Morphine Reward in Dopaminedeficient Mice

1
Morphine Reward in Dopamine-deficient Mice

• Hnasko TS, Sotak BN, Palmiter RD (2005). Nature
  438854-857.
• Presented by Mattia M. Migliore
• March 30, 2006

2
Introduction

• First recorded reference to opium use occurred
  around 300 B.C.
• Morphine is an analgesic first isolated from
  opium in 1806.
• Morphines name came from Morpheus, the Greek god
  of dreams.
• Opioids exert their effects by binding to opioid
  receptors (µ, d, and ?) which then couple to
  G-proteins, inhibit adenylate cyclase, activate
  K currents, and decrease Ca2 currents.
• Much like other opioid analgesics, morphine has
  the potential to cause addiction.

http//www.heroin.org/papaver.jpg
http//www.sciencebase.com/images/ structure_of_mo
rphine.jpg
3
Introduction (cont)

• Addiction can be defined as uncontrolled,
  compulsive use of a substance despite adverse
  consequences resulting from its use.
• Addiction can result from repeated exposure to a
  substance, which then results in neurochemical
  adaptations in the reward system of the brain.
• People can become addicted to drugs, alcohol,
  tobacco, gambling, sex, and even food.
• Addiction is extremely difficult to study because
  no animal model of addiction even comes close to
  the complexity of the human condition.

www.hrmvideo.com/ resources/docs/2662.gif
4
Introduction (cont)

• Drug addiction (also termed substance dependence)
  affects millions of people world wide.
• The degree of drug abuse ranges from just
  occasional use to compulsive use ultimately
  resulting in fatal consequences.

www.WHO.org
5
Figure 1                                        
                                                  
                                                  
                                                  
                                                  
                                                  
  
(Nestler and Aghjanian, 1997)
6
(Goldstein and Volkow et., 2002).
7
Neurobiology of Addiction

• Drugs of abuse illicit a feeling of euphoria or a
  high by activating the brains reward
  circuitry.
• Dopamine has been believed to be responsible for
  feelings of reward for the last 30 yrs, and has
  been called the feel good neurotransmitter.
• DA has long been implicated in the development of
  addiction.
• Most drugs of abuse have been shown (via
  microdialysis studies) to increase extracellular
  DA levels and/or DA cell firing in the nucleus
  accumbens.

Goodman and Gilmans 11th edition.
8
Synthesis of Dopamine
http//web.indstate.edu/thcme/mwking/catecholamine
synthesis.jpg
9
Evidence supporting DAs role in reward

• In 1954, Olds and Milner showed that direct
  electrical stimulation of the brain had powerful
  rewarding effects. Later, Olds et al. used
  intracranial self-administration of various
  substances to try to identify the
  neurotransmitters involved in reward.
• Studies showed that DA receptor antagonists can
  inhibit the rewarding effects of food, and of
  intracranial self-stimulation (Zhou and Palmiter,
  1995).
• Dopamine agonists and drugs that inhibit the DAT
  have been shown to cause animals to self
  administer these agents, and to develop a
  conditioned place preference (CPP) for these
  drugs.

10
Evidence supporting DAs role in reward (cont.)

• Bilateral 6-OHDA lesions result in a severe
  decrease of activity, and the animals will refuse
  to eat or drink (an obliteration of the natural
  reward cues).
• Schultz et al. showed that the anticipation of a
  reward (juice) in monkeys caused an increase in
  firing, and a change in the pattern of DA neuron
  firing.
• Maldonado et al. showed that D2 receptor knock
  out mice do not show a CPP in response to
  morphine.
• Volkow et al. used neuroimaging studies in humans
  to show that cocaine and methylphenidate increase
  brain dopamine levels, and this increase was
  associated with the feeling of a high.

11
Molecular Mechanism of Drug Addiction
12
(Nestler and Aghajanian, 1997)
13
Brain regions involved in drug addiction
(Golstein and Volkow, 2002).
14
(Volkow et al, 2003)
15
(Golstein and Volkow, 2002).
16
What makes some people become substance dependent?
17
Hypothesis

• Dopamine is not an essential component of opiate
  responses, and that dopamine is not required for
  opioid mediated reward.

18
Methods

• Dopamine deficient mice a complete deletion of
  the tyrosine hydroxylase (TH) encoding gene
  results in a deficiency in both DA and NE. In
  order to create only DA deficient mice, Hnasko
  et. Al used the TH encoding sequence to target
  the dopamine ß-hydroxylase (DBH) promoter in
  embryonic stem cells. Then DBH-TH /- mice were
  crossed with TH /- mice to yield TH /- DBH-TH
  /- which were then crossed with TH /- mice to
  yield dopamine deficient mice capable to still
  producing NE.

Zhou Q-YP, Richard D. (1995) Dopamine-deficient
mice are severely hypoactive, adipsic, and
aphagic. Cell 831197-1209.
19
Synthesis of Dopamine
http//web.indstate.edu/thcme/mwking/catecholamine
synthesis.jpg
20
Methods (cont.)

• Mice required daily L-Dopa administration to
  induce them to eat. Morphine was administered
  18-24 hrs after L-Dopa .
• Virally Rescued Dopamine Deficient Mice (vrDD)
  In order to perform the behavioral tests, they
  used a viral gene transfer to restore DA in the
  striatum (because DA deficient mice are slow and
  hypoactive).
• Behavioural tests
• 1. Locomotor tests were done using photo-beam
  activity cages. Morphine was administered IP at
  0,0.25,2.5,12.5, and 25 mg/kg.
• 2. Tail flick tests were perfomed by
  using warm water baths. Briefly, the animals
  tail was submerged 0.5-1 cm in the water bath,
  and the latency to withdraw the tail was recorded
  (with a cut off time of 15s). The animals were
  tested three times/treatment and average used.
  Morphine was administered 30 min. Prior to test
  IP at 0,3,6,12, and 24 mg/kg.

21
Methods (cont.)

• 3. Conditioned Place Preference (CPP) was
  performed using clear plastic boxes with 3
  chambers (1 neutral grey compartment in the
  middle, and 2 compartment with different colored
  walls, different textured flooring, and different
  scents). First, the mice were administered
  caffeine (18-24 hrs after L-Dopa treatment) and
  placed in the center and allowed to explore for
  25 min. On days 3-5 (conditioning phase), the
  animals received saline SQ in the morning and
  restricted to one compartment for 25 min, and
  then received morphine SQ and restricted to the
  opposite compartment for 25 min in the
  afternoons. Preference was tested on the sixth
  day. In the L-Dopa rescue, L-Dopa was
  administered similarly to the caffeine.

(Cami and Farre, 2003).
22
Results
23
(No Transcript)
24
(No Transcript)
25
(No Transcript)
26
(No Transcript)
27
www.unifr.ch/.../ Neurotransmitters/peptides.htm
28
(No Transcript)
29
(No Transcript)
30
Conclusions

• Dopamine appears to be essential in the
  development of locomotor response to morphine.
• Dopamine appears to play an important role in the
  level of analgesia experienced after morphine
  administration.
• Dopamine may be required for reward seeking, but
  does not appear to be indispensable for
  morphines rewarding effects.