ACETYLCHOLINE ( ACh) modulation of circuit excitability
Bio 337 Neurotransmission and Neuromodulation
Learning objective: If someone asks you How does Acetylcholine effect excitability ? Your answer will be: That depends on ..
Cholinergic Circuits in the CNS: under-rated & problematic! Acetylcholine ( ACh) is important in the Brain:
ACh modulates the activity of circuits that underlie attention, memory & motivated behaviors
The Challenges:
BASAL FOREBRAIN CHOLINERGIC NUCELI PTG CHOLINERGIC NUCELI
1. Gentle introduction to Acetylcholine: (ACh)
the ACh synapses and receptors you probably already know
2. ACh neurons and their projections in the brain
Distribution & why should you care?
3. Brain ACh receptor types
focus on neuronal type nAChRs ( but dont forget mAChRs)
4. Release of ACh in the brain
5. ACh as a modulator : Location, Location, Location
6. Data based examples : ACh as modulatory transmitter in circuits of motivation & emotional memory
Acetylcholine: (ACh) what you may already know: 1. ACh basics 2. muscle-type nicotinic AChRs and the NMJ 3. Peripheral muscarinic AChRs
Acetylcholine Basics
From lecture by Professor Wollmuth (fonts changed)
Synthesis and Packaging of Acetylcholine (ACh)
+ (CH3)3-N-CH2CH2-O-C-CH3
=
mitochondria
Na+ choline [choline] in serum ~ 10 M
choline + acetyl-CoA
Axon
Choline Acyltransferase ACh
Pre-synaptic Terminal
The NMJ : the archetypal synapse
motoneuron
Muscle fiber
myelin
The NMJ
axon
Active zone Syn cleft nAChRs NMJ fold
Presynaptic terminals
NMJ/ MUSCLE-TYPE NICOTINIC AChRs: The archetypal ligand-gated ion channel (= ionotropic receptor)
Studies of the NMJ nAChR have guided the structural & biophysical dissections of other ionotropic channels
g subunit
ACh binding sites
a subunit
Acetylcholine
d subunit
Muscarinic ACh receptors in cardiac & smooth muscle
--- 7TMs, 2nd messenger cascades ..indirect mechanisms of channel gating..
ACh binds to M2 muscarinic acetylcholine receptor activates Gi protein bg subunits bind to IK,ACh channel activate channel hyperpolarize membrane potential HR
ACh in the brain :
Key concepts:
1. There are &*(^% few cholinergic neurons in the brain. 2. The projections/ terminal fields are REALLY extensive 3. There is reciprocal innervation of cholinergic nuclei with numerous brain regions ( many related to emotional memory)
ACh in the Brain
Ok.. As brain transmitters/ modulators go .. ACh IS a bit of a sports challenge..
BASAL FOREBRAIN CHOLINERGIC NUCELI
PTG CHOLINERGIC NUCELI
Cholinergic Circuits in the CNS (AChE)
VAChT Ab
Cholinergic Circuits in the CNS (VACh Transporter Ab)
B = Nucleus Basalis MAJOR cholinergic nucleus
VAChT Ab
BLA Lat CeA
BLA BLA VAChT Ab
JCN 519: 790805 (2011)
Cholinergic projections to the BLA (highest density terminal field)
RECIPROCAL REGULATION of CHOLINERGIC, GABA-ergic, GLUTAMATergic & DOPAMIN-ergic CIRCUITS
PFC
Ac
Basal Forebrain ACh nuclei
VTA
ACh in the brain -- why should you care?
ACh and AChRs as therapeutic targets
a. neurological ALS, My Gravis, AD, PD b. neuropsychiatric SZ, ADHD, depression, addiction c. inflammation
Role of AChR signaling in neuropsychiatric disorders Alzheimers Disease
Impaired short term memory, dysnomia, progressive aphasia, disorientation, dementia Auguste Deter, 51 yo , admitted: 1901 d.1906
Nicotine
Alois Alzheimer 1864-1915
psychosis (hallucinations, delusions) Affective/mood dysregulation (depression, mania) Thought disorder Altered information processing (cognitive impairment, sensory gating deficits)
Schizophrenia
Acetylcholine Receptors in the CNS: (AChRs)
Key concepts:
1. There are multiple classes of neuronal nicotinic AChRs ( muscle-type nicotinic AChRs ) *altho all cationic, net effect depends on location * 2. There are multiple classes of neuronal muscarinic (7TM) AChRs with different effector channels * Some hyperpolarize, some depolarize, net effect depends of type and location *
: Two types of ACh receptors in brain
Ligand-gated ion channels = nicotinic AChRs
G-protein coupled receptors = muscarinic AChRs
neuronal nicotinic AChRs (nAChRs)
Heteropentameric nAChRs
Homopentameric nAChRs
neuronal nicotinic AChRs
Agonists: nicotine, acetylcholine carbachol Antagonists: mecamylamine, atropine! (aBgTx a7 specific)
neuronal nicotinic AChRs
(so much more than you wanted to know and NO you do not have to)
: Two types of ACh receptors in brain
Ligand-gated ion channels = nicotinic AChRs
G-protein coupled receptors = muscarinic AChRs
From lecture by Professor Wollmuth
Muscarinic (M2) acetylcholine receptor
bg-Subunits of G proteins may have regulatory activity, too
Kir
AC inactive
g b K+ ai
GTP
Muscarinic Acetylcholine receptors (mAChRs)
ACh activation of many 4/5 mAChRs leads to a decrease in excitability &/or release due to increased K (gK) conductance
BUT M1 receptors: are unusual because they gK & increase excitability
+5 mV
-80 mV -80 mV
1 nA 5 ms
mAChR
control
M2 and M4 receptors can also be coupled to inhibition of N type Ca channels to decrease release
(so much more than you wanted to know and NO you do not have to)
Gene Function Effectors Agonists acetylcholine oxotremorine muscarine carbachol McNA343 77-LH-28-1 Antagonists atropine scopolamine dicycloverine Thorazine tolterodine ]pirenzepine EPSP in autonomic ganglia secretion from salivary glands and stomach
In CNS (memory?) cortex, hippo, striatum
neuronal muscarinic AChRs
(Gi), (Gs )
Gq
M1
CHRM1
K+ conductance
(aka M current)
induces slow EPSP
M2
*slow heart rate; reduce contraction atrium reduce conduction velocity of AV node
CHRM2 In CNS: homotropic inhibition, basal forebrain, thalamus
Gi
K+ conductance
(aka GIRK)
Ca2+ conductance
acetylcholine methacholine carbachol Oxotremorine muscarine
atropine dicycloverine Thorazine Diphenhydramine tripitramine Gallamine
smooth muscle contraction Increase intra Ca vascular endothelium M3 CHRM3 increased endocrine and exocrine secretions, In CNS coretex hippocampus ,thalamus
Gq
acetylcholine bethanechol carbachol oxotremorine pilocarpine
atropine Diphenhydramine dicycloverine Tolterodine oxybutynin ipratropium darifenacin tiotropium atropine Dicycloverine oxybutynin mamba toxin
atropine Diphenhydramine dicycloverine ipratropium
M4
CHRM4
decreased locomotion In CNS; cortex , striatum, hippocampus
Gi
K+ conductance Ca2+ conductance
acetylcholine carbachol oxotremorine acetylcholine carbachol oxotremorine
M5
CHRM5
In CNS substantia nigra, VTA?
Gq
Acetylcholine (ACh) release
Q: Is ACh released via classic point to point transmission at axo-dendritic synapses or by volume transmission ? A: Yes (& no). phasic release of ACh and a role for overflow ** Axo-axonic synapses ***
other
BASICS OF CNS SYNAPTIC STRUCTURE
Type I
Type I
Type I Type II Type II
Type II
ACh release near soma & dendritic shaft (as in PNS & some CNS)
ACh release at spines (?)
spine
soma
By far the most common situation in the CNS: ACh is released at Axo-axonic synapses to modulate release of other neurotranmitters
ACh input Glutamate or GABA-ergic synapse Ca2+
Na
Ca2+
CICR
[Ca2+]int
G protein Ligand gated coupled AChRs AChRs (nicotinic) (muscarinic)
ACh in the brain:
SO: How does the activation of pre &/or
post synaptic nAChRs &/or mAChRs add up to modulate excitability? That depends on.. which TYPES of AChRs
& LOCATION
LOCATION
LOCATION
Studies of cholinergic modulation in vitro
Hipp or cortical input from YFP mice
Optical (SDCM) recording of Ca++ signaling along individual axons
A B
WT or +/-
C
D
Electrophysiological recording of synaptic interactions
nAcc or Amyg (WT or +/-)
Synaptic facilitation by nicotine at inputs from Hipp nAcc
NICOTINE APPLICATION PIPETTE
+NIC
In vitro analysis vs in vivo reality
Sotell me, what would be the effect of (just) nicotine on this motivational circuit?
& what about the effects of ACh?
Hipp
mAChRs
a7, a4 b2
mAChRs
mAChRs mAChRs
mAChRs
Understanding the effects of nicotine
(tho very relevant to the physiology, morbidity and mortality of zillions of people world wide)
is still only part of the picture..
How do we figure out how ACh per se influences circuit activity ?? so we could have a way to FIX it when it goes wrong?
(NB! the next 6 slides are for enjoyment and personal edification only)
: major obstacles to studying effects of endogenous
cholinergic circuits..
Cholinergic neurons are few in number & at low density within disperse nuclear groups
NBM
NBM
ChAT Tau GFP line from SJ. Vijayaraghavan
Cholinergic Neurons in the basal forebrain 2011 (Chat GFP transgenic)
ChAT Tau GFP line from SJ. Vijayaraghavan
Dorsal and ventral striatum (emotion motion)
Cholinergic projections in the CNS (Chat GFP transgenic)
Amygdala (BLA) emotional memories
ventral hippocampus ChAT Tau GFP line from SJ. Vijayaraghavan
OPTOGENETIC LABELING OF CHOLINERGIC NEURONS ChAT-Tau GFP; ChAT- ChR2 AAV- DiO- floxed ChR2- RED into NBM
ChAT Tau GFP x ChAT-CRE double transgenic
Cholinergic neurons in NBM OPTO probe labeled Cholinergic neruons NBM
2 weeks
Cholinergic axons in terminal fields PTO probe labeled Cholinergic axons
Direct, electrophysiological stimulation of cortical projections + recording in BLA -/+ optogenetic stimulation of cholinergic inputs (L.Jiang) Light Stim Cortical Input Stimulating
Recording electrode Light electrode Record BLA
ELEC STIM
OPTO +
CORTICAL INPUT
OPTO TAGGED CHOLINERGIC INPUT Target neuron
OPTO stimulation of NBM inputs to BLA, like nicotine, elicits LTP (Li Jiang)
Stim Cortical Recording Input, 1 Hz electrode
Record BLA
1. Gentle introduction to Acetylcholine: (ACh)
the ACh synapses and receptors you probably already know
2. ACh neurons and their projections in the brain
Distribution & why should you care?
3. Brain ACh receptor types
focus on neuronal type nAChRs ( but dont forget mAChRs)
4. Release of ACh in the brain
5. ACh as a modulator : Location, Location, Location
6. Data based examples : ACh as modulatory transmitter in circuits of motivation & emotional memory
