve comes in multiples of the quantal content
- Quantal content is the amount of postsynaptic depolarization caused by release of neurotransmitters from a single synaptic vesicle
- The amplitude of the mini caused by a single SV is proportional to the strength of the synapse
how do MINIs change frequency?
The frequency of minis is proportional to the numbers of synapses between pre and post synaptic cell
how does chemical synaptic transmission occur? (11)
- Transmitter is synthesized and stored in vesicles
- An action potential invades the presynaptic terminal
- Depolarization of presynaptic terminal causes opening of voltage gated Ca2+ channels
- Influx of Ca2+ through channels
- Ca2+ causes vesicles to fuse with presynaptic membrane
- Transmitter is released into presynaptic membrane
- Transmitter binds to receptor molecules in postsynaptic membrane
- Opening or closing of postsynaptic channels
- Postsynaptic current causes excitatory/inhibitory postsynaptic potential that changes the excitability of the postsynaptic cell
- Removal of NT by glial uptake or enzymatic degradation
- Retrieval of vesicular membrane from plasma membrane
properties of VG Ca2+ channels
1 protein required with 4 domains ⇒ each is similar to alpha subunits in VG K+ channels
- 24 membrane spanning helices
- 1 pore loop per domain for Ca2+ selectivity
- Voltage dependence ⇒ 1 voltage sensor per domain
- Beta subunit regulates and is encoded by a different gene
what is BAPTA?
calcium based indicators when calcium binds it causes a photon release so we can measure light
- Calcium chelator
- Homolog of EGTA
- pH insensitive
- High Ca2+ selectivity ⇒ Kd ~ 100 nM
- can be loaded into a cell by a microelectrode or encoded with certain cells in the NS
chemical calcium indicator
calcium sensitive dye
- inject/load indicator into the cells and not in the bath as a binding site for free calcium
- 2 fluorescent molecules ⇒ donor and acceptor
- Conformational change in the indicator causes it to fluoresce
what happened when calcium is not bound to indicator molecules vs when it is?
- when unbound, the two fluorescent molecules will only be weakly activated by light
- When calcium is bound then the donor and acceptor are close to one another so that you get fluorescence resonance energy transfer to emit another wavelength of light
FRET
excitation from first molecule which causes enhanced emission in the second molecule of the calcium binder
- the presynaptic cell has a huge calcium influx during the action potential which triggers the indicator to release photons we can measure
what do we measure when we inject calcium in the presynaptic cell?
we measure the postsynaptic response
- This tells us it is a calcium dependent mechanism from the presynaptic cell causing the depolarization of the postsynaptic cell
- injection of Ca2+ into presynaptic neuron is sufficient for chemical synaptic neurotransmission
what are increases in intracellular Ca2+ necessary for?
chemical synaptic neurotransmission
what would happen if we inject calcium chelator into the postsynaptic neuron instead of the presynaptic neuron?
Nothing would happen since it is required on the presynaptic side for chemical release
where does calcium come from on the presynaptic side? (3)
- Extracellular CA2+ entry
- VG Ca2+ channels
- Ligand gated Ca2+ channels - Intracellular Ca2+ stores
- IP3 gated CA2_ channel
- Ryanodine gated Ca2+ channel - Mitochondria via unknown channel
where does most of the calcium come from?
increase in intracellular Ca2+ from external source is necessary for chemical synaptic neurotransmission
extracellular cadmium
blocks calcium channels
- if there is no inward calcium current coming in then there is no postsynaptic response
- Most of it comes from external calcium
how long does the life cycle of a synaptic vessel take from endocytosis and exocytosis? (4parts)
about 1 minute
- the vesicle buds off and goes into the reserve pool ⇒ fills the entire synaptic bouton
- They are inter-connected through a web of protein
- They mobilize toward the synaptic site ⇒ active zone at the terminal
- The vesicle is coated in clathrin and then buds off again to go back to the uncoated reserve pool
what steps occur at the active zone of the terminal? (3)
- Via protein-protein interactions they will dock at the terminal but don’t release anything yet
- Then they go through a priming phase which gets them ready to release their content when the signal comes in
- Fusion (go signal) occurs from increases in intracellular calcium at the plasma membrane ⇒ this leads to exocytosis where the NT is sent to the synaptic cleft
SNARE proteins
soluble N-ethylmaleimide sensitive factor attachment protein receptor ⇒ attachment protein receptor
vSNARE
snare proteins located on vesicle membranes
vSNARE examples (2)
- Synaptobrevin
- Synaptotagmin: Ca2+ sensor
tSNARE
snare proteins located on target membranes ⇒ usually plasma membranes
tSNARE examples (2)
- Syntaxin
- SNAP-25
what are the mechanisms of exocytosis? (5)
- Free snares are in the reserve pool but then they make complexes with other vesicles soon after ⇒ vSNARES are on the vesicle while tSNARES are on the plasma membrane
- (docked) The snares form a complex initially of synaptobrevin, syntaxin, and snap 25 which docks the vesicle to the plasma membrane
- (primed) Synaptotagmin binds later ⇒ this is a calcium binding protein and is not activated with low intracellular calcium
- The calcium will bind to synaptotagmin and cause it to change its shape pulling the vesicle really close to the plasma membrane
- (fusion) when the two lipid bilayers are very close, they form an invagination and bind together which opens the vesicle ⇒ exocytosis occurs
Botulinum toxin A (BoTX-A)
cleaves SNAP 25 (plasma membrane) and when injected into facial muscles (neuromuscular junction) it blocks skeletal muscle contraction which reduces wrinkles and facial expression
- Interferes with the synaptic vesicles ability to fuse
Tetanus toxin (TeTX)
cleaves synaptobrevin and when dosed (depending on the neuron) the effect is inhibitory interneurons will inhibit motor neurons and muscle contractions ⇒ prevents any muscle from being overactivated
- No inhibitions = permanent contraction (tetanus) ⇒ lockjaw
- The effect on neuromuscular junctions is similar to botox but the effect on the motor neurons overwhelms any diminution of function at the neuromuscular junction
outside out patch recording
attach to the cell membrane and break the membrane like in whole cell ⇒ move the electrode away and it tears off a portion of the membrane
- The outside of the membrane has the saline of the electrode
- You can study a single chemically gated or ligand gated ion channel
how can outside out recording be used to change permeability
you can add the ligand at different concentrations to the electrode ⇒ high or low
- The ligand channel will open and close when it binds
- There are unitary deflections of inward currents (depolarizing) and vary in duration from short to long
what information does outside out recording give us?
this is evidence of a single ligand gated channel opening and closing in the presence of its ligand
- You can also do this when attached to the cell membrane with a voltage clamp
what happens during outside out recording when you stimulate more?
the scale becomes much larger (inward current) correlating with more channels opening
- There is more ligand present to bind to the receptors
- Almost all channels open simultaneously in sync ⇒ channels are highly localized
- if we also have a recording electrode we see that we get a depolarizing current going into the cell
nAChR
nicotinic acetylcholine receptor ⇒ gated by acetylcholine (ligand)
- Ligand gated channel in muscles and some neurons
when is nAChR activated?
closes when ACh comes off
is nAChR voltage gated?
not voltage dependent
what goes through nAChR?
Na+ and K+
- Ions pass according to their DF
- When a channel lets more than one ion through, it is important to think in terms of NET current
what is the reversal potential for nAChR?
Erev will be somewhere in between the equilibrium potentials for the ions that pass through => sodium rushing in and potassium leaving the cell
- At resting membrane potential if the channel is activated there will be sodium coming in more than potassium leaving
which ion is nAChR biased towards?
Biased for sodium to come in
- cholinergic receptor tends to be depolarizing
Vm < Erev
inward current
- less than 0 mV
Vm = Erev
no current
- 0 mV
Vm > Erev
outward current
- over 0 mV
Erev
reversal potential of the channel ⇒ at this point there is no net current
- We can measure the current that comes through the channel when acetylcholine is released where the channels are located at the endplate of nAChR
what happens when nAChR currents are plotted including Erev?
the currents with the membrane potential there is a linear line
- the reversal potential lies for sodium, chloride, and potassium
- Sodium has a higher conductance than potassium for these channels but this also depends on DF and membrane potential
what is the equation for EPC
EPC = gAChR(Vm - Erev)
- gAChR remains essentially constant after the channel is open because the AChR is not voltage sensitive
what happens if there is less extracellular sodium
the reversal potential will shift more toward a negative Vm
- The driving force on sodium is less which shifts the reversal potential toward potassium more
what happens if you increase extracellular potassium
you are lessening the driving force on K+ which shifts reversal potential toward sodium
what occurs for EPCs and EPPc at -100 mV?
-100 mV is closer to potassium so inward current and excitatory endplate potential (EPP) => depolarization from sodium
- At -90mv the driving force inward is less so the EPP is also less
what occurs for EPCs and EPPc at 70 mV?
there is an outward current since sodium is closer to Ex which causes an outward movement of ions and the inside of the cell will become more negative (inhibitory) => hyperpolarization from potassium
Erev > threshold
excitation (depolarization)
Erev < threshold
inhibition (hyperpolarization)
If Erev > Vrest, but < threshold
will get a depolarization but not one that reaches threshold
excitatory NT activity
when a bunch of glutamate activates at once you get an action potential vs only a few activating will give you an EPSP but not an action potential
inhibitory NT activity (2) (Erev < Vrest & Erev > Vrest but < threshold)
- when the reversal potential is much lower than resting potential and threshold potential, then you get a downward deflection inhibitory postsynaptic potential ⇒ chloride will rush in to bring it to the reversal potential for chloride
- Inhibitory channels allows chloride to flow through which has an Ex = -70 mV and are selective - Sometimes there are depolarizing inhibitory potentials when the resting membrane potential is the same and the threshold is the same, but the reversal potential is in between these two points
- When the synapse gets activated there is a depolarization where chloride will rush out but it gets stopped at its reversal potential which is still under threshold so no action potential occurs ⇒ it shunts the membrane potential which makes it inhibitory
summation
dendrites push the excitatory and inhibitory potentials to the axon hillock where they are summed
what happens when we activate two or more excitatory synapses?
we get an action potential
what happens when there are excitatory and inhibitory synapses at the same time?
they summate towards only a small excitatory potential
what happens if multiple excitatory potentials are activated with an inhibitory synapse?
it would usually result in an action potential, but the inhibitory potential pushes it under threshold
end card
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