Download Russ Carpenter

Prenatal Cannabis Exposure
Increases Heroin Seeking with
Allostatic Changes in Limbic
Enkephalin Systems in Adulthood
M.Sabrina Spano, Maria Ellgren,
X. Wang, Yasmin L. Hurd
• THC exposure in prenatal development:
– Higher rates of fetal distress
– Growth retardation
• Transferred from mother to offspring via
placental blood during gestation and via
maternal milk during lactation
• CB1 receptors mediate neural actions of
cannabinoids & emerge early in development
– synaptogenesis, proliferation and migration of
neuronal cells
• Clinical studies indicate in utero exposure is
associated
– impulsive behavior, cognitive impairment,
psychiatric disorders (schizophrenia, anxiety) in
later life
• Kandel 03 suggested potential for
cannabis to increase the risk of
consumption of other drugs of abuse
• hypothesized that long lasting
neurobiological changes in neuronal
systems linked with limbic function might
be affected by in utero THC exposure
– Gateway….?
Opioids
• Endogenous opioid system shares
neuroanatomical and neurochemical
characteristics with the cannabinoid
system; tightly linked
• 3 families
– Enkephalin from preproenkephalin (PENK)
– Dynorphin from preprodynorphin
– Endorphin from proopiomelanocortin (POMC)
• Cannabinoids stimulate release of
enkephalin in the NAc and VTA
– NAc is part of the mesocorticolimbic circuit
– VTA is origin of DA mesocorticolimbic circuit
• PENK widely dist. (NAc, amygdala, PFC)
• Previous human studies show sig.
impairment of PENK and meso-limbic DA
genes in association with in utero CB
exposure
– Enduring effects into adulthood?
– Influence adult behavior relative to addiction
disorders?
• In rats, perinatal THC exposure alters
opioid gene expression, opioid receptor
binding and morphine self-administration
– Differs between males and females
• treated with THC through gestation &
entire lactation period
– Dams treated with THC orally, different
pharmacokinetic property than alternate
routes of ingestion e.g. smoking, iv
Methods
Fear Leads to Anger---Anger Leads to Stress
Stress Leads to Doobies—Doobies lead to Twinkies
Test animals
•
•
•
•
Female Long-Evans rats
Reversed light/dark cycle
Permanent right jugular catheter for IV
Following surgery were mated with a male
to induce pregnancy
Admin of drug
• Drug exposure limited to prenatal period
– Gestational day5 to post-natal day 2
– Corresponds with mid-gestation period in
humans ~ week 20
• THC given IV to better mimic the
pharmacokinetics of smoking cannabis in
pregnant human females
– THC injected .15mg/kg daily
During Drug Treatment
• Maternal weight gain, gestational length and
fetal weights were recorded
• PND 2 pups from both groups cross fostered;
some brains taken at this time
• PND 21 males weaned; housed 4/cage
• PND 55 iv catheter implanted
– Housed individually, 7 day recovery
• These are the test animals
– More brains taken PND 62 prior to behav studies
Heroin Self Admin
• Active lever press = 15ug/kg in 85 ml fluid
• Fixed Ratio 1, 10s timeout, 3 hour
sessions during dark cycle
• After 6 days, dose increased to 30 mg/kg
• Baseline= <15% change in # bar presses
for 3 consecutive days
• During training food=20g/day
– Ad libitum after stable response reached
Dose Response
• Between session dose response test
• Rats now receive either higher 60,100mg
or lower 7.5, 15 mg doses
– Self admin as normal for 3 days, order of
presentation was random
• Following test, back to 30mg maintenance
for 5 more days
Food Stress
• Food deprived for 24 hours
• Response measured following day
Extinction and Reinstatement
• First week of extinction = saline
• Following days = no fluid injected
• Decrease in bar presses of 85% baseline
for 3 days = extinction
• Priming:
– saline
– heroin 0.25 mg/kg sc 10 min prior
– CB1 antagonist 3 mg/kg ip + heroin
Results
• No significant change between groups in
– % weight gain in pregnant dams
– Gestational length
– Pup length at PND 2
– Pup weight
– Weight between groups at PND 62
• Start of heroin self adminstration training
15 mg/kg/infusion
30 mg/kg/infusion
• No self admin at 15 mg/kg dose
• Both THC and Vehicle groups self admin
at 30 mg/kg dose
• THC animals show shorter latency to first
active response 34 ± 0.84s vs 115 ± 0.91s
Dose Response Test
• Dose-dependant decrease in responding
with increasing heroin dose
• THC animals show higher responding at
lower doses (7.5 & 15 mg/kg)
• No diff in responding at 30, 60 or 100 mg
doses between groups
Response after Stress Test
• THC animals respond to food deprivation
stress with more active lever presses
Extinction, Priming, SR Treatment
• THC group responded to active bar more
during first day of extinction (more than
maintenance, more than vehicle group)
• Heroin priming reinstated active bar
presses
– THC group responded to active bar sig more
than they had during maintenance
• SR treatment abolished priming induced
reinstatment
Locomotion Response
• THC group showed less locomotion during
acquisition and maintenance phases
– No diff during extinction phase
• Differences are due to heroin intake
CB1 and m Opioid Receptor G-Protein
Coupling
• Agonist-stimulated GTPgS binding
• No change in m or CB1 binding at PND2
• In adults, prenatal THC exposure
significantly associated with
– decreased m opioid binding in NAc shell
– Increased binding in SN
– No change in CB1 receptor coupling
1=NAc shell, 2=NAc core, 3=CP, 4=VTA, 5=SN
PENK mRNA Expression
• Prenatal THC Exposure decreased PENK
mRNA expression in NAc at PND2
– No reliable measurements from amygdala
• No change in preprodynorphin which is colocalized in the
NAc
• HOWEVER…
• Prenatal THC exposure increased PENK mRNA
in Adults at PND 62 in
– NAc core and shell
– Central and medial amygdala nuclei
• Compensation?
1=NAc shell
2=NAc core
3=CP
4=m.amy
5=c.amy
Increased
PENK
mRNA in
cAmy
Discussion
• Hyperactivity of the mesocorticolimbic
enkephalinergic system in adult animals
may be due to the blunted PENK gene
expression during early development
Self Administration
• No diff in self admin of heroin between
groups in adulthood
– THC group showed
•
•
•
•
Shorter latency to first active lever press
Higher response to lower doses of heroin
Increased response following food stress
Higher level of seeking during extinction
• Suggests long term vulnerability in the
motivation to self-administer heroin
Relapse!
• Following 21 days of drug extinction
– Heroin primed THC group responded with
higher number of bar presses than
maintenance (increased seeking)
– CB1 antagonist SR141716A completely
blocked heroin primed relapse in both groups
– Cross talk between opioid and cannabinoid
systems
m opioid receptor binding
• THC group show less mOR binding in NAc
– Key region in reward processing
– also modulates locomotion
• THC group showed less loco when on heroin
• Similar to mOR deficient animals
Take-Home Message
• THC administered during prenatal period
significant affects PENK mRNA expression
during prenatal period and adulthood
• Alterations of the opioid system last into
adulthood and enhance vulnerability to
opiate-seeking behavior