Bi 202 October 2010 Drug Addiction Henry Lester

This lecture is at www.its.caltech.edu/~lester/Recent-lectures 1

NH2

CH3

amphetamine

N

N CH3

CNC2H5

C2H5

O

LSD

H3C

H2C

OH

ethanol

N

CH3N

nicotine

N

O

HO

HOCH3

morphine

N

phencyclidine

OH3C

H3C

CH3

OH

C5H11

tetrahydrocannabinol

NOC

OCO

CH3OH3C

cocaine

N

N N

N

O

O

H3C

CH3

CH3

caffeine

Recreational drugs Addictive drugs

2

Source

morphine-heroinPapaver

somniferum

tetrahydrocannabinolCannabis

sativa

nicotineNicotiana tabacum

cocaineErythroxylum

coca

amphetamine synthetic

ethanolSaccharomyces

cerevisiae (fermentation)

LSD synthetic

caffeineCoffea sp.,

Camellia sinensis

phencyclidine synthetic

“poppy that brings sleep”(opium)

marijuana, hemp

tobacco

coca

coffee

tea

yeast

ergot

wheat fungus;Salem witch trials?

Based on plant

3

H+

blood

lungs,nose,stomach

NOC

OCO

CH3OH3C

+HNOC

OCO

CH3OH3C

H+NOC

OCO

CH3OH3C

+HNOC

OCO

CH3OH3C

cocaine base(crack)

cocaine in the body exemplifies permeation by weak acids & bases

cocaine hydrochloride

Cl-

South American Indians use Ca(OH)2 from limestone to shift this equilibrium

Lipid barrier,e. g. membrane(s)

4

Neurons that make dopamine: “pleasure-reward” system highlighted in a sagittal view (human

brain).Most addictive drugs affect this system

5

Nature 2002 417:37

Examples of guided rat navigation using brain microstimulation. a, Route followed by a rat guided through a slalom course. Inset, detail of the events that took place inside the dashed enclosure. b, Route taken by a rat guided over a three-dimensional obstacle course. The animal was instructed to climb a vertical ladder, cross a narrow ledge, descend a flight of steps, pass through a hoop and descend a steep (70°) ramp. Two rounds of high-density dopamine cell stimulation were required to guide the rat successfully down the ramp, demonstrating the motivational qualities of stimulation.

Green dots indicate positions at which reward stimulations were administered to dopaminergic cells

Blue arrows indicate positions where the experimenter gave right (R) and left (L) directional cues, by stimulating the part of the brain that receives left or right whisker signals.

Red dots indicate rat head positions at 1-s intervals.

Black arrows indicate positions 0.5 s after directional commands.

Goal-seeking behavior controlled by the dopamine pleasure / reward system

6/30

7

“Network Model of Pathways by which Acute Exposure to Addictive Drugs Uncouples Behaviorally Relevant Control of DA

Neurotransmission”(Sulzer, 2011)

VTA GABAergic and DA neurons have contrasting responses to nicotine in vivo

DA neuron, ~ 1700 spikes

Nicotineinjection

GABAergic neuron (5 s smoothing), ~ 8300 spikes

0.05 m V2 m s

0.05 m V2 m sF

requ

ency

, H

z

0 100 200 300 400 500 600 700

0

2

4

6

0

5

10

15

20

25

s

Fre

quen

cy,

Hz

0.1 m V

0.5 m s

0.1 mV0.5 ms

A B C D0.05 m V2 m s

0.05 mV2 ms

4*, 6*, and/or 7

4* only

V

GABAergic

DAergic

VTA

WT mouse

8

Tolerance

a. Metabolic tolerance:

Metabolism of the drug proceeds more efficiently.

This occurs primarily in the liver.

It occurs for many types of drugs, including aspirin and penicillin.

b. Cellular tolerance:Individual neurons or neuronal circuits become less responsive to the drug.

1. Tolerance2. Dependence3. Goal-seeking behavior

Three general components of addiction

9

10

Rats were exposed to morphine for 5

days and then observed for “withdrawal

behavior”: irritability, jumping, wet-dog

shakes, head-bobbing, sweeping tail

movements, yawning.

Action potential frequencies were

recorded in the noradrenergic neurons

of the locus coeruleus.

When morphine was withdrawn from the

receptors, with the help of a morphine

antagonist, the firing frequency in the

neurons increased in parallel with the

withdrawal behavior.

Rasmussen et al J Neurosci 10, 238

(1990)

withdrawal behavior

firing frequency in

locus coeruleus

morphineantagonist

(naltrexone)

ratsmorphine-treated

control

Morphine dependence at the single-cell level

11

kinase

P P. . .other proteins

bind to the phosphates . . .

Activated GPCRs are sometimes phosphorylated and endocytosed. This “downregulation” terminates signalling.

P P

But continual signalling can activate genes

During activation, the G protein leaves . . .

. . . revealingphosphorylationsites . . .

(not a synaptic vesicle)

. . . triggeringendocytosis.

12

kinase

phosphorylatedprotein

cAMPCa2+

intracellularmessenger

receptor

tsqiG protein

enzymechannel effector

NMDA receptors

and

nAChRs

are highly permeable to Ca2+

as well as to Na+.

Possible molecular mechanism #1 for changes with chronic nicotine:

Signal transduction triggered by a ligand-gated channel

Brunzell, Russell, & Piccotto, 2003

13

The nicotine video

This summarizes knowledge in ~ 2004.

“physical” addiction vs “psychological” addiction.

Desensitization and “Upregulation”

Produced for Pfizer to explain varenicline (Chantix) to physicians

nicotine20 seconds

1 millionchannels

Closed states(s) more stable than open states

14

1. Nicotine is highly membrane-permeant. ACh is not.

Ratio unknown, probably > 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: ~105

3. EC50 at muscle receptors: nicotine, ~400 μM; ACh, ~ 45 μM.

Ratio, ~10. Justified to square this because nH = 2.

Functional ratio, ~100.

HAL & Dennis Dougherty (Chem) study this difference

Nicotine and ACh act on many of the same receptors, but . . .

15

W149BY93

A

non-W55D

Y198C2

Y190C1

(Muscle Nicotinic numbering)

H-bond 12-fold tighter binding vs muscle

Cation-π interaction16-fold tighter binding vs muscle

Additional H-bond to non-α subunit

The AChBP interfacial “aromatic box” occupied by nicotine (Sixma, 2004),Probed functionally by unnatural amino acid mutagenesis

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Changes in the Brain during Chronic Exposure to Nicotine

BehaviorBehavior

CircuitsCircuits

SynapsesSynapses

NeuronsNeurons

Intracell.Intracell.

BindingBinding

Nic vs AChNic vs ACh

ProteinsProteins

RNARNA

GenesGenes

Components of nicotine dependence

Reward

Cognitive Sensitization

Stress Relief

Weight control

Self-medication in schizophrenia (HAL, PHP project)

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α

}α4 α4 α4 α6 α6

│││

(α7)5

│││

β2 β2 β2 β2 β2

α}

α4 α4 α4 α6 α4

β2 β2 β2 β2 β2

aux α4 β2 α5 β2 or β3 β2 or β3

Expression WS WS WS DA NA RGC DA RGC WS

Smoking level required for activation / desensitization

Heavy ────────────Moderate─────────────── Heavy

Upregulated by nicotine at moderate smoking levels?

N Y ? N N N

Lynx binding ──────Y───── ? ? ? Y

lynx

agonist

aux

α

α

agonist

DA

18

Complexity of nicotine dependence May arise from the

widespread distribution of Highly nicotine-sensitive nAChRs

19

The tactic of fluorescent knock-in mice for evaluating cellular and subcellular specificity of nAChR upregulation

1. Generate knock-in mice with fully functional, fluorescent nAChRs

2. Expose the mice to chronic nicotine

3. Find the brain regions and cell types with changed receptor levels

YFP

4. Perform physiological experiments on these regions and cells to verify function

VACh, nicotine puffs

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Cellular and subcellular specificity of Selective α4* nAChR Upregulation

Thalamus,

superior colliculus

SNc, VTA

SNr,VTA

Striatum

Upregulation?

Transmitter Soma Term. Region / projection

Glu Yes Yes Entorhinal cortex → dentate gyrus

ACh No No Medial habenula → Interpeduncular nucleus

DA No Yes Ventral tegmental area, substantia nigra pars compacta → Striatum

GABAA Yes Yes SN pars reticulata, VTA → SNC, VTA

Glu Yes ?? Subthalamic nucleus → SNR

CA

DG

EC

Medial Perforant Path

Nashmi et al J Neurosci 2007; Xiao et al, J. Neurosci 2009; Xiao et al, in review21

MH

IPN

STN

~ 2 min

nAChRactivation

Nicotine activates quiescent nAChRs

Nicotine desensitizes ongoing activation

upregulated nAChRs

naïve nAChRs

nAChRactivation

Either activation and/or desensitization can be amplified by upregulation

22

BloodLungs

H+

Like most drugs, nicotine is a weak base.Its neutral form passes through 6 plasma membranes in ~ 20 s

logP = 1.1 = log (solubility in octanol / water) 23

NH+

N

N

N

N

N

CSF

Alvelolarepithelium

Brain capillary

24

Nucleus

UPRE

PlasmanACh

R

Nicotine in CSF

Classical Pathway:Channel

activation & desensitization

→ Do neurons survive

Despite stressors?

Unfolded protein response

membrane

COPII vesicleSec 13/31

Sar1Sec24Sec23

ATF6

Golgi

Pharmacological

Chaperoning→ upregulation

M3-M4 loop

H+ +

ERBiP

PERKIRE1

Clathrin

Secretory vesicle

COPII

Golgi complex

COPI

Early endosome

COPI

Lysosome

Ca2+

Na+

“Inside-out” Drug Action by Nicotine at α4β2 nAChRs

NH+

N

H+

N

N

Endoplasmic reticulum

nAChR

ATF6

IRE1

XBP1

eIF2α

PERK

ATF4

1. Agonist binding eventually favors stable,

high-affinity states (a “chaperone”)

106

channels

nicotine20 sec

Three possible results of nicotine-nAChR binding in the endoplasmic reticulum

“closed”

AC Highest affinity

Fre

e E

ne

rgy

Reaction Coordinate

“activated”

“desensitized”

Bound states with increasing affinityunbound

agonist

?

2. Nicotine binding at subunit interface favors assembled nAChRs (a

“matchmaker”)

3. Nicotine may displace lynx, directing nAChRs toward cholesterol-poor domains (an “escort”)

nicotine

lynx

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Chronic nicotine causes tolerance of dopamine release

Master animal

Yoked animal

Rahman, Zhang, Engleman, & Corrigall, 2004

0.0

0.5

1.0

1.5

2.0

2.5

3.0

3.5

4.0 Yoked salineYoked nicotine

Saline Nicotine

-40 0 40 80 120 160

Time (min)

Dia

lysa

te D

A (

nM)

26

Chronic Saline

1A

Endogenous ACh

1A

2A

1B

2B

0.0

0.5

1.0

1.5

2.0

2.5

3.0

3.5

4.0 Yoked salineYoked nicotine

Saline Nicotine

-40 -20 0 20 40 60 80 100120140160180

Time (min)

Dia

lysa

te D

A (

nM

)

Rahman et al, 2004

2BDecreased Reward

Plus Acute Nicotine(repeated exposure)

Chronic nicotine cell-specifically up-regulates functional 4* receptors:

Basis for circuit-based tolerance in midbrain(Nashmi et al, 2007)

Endogenous ACh VTA

LDT

Cholinergic

NAc

DAergic

GABAergic

Chronic Nicotine Tolerance

2A

Upregulated 4* nAChRs

Craving

Endogenous ACh

1B Reward

Plus Acute Nicotine(1st expsoure)

+ acute nicotine27

200 m

Medial Perforant Path

Py Or Rad

LMol

Alveus

Temperoammonic Path

Humans: Some smokers report that they think better when they smoke; smokers who smoke nicotine cigarettes (but not nicotine-free cigarettes) display certain cognitive enhancements (Rusted and Warburton, 1992; Rusted et al., 1995).

Rodents:Mice show more contextual fear conditioning if, one day after withdrawal from chronic nicotine, they receive an acute nicotine dose (Davis et al., 2005); this is α4β2* dependent.Also chronic nicotine produces better spatial working memory performance in the radial arm maze (Levin et al., 1990; Levin et al., 1996).

Chronic nicotine increases medial perforant path 4 fluorescence ~ 2-fold.Relevant to cognitive sensitization?

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Changes in the brain during chronic exposure to nicotine

1. The modern hypothesis: selective upregulation of nAChRs (via SePhaChARNS) is necessary and sufficient for the early stages of nicotine dependence (hours, days, and weeks)

2. Selective upregulation thus instantiates some phenomena typically invoked to explain the neuroscience of drug abuse:

adaptation, neuroadaptation, plasticity, compensation, and homeostasis

3. We do not yet understand several processes, including somatic signs of withdrawal and stress-induced nicotine use.

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