Bi / CNS 150 Lecture 8
Synaptic inhibition; cable properties of neurons;
electrical integration in cerebellum
Wednesday, October 15, 2013
Henry Lester
Chapter 2 (p. 28-35);
Chapter 10 (227-232)
1
Nicotinic ACh, GABAA , and glycine receptors look alike at this resolution (prev. lecture)
~ 2200
amino acids
in 5 chains
(“subunits”),
Binding
region
MW
~ 2.5 x 106
Membrane
region
Colored by
secondary
structure
Colored by
subunit
(chain)
Cytosolic
region
2
The pentameric GABAA and glycine receptors look like ACh receptors;
but they are permeable to anions (mostly Cl-, of course)
1. -amino-butyric acid (GABA) is the principal inhibitory transmitter in the brain.
2. Glycine is the dominant inhibitory transmitter in the spinal cord & hindbrain.
GABAA receptors are more variable than glycine receptors in subunit
composition and therefore in kinetic behavior.
. . Cation channels become
anion channels with only
one amino acid change per
subunit, in this approximate
location
Like a previous lecture
A Synapse “pushes” the Membrane Potential
Toward the Reversal Potential (Erev) for the synaptic Channels
ACh and glutamate receptors flux
Na+ and K+,
(and in some cases Ca2+),
and Erev ~ 0 mV.
At GABAA and glycine
receptors,
Erev is near ECl ~ -70 mV
Membrane potential
+80
At Erev , the current through
open receptors is zero.
ENa
+60
Positive to Erev, current flows
outward
+40
Negative to Erev, current
flows inward
-5
+20
-20
Resting -50
potential
-80
EK
-100
Like Figure 10-11
4
Pharmacology of GABAA Receptors: activators
Benzodiazepines (= BZ below):
Valium (diazepam),
(Ambien, Lunesta are derivatives)
The natural ligand binds at
subunit interfeces
(like ACh at ACh receptors)
phenobarbital site is unceratin
5
The AChBP interfacial “aromatic box” occupied by nicotine (prev. lecture)
. . . GABA and glycine also make cation-p interactions
aY198
C2
aW149
B
aY93
A
aY190
C1
non-aW55
D
(Muscle Nicotinic numbering)
6
GABAA and Glycine blockers bind either at the agonist site or in the channel
Strychnine
Bicuculline
(glycine
receptor)
(GABAA receptor
Agonist
site
Picrotoxin
(GABAA & glycine
receptors)
How does the receptor transduce binding into channel gating? (prev. lecture)
. . . Both ideas are
also in play for
GABA or glycine
receptors
Swivel?
Miyazawa,
Nature
2003
CLOSED
OPEN
Twist?
Corringer,
J Physiol
2010
8
We have Completed our Survey of Synaptic Receptors
A. ACh, Serotonin 5-HT3, GABA,
(invert. GluCl, dopamine, tyrosine)
receptor-channels
Most
^
Figure 10-7
9
Parts of two generalized CNS neurons
Postsynaptic
neuron
Presynaptic
neuron
Excitatory Inhibitory
terminal terminal
axon
node of
Ranvier
initial
myelin
segment
apical
dendrites
(apex)
little hill
axon hillock
cell
(base)
basal
body
dendrites
(soma)
presynaptic
nucleus
terminal
presynaptic
terminal
postsynaptic
dendrite
synaptic cleft
direction of information flow
Like Figure 2-1
(rotated)
10
The cerebellum: a famous circuit in neuroscience.
In today’s lecture,
it exemplifies pre- and postsynaptic structures.
Molecular
layer
Purkinje
cell layer
Ganule
cell layer
White
matter
10% of the neurons
in the CNS are
cerebellar granule cells
Figure 42-4
11
A plurality of synapses in the CNS (> 1013 ) occur between
parallel fiber axons and Purkinje cell dendritic spines
Molecular layer
500 nm
12
Types of synapses
(Don’t mind the Type I, Type II stuff)
Figure 10-3
13
Types of synaptic integration
1. Temporal
A. Molecular lifetimes
B. Capacitive filtering
2. Spatial
3.
Excitatory-inhibitory
14
Previous lecture
Synaptic integration 1A.
Molecular lifetimes
Concentration of
high
acetylcholine at
NMJ
(because of
0
acetylcholinesterase,
turnover time
~ 100 μs)
State 1
closed
State 2
k21
all molecules
begin here at
t= 0
open
units: s-1
Number of open
channels
ms
15
At the nerve-muscle synapse, acetylcholinesterase is present
at densities of > 1000 / μm2 near each synapse,
high enough to destroy each transmitter molecule
as it leaves a receptor
What causes the ~ δ-function of glutamate & GABA at CNS synapses?
Na+ -coupled transporters for glutamate & GABA
are present at densities of > 1000 / μm2 near each synapse,
probably high enough to sequester each transmitter molecule
as it leaves a receptor
(more in a few slides).
16
Synaptic Integration 1B. Capacitive filtering
I C  C dV
Figure 9-6
dT
;
V  C  IC
17
1B. Temporal Summation
2. Spatial summation
Recording
Recording
Axon
Axon
Synaptic
Current
Synaptic
Current
~ 100 pA
Synaptic
Potential
Synaptic
Potential
Long time constant
(100 ms)
Vm
Short time constant
Vm
(20 ms)
I C  C dV
dT
;
V  C  IC
Long length constant
(1 mm)
Short length constant
(0.33 mm)
2 mV
25 ms
Improved from Figure 10-14
18
1. If dendrites were passive, they would act like leaky cables . . .
Excitatory synapses
V
EPSP measured in soma
Gulledge & Stuart (2005) J. Neurobiol 64:75,
V
EPSP measured in dendrite
19
. . . and passively integrate inputs . . .
V
V
V
Δt = 0
Δt = 0
Δt = 5 ms
Prolonged
rising phase
Simultaneous,
colocalized
EPSPs
(two individual trials)
Nearly simultaneous,
colocalized
EPSPs
(two individual trials)
Simultaneous,
Spatially distinct
EPSPs
Inspect the simulation, and run the movie, at
http://www.neuron.yale.edu/neuron/static/about/stylmn.html
Gulledge & Stuart (2005) J. Neurobiol 64:75,
20
. . . but two-photon
microscopes allow
researchers to visualize
patch-clamped dendrites in
living animals . . .
Gulledge & Stuart (2005) J. Neurobiol 64:75,
21
. . . dendrites are not passive. They have Na channels
Now break the patch,
to fill the cell with dye:
* = axon hillock
Averaged traces
25 μm
Magee & Johnston, J Physiol (1995)
immunocytochemistry
Whitaker, Brain Res, 2001
22
brain slice
. . . voltage-gated Na+ and Ca2+ channels
in dendrites
lead to
partial “backpropagation”
of
action potentials,
implying
that parts of cells
can process signals
semi-independently.
Stay tuned!
Gulledge & Stuart (2005) J. Neurobiol 64:75,
23
3. Excitatory-inhibitory integration:
The “veto principle” of inhibitory transmission
Inhibitory synapses work best when they are “near“ the excitatory event they will inhibit.
“Near” means < one cable length.
A. Inhibitory synapses on dendrites
do a good job of inhibiting EPSPs on nearby spines
B. Inhibitory synapses on cell bodies and initial segments
do a good job of inhibiting spikes
24
“Veto” inhibition at the axon initial segment:
Schematic of a GABAergic “chandelier cell” in
human cerebral cortex
Inhibitory
Chandelier
Cell
Ch terminals
Ch.
axon
Pyramidal
Cells
Ch terminals
from Felipe et al, Brain (1999) 122, 1807
25
Now we localize the inhibitory “vetos”
of cerebellar Purkinje cells
by “pinceaux” (paintbrushes) of basket cells
Molecular
layer
Purkinje
cell layer
Ganule
cell layer
White
matter
Figure 42-4
26
How to localize and quantify inhibitory synapses
NH2
A fusion protein:
GABA transporter (GAT1)-GFP
27
cerebellum
28
<Immunocytochemistry
For GABA transporter
Molecular layer (basket cells stain)
Purkinje cell layer
“pinceux” (paintbrushes)
stain heavily
Granule cell layer
29
<Immunocytochemistry
For GABA transporter
mGAT1 GFP
knock-in fluorescence >
Molecular layer
(basket cells stain)
Purkinje cell layer
“pinceaux” stain
heavily, showing
soma-hillock “veto”
Granule cell layer
30
GAT1-GFP expression in cerebellum:
basket cell terminals in molecular layer,
Showing dendritic “veto”
GABA transporter density is ~1000/(μm2)
50 mm
31
End of Lecture 8
32