Social stress is as effective as physical stress in reinstating morphine-induced place preference in mice

1
Social stress is as effective as physical stress
in reinstating morphine-induced place preference
in mice

• B. Ribeiro Do Couto, M. A. Aguilar,
• C. Manzanedo, M. Rodriguez-Arias,
• A. Armario, J. Minarro

2
Introduction

• Human heroin addicts that have been clean for
  years relapse when
• Confronted with context or cues associated with
  heroin (old hangout, paraphernalia, etc.)
• Given heroin or opportunity to use heroin
• Experience stressful life events

3
Introduction

• In animal studies when trained to
  self-administer or conditioned for a place
  preference and then undergo extinction will
  relapse also when
• Presentation of drug associated cues (light,
  odor, drug paired chamber, etc.)
• Exposure to the drug itself (injection)
• Exposure to a stressor (foot shock, restraint,
  SD,etc.)

4
Introduction

• Timeline of
• extinction-reinstatement model of
  self-administration

Chambers Self (2002) Neuropsychopharmacol.
27889-5
5
Introduction

• Using the self-administration paradigm it has
  been reported that stress can be even more
  effective in reinstating drug seeking behavior
  than re-exposure to drugs.
• Shaham, etal. 1997 Psychopharmacol. 119334-341

6
Introduction

• Self-administration of opioids and
  extinction-reinstatement experiments reveal that
  reinstatement is demonstrated by
• Food deprivation
• Administration of CRF
• Not restraint stress

7
Introduction

• Using conditioned place preference (CPP)
  procedure of extinction-reinstatement, Foot
  shock
• Delays the extinction of morphine CPP
• Reinstates morphine CPP after extinction
• Reactivates morphine seeking following drug free
  periods even when not exposed to extinction
  conditions

8
Introduction

• Psychological distress and provoking relapse to
  illicit drug use
• In humans, not only subordination stress is
  responsible for relapse
• Emotional stressors are the primary activators of
  social stress

9
Introduction

• Social defeat in rodents could be considered as
  a stressor with essential ethological relevance
• Previous studies reveal that morphine-induced CPP
  can be influenced by prior social defeat
• Effects of social defeat on reinstatement of
  morphine seeking have not been studied

10
Objectives of this Study

1. Evaluate the effects of two physical stressor
   (restraint and tail pinch) on the reinstatement
   of morphine-induced CPP
2. Determine the effects of a social defeat on
   reinstatement of morphine-induced CPP
3. Compare the effects of physical and social
   stressors on reinstatement of morphine-induced CPP

11
Methods

• OF1 strain of mice, 42 days old
• Housed in groups of 4 for 10 days before training
• All animals lived in a vivarium (21ºC)
• Reversed light schedule (animals were tested when
  they are normally active)
• Procedures in accordance with the European
  Communities Council Directives

12
Conditioned Place Preference Examples
13
Methods

• 8 identical Plexiglas boxes were used
• Black and side were separated by a gray central
  compartment
• Black side had fine grid floor (preferred?)
• White side had wide grid floor
• 4 beams in each black and white and 6 in central
  area recorded position of mouse

14
Methods

• Pre-Conditioning (Pre-C) Manzanedo et al., 2001
• Mice given access to both compartments for 900 s
  (15 min) for 3 days
• Recorded on 3rd day to determine unconditioned
  place preference
• Animals that showed aversion (lt33) or preference
  (lt66) for any compartment were discarded (n10)

15
Methods

• Pre-C notes
• In each group half of animals were assigned
  morphine or control injection in black
  compartment, while the other half were assigned
  the injection in white compartment
• No significant differences between in time spent
  in the drug-paired and vehicle paired
  compartments during Pre-C

16
Methods

• Second phase
• 4 days of conditioning to each groups dose
• Mice first received saline injection in the
  saline paired compartment for an amount of time
  dependent on dose
• 4 hour interval (home cage?)
• Mice received morphine injection before
  confinement to drug-paired compartment
• Central area was closed

17
Methods

• Third phase Post-C
• Test the rat for preference by removing
  guillotine doors on 8th day
• Place rat in CPP enclosure for 15 min
• Recorded which chamber rat preferred.
• Only animals conditioned with morphine presented
  CPP in all experiments.

18
Methods

• Extinction
• Daily extinction sessions (15 min) were performed
  
• When the time spent in the drug-paired
  compartment was not significantly different from
  Pre-C levels extinction.
• Apparently all animals received the same number
  of extinction session independent of individual
  scores.
• Saline conditioned groups only went through one
  extinction session to confirm lack of CPP.
• Morphine-conditioned mice with weak CPP (gt60 s)
  were discarded.

19
Methods

• Reinstatement
• Reinstatement sessions were the day after the
  last extinction session
• Respective stress protocol was administered in
  the vivarium
• Mice were put back into the CPP chamber with
  doors open (15 min)
• This occurred immediately after stress or was
  delayed for 15 min.

20
Table 1
21
Experiment 1

• Effects of restraint on the reinstatement of
  morphine-induced CPP
• Extinction sessions (3-10 sessions) and after 24
  h after extinction
• 15 min of immobilization-induced stress
• impossible for mouse to turn around
• Immediately or 15 min after (delayed) restraint
  stress, the reinstatement test was performed.

22
Figure 1 - Immediate
23
Figure 1 Delayed Reinstatement
24
Table 1
25
Experiment 2

• Effects of tail pinch on morphine-induced CPP
• One day after the last extinction session, mice
  were submitted to a modified tail-pinch schedule
  (15 min) to evaluate physical/tactile stress.
• Tail-pinch performed with a plastic clothespin
  (800 g pressure)
• Fastened on tail at 1-1.5 cm from body
• Performed in different cage
• Immediately or 15 min after (delayed) tail-pinch,
  the reinstatement test was performed.

26
Figure 2 Immediate
27
Figure 2 Delayed Reinstatement
28
Experiment 3

• Effects of an agonistic encounter on the
  reinstatement of morphine-induced CPP
• 24 h after the last extinction session, mice were
  socially stressed by an aggressive opponent of
  equal age and body weight for 15 min
• Aggressive opponent was selected for high
  aggression
• Experimental mice were defeated in lt 30 s,
  because opponent initiated attack immediately
  after seeing experimental mouse.
• Defeat posture was an upright submissive position
  with limp forepaws, upwardly angled head, and
  retracted ears.

29
Experiment 3

• Agonistic encounter without social defeat
• Opponents were made temporarily anosmic by
  intranasal lavage with 4 zinc sulfate solution
  24 h before encounter (Smoothy et al., 1986)
• These mice elicit attack but never initiate
  attack
• No aggressive behaviors were observed
• These controlled for the extraneous variable of a
  social interaction.
• Furthermore, another control was exposure to the
  cage without any social interaction.

30
Table 1
31
Figure 3 Social Defeat immediate reinstatement
32
Figure 3 Social Defeat Delayed Reinstatement
33
Figure 4 Non aggressive Social interaction
34
Figure 4 Social interaction Delayed
Reinstatement
35
Assessment of corticosterone Concentration

• Immediate and delayed (30 min after) obtention of
  blood sample.
• Immediate CORT release after stress is not a
  reflection of the actual stressor or of ACTH
  release.
• This is because adrenocortical synthesis is
  saturated.
• Severe stressors are characterized by a slower
  return to basal hormone levels.
• Corticosterone levels were determined using a
  radioimmunoassay.

36
Corticosterone levels after stress exposure

• Stressed groups presented higher CORT levels than
  controls.
• Animals exposed to social defeat had higher
  levels of CORT than animals exposed to restraint
  or tail pinch in the 30 min groups.
• Defeat stress causes CORT increase to remain
  elevated?

37
Summary

• Physical stressors (restraint, tail pinch)
  administered immediately or 15 min before
  reinstatement tests are capable of reinstating
  CPP after extinction.
• In general agreement with foot shock results.
• Psychological stress (social defeat) suffered
  immediately or 15 min before reinstatement tests
  are capable of reinstating CPP after extinction.
• Social stress is as effective as physical stress
  in reinstating drug-seeking behavior.

38
Discussion

• Pavlov first described the reinstatement of
  learned behaviors after extinction in his
  classical conditioning studies with dogs.
• Social defeat induces activation of the
  mesocorticolimbic system which is involved in
  social stress-induced reinstatement.
• Social defeat stress also increases µ-opioid
  receptor mRNA in the VTA, also involved in
  stress-induced reinstatement.

39
Discussion

• Both physical stressors and social defeat
  increase extracellular levels of DA in the
  nucleus accumbens and prefrontal cortex.
• Foot shock induces less DA release than drug
  priming but can be a more effective stimulus for
  reinstatement.
• Stressors may induce a withdrawal-like state that
  lead to relapse.
• But, withdrawal from heroin does not reinstate
  heroin seeking as stressors do.

40
Discussion

• Stress may provoke relapse by interfering with
  neuronal inhibitory processes, which inhibits
  responding when reinforcers are not available.
• Foot shock stress increases resistance to
  extinction.
• Footshocks reinstating effects depend on the
  medial septum, brain area involved in response
  inhibition.

41
Discussion

• Repeated exposure to drug might sensitize or
  induce neuroadaptations in brain systems involved
  in the stress response.
• Foot shock stress increased CRF levels in the VTA
  and blockade of CRF receptors in this area
  attenuated the stress-induced reinstatement of
  cocaine seeking.
• Experiments in this area are limited.
• Neuroadaptation factors (BDNF, MAPK signal
  transduction, glutamate receptors) have yet to be
  explored in stress-induced reinstatement
  experiments.

42
Discussion

• One study (foot shock, restraint, heroin)
  suggested that the effects of stress on
  reinstatement are context and time dependent.
• This study shows contradictory results.
• Restraint and tail pinch were administered in a
  different environment from CPP conditioning.
• 2 temporal intervals (0,15 min) before
  reinstatement test.
• Here, all mice produced a clear reinstatement in
  morphine-induced CPP.

43
Discussion

• An encounter with a non-aggressive opponent does
  not reinstate CPP.
• Suggests that social interaction alone is not
  stressful.
• A state of arousal induced by exposure to an
  appetitive stimulus (receptive female) has no
  effect on the reinstatement of heroin seeking.
• Suggests that the neurochemistry of arousal from
  a social interaction might be the same as a
  negative stressor but yields distinguishable
  behavioral outcomes.

44
Conclusion

• Exposure to social stress produces the same
  effects as physical stressors on the
  reinstatement of morphine-induced CPP.
• Mechanisms involved are yet to be elucidated.
• Further conclusions?

45
THE END

• Thank you
• Andrew R. Burke