Transcript RNA interference as a resistance mechanism against crop parasites

Orexin and binge-like
consumption:
Sucrose, Saccharin, Ethanol
ANDY DEEMER
Bingeing
• Eating, Drinking, Drugs
• Orexin plays a role but…
•
•
•
•
Caloric Status? (Non-)Caloric reward?
Conditioning? Cue-induced reward seeking
♂ ♀ differences?
Inconsistencies in several studies
• Methodology related? Gender related?
• Gender Differences
• Orexin system
• ♀ rat - higher Orexin A and OXR1 in HL than ♂
• Responses to food restriction
• More studies = more differences revealed.
Economic, Social and Therapeutic
Potential of Orexin Research
• Overconsumption of ______________ is a
significant public health/economic issue.
• Potential applications in obesity, diabetes, drug
abuse
• Understanding factors at play = better therapy
• Caloric restriction important?
• Gender diffs?
• Bingeing substance (caloric/non, hedonic)
Cason & Aston-Jones 2014
Role of orexin/hypocretin in conditioned
sucrose-seeking in female rats
• Orexin system sexual dimorphism
• Eating disorders more common in women
• OX system gender differences may play a role
• Hedonic vs Homeostatic
• Saccharin vs sucrose
Cason & Aston-Jones 2014
Methods
• Female Sprague-Dawley rats
• Two Feeding Treatments
• Ad libitum (AL) = free access
• Food-restricted (FR) =
"SpragueDawleyRat" by Jean-Etienne Minh-Duy Poirrier - http://www.flickr.com/photos/jepoirrier/422469518/in/set-72157594329856603/. Licensed under CC BY-SA 2.0 via Wikimedia Commons - http://commons.wikimedia.org/wiki/File:SpragueDawleyRat.jpg#/media/File:SpragueDawleyRat.jpg
• 1 daily feeding - maintained at 85% of AL body mass)
• Drug = SB-334867 (OXR1 antagonist)
• 3 doses used in this study 10, 20, and 30 mg/kg
• Intraperitoneal (4 mL/kg)
Cason & Aston-Jones 2014
Methods
• Training – self-administer sucrose pellets
• Press lever = get sucrose pellet
• Compound cue (light and sound)
• Fixed ratio (FR) 1 responding
•
Until 10 sessions earning > 9 sucrose pellets
• Progressive ratio (PR) responding
•
•
Using mice previously trained from FR experiment
Find breakpoint for lever presses
Images adapted from: http://rnpsychology.org/demo/index.htm
Cason & Aston-Jones 2014
Fixed vs Progressive Ratio Schedule
Progressive Ratio = 5ePelletNumber*0.2 – 5
Point at which no more
rewards are earned over an
hour = breakpoint.
Cason & Aston-Jones 2014
Methods - Ratio Experiments
Fixed Ratio:
Train
Vehicle/Drug injection
Self-administration session
Progressive Ratio:
Use rats from fixed ratio experiment
Train
Self-administration session
Each rat was tested at multiple doses (2 doses)
Vehicle/Drug injected 30 minutes prior to
test sessions (all 3 of today’s papers)
Injection
Cason & Aston-Jones 2014
Cue-induced reinstatement of
sucrose-seeking
• Take mice used for FR1 (but not PR)
• Daily extinction sessions
• Lever presses = nothing happens
•
•
Untraining
Until 2 consecutive sessions with < 25 active lever
presses
• Bring cues back (but no sucrose)
• Measure SB effect on presses
• 2 late extinction and 2 reinstatement sessions
Cason & Aston-Jones 2014
Self-Admin Training
• # days to train similar for
FR and AL rats
• Food-restricted
•More active and inactive
presses + pellets earned
Cason & Aston-Jones 2014
Fixed Ratio
Experiment
• FR more active presses +
pellets earned
• SD sig. effect on active
presses only at high dose
• No sig. effect of SD on
pellets earned
Cason & Aston-Jones 2014
Progressive Ratio Experiment
• No effect of group
or SB dose on
breakpoint
Cason & Aston-Jones 2014
Cue-induced reinstatement of
sucrose-seeking
• AL and FR rats met extinction • Food group effect
criteria < 5 extinction sessions • No sig. SB effect on
CIRSS
• SB attenuation of active
• Contrary to their previous
presses - late extinction
findings in ♂ rats
Cason & Aston-Jones 2014
Cason & Aston-Jones Take Homes
• Cue-induced reinstatement and SB-334867
• Sex-dependent story
• ♀ - OXR1 unnecessary; important in ♂
• Increased extinction responding
•
Orexin role in learning/reward valuation in females?
• Future Research:
•
•
Systems at play in ♀ cue-induced seeking (e.g. leptin, insulin,
ghrelin, estrogen)
Untangling ♂ ♀ circuitry differences
• SB-334867 effects on Fos expression in orexin targets
Cason & Aston-Jones 2014
Cason & Aston-Jones Take Homes
• Agree with conclusion?
• “Our findings show that OxR1 regulates operant
responding for sucrose reinforcement, but not
motivation to work…”
• Fig 4, Panel 1:
With OXR1 unblocked, try
less = regulates motivation
to work?
Alcaraz-Iborra et al. 2014
Binge-like consumption of caloric and noncaloric palatable substances in ad libitumfed C57BL/6J mice: Pharmacological and
molecular evidence of orexin involvement
• Objectives:
• SB-334867 effect on binge consumption of sucrose
and saccharin in ad libitum-fed mice
• Effect of repetitive sucrose and saccharin bingeing
on OX mRNA expr in LH
•
Mirror chronic morphine, cocaine, ethanol?
Alcaraz-Iborra et al. 2014
Methods
• Male C57BL/6J (8 wks old @ start)
• Drug = SB-334867 (OXR1 antagonist)
• Intraperitoneal (10 mL/kg)
• Drinking-in-the-Dark (DID) procedure
• High voluntary bingeing on ethanol, sucrose,
saccharin during early part of dark cycle
• Provide sucrose/saccharin ~ 3 hrs. into dark cycle
• Open-field activity monitoring
• qPCR
Alcaraz-Iborra et al. 2014
Experiment 1:
Effect of SB on sucrose and saccharin binge
drinking
• Binge-training / screening
• 3 days of DID (2 hrs bottle access)
•
ip injection w/ vehicle - habituation
• 4th day = test day (4 hrs bottle access)
•
Injection of SB (10, 20 or 30 mg/kg) or vehicle 30 min
prior to test
• Measure:
•
•
Liquid imbibed
Calories consumed (chow included)
Alcaraz-Iborra et al. 2014
• ip SB-334867 ↓
sucrose and
saccharin bingeing
• Higher doses ↓
sucrose bingeing
more than lowest
dose
• Strange calories
consumed data in
group
Totalsaccharin
calories consumed (kcal/g/4
h) on test day
SB (10 mg)
Sucrose group
(kcal/g)
3.47 ± 0.20
2.24 ± 0.14**
Saccharin group
(kcal/g)
2.74 ± 0.42
0.20 ± 0.09**
SB (20 mg)
1.11 ± 0.28**
SB (30 mg)
0.97 ± 0.19**
1.24 ± 0.49**
0.64 ± 0.39**
VEH
*
Alcaraz-Iborra et al. 2014
Experiment 2:
Effect of SB on locomotor activity
• Open-field locomotor activity monitoring
•
•
•
Days 1-3: Habituate to injection (vehicle) and activity
chamber & record behavior
Day 4: 30 mg/kg SB → activity chamber
Evaluate difference
• Purpose: SB-334867 impact distance traveled and
movement time?
•
Explanation for reduced fluid/food intake?
• Results: No sig. diff.
•
The means they present seem fairly different, but no sig.
diff
Alcaraz-Iborra et al. 2014
Experiment 3:
Repeated sucrose/saccharin bingeing and
mRNA expression in the HL
• Repeated bingeing (DID – i.e. voluntary)
•
Four 2-hour daily binge sessions
•
Sucrose, Saccharin, or Water group
• Brain dissection → LH removed → RNA extracted
• qPCR with GAPDH for comparison
Alcaraz-Iborra et al. 2014
• Repeated bingeing
↓ HL OX mRNA
Consumption data (ml/kg/2 h) associated with the
mRNA study.
Water
Sucrose
Saccharin
consumption consumption consumption
(ml/kg/2 h)
(ml/kg/2 h)
(ml/kg/2 h)
• Sucrose group: 5x >
water intake
Day 1
34.40 ± 0.64
72.42 ± 8.78
67.70 ± 6.86
Day 2
23.42 ± 3.50 117.72 ± 10.46 94.41 ± 15.24
Day 3
22.14 ± 0.87 125.80 ± 4.16 77.88 ± 6.87
Day 4
21.12 ± 2.29 103.74 ± 8.07 86.69 ± 3.64
Alcaraz-Iborra et al. 2014:
Take-homes
• OXR1 role in caloric bingeing in AL-fed animals
• SB-334867 ↓ caloric & non-caloric bingeing in
AL-fed ♂ mice
• Unclear why in AL-fed mice (ip injection)
•
Future Research: Site-directed SB studies
• 1st evidence of OXR1 role in non-caloric
bingeing/hedonic overconsumption
• Repeated daily bingeing ↓ HL OX expr (qPCR)
Alcaraz-Iborra et al. 2014:
Take-homes
• SB effect on bingeing not caused by SB-altered
locomotion.
• Why not show more data?
• DID potential issue:
• Energy status at time of DID
• Future Research: Alter energy status during DID
Olney et al. 2015
Binge-like consumption of Ethanol and
Other Salient Reinforcers is Blocked by
Orexin-1 Receptor Inhibition and Leads to a
Reduction in Hypothalamic Orexin
Immunoreactivity
Illustration by Emily Coren.
Olney et al. 2015
Purpose
• Characterize OXR1 role in “binge” drinking
• Ethanol, sucrose, saccharin
• Binge-like EtOH and Sucrose drinking
• Effect on OX immunoreactivity in HL
• Effect of ip SB
• SB specificity for EtOH modulation
• Compare with saccharin bingeing
Olney et al. 2015
Methods
• C57BL/6J male mice (7-9 wks old)
• SB-334867 (0, 5, 10 mg/kg)
• Bingeing cycles:
• 1 or 3 cycles; EtOH or Sucrose
• DID (modified 2 hrs not 4 – short drug action)
• Except experiment 1 – used 4 hrs.
• Blood alcohol content
• Brain dissection
• Orexin A immunoreacitivy (LH and PFA)
Olney et al. 2015
Methods
• SB ip injection
• Reduced EtOH consumption? Saccharin?
• Each mouse, all doses
•
4 day cycle with 3 days rest between doses
• Locomotion
• Open field test with Saccharin group mice
• 2 hours
• 0 or 10 mg/kg SB
Olney et al. 2015
• No cycle effect on:
•EtOH Consump.
•Sucrose
Consump
•BEC
• EtOH bingeing ↓
LH OX levels
Olney et al. 2015
EtOH Bingeing reduced HL OX
Olney et al. 2015
• SB ↓ EtOH consumption
• Short-lived (Hr 1 vs. Hr 2)
• BEC levels parallel total consumption
• SB ↓ Saccharin consumption
• Only signif. largest dose over 2 hrs.
• Take Home: SB effect on ↓ bingeing not
specific to EtOH or caloric foods
Olney et al. 2015
Locomotion
• Like Alcaraz-Iborra:
• SB-induced general lethary
cause of reduced bingeing?
• Open field test:
• No impairment of
locomoter activity
Olney et al. 2015: Take-homes
• OXR1 role in caloric and non-caloric bingeing
• Not EtOH specific
• Bingeing ↓ HL OXR1
• SB effects not due to altered locomotion
• DID 4hr vs 2hr
• Duration of drug effect may be less than 4 hrs
• Inconsistent dose effects between researchers
•
SB variability: Supplier effects, batch effects
• Future Research:
• Mechanisms responsible for post-binge OX and
OXR1 levels
• Interplay of dynorphin and OX w/in VTA
Overall Take-homes
Site-specific studies necessary (not ip injection)
- ID OX circuits underlying phenomena
- VTA likely involved
Thank You!