Relapse to food seeking Role of CRF PYY

Relapse to food seeking: Role of CRF, PYY 3 -36, and hypocretin Yavin Shaham Neurobiology of Relapse Section Behavioral Neuroscience Branch IRP/NIDA/NIH/DHHS, Baltimore Post-doctoral fellows: Sunila Nair, Udi Ghitza Students (NIH training program): Sarah Gray, Sam Golden, Tristan Adams-Deutsch CRF = Corticotropin-releasing factor PYY 3 -36 = Peptide YY 3 -36 Hypocretin = Orexin

Neurobiology of Relapse Section Shaham laboratory Staff scientist: Jennifer Bossert Post-doctoral fellows: Sunila Nair, Charles Pickens Post-baccalaureate students: Sam Golden, Kristina Wihbey, Tristan Adams-Deutsch Hope laboratory Staff scientist: Bruce Hope Post-doctoral fellow: Eisuke Koya Graduate student: Danielle Guez (Yale U) Post-baccalaureate student: Alex Berkow

Ongoing research projects 1. Relapse to food seeking 2. Context-induced reinstatement of heroin seeking 3. Incubation of cocaine craving (time-dependent increases in cue-induced cocaine seeking) 4. Incubation of conditioned fear (time-dependent increases in conditioned fear) 5. Neuronal mechanisms of context-dependent cocaine sensitization (Hope lab) Extramural collaborations: AD Le (Toronto U): Stress-induced relapse to alcohol (NIAAA) Geoff Schoenbaum (U of Maryland): Effect of cocaine self-administration on learning and memory (NIDA) Abraham Zangen (Weizmann Inst. , Israel): Developing a novel conflict model of drug (NIDA) Marina Wolf and Micky Marinelli (Rosalind Franklin University): Accumbens synaptic plasticity and incubation of craving (NIDA)

Outline • A brief description of a reinstatement model—an animal model of relapse to drug-taking behavior • The adaptation of the reinstatement model to study: 1. Role of CRF in reinstatement of food seeking induced by: * A pharmacological stressor (yohimbine) * Acute re-exposure to food pellets (pellet priming) 2 -3. Effect of PYY 3 -36 and hypocretin 1 receptor antagonist on reinstatement induced by: * Yohimbine * Pellet priming * Cue (a tone-light cue previously paired with pellet delivery)

The reinstatement model of drug relapse Self-administration training (Drug is NOT available) X TESTING (Drug is NOT available) X Drug priming Drug cues Stress Experimental day Cumulative # of publications Lever presses (Drug is available) Extinction 500 400 Reinstatement studies (1971 -2007) 478 332 300 200 100 0 0 3 4 10 13 96 20 70 75 80 85 90 95 00 05 07 Year

Study 1 The role of CRF in reinstatement of food seeking induced by the pharmacological stressor yohimbine Hypothalamic-pituitaryadrenal (HPA) axis Extra-hypothalamic CRF systems From Behan et al. 1996

Stress and relapse to maladaptive eating habits The clinical problem: Excessive eating of unhealthy food is a major health problem Stress can provoke relapse to unhealthy eating habits, and humans are particularly vulnerable to this effect when dieting The preclinical situation: This issue has not been explored in preclinical laboratory studies, which have primarily focused on the effect of stress on ongoing feeding behavior

Why yohimbine? Yohimbine (an alpha-2 adrenoceptor antagonist) increases brain noradrenaline release and induces stress-like responses in humans and laboratory animals Yohimbine induces heroin craving in human addicts Yohimbine reinstates drug seeking in monkeys and rats Heroin Lever presses 100 * 80 60 * * 75 0 2. 5 15 0 0 1. 25 2. 5 * 120 80 30 0 Nicotine * 45 25 20 * 60 50 40 0 Alcohol Methamphetamine * 40 0 1. 25 2. 5 0 0 0. 625 1. 25 Yohimbine dose (mg/kg, i. p. ) Yap & Shaham. 2000 (unpublished) Shepard et al. 2004 Le et al. 2005 Biol. Psychiatry Psychopharmacology Le et al. 2007 (Unpublished)

Why CRF? CRF injections into the ventricles and several extrahypothalamic sites (BNST, VTA, median raphe) reinstate drug seeking Non-selective and selective CRF 1 receptor antagonists attenuate stressinduced reinstatement of drug seeking Heroin Lever presses 60 Cocaine No stress Intermittent footshock 45 * 30 * 0 15 * * 15 0 Alcohol 30 0 15 CP-154, 526 dose (mg/kg) Shaham et al. Psychopharmacology, 1998 * 30 * 0 15 30 CP-154, 526 dose (mg/kg) Le et al. Psychopharmacology, 1998 * 0 15 30 MTIP dose (mg/kg) Gehlert et al. J Neurosci, 2007

Experimental procedure Pellet self-administration Three 3 -h sessions/d every other day (FR-1; 20 sec timeout) Limited access to regular food: ~75% of daily intake Extinction pellets NOT available Tests for reinstatement pellets NOT available Limited access to regular food: ~75% of daily intake (diet condition) Exposure to: Yohimbine Non-contingent pellet (priming) 45 mg pellets: 25% fat 48% carbohydrate

Changes in body weight during training, extinction, and reinstatement (n=35) Pellet self-administration Extinction Three 3 -h sessions/d every other day (pellets NOT available) (FR-1; 20 sec timeout) Limited access to regular food: ~75% of daily intake Weight change (g) Training 10 0 -10 -20 (pellets NOT available) Limited access to regular food: ~75% of daily intake (diet condition) Daily weight fluctuations 20 Tests for reinstatement Extinction & Reinstatement Pellets available No pellets 10 Ghitza et al. Neuropsychopharmacology, 2006 20 Day 30 40

Development of “compulsive” food-taking behavior during training Pellet self-administration Extinction Three 3 -h sessions/d every other day (FR-1; 20 sec timeout) Responses or pellets (9 h) Limited access to regular food: ~75% of daily intake Limited access to regular food: ~75% % of daily intake (diet condition) Timeout responses Pellets 1200 900 600 300 0 2 4 6 8 Tests for reinstatement 10 Training day 12

The CRF 1 receptor antagonist antalarmin decreases yohimbine-induced reinstatement of food seeking Pellet self-administration (pellets NOT available) Limited access to regular food: ~75% of daily intake Reinstatement 450 300 0 3 h/day 1 Vehicle 20 mg/kg 40 mg/kg 100 2 3 4 5 6 7 8 9 10 Extinction day Lever presses (3 h) Lever presses Antalarmin dose 9 h/day 150 (pellets NOT available) Limited access to regular food: ~75% of daily intake (diet condition) Extinction 600 Tests for reinstatement Extinction 75 50 * 25 0 0 * 2. 0 Yohimbine dose (mg/kg, i. p. ) Pellet priming Antalarmin was synthesized by Dr. Kenner Rice

Antalarmin decreases yohimbine-induced reinstatement of alcohol seeking 80 60 40 20 0 Vehicle Yohimbine (1. 25 mg/kg) * * 0 10 20 Antalarmin dose (mg/kg, i. p. ) Corticosterone (ng/ml) Lever presses (1 h) Yohimbine-induced reinstatement Yohimbine-induced plasma corticosterone release 750 Vehicle Yohimbine (1. 25 mg/kg) 500 250 0 0 20 Antalarmin dose (mg/kg, i. p. ) Marinelli et al. Psychopharmacology, 2007

The CRF 1 antagonist antalarmin reverses yohimbine’s effects on social and “antisocial” behaviors “Antisocial” behavior 100 Antalarmin dose 75 Vehicle * 50 20 mg/kg 25 0 0 2. 0 Yohimbine dose (mg/kg, i. p. ) Antisocial bouts (10 min) Social behavior 40 * 30 20 10 0 0 2. 0 Yohimbine dose (mg/kg, i. p. ) Social and “antisocial” behaviors were evaluated using a social interaction test: Social behavior: crawling together, grooming, sniffing, licking “Antisocial” behavior: wrestling, distress vocalization, confrontation

Conclusions: Study 1 As in the case of drug relapse, the reinstatement model can be used to study mechanisms underlying relapse to maladaptive eating habits The effect of antalarmin on yohimbine-induced reinstatement of food and alcohol seeking is likely mediated by extrahypothalamic CRF 1 receptors

Project 2 Effect of PYY 3 -36 on yohimbine-, pelletpriming- and cue-induced reinstatement of high -fat food seeking Background The gut peptide PYY 3 -36 is an endogenous Y 2 NPY receptor agonist that decreases home-cage feeding after systemic or arcuate nucleus injections PYY 3 -36 systemic effects are reversed by systemic or arcuate injections of the Y 2 receptor antagonist BIIE 0246.

Batterham + 9 authors (2002) Gut hormone PYY(3 -36) physiologically inhibits food intake. Nature 418: 650 -654. Batterham + 7 authors (2003) Inhibition of food intake in obese subjects by peptide YY 3 -36. N Engl J Med 349: 941 -948. Pittner + 9 authors (2004) Effects of PYY(3 -36) in rodent models of diabetes and obesity. Int J Obes Relat Metab Disord 28, 963 -971 But…. Tschop + 42 authors (2004) Does gut hormone PYY 3 -36 decrease food intake in rodents? Nature 430: 1 p following 165; discussion 162 p following 165. Boggiano + 36 authors (2005) PYY 3 -36 as an anti-obesity drug target. Obesity Reviews 6: 307 -322 * The effect of PYY(3 -36) on operant food self-administration and reinstatement has not been assessed

Experimental procedure Pellet self-administration Two 3 -h sessions/d every other day (FR-1; 20 sec timeout) Limited access to regular food: ~65% of daily intake Extinction pellets NOT available Tests for reinstatement pellets NOT available Limited access to regular food: ~65% of daily intake (diet condition) (Tone-light cue NOT available) (Tone-light cue available) XX 45 mg pellets: 35% fat 45% carbohydrate Exposure to: Yohimbine Non-contingent pellet Cue

PYY 3 -36 has a minimal effect on high-fat pellet selfadministration two 3 -h sessions/d every other day (FR-1; 20 sec timeout) Limited access to regular food: ~65% of daily intake Food pellets (3 h) Pellet self-administration 225 Session 1: Total intake Session 2: Total intake 150 75 0 0 100 200 0 100 PYY 3 -36 dose (µg/kg, i. p. ) Ghitza, Nair et al. The Journal of Neuroscience, 2007 200

PYY 3 -36 inhibits pellet-priming- and cue-induced reinstatement, but not yohimbine-induced reinstatement Cue Lever presses (3 h) 100 No pellet Pellet 75 50 * 25 0 0 * 100 200 PYY 3 -36 dose (µg/kg, i. p. ) Lever presses (3 h) Pellet priming 250 100 No cue Cue 75 50 * 25 0 0 100 200 PYY 3 -36 dose (µg/kg, i. p. ) Yohimbine Vehicle Yohimbine (2. 0 mg/kg) 200 150 100 50 0 * 0 100 200 PYY 3 -36 dose (µg/kg, i. p. )

Conclusions: PYY 3 -36 Systemic PYY 3 -36 decreased pellet-priming-induced reinstatement at doses that had minimal effect on ongoing high-fat food selfadministration The systemic effect of PYY 3 -36 on reinstatement generalizes to cuesbut not to yohimbine-induced reinstatement of food seeking The systemic effect of PYY 3 -36 is dependent on Y 2 receptors Systemic PYY 3 -36 had no effect on reinstatement of heroin seeking, suggesting that its effects are specific to food relapse

Project 3 Effect of the hypocretin 1 receptor antagonist SB 334867 on high-fat food self-administration and relapse to food-seeking in rats The hypocretins (orexins) Prepro-hypocretin Hypocretin 1 receptor From Sakurai, 2006 and 2007 Hypocretin 2 receptor

Why hypocretin 1 receptors? Systemic injections of SB 334, 867 decrease home-cage food intake and reverse hypocretin 1 -induced increases in food intake (Rodgers et al. 2002) Systemic injections of SB 334, 867 decrease: Reinstatement of morphine conditioned place preference induced stimulation of lateral hypothalamus hypocretin neurons (Harris et al. 2005) Footshock stress-induced reinstatement of cocaine seeking (Boutrel et al. 2005) Cue-induced reinstatement of alcohol seeking (Lawrence et al. 2006)

SB 334, 867 decreases high-fat pellet self-administration Pellet intake Time course 50 300 40 * * 200 100 0 Pellets (3 h) 400 Vehicle 10 mg/kg 20 mg/kg 30 20 10 20 0 30 60 90 120 150 180 Session minutes 1500 Training Timeout responses Pellets 1000 500 0 1 2 3 4 5 6 7 8 9 Training session Nair et al. British Journal of Pharmacology, 2008 Timeout responses/pellet Pellets or lever presses SB 334867 dose (mg/kg, i. p. ) Timeout lever presses per pellet 6 4 2 0 0 10 20 SB 334867 dose (mg/kg i. p. )

SB 334, 867 has no effect on hypocretin 1 -induced reinstatement of food seeking 100 * 75 50 25 0 0 3. 0 6. 0 Hypocretin 1 dose (µg, icv) Lever presses (3 h) Effect of SB 334, 867 100 Lever presses (3 h) Hypocretin 1 -induced reinstatement 75 Vehicle 10 mg/kg 20 mg/kg 50 25 0 0 6. 0 Hypocretin 1 dose (µg, icv)

SB 334, 867 has no effect on pellet-priming-, pellet-cue- or yohimbine-induced reinstatement of food seeking Pellet priming Yohimbine Cue Lever presses (3 h) 100 75 Vehicle 20 mg/kg 50 25 0 No pellet Pellet No cue Cue 0 2. 0 Yohimbine (mg/kg, i. p. )

Conclusions: hypocretin 1 receptor The hypocretin 1 receptor plays a role in the self-administration of highfat pellets The hypocretin 1 receptor likely plays a minimal role in reinstatement induced acute re-exposure to hypocretin 1, food priming, cues and stress

Implications of the findings Our data suggest: 1. A dissociation between the neuronal mechanisms mediating stress-induced relapse to food seeking versus food priming- or cue-induced relapse 2. A potential dissociation between the neuronal mechanisms mediating food selfadministration versus food priming- or cue-induced relapse From Shalev et al. Neurobiology of relapse to heroin and cocaine seeking: a review. Pharmacol Rev, 2002 “Taken together, it appears that multiple and dissociable brain systems are involved in relapse to heroin and cocaine seeking induced by drug priming, conditioned cues and stress. Somewhat surprisingly, it also appears that the neuronal events that mediate heroin- or cocaine-induced reinstatement are to some degree different from those involved in their reinforcing effects. ”

Clinical implications CRF 1 receptor antagonists (currently in clinical development for the treatment of anxiety, depression and drug abuse) should be considered as pharmacological adjuncts in dietary treatments of maladaptive or excessive eating habits PYY 3 -36 (currently in clinical development for the treatment of obesity) may be a more effective treatment for relapse prevention than for decreasing ongoing high-fat food intake Hypocretin 1 receptor antagonists may be a more effective treatment for decreasing ongoing high-fat food intake than for relapse prevention Acknowledgments Dr. Jennifer Bossert Jamie Uejima Kristina Wihbey Dr. Kenner Rice “The best material model for a cat is another, or preferably the same cat” Norbert Wiener, 1945