Purves et al. Chapter 5
Cell theory (cells being separated) implies that cells must communicate
with each other through extracellular connections and most communication
is through chemical messages
- "synapse" - = "clasp" (sherrington)
Fig. 5.1AB p. 78 "electrotonic synapse"
Figure of chemical synapse is like Fig. 5.3 p. 80 (only simpler)
Vesicles, spine, receptors (the ionotropic type, i.e. channels) are shown
Johnnie Moore took this course in 2002. She published this paper: Moore
et al., Stable gene silencing of synaptotagmin I in rat PC12 cells inhibits
Ca 2+-evoked release of catecholamine, Am J. Physiol Cell Physiol 291, 270-281,
Topics to discuss:
Fig 5.1A B p. 78
Gap junction is an exception to the above generalization in that cells are
coupled electrically with cytoplasmic continuities (small ones).
Gap junctions are used in crayfish escape,
"Landmark" paper EJFurshpan and DDPotter, Mechanism of
nerve-impulse transmission at a crayfish synapse, Nature 180, 342, 1957
and J. Physiol. 145, 289, 1959. "Landmarks" was a department in
The Journal of NIH Research, includes interviews. (ref=vol 8, Oct 1996,
They are also used to connect myocardial cells electrically at intercalated
Conductance is high - 120 pS.
Fig 5.1C p. 78
Gap junction is patch of hexamers forming big channel in register with adjacent
Proteins, called connexins, are very diverse.
They are often named with a number that represents the molecular weight.
Recent paper CDLandisman & BWConners, Long-term modulation of
electrical synapses in the mammalian thalamus, Science 310, 1809-1813, 2005.
There are gap junctions with connexin36 (Cx36) in thalamic reticular nucleus
(TRN). There are chemical synapses using metabotropic glutamate receptors
(mGluRs) from neocortex. Activating this corticothalamic path causes long-term
decrease in electrical synapse strength. This is like long term depression,
a model for learning at the level of the chemical synapse.(Each item of
terminology that might be unfamiliar to you now will be covered later.)
Pore is big enough to give cytoplasmic continuity for medium sized molecules
In addition to electrical coupling, there can be communication by molecules.
In EM, membranes appear very close but not fused
Extracellular tracers (heavy metal Lanthanum) proves there is extracellular
In 1906 Sir Charles S. Sherrington (England) published Integrative Action
of the Nervous System and later (1932) won the Nobel
prize for "functions of neurons." He coined the term "synapse."
In studies of the spinal reflex, he determined that the spinal motor neuron
was the "final common pathway" (for integrative action of the
nervous system). Spinal reflexes were studied in spinal animals in which
the spinal cord was transected to prevent descending cortical influence.
There are no inhibitory neuromuscular junctions in vertebrates, so whether
the spinal motor neuron fires, based on summed inhibitory and excitatory
influences, is the last chance to integrate neural influences.
Fig. 5.4 p. 81
1926 Otto Loewi (Austria) experiment he dreamed, stimulate vagus (10th cranial
nerve, parasympathetic), take substance and show that it slows a heart in
another dish, vagus substance = acetylcholine (ACh) a monamine transmitter.
1930's Sir Henry H. Dale (England) acetylcholine
share 1936 Nobel
"chemical transmission of nerve impulses"
Fig. 5.22AB p. 103
Sir John C. Eccles 1963 Nobel
(with Hodgkin & Huxley) EPSP & IPSP
Excitatory or Inhibitory PostSynaptic Potentials (Eccles, using spinal motor
Excitatory and Inhibitory integrate in cell, and axon hillock "decides"
whether to fire.
Fig. 5.21A p. 102
glutamate is the excitatory transmitter
EPSP - depolarize (unless clamped positive to reversal potential)
increase sodium and potassium conductance
inferred because reversal potential is near zero (in voltage clamp)
Fig 5.21BC p. 102
GABA (gamma amino butyric acid) is the inhibitory transmitter
IPSP - hyperpolarize (unless clamped negative to reversal potential)
increase potassium and chloride conductance
inferred because reversal potential negative to resting potential (in voltage
and by changing Cl- gradient by using ion specific electrodes to inject
Sir Bernard Katz (England) Ulf von Euler (Sweden) Julius Axelrod (US) "humoral
transmitters...nerve terminals....storage release inactivation"
Fig. 5.7A p. 86
classic experiment by Katz showing that the transmission at the neuromuscular
junction is "quantal." Quantum is one vesicle. EPP (end plate
potential) is reduced to meep's (miniature end plate potentials, 0.4 mV)
by lowering extracellular Ca2+ ion, and nerve stimulation elicits responses
the size of 0, 1, 2, or 3 meep's according to the Poisson distribution.
"end plate" potential is big and effective in generating muscle
action potential. Usually 200 vesicles give 40 mV potential.
Fig. 5.8 A p. 87
Here is a classic pictures, work by Hueser and Reese, of vesicle release
at the neuromuscular junction, a freeze
fracture electron micrograph
also a transmission electon micrograph
(Heuser) where the vesicle release is called an omega figure because it
is shaped like the Greek letter.
Just to put into perspective the degree to which much of this information
TRANSPARENCY shows the version of this picture the Biology Department teaches
Specifics shown in this figure:
Ca2+ enters presynaptic terminal upon arrival of the action potential.
The receptor shown is a channel passing Na+ (this situation can vary).
Neurotransmitter is broken down (true for acetylcholine, but this situation
Fig. like Fig 5.3 p. 80 (only simpler)
Presynaptic membrane, cleft, Postsynaptic membrane (intracellular density
seen in EM [electron microscopy]), vesicle
Specifics in this figure
Note that the post-synaptic membrane is up on a spine.
Membrane is recycled, and endocytotic pits and vesicles are coated (with
clathrin); coated pit
from my work another
(not related to synapses).
Fig 5.13B p. 91
There is a protein called dynamin that helps pinch off vesicles. It is the
product of the temperature sensitive Drosophila paralytic mutant called
shibire. At restrictive temperature, there is a block
in endocytosis of vesicles (another view).
vesicles and T-shaped ribbons in Drosophila
Here is a transmission electron micrograph of a synapse
Vertebrate - inputs to cell or dendrite (spine)
Invertebrate - cell is usually away from action surround "neuropil(e)."
Here is a picture from
my work on Drosophila, retina (compound eye is off top, cartridges of synaptic
connections are at bottom, cell bodies of post-synaptic neurons are between.
General: vesicles are interesting, transmitter is very concentrated, there
are pumps to move transmitter "up hill" (against gradient) into
vesicle, sometimes part of synthesis is in vesicle.
Fig 5-13, p. 91
very modern, interesting and detailed
also, interesting Box 5B, pp. 93-95, on diseases and toxins that affect
there are vesicle membrane proteins, target (presynaptic) membrane proteins,
and cytoplasmic proteins
Ca2+ in through Q or N type voltage gated channel
(N stands for "neither," as opposed to T=transient or L=long lasting,
the N channel is blocked by omega toxin from Conus [snail genus])
Synaptobrevin / VAMP (vesicle-associated membrane protein) = v-SNARE (SNAP
Botulinum and Tetanus toxin (clostridial toxins) are proteases which cleave
Botulism (Clostridium botulinum) anaerobic, improper canning (need to heat
to kill spores) - block release
When I ws 10, in the Cold War, we discussed, at the dining room table, how
1 teaspoon in the reservoir would kill the city. Now. 45 years later, people
take it (injected) to get rid of face wrinkles.
Tetany is term for sustained muscle contraction based on twitches adding
Tetanus toxin cleaves synaptobrevin in inhibitory interneurons.
The disease is contracted in deep (because it is an anaerobic bacterium)
dirty puncture wounds.
You would die with muscles contracted, called "lock-jaw."
There is a vaccine and boosters every 10 years are suggested.
Synaptotagmin - binds calcium
synapsins get phosphorylated (by CaM Kinase II and PKA) interact with actin
Target membrane proteins:
Syntaxin = t-SNARE = unc-18 (uncoordinated C. elegans roundworm mutant)
Neurexin - black widow spider venom (alpha Latrotoxin) causes too much release
Neuroexins bind to synaptotagmin
NSF - N-ethylmaleimide sensitive factor (ATPase activity when complex dispersed)
SNAP - soluable NSF associated protein
Rab3 (like ras, small GTP binding protein) (lots of rab's, specific for
Fig. 5.14 AB p. 92
SNAREs and SNAP (docking)
In addition to SNAREs and SNAP, Ca binding synaptotagmin is for fusion
Fig. Box 5B pp. 93-95
BoTX and TeTX sites.
Test questions from 2005 -2012 that relate to this outline
Why would pheochromocytoma cells be a reasonable model for studying neurotransmitter
can culture adrenal medulla cancer cells which release catecholamines
"Ramon y Cajal's 'cell theory' implies that something like a chemical
synapse MUST exist (even though they has not been demonstrated yet)"
the alternate, Golgi's reticular theory, has continuous processes, while
separate cells require communication between cells
Describe the structural or molecular specializations that form an "electrical
in gap junction, connexons are formed from hexamers of connexins in register
from one cell to the next
"There are no inhibitory neuromuscular junctions in the vertebrate."
What does that information tell us about "the final common pathway
in the integrative action of the nervous system?"
it must be in the spinal motor neuron since there is no integration after
What kind of technique would be needed to determine the reversal potentials
for the IPSP and the EPSP?
Activation of the nerve elicited (what? - be specific) at the end plate
when Katz decreased the extracellular calcium ion concentration.
miniature end plate potentials, 0, 1, 2, or 3 of them
Describe either the freeze-fracture or the transmission electron microscope
image of a vesicle in the process of release.
holes (pits), omega figures
Why would there be transporter molecules in the vesicle membrane?
to maintain a steep gradient against leakage
On the spinal motor neuron, there are synapses onto the dendrites and cell
body. A picture was shown of the first synaptic area behind the Drosophila
compound eye, and the point was made that invertebrate neuropils we would
see later in the semester are similarly organized. How does this synaptic
organization differ from that of the vertebrate?
the cell body of the postsynaptic neuron is on the outside of the ganglion,
aloof from synapses
How could Shibire mutants have temperature sensitive paralysis when the
mutation does not block vesicle release?
it blocks recycling of vesicle membrane
Why would a doctor give a patient a clostridial protease that cleaves synaptobrevin?
cosmetic dermatologists would discourage wrinkles in the face by blocking
What is the target membrane that the vesicle-SNARE latches onto?
What nerve did Otto Loewi from Austria use in the first demonstration that
a neurotransmitter substance was used in signaling?
A patch with hexamers in register between two adjacent cells describes what
kind of intercellular communication structure?
"The spinal motor neuron is the final common pathway in the integrative
action of the nervous system." Why is the spinal motor neuron (as opposed
to the muscle cell) the final place where integration of signals can take
there is excitation, no inhibition, on the muscle cell
Starting at a normal resting potential (not voltage-clamped), what happens
to the membrane potential of the postsynaptic cell if a GABA-releasing presynaptic
cell is activated?
A micropipette that was used to selectively inject chloride ions into neurons
told us (what) about postsynaptic potentials?
IPSPs are based in part on an increase in chloride conductance
In Sir Bernard Katzís Nobel Prize-winning research, he turned end
plate potentials into 0, 1, 2, or 3 miniature end plate potentials. Answer
either (1) How? Or (2) Why?
(1) by decreasing calcium ions (2) to show that vesicles were the unit of
Name a protein involved in retrieval of vesicle membrane.
After calcium ions come in through channels in the presynaptic membrane,
it binds to a calcium-binding protein to mediate vesicle release. Answer
either (1) What is its name? or (2) Where is it (which specific compartment
involved in vesicle release)?
(1) synaptotagmin (2) bound to vesicle membrane
Why do they refer to "omega figures" in the ultrastructure of
vesicles in the process of release are shaped like Greek letter omega
What kind of synapse involves opening of a chloride channel?
Suppose you voltage clamp a postsynaptic neuron to a level negative to the
reversal potential. What would you record (postsynaptically) if you activated
a cell making a GABAergic
in this case it would depolarize
Name a protein involved in recycling of synaptic vesicle membrane.
What is the quantum of transmission that Katz found to elicit a miniature
end plate potential of 0.4 mV?
Compared with the conductance of a voltage-gated ion channel, what is the
conductance of a one gap junction channel?
What happens to the conductance of the postsynaptic menbrane during the
What is a connexin used for?
How does Ca2+ get into the cell to affect vesicle release?
through a calcium channel
What do the clostridial toxins from anaerobic bacteria do?
cleave Synaptobrevin / VAMP (vesicle-associated membrane protein) = v-SNARE,
inhibit vesicle release
Katz was interested in the end plate potential. Where is the end plate?
the "post-synaptic membrane" of the muscle cell
Two SNAREs hook onto eachother, v-SNARE and t-SNARE. What are v- and t-?
vesicle and "target" (presynaptic membrane)
What happens to the end plate potential when extracellular Ca2+ is decreased?
Why do tetanus and botulinum toxins have opposite effects on motor activity?
both inhibit vesicle release, tetanus is in inhibitory interneurons
What would not happen at the restrictive temperature in a temperature sensitive
mutation affecting the protein dynamin?
coated pits would not pinch off to vesicles
Why did Sherrington consider the spinal motor neuron rather than the muscle
cell itself to be the "final common pathway" of "the integrative
action of the nervous system?"
because it receives + and - inputs while the muscle only gets + input
What happens to the conductance at the postsynaptic ionotropic receptors
for K+ and Cl- for the IPSP?
they go up
Not just any old Ca2+ channel would work on the presynaptic membrane. It
has to be voltage gated. Why?
to detect the arrival of the action potential
What are connexin proteins, such as Cx36, used for?
With extremely low extracellular Ca2+, why would the potential recorded
at the end plate be several different sizes for several different nerve
because you might get o, 1, 2, ... miniature end plate potentials
What does parasympathetic output via the vagus do to the heart rate?
The toxin from Clostridium botulinum (BoTX) cleaves an important
protein. Precisely where is this protein localized?
on the vesicle
Why are vesicles in the process of release sometimes called omega figures?
a TEM of it is shaped like the Greek letter
Electrical "synapses" use what membrane specialization in common
with heart muscle cells?
gap junctions with connexons made up of connexin protein
What happens to the conductance of the postsynaptic membrane when GABA (gamma
amino butyric acid) elicits an IPSP (inhibitory postsynaptic potential)?
increases (for potassium and chloride)
"Activating the corticothalamic path causes long-term decrease in electrical
synapse strength." These electrical synapses are made of what protein?
Why did the Nobel prize winning Sir Charles Sherrington refer to the vertebrate
spinal motor neuron as "the final common pathway in the integrative
action of the nervous system?"
there can be no integration (of + & - inputs) further out since each
muscle cell has only (one) + connection
Graded depolarizations and hyperpolarizations on the dendrites and cell
body pool. As a result, an action potential is initiated (where)?
Determining reversal potentials was instrumental in establishing the conductances
mediating the EPSP and IPSP (excitatory and inhibitory postsynaptic potentials).
What was done, in addition to Voltage clamping and recording from the postsynaptic
membrane, to determine the reversal potentials?
stimulate the presynaptic neuron
With very low Ca2+, upon stimulating the motor neuron's axon, the end plate
potential (EPP) was usually 0.4, 0.8, 1.2, etc mV. What other size was observed?
Recycling of vesicle membrane begins with a clathrin coated pit. A mutation
of what protein prevents the pit to pinch off to a vesicle?
dynamin (the shibire gene product)
There are synaptic connections to the dendrites and cell bodies of vertebrates.
How is the situation strikingly different for invertebrates?
cell bodies are on outside of neuropil, away from where connections are
If heat resistant bacterial endospores survive improper canning, by what
mechanism would eating those tomatoes affect transmitter action?
bacterial botulism toxin would cleave synaptobrevin (v-SNARE)
Give one of the several reasons why alumni Michelle Li, also Johnnie Moore,
used PC12 cells in their published research.
they release catecholmines
Compare the conductance of a gap junction channel with that of a potassium
What's with the "36" when they call the gap junction protein in
the thalamus "Cx36?"
How did Nobel Prize winner Loewi prove that a substance released by the
vagus slows the heart?
solution bathing heart slowed by vagus stimulation slows another heart
What property of the neuromuscular junction rationalizes Sir Sherrington
referring to the spinal motor neuron as "the final common pathway"
of "the integrative action of the nervous system?
since there is only excitation at the vertebrate n.m.j, the motor neuron
is the last place excitation and inhibition can integrate
If you observed a coated vesicle near a synapse, say whether it is in the
process of exocytosing and justify your answer.
no. endocytosing, those are the pits with clathrin
Relate vesicle release with either (1) improperly canned goods, (2) dangers
of terrorism against civilians, or (3) facial cosmetic treatment.
BoTox decreases release, form heat resistant endospores turning into anaerobic
bacteria that make a potent toxin that can be injected to decrease face
Why, in Sir Bernard Katz's Nobel Prize-winning experiment, did they refer
to "end plate potentials" instead of "postsynaptic potentials?"
neuromuscular junction was used to model a synapse
"Silencing of synaptotagmin in PC12 cells inhibits Ca2+-evoked catecholamine
release." Why does calcium elicit release if synaptotagmin is not inhibited?
Calcium, coming in through calcium channels, mediates vesicle release
Consider the mepp (miniature end plate potential) and answer either (1)
What cellular mechanism delimits it to be way smaller than the full end
plate potential? Or (2) Suppose your preparation were giving you just a
few mepps on the average ­p; what would you do to the preparation to
restore the full end plate potential?
Fewer vesicles are released with low extracellular calcium, add calcium
There are synaptic connections to the dendrites and cell bodies of vertebrates.
How is the situation strikingly different for invertebrates?
Synapses are in a neuropil(e) and cell bodies are on the outside of ganglia
At first they were surprised that a mutation in dynamin (shibire) would
exhibit paralysis when moved to the restrictive temperature. Rationalize
If vesicle membrane fails to recycle, eventually vesicles would not be released
"Botulism and tetanus toxins cleave synaptobrevin." Precisely
what membrane is synaptobrevin on?
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