• NMJ strength is much larger (extremely strong syn)
• Neuronal responses are much smaller
• Muscle do not need to compute, no decisions necessary

NMJ vs. Central Synapse

• acetylcholine vs. glutamate and many other neurotransmitters
• End Plate Potentials and Currents vs. Postsynaptic potentials and Currents
• acetylcholine binding = contraction vs. more complicated relationship between neurotransmitters and response @ central synapse
  • synaptic strengthening
  • can be inhibitory
  • metabotropic actions
• neuron to muscle vs. neuron to neuron
• one-to-one vs. summation of inputs

Central Synapses

• fast neuronal excitation
  • EPSP and EPSC
• fast neuron inhibition
  • IPSP and IPSC

NMJ

• endplate potential and current (EPP and EPC)

• family of similar receptors all having 5 subunits or staves w/ the requirement of 2 α subunits for transmitter binding
• each subunit made up of 4 transmembrane regions
• common family of receptors covering a wide variety of neurotransmitters including GABA, serotonin, glycin, and obviously acetylcholine
• does not include the glutamate ionotropic receptors which are in their own family
• response is neither necessarily excitatory or inhibitory but is actually based on the particular subtype of receptor
  • GABA is almost always inhibitory
  • acetylcholine is almost always excitatory

• acetylcholine and dopamine were discovered to be highly specific for neurotransmission with little to no use in other bodily/cellular functions
• glutamate is an amino acid - how can it also be neurotransmitter
• it was initially proposed to be involved in some sort of nonspecific action
• despite being most common neurotransmitter, accepted evidence for glutamate as a neurotransmitter not discovered until 1970s-80s

• molecule must be packaged into vesicles as mechanism of release
• must have receptors to bind as mechanism for action
• extremely simplified, bare-bones definition that still does not hold true for all potential neurotransmitters (i.e. nitric oxide)

• VGLUTs - vesicular glutamate transporters
  • bring glutamate into vesicles
• EAATs - excitatory amino acid transporters
  • bring glutamate (as well as some other amino acids) into neuron
• ionotropic receptors
  • named for the pharmacological agonists they bind other than glutamate
    • kainate receptors - named for kainic acid
      • GluK1-5
    • AMPA receptors - named for AMPA
      • GluA1-4
    • NMDA receptors - named after N-methyl-D-aspartic acid
      • various GluN#X receptors
• there are also a wide variety of slow-acting metabotropic receptors

• 4 staves/subunits each made up of 4 transmembrane regions
  • each stave is separate subunit
• Q/N/R site determines ion selectivity
  • Q = glutamine (neutral)
  • R = arginine (positive)
  • N = asparagine (neutral)
• differences from nAchR's
  • 4 staves instead of 5
  • while having same number of transmembrane regions/stave, M2 region is truncated
  • p-region is more similar to that of voltage-gated channels 

• ionotropic glutamate receptor
• equally permeable to K and Na
• reversal potential near 0
• responsible for bulk of neuroexcitatory transmission in brain
• inwardly-rectifying channels as intracellular polyamines block AMPA pore when cell is depolarized
• have variable permeability to calcium
  • Q/N/R site in P-region determines the ion selectivity of channel
• in clacium-impermeable subunits like GluR2, presence of R (arginine) blocks passage of larger ions and reduces conductance
• in calcium-permeable subunits have no R @ QNR site in pore allowing passage of Ca⁺⁺
• AMPAR current
  • extremely fast influx of cations followed by fast desensitization
  • desensitization is the result of conformational change in continuous presence of ligand

• more modulatory, smaller role than AMPA receptors
• electrophysiologically very similar to AMPA receptors
• many times kainate and AMPA are grouped together as non-NMDA receptors

• ionotropic glutamate receptor
• must have at least one GluN1 subunit
• GluN2 required for binding of glutamate
• commonly contain a mixture of GluN1 and GluN2 subunit types
• GluN1 and GluN3 can bind coagonists glycine and D-serine
• subunit composition determines functional properties
• Important properties
  • both glutamate and glycine are required for channel to open
  • highly permeable to calcium (5-10x more than most non-NMDA receptors)
  • strongly voltage-dependent as well
    • @ hyperpolarized potentials Mg⁺⁺ can enter pore and block conductance
    • w/ depolarization Mg⁺⁺ can be pushed out of pore allowing passage of ions
  • To open...
    • glycine and glutamate must be bound
    • cell must be depolarized
    • p(open) is both ligand- and voltage- dependent
    • therefore, NMDA IV plots have biphasic shape
  • IV plot

[image]

• coincidence detection
  • because, NMDA conduction relies on cell being depolarized, it makes it less likely to activate randomly w/ion release
• Long-term potentiation
  • proposed mechanism for many processes including memory storage
  • NMDA activation causes increased Ca⁺⁺ conductance and concentration resulting in CAMKII activation and phosphorylation of AMPA receptors + insertion of AMPA receptors = increased AMPA conductance
  • this increases synaptic strength following high-frequency stimulation
  • leads to greater postsynaptic response

1.     Brain insult occurs, such as a stroke

2.  Blood flow is cut off to region of brain

3.  Insufficient glucose reaches neurons

4.  ATP concentration decreases

5.  Na/K pump is inhibited

6.  Neurons depolarize

7.  Ca2+ channels are activated

8.  Neurotransmitters, including glutamate, 

are released

9.  Glutamate receptors are activated

10. Mg2+ unblocks NMDA receptors due to 

depolarization

11. Massive Ca2+ influx occurs through NMDA 

receptors

12. High intracellular [Ca2+] leads to cell death 

• rule of thumb: fire together, wire together
• as synapses begin to form NMDAR's can strengthen synapses in a way that both imparts specificity and associativity of action
  • specificity
    
    • neuron may be actively projecting to glutamatergic neuron @
    • one part of cell while another neuron is not actively projecting
    • this results in strengthening of stimulus individually from other point of contact
  • associativity
    • if one neuron is firing a strong stimulus to one part of neuron while another neuron fires a weak stimulus elsewhere, both synapses will be strengthened together
    • results in an association being made between synapses 

• allows passive flow of current through gap junctions
  • gap junctions made up of aligned paired channels called connexons that create passages for ion diffusion between separate cells
  • each connexon has 6 connexins
  • each connexin has 4 transmembrane regions
  • cause extracellular space separating cells to shrink from ~20nm to 3.5nm
• cytoplasm is basically continuous between cells
• this allows for flow of current in either direction and current will flow regardless of amplitude (AP's not required)
• junctions will also pass both positive and negative currents
• this also increases the speed of propagation from one cell to another minimizing the delay as current freely flows into the next cell

[image]

[image]

• Fast transmission of current.
• Synchronize networks of neurons.
• Communication between glial cells.

• @ electrical synapses, free ion flow from one cell to another typically allows for current flow and potential changes in bidirectionally
• @ rectifying synapses, the synapse is designed in such a way that the current flow is unidirectional
  • accomplished through a rectifying current that prevents back-propagation of potential change
  • important as bidirectional flow can decrease efficiency and cause unwanted information flow 

• agonists
  • acetylcholine
  • nicotine
• potentiators/ cholinesterase inhibitors
  • neostigmine and physostigmine
• antagonist
  • turbocurrarine
  • currare

• NMJ is a chemical synapse (i.e. Ach binds to receptors causing increased conductance and current flow)
• end plate potentials are almost always suprathreshold
  
  • AP is almost always elicited
• if curare is added, we can see that muscle fibers exhibit passive cable properties similar to dendrites in that the AP dissipates over distance
  • if curare is removed the AP will propagate down the muscle fiber
  • shows the cable properties of the fiber as well as the fact that muscle contraction must be elicited by AP
• through studies of direct addition of Ach to muscle fiber, it has been shown that AchR's are localized @ NMJ's
  • as Ach is added farther and farther from NMJ, response/EPP is less and less
  • allows us to break fiber into synaptic regions (craters) of highest concentration of AchR's, rim, and extrasynaptic regions w/ lowest concentration

• chemical that permanently binds nAchR's
• extremely useful for visualizing NMJ's and receptors for this reason

• using patch-clamp technique, hold voltage @ various potentials while adding acetylcholine or neurotransmitter of interest to channel/muscle fiber/neuron of interest
• find the potential @ which the current passed is equal to 0mV

• synaptic bouton projects onto muscle fiber end plate
• end plate lined w/ junctional folds which maximize the surface area and penetrate into the fiber
• nAchR's line the top of the junctional folds allowing for EPP to occur while the Na⁺ voltage-gated channels line the bottom 

[image]

• 5 subunits
• 4 transmembrane regions/subunit
• Acetylcholine binding is *permissive* for channel opening
• In other words, 2 Ach molecules must be present for the channel to open, but the channel will not *always* be open when ACh is present
• Like all ion channels, the precise timing of channel opening and closing is stochastic and is due to random vibrations of the molecule
• It is impossible to know which one will result in a change in channel confirmation, but we can calculate average behavior of the channel by recording many, many channel openings.

General Mechanisms of Termination

• Diffusion away from the binding site
• Enzyme degradation
• Transport across a membrane (reuptake)

@ NMJ

• ACh – degraded by enzymatic degradation (no reuptake – makes ACh unique)
  • acetylcholinesterase
  • At high [ACh] AChE is not as effective.

• autoimmune disease in which body produces abnormal antibodies that attack and degrade or destroy nAChR's on skeletal muscles
• this leads to muscle weakness due to decreased active receptor sites
  • Fewer AChRs results in smaller amplitude EPPs.
  • Over time, there is a decrease in force of muscle contraction as individual muscle fibers no longer reach threshold 
• Symptoms
  • drooping one or both eyelids
  • double vision
  • altered speaking
  • difficulty swallowing
  • problem chewing
  • limited facial expressions
  • weakness in arms, legs, neck, fingers, etc.
  • weakness in chest muscles sometimes
    • if severe, may result in myasthenic crisis

• acetylcholinesterase inhibitors are first line of defense 
  • physostigmine 
    • slows the breakdown of acetylcholine at the neuromuscular junction and thereby improves neuromuscular transmission and increases muscle strength
    • also worsens weakness w/ sustained activity
• Treatment allows significant relief from weakness.
• Most patients live a full life with normal life span
• Occasionally, emergency intervention is required if breathing muscles become too weak.

• GABA receptors
  • GABA_A
  • GABA_C
• Glycine and its inhibitory glycine receptor
• All of these receptors are in the nAchR family and therefore share similar structure

• depolarizing or hyperpolarizing currents are not what determine whether a channel is excitatory or inhibitory
  • @ any given time, depending on Vm, the direction and therefore type of current can switch based on DF
• it is the reversal potential of the channel that matters to this denotion
• excitatory
  • excitatory channels are supposed to make AP firing more likely -> bring cell closer to AP threshold when below threshold
  • excitatory channels, therefore, can be denoted by a V_rev that is more depolarized than the AP threshold
• inhibitory
  • inhibitory channels are supposed to make AP firing less likely -> bring cell farther from AP threshold when below threshold
  • inhibitory channels, therefore, can be denoted by a V_rev that is more hyperpolarized than the AP threshold

• GABA reversal potential changes during development
• E_GABA becomes hyperpolarized w/ development
  • given AP threshold of -50mV, GABA is actually excitatory for the first 7 days of development 
  • by day 14, GABA becomes inhibitory as its V_rev becomes more hyperpolarized
• Why?
  • During development, intracellular levels of chloride decrease, resulting in a more hyperpolarized ECl (and therefore EGABA)
  • increased intracellular chloride concentration achieved by increasing expression of the KCC2 transporter (potassium-coupled, chloride transporter)

• spatial summation - summation of graded potentials conveyed from different points of neuron
  • The amplitude of PSPs at the “trigger zone” (i.e. axon hillock) will depend on the input resistance of the cell and the length constant
  • The larger the resistance, the greater ΔV will be produced by a given current (V = IR)!
  • The farther the PSP has traveled, the more it will have decayed.
  • The greater the length constant of the dendrite, the farther the PSP will travel
• temporal summation - summation of graded potentials conveyed @ different times
  • Slower time course of distal events allows for greater degree of temporal summation because a greater percentage of the potential will be extant at the axon hillock
  • The time constant of the cell will determine decay of PSPs, thus affecting summation (tau = R*C)
• IPSPs can also summate both spatially and temporally
• axon hillock
  • The membrane potential at the axon hillock is the summation of all EPSPs and IPSPs received by the neuron.
  • It is the membrane potential at the axon hillock that determines if an action potential will be fired
• synapses closer to the axon hillock have a larger effect than those @ a distance due to the decremental nature of PSPs

• summation of inputs occurring close together
  • the total EPSP experienced from two inputs, A and B, are not the same as the sum of A+B combined
  • one possible reason is channel availability and that when together, A and B have to share the channels
  • the more likely reason especially in consideration of greater distances is that the DF of one input affects the ion entry due to the other input
• when an input from farther away is considered, the total EPSP created by two inputs, A and C, are much closer to the sum of A+C than when close together
  • some effect of DF will likely be experienced, but the total EPSP should be nearly equal to the sum of A+C, especially when the other input comes from a different branch non-adjacent to the other stimulus