Synapses and Signalling Flashcards
electrolytic theory (Watter Nernst)
explains existence of membrane potential in living cells
net movement of solute is comb. of electrical potential and chemical across membrane
membrane ions on either side at resting potential
high conc. sodium outside, low inside
high potassium inside, low outside (potassium poores open without stimulation)
excitable cells
non excitable cells
can generate AP
do not generate action potentials but exhibit dynamic changes in resting membrane potential - glial, immune, epithelial, tumour
history of nerve discoveries
lecture 6 bottom of page
patch clamp
& equipment
cell in bath chamber surrounded by extracellular medium
glass pipette contact cell body
record activity in patch of membrane
(can record single channels)
sodium into cell measured as -ve current, potassium out is +ve
need micromanipulators to position pipette, faraday cage to reduce electrical noise, air table reduce vibration, clean solutions, smooth pipette tip
excised patches
take out bit of membrane and expose outer/inner to extracellular liquid which can change rapidly
can control voltage and change inner/outer solutions
but can change channel properties so not good
whole cell patch clamp
records from all ion channels
control potential and current
can change inside/out solutions
but leakage current can damage membrane
command voltage
set membrane potential in patch clamp
if change from -60mV to 30mV then currect response changes and peak increases
transient current (shortlived)
the big AP dip can see is mediated by sodium because still see effect if block potassium, rapidly inactivates
if block sodium - isolate outward SLOW current
Hodgkin-Huxley model significance
introduced concepts of ionic channels as separate molecular structures
confirmed by molecular bio
provide good numerical description by experimental data, foundation for computational neuroscience
open probability
of sodium channels - v high initially then almost 0 when inactivation (similar as predicted by H-H)
potassium probability doesn’t change with time because don’t inactivate
potassium channel types
delayed rectifier A-type/transient inward rectifier BK calcium dependent (large conductance) SK calcium dependent (small conductance) K-ATP (ATP dependent)
delayed rectifier (K channel)
strong dependence on membrane voltage
slow kinetics
sodium channel types (and why they’re diff/similar)
I II IIA III VI
more similar to each other, diff sesitivity to antagonists and diff thresholds
voltage activated calcium channels
if block sodium and potassium channels, the current measured resembles inward sodium but smaller amplitude
so this calcium activity is normally masked by sodium activity
mostly in pre-synaptic terminal for calcium entry for NT release
calcium channel types
LVA/T-type (low voltage activated, below -30mV) - rapid
HVA (high) - above -30mV, split into more types depending on pharmacology/location/threshold/kinetics
structure of ion channels
all same except inward rectifies and K-leaks
6 subunits
4 homologous subunits forms transmembrane pore
each 4 subunits contains 6 transmembrane segments S1-S6
all 4 S5 and S6 segments form internal pore of channel with ion selectivity filter
S4 sensitive to voltage - responsible for gating
S4
12 +ve AAs move in response to voltage, relax pressure on S5 and S6 so open channel
so 4 gates but 3 enough to open channel and 1 inactivation gate
how do ion channels close?
polypeptide structure at bottom closes channel
inactivation gate
1 per channel
between S6 of subunit III and S1 of subunit IV
gating and opening of Na vs K
Na - rotation sliding movement of subunits and +ve residues move outward so widen pore
K - more like paddle movement of S1-S4 alpha helices
inactivation and activation of Na
need to be open and activated before inactivating gate can go inside pore
why is ion selectivity filter a thing?
if were not selective, AP wouldn’t work
ion selectivity filter
small no. AAs in p-loop chains of S5-P-S6 (ring of selectivity filter)
inner filter rings are -ve so repels -ve attracts +ve
Na is slightly smaller than Ca so….
can pass Ca channels in absence of Ca
K ion selectivity filter
narrow pore of neutral AAs so no net charge
filters out large ions
what do K ions have to lose to pass small pores?
hydration shells of water molecules
oxygen atoms of AAs in filter region help strip shells of ions
hydration shell of Na?
Na too small so can’t strip shell so effectively larger in respect to channel
why is correct resting potential important?
AP requires depolarisation from certain level because if shift, it can change firing rate, change patterns, change ease of reaching threshold
higher depolarisation –> higher firing frequency
hyperpolarisation makes cell less excitable
depolarise too far inactivates AP
how does the circadian rhythm change excitability of SCN neurones?
master clock in SCN of hypothalamus
AP firing freq changes with cycle from changes of melatonin secretion from pineal gland
in daytime increase Ca so affect K conductance
gating-pore channelopathies
diseases with ion channel malfunction
current leak through
displacement of S4 affected - leaves space for water
non-selective, fast activating, non-inactivating, low conductance
leads to permanent depolarisation
diseases associated with altered resting potential and ions in NS
end of lecture 7
electrical synapses
gap junction
no delay, 2-way, little flexibility and plasticity, not great role
connexins in vertebrates
innexins in invertebrates
are ion channels (6 subunits) which form gap junctions, allow current flow between 2 neurones
chemical synaptic transmission
neurones isolated from each other, across cleft, short delay (0.5-1ms), 1-way, great flexibility and plasticity, main role in brain
driven by conc gradient 2 responses (dep./excitation or hyperp./inhibition)
synapses everywhere but mostly on dendrite/axon spine/synapse shaft on dendritic process
snap
SNARE-associated protein
SNARE
v-SNARE (vesicle)
t-SNARE (target membrane)
v and t needed for docking
docking can be blocked by tetanus and botulinum toxin
vesicles fusion
calcium increases, changes conformation so SNARE twisted (energy from ATP), cause vesicles to fuse
calcium dependence not linear (3-4 needed to release 1 vesicle)
stochastic
Ca increases probability of NT release but not guarantee
measuring synaptic transmission
HPLC, amperometry, muscle contraction, electrophysiology, biosensors, FM -dyes (direct visualisation of vesicles)
NT release without AP
miniature synaptic current/potential
by single vesicle
vesicular release of NT is, and therefore synaptic response should be….
quantal
size of response should depend on amount of NT
quantal content
no. vesicles released by response to single AP
quantal content of miniature response - typically 1 vesicle
of evoked response - over 1
quantal content = amplitude evoked divided by amplitude miniature
amplitude = no. vesicles x quantal size
probability of release and docking
not constant but depends on last event and affects recycle
vesicles retrieval (endocytosis)
full fusion - vesicle fusion and then get whole back
kiss + run - small amount NT released then vesicles closes and stays inside
bulk retrieval - many vesicles fused in at same time
vesicles recycling (to reserve pool)
proteins form coat on membrane then around vesicles as fuses in so can bud off membrane, then coated with clathrin cage
synaptic strength
is the average response triggered by single AP
varies large scale among diff synapses (changes during development, plasticity)
depends on presynaptic no. vesicles, release probability
and post-synaptic receptor density to NT and efficiency (phosphorylation by kinases can affect onductance and open time)
(amount of NT in vesicles - not strong influence)
Ribbon synapse
mainly in receptor cells
conveyor belt of vesicles
persistent high f synaptic transmission
e.g. cone cells of retina, inner hair cells of ear
Calyx of Held synapse
big synapse in mammalian auditory central nervous system
many active zones
large vesicle pool
large quantal content
fire high f for long time
vesicles filling
2 kinds of transporter in vesicles
- vH+ - ATPase : creates high gradient of protons for NT transport
- transporter for specific NT, use proton gradient (H out and NT in)
cationic NTs vs glutamate (/other -ve)
monoamines, ACh, depend on change in pH
use electrical components
vesicle transporter proteins
and examples
is a criteria for a substance to be a NT
examples lecture 9 top (e.g. vAChT)
NT criteria
present in presynaptic with synthesis machinery (should be made in cell body) and specialised vesicular transporter
released and con. increases when presy. stimulation
mimic affects of presy. stimulation when added to extracellular fluid
mechanism of removal exist
NT types
classical - small, some AAs (ACh, dopamine, serotonin, adrenaline, histamine)
neuropeptides - larger, 3-20 AAs (endorphins, oxytocin)
other - NO, ATP, adenosine
co-localisation
each synapse has main classical NT but also some release co-transmitter in same/diff vesicles
so can’t always classify synapse w/ NT
e.g. (lecture 9 bottom)
synapse specialisation
specific neurone make specific NT but short-lived event because can change what NT make
ACh synthesis
acetyl coenzyme A + choline by choline acetyltransferase (ChAT)
cholinergic synapse
action of NT determined by…..
receptor
speed, facilitation, depression of firing, modulation of activity
temporal characteristics
speed/duration depends on kinetics of receptor (ligand binding and receptor properties e.g. don’t desensitise so always active when ligand bound)
why can’t we divide NTs into excitatory/inhibitory
action depends on receptors
so can have diff roles in diff tissues
types of receptors
ionotropic metabotropic (ion channel separate from receptor)
many NTs have both
ionotropic
ligand-gated channels important in direct electrical to chemical signal 3 families (trimeric, tetrameric, pentameric) e.g. lecture 9 2nd page
metabotropic
G-protein coupled
uncoupling of receptor (a from b y) when activated
alpha interacts with effector protein
increase CAMP and Ca conc. (learning, memory)
acetylcholine receptors
nicotinic (ionotropic)
muscarinic (metabotropic)
nicotinic (ACh)
properties, structure, mechanism, selectivity, types
ionotropic increase cations (Na), fast excite
hetero-pentamer of 4 subunits (2a,b,y,d) each w/ transmembrane alpha-helix (M2) and the 5 M2 helices form the pore
2 alpha bind ACh so rotation of alpha subunits cause rotation of all and open receptor
twisting so smaller polar residues line channel instead of bulky hydrophobic Leu side chains in closed channel
3 rings of -ve residues so attract +ve
a2,b,y,d skeletal muscles
a2,b3 neurones
but always 2a
muscarinic
(properties, subtypes
metabotropic
K permeability, slow excite/inhibit
5 subtypes
M1,3,5 (Galphaq) activate protein kinase C/phospholipase C so produce IP3 and release Ca
M2,4 (Galphai) inhibit adenylate cyclase so inhibit cAMP/regulate K channels
ACh pathways in brain
axons releasing ACh in brainstem to hypothalamus to modulating activity of neurones in brain but in PNS is an excitatory NT
glutamate receptors (types, glutamate)
ionotropic: fast AMPA - increase Na, main excitatory in most brain neurones, high glut to activate
slow NMDA - increase Ca, not active at resting pot. because Mg block, excess Glu means cell damage (excitoxicity)
metabotropic: increase IP3, modulatory
glutamate: side effect of Krebs cycle, coupled with synthesis of glutamic acid
glutamic acid converts to GABA which antagonises action of glutamate
GABA receptors (types, structure, allosteric)
inhibitory (excite in embryo)
GABAa is ionotropic, increase Cl, fast, hyperpolarise
GABAb metabotropic, increase K, decrease Ca, slow
5 subunits, 2 bind GABA
allosteric modulation - increase open time so increase inhibitory current (sedation by benzodiazepine, barbiturate)
Serotonin (origin, types)
from tryptophan AA in brainstem nuclei
widespread
3 types: 5-HT1 GPCR in brain, 5-HT2 GPCR in periphery + brain (LSD), 5-HT3 ion channel in sensory neurones and brain (vomit)
Dopamine/adrenaline (origin, adrenergic synapses, receptors)
from tyrosine AA, released from 2 brainstem nuclei mostly from substantia nigra
noradrenaline from nucleus locu coeruleus, widespread to cerebellum/neocortex
no adrenergic synapses in brain - instead, adrenaline released from noradrenergic presynptic terminals and diffuse towards target
all GPCR receptors, diverse effects
drugs
agonists/antagonists for receptors
e.g. nicotine agonist for AChR
tubocuraine poison antagonist for AChR
NT uptake pathways
degradation (ACh)
re-uptake by glial cells (glutamate)
important things regarding integration of the NS
signalling is via AP
amount of info = AP fired
AP don’t decrement (all or none) so record frequency when investigating
input
integrative
conductive
output
from env. to sensory, from other neurones to motor/other
at dendrites but some at soma
AP, myelinated or passive propagation in small neurones
presynaptic terminals
frequency encoding
convert amplitude of stimulus to freq of AP
factors determine firing
synaptic input (temporal/spatial summation) position of synapses proportion of inh/exc synapses modulation facilitation/depression
depend on ion channel properties and electrical properties of cellular membrane
membrane properties
insulated (separate charges)
acts like capacitor plates (larger neurone bigger capacitance because more charge store but greater time constant takes longer to charge)
greater resistance = bigger voltage change because less charge leaking
capacitance of membrane equations
lecture 10 top 2nd page
summation
combine info before deciding to fire
temporal/spatial
high chance pass threshold if w/o inhibition (IPSPs stop APs)
temporal summation
membrane has time constant and holds charge, previous AP don’t disappear so build
higher f means shorter time between and larger change of AP
synapse position affects synaptic potential
where dendrites are
dendrites depolarisation spreads passively down to soma and potentials will decrement (distance a signal decrements is 1/e = lambda length constant)
synapse on soma/near dendrites increase inhibiting
diameter of dendrites
thicker means longer lambda (length constant, distance a signal decrements)
thin means large SA for more synapses and more info
integration process in nerve terminal
AP activates Ca channels in presynaptic
amplify Ca from ER/IP3
more AP means Ca is more elevated and prolonged
higher Ca means increased NT (more vesicles)
presynaptic receptors influence Ca level and release probability
NT released affected by depletion and replenishment of vesicles
facilitate
freq firing converted to more NT release
depress
decrease NT, by depletion of vesicles is small pool/modulation
modulation of glutamate release in neocortical and hippocampal nerver terminals
AP excite, GABAb receptors inhibit
calcium still elevate
ATP co-transmitter activate Ca channels so increase Ca
but also ATP to adenosine so inhibit
neurones are organised in…
spiny vs non-spiny
networks - activity expressed as overall firing rate of APs
principle neurones - excitatory, pyramidal shape, long range inputs, local inputs from interneurones, send axons to other networks
interneurones - inhibitory, inputs from local principle neurones, outputs locally
dendritic spines
protrusion from stalk of dendrite, synapses with single axon
bulbous head and thin neck connects spine to stalk of dendrite
principle neurones multiple synapses on spines
principle/inter synapse on dendritic stalk
interneurones inhibitory effects
feed-forward - enhances by inhibit opposing
feed-back - self-regulating, excitatory act on inhibitory to act on primary excitatory neurones to dampen activity, prevent over-excitation
bottom-up pathway
top-down pathway
sensory N input to relay N (several levels)
motor cortex to muscles (output)
parallel processing
distributed processing
sensory info sent to specific zones in cortex which consists of columns innervated by parallel streams
cortical columns/local network may be part of diff pathways of analysis
neurone-glia communication
astrocytes enwrap synaptic terminals and monitor 90% brain tissue
astrocyte Ca signal transducer - spill over glutamate/ATP activates electrical/Ca signalling that spill out synapses
