What changes in the brain chronic nicotine exposure to nicotine

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What changes in the brain chronic nicotine
exposure to nicotine? Studies on genes,
receptor binding, proteins, drugs, cells,
circuits, and behavior
. . . and some ideas for therapeutic drugs
Henry Lester
May, 2008
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1. How does one explain nicotine addiction?
Does it matter? Wont everyone stop smoking
soon? Smokeless tobacco?
2. Nicotine as an imperfect therapeutic
drug Best example Parkinsons disease
3. Cellular / molecular approaches to better
therapies
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Progress on smoking cessation is very slow
Prevalence of current smoking among adults aged
18 years and over
Self-medication may be the reason
In 2002, individuals with a current psychiatric
disorder comprised 7 of the US population, but
they smoked 34-46 of all cigarettes in the US.
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The nicotine video
Produced for Pfizer to explain varenicline
(Chantix) to physicians This summarizes
knowledge in 2004. ligand is a molecule that
binds to another. physical addiction vs
psychological addiction. Desensitization and
Upregulation
Some abbreviations on future slides ACh,
acetylcholine nAChR, nicotinic acetylcholine
receptor DA, dopamine
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Focus on a4ß2 receptors
Conclusions from knockout and hypersensitive mice
(2005) Activation of a4b2-containing (a4b2)
receptors by nicotine Is necessary and sufficient
for sensitization, tolerance, reward, (but
withdrawal?)
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Nicotine and ACh act on many of the same
receptors, but . . .

• 1. Nicotine is highly membrane-permeant. ACh is
  not.
• Ratio unknown, probably gt 1000.
• 2. ACh is usually hydrolyzed by
  acetylcholinesterase (turnover rate 104 /s.) In
  mouse, nicotine is eliminated with a half time of
  10 min.
• Ratio 106
• Nicotine is 400 times more effective at a4ß2
  receptors than at muscle. What causes this
  difference?

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The aromatic box occupied by nicotine . . .
aC2
aB
aA
non-aD
aC1
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Nicotine EC50 values (half-maximally activating
concentrations)
Muscle nAChR single component 400
µM a4ß2 two components 1 µM
Underlying the 400-fold higher nicotine
sensitivity of neuronal vs muscle
receptors Factor of 16 for the cation-p
interaction Factor of 12 for H-bond 16 x 12
192. We still cant explain a factor of
400/192 2.
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Possible molecular mechanism 1 for changes with
chronic nicotine Signal transduction triggered
by a ligand-gated channel
NMDA receptors and nAChRs are highly permeable
to Ca2 as well as to Na.
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Possible Mechanism 2 for changes with chronic
nicotine Upregulation
Chronic exposure to nicotine causes upregulation
of nicotinic receptor binding (1983 Marks
Collins Schwartz and Kellar) Upregulation 1)
Involves no change in receptor mRNA level 2)
Depends on subunit composition.
Shown in experiments on clonal cell lines
transfected with nAChR subunits Nicotine seems
to act as a pharmacological chaperone
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Upregulation may be a thermodynamic consequence
of nicotine-receptor Interactions
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Stolen photons tell us which subunits are near
each other
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The ultimate reductionist approach, studying
nAChR traffic/regulation at the single molecule
level. TIRF microscopy of nAChR geGFP in oocytes
1
3
2
2
3
1
4
4
12 µm
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Strategy to choose between the bored or
exuberant receptors in the response to chronic
nicotine exposure
1. Generate mice with fully functional,
fluorescent a4 receptors. (Why mice?)
2. Chronically expose the mice to nicotine (2
weeks).
3. Find the brain regions and cell types with
changed fluorescence.
4. Perform experiments on these regions and
cells to decide whether the new receptors are
bored or exuberant.
5. Model the cellular and circuit changes
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Chronic nicotine increases a4 fluorescence
2-fold in hippocampus --a brain area that
provides a good model for cognition.
Alveus
Py
Or
Rad
LMol
200 mm
Medial Perforant Path
Temperoammonic Path
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Chronic nicotine exposure causes cognitive
sensitization
In the human context, cognitive sensitization is
epitomized by smokers reports that they think
better when they smoke this anecdotal
observation is confirmed by data that smokers who
smoke nicotine cigarettes (but not nicotine-free
cigarettes) display several cognitive
enhancements. In the rodent context, rats show
more contextual fear conditioning if, one day
after withdrawal from chronic nicotine, they
receive an acute nicotine dose also chronic
nicotine produces better spatial working memory
performance in the radial arm maze.
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Midbrain dopaminergic cells (tyrosine hydroxylase
stain) Substantia nigra pars compacta (SNc,
controls motion) Ventral tegmental area (VTA,
controls reward)
Substantia nigra pars reticulata (SNr, GABAergic)
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VTA GABAergic and DA neurons have contrasting
responses to nicotine in vivo
WT mouse
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a4-YFP knock-in substantia nigra pars compacta
neurons
Spectrally unmixed background autofluorescence
Spectrally unmixed a4YFP
10 mm
10 mm

a
4YFP
1500
Background
1000
YFP Intensity
500
0
500
520
540
560
580
600
Wavelength (nm)
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Substantia nigra data also support the exuberant
receptor idea
Chronic nicotine does not change a4 levels in
dopaminergic neurons . . .
Substantia Nigra Pars Compacta
a4 intensity per TH neuron
. . . but does upregulate a4 levels in
GABAergic inhibitory neurons.
Substantia Nigra Pars Reticulata
a4 intensity per GAD neuron
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• Chronic nicotine exposure causes tolerance of
  dopamine release
• The yoked self-administration experiment

4.0
Yoked saline
3.5
Yoked nicotine
3.0
2.5
Dialysate DA (nM)
2.0
1.5
1.0
0.5
0.0
Saline
Nicotine
Yoked animal
0
40
80
120
160
-40
Master animal
Time (min)
Rahman, Zhang, Engleman, Corrigall, 2004
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Chronic nicotine cell-specifically upregulates
a4 receptors Basis for circuit-based tolerance
in midbrain via exuberant inhibition
Chronic Saline
Endogenous ACh
VTA
NAc
LDT
1A
DAergic
Cholinergic
GABAergic
4.0
Yoked saline
3.5
Yoked nicotine
1B
3.0
2.5
1A
Dialysate DA (nM)
2.0
1.5
1.0
2B
2A
0.5
Saline
Nicotine
0.0
0
20
40
60
80
120
140
160
180
-40
-20
100
Time (min)
Rahman et al, 2004
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Inverse correlation between long-term tobacco
smoking and Parkinsons disease
In identical twins discordant for both
Parkinsons disease smoking, the unaffected
twin smoked at a significantly higher rate. In
those twins where one or both smoked, The
unaffected twin smoked 12 pack-years more.
There are good indications that nicotine itself
is a protective agent. Clinical trials of
nicotine patches have given mixed results because
of side effects
Beneficial results of short-term nicotine
exposure Pain reduction. Increased
concentration ADHD, Schizophrenia. Alzheimer
(Aricept donepezil, a cholinesterase inhibitor
Reminyl galantamine) Decreased inflammation.
Antidepressant actions.
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Hypothesis Circuit-based neuroprotection by
chronic nicotine in substantia nigra via Cholinerg
ic, Dopaminergic, and GABAergic neurons in
Hindbrain Midbrain
Upregulated a4 nAChRs
GABAergic neurons may have increased or more
regular firing in chronic nicotine. . .
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1. Assess candidate drugs for changed
upregulation
An inhibitor of upregulation may show us how to
prevent or reverse nicotine addiction an
enhancer of upregulation may show us how to
develop better therapies for Parkinsons Disease.
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2. New mouse strains that highlight action on
one receptor type
. . . but, unlike selective a4 activation, shows
no sensitization
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Some changes in the brain during chronic exposure
to nicotine
1. Nicotine potently activates some neuronal
nAChRs because it participates in both cation-p
and H-bond interactions within the conserved
aromatic box.
Genes Binding Proteins Cells
Circuits Behavior Therapies

• Chronic exposure to nicotine chaperones a4ß2
  number and stoichiometry.

3. These processes lead to cell-specific a4ß2
upregulation.

• a. Upregulation explains tolerance to chronic
  nicotine, via a GABAergic-DA circuit in the
  midbrain.
• b. Upregulation also explains enhanced LTP in
  the perforant path, via a direct presynaptic
  mechanism. This is a simple model for cognitive
  sensitization

5. Cell-selective upregulation may act as a form
of pharmacological deepbrain stimulation to
explain the inverse correlation between smoking
and Parkinsons disease
6. These ideas provide a basis for discovering
new drugs that could treat nicotine addiction or
prevent Parkinsons disease.
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Xiu, Nyssa Puskar, Jai Shanata, Shawna Frazier,
Professor Dennis A. Dougherty Sarah
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Al Collins, Mike Marks, Jeremy Owens,
Tristan McClure-Begley, Paul
Whiteaker Jim Boulter, Istvan Mody, Oliver
Dorigo, Arnie Berk, Max Shao, Jack Feldman Jon
Lindstrom Julie Miwa, Nathaniel Heintz Uwe
Maskos, Jean-Pierre Changeux
Unnatural Amino Acid Club
Univ of Cambridge
Univ Queensland Univ of Colorado, Boulder
UCLA Univ. Pennsylvania Rockefeller
Univ Institut Pasteur
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1962 Smoking and health
2007 Harm reduction in nicotine
addiction Helping people who can't quit. A
report by the Tobacco Advisory Group of the Royal
College of Physicians
We demonstrate that smokers smoke predominantly
for nicotine, that nicotine itself is not
especially hazardous, and that if nicotine could
be provided in a form that is acceptable and
effective as a cigarette substitute, millions of
lives could be saved. We also argue that the
regulatory systems that currently govern nicotine
products in most countries actively discourage
the development, marketing and promotion of
significantly safer nicotine products to smokers.
Harm reduction is a fundamental component of
many aspects of medicine . . . yet . . .
effective harm reduction principles have not been
applied to tobacco smoking. This report makes the
case for radical reform of the way that nicotine
products are regulated and used in society. The
ideas we present are controversial, and challenge
many current and entrenched views in medicine and
public health.
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