Faculty Bibliography

RATIONALE: Food restriction (FR) enhances learned and unlearned behavioral responses to drugs of abuse and increases D-1 dopamine (DA) receptor-mediated activation of extracellular signal-regulated kinases (ERK) 1/2 MAP kinase in nucleus accumbens (NAc). While a role has been established for ERK signaling in drug-mediated associative learning, it is not clear whether ERK regulates unconditioned behavioral effects of abused drugs. OBJECTIVES: The purpose of this study was to determine whether blockade of ERK signaling, using the brain-penetrant MEK inhibitor, SL-327, decreases behavioral or NAc cellular responses to acute drug treatment and their augmentation by FR. MATERIALS AND METHODS: Separate experiments assessed the effects of SL-327 (50 mg/kg, intraperitoneally) on (1) the reward-potentiating effect of D-amphetamine in an intracranial self-stimulation protocol, (2) the locomotor-activating effect of the D-1 agonist, SKF-82958, and (3) Fos-immunostaining induced in the NAc by SKF-82958. RESULTS: FR rats displayed enhanced responses to drug treatment on all measures. SL-327 had no effect on sensitivity to rewarding brain stimulation or the reward-potentiating effect of D-amphetamine. The MEK inhibitor, U0126, microinjected into the NAc was also without effect. The locomotor-activating effect of SKF-82958 was unaffected by SL-327. In contrast, SL-327 decreased NAc Fos-immunostaining and abolished the difference between feeding groups. CONCLUSIONS: These results support the conclusion that ERK signaling does not mediate unlearned behavioral responses to drug treatment. However, the upregulation of ERK and downstream transcriptional responses to acute drug treatment may underlie the reported enhancement of reward-related learning in FR subjects

Chronic food restriction (FR) increases behavioral sensitivity to drugs of abuse in animal models and is associated with binge eating, which shares comorbidity with drug abuse, in clinical populations. Behavioral, biochemical and molecular studies conducted in this laboratory to elucidate the functional and mechanistic bases of these phenomena are briefly reviewed. Results obtained to date indicate that FR increases the reward magnitude and locomotor-activating effects of abused drugs, and direct dopamine (DA) receptor agonists, as a result of neuroadaptations rather than changes in drug disposition. Changes in striatal DA dynamics, and postsynaptic cell signaling and gene expression in response to D-1 DA receptor stimulation have been observed. Of particular interest is an upregulation of NMDA receptor-dependent MAP kinase and CaM Kinase II signaling, CREB phosphorylation, and immediate-early and neuropeptide gene expression in nucleus accumbens (NAc) which may facilitate reward-related learning, but also play a role in the genesis of maladaptive goal-directed behaviors. Covariation of altered drug reward sensitivity with body weight loss and recovery suggests a triggering role for one of the endocrine adiposity hormones. However, neither acute nor chronic central infusions of leptin or the melanocortin 3/4 receptor agonist, MTII, have attenuated d-amphetamine reward or locomotor activation in FR rats. Interestingly, chronic intracerebroventricular leptin infusion in ad libitum fed (AL) rats produced a sustained decrease in food intake and body weight that was accompanied by a reversible potentiation of rewarding and locomotor-activating effects of d-amphetamine. This raises the interesting possibility that rapid progressive weight loss is sufficient to increase behavioral sensitivity to drugs of abuse. Whether weight loss produced by leptin infusion produces the same neuroadaptations as experimenter-imposed FR, and whether any of the observed neuroadaptations are necessary for expression of increased behavioral responsiveness to acute drug challenge remain to be investigated

Adenosine A(2A) receptors are preferentially expressed in rat striatum, where they are concentrated in dendritic spines of striatopallidal medium spiny neurons and exist in a heteromeric complex with D(2) dopamine (DA) receptors. Behavioral and biochemical studies indicate an antagonistic relationship between A(2A) and D(2) receptors. Previous studies have demonstrated that food-restricted (FR) rats display behavioral and striatal cellular hypersensitivity to D(1) and D(2) DA receptor stimulation. These alterations may underlie adaptive, as well as maladaptive, behaviors characteristic of the FR rat. The present study examined whether FR rats are hypersensitive to the A(2A) receptor agonist, CGS-21680. In Experiment 1, spontaneous horizontal motor activity did not differ between FR and ad libitum fed (AL) rats, while vertical activity was greater in the former. Intracerebroventricular (i.c.v.) administration of CGS-21680 (0.25 and 1.0 nmol) decreased both types of motor activity in FR rats, and returned vertical activity levels to those observed in AL rats. In Experiment 2, FR rats given access to a running wheel for a brief period outside of the home cage rapidly acquired wheel running while AL rats did not. Pretreatment with CGS-21680 (1.0 nmol) blocked the acquisition of wheel running. When administered to FR subjects that had previously acquired wheel running, CGS-21680 suppressed the behavior. In Experiment 3, CGS-21680 (1.0 nmol) activated both ERK 1/2 and CREB in caudate-putamen with no difference between feeding groups. However, in nucleus accumbens (NAc), CGS-21680 failed to activate ERK 1/2 and selectively activated CREB in FR rats. These results indicate that FR subjects are hypersensitive to several effects of an adenosine A(2A) agonist, and suggest the involvement of an upregulated A(2A) receptor-linked signaling pathway in NAc. Medications targeting the A(2A) receptor may have utility in the treatment of maladaptive behaviors associated with FR, including substance abuse and compulsive exercise.

The present experiments were undertaken to clarify the role of central alpha(1)-adrenoceptors in reward processes. Rats, trained to self-stimulate via electrodes in the medial forebrain bundle of the lateral hypothalamus, were administered alpha(1)-selective drugs near the locus coeruleus (LC), a site of a dense concentration of alpha(1)-receptors. Effects on reward potency were assessed from shifts in rate-frequency curves while effects on motor response capacity were judged from changes in the maximal rates of responding. It was found that local blockade of LC alpha(1)-receptors with terazosin produced a significant dose-dependent and site-dependent rightward shift of 0.08 log units and a significant decrease of 16.3% in the maximum response rate. Both effects were completely reversed by coadministration of the alpha(1)-agonist, phenylephrine and were not attributable to terazosin's weak action at alpha(2)-adrenoceptors. It is concluded that LC alpha(1)-adrenoceptors are involved both in reward/motivational processes and operant response elaboration which are postulated to work together to facilitate goal attainment.Neuropsychopharmacology advance online publication, 5 July 2006; doi:10.1038/sj.npp.1301145

Chronic food restriction (FR) enhances the rewarding and motor-activating effects of abused drugs, and is accompanied by changes in dopamine (DA) dynamics and increased D-1 DA receptor-mediated cell signaling and transcriptional responses in nucleus accumbens (NAc). However, little is known about effects of FR on DA synthetic activity in the mesoaccumbens and nigrostriatal pathways. In Experiment 1 of the present study, tyrosine hydroxylase (TH) gene expression was measured in ventral tegmental area and substantia nigra, using real-time RT-PCR and in situ hybridization; no differences were observed between FR and ad libitum fed (AL) rats. In Experiment 2, TH protein levels, determined by Western blot, were found to be elevated in NAc and caudate-putamen (CPu) of FR relative to AL rats. In the absence of increased transcription, this may reflect a slowing of TH degradation. In Experiments 3 and 4, DA synthetic activity was assessed by Western blot measurement of TH phosphorylation at Ser40, and HPLC measurement of in vivo tyrosine hydroxylation rate, as reflected by DOPA accumulation following administration of a decarboxylase inhibitor (NSD-1015; 100 mg/kg, i.p.). Basal phospho-(Ser40)-TH levels did not differ between groups but DOPA accumulation was decreased by FR. Decreased DOPA synthesis, despite increased levels of TH protein, may reflect the inhibitory effect of increased DA binding to TH protein or decreased concentrations of cofactor tetrahydrobiopterin. Finally, in response to D-amphetamine (0.5 and 5.0 mg/kg, i.p.), phospho-(Ser40)-TH was selectively decreased in NAc of FR rats. This suggests increased feedback inhibition of DA synthesis-a possible consequence of postsynaptic receptor hypersensitivity, or increased extracellular DA concentration. These results indicate that FR increases TH protein levels, but may decrease the capacity for DA synthesis by decreasing TH activity. According to this scheme, the previously observed upregulation of striatal cell signaling and transcriptional responses to DA receptor agonist administration may include compensatory neuroadaptations

It was previously reported that chronic food restriction and maintenance of rats at 75-80% of initial body weight enhanced the reward-potentiating effect of D-amphetamine in the lateral hypothalamic self-stimulation (LHSS) paradigm. Moreover, the enhancement reversed in parallel with body weight recovery when ad libitum access to food was reinstated. The present study tested the hypothesis that hypoleptinemia during food restriction is necessary for expression of enhanced drug reward. In Experiment 1, intracerebroventricular (i.c.v.) infusion of leptin (0.5 microg/0.5 microl/hr for 8 days) in food-restricted rats did not alter the rewarding effect of D-amphetamine (0.5 mg/kg, i.p.). Considering that i.c.v. leptin may not diffuse into deep brain regions where direct effects on drug reward sensitivity may be exerted, effects of acute bilateral microinjection of leptin (0.5 microg) in ventral tegmental area and nucleus accumbens were tested in Experiment 2 and found to have no effect. In Experiment 3, chronic i.c.v. leptin infusion in ad libitum fed rats decreased food intake and body weight and enhanced the rewarding effect of D-amphetamine. Sensitivity to D-amphetamine returned to normal as body weight recovered following cessation of leptin infusion. This result suggests that weight loss, whether from hormone-induced appetite suppression or experimenter-imposed food restriction, is sufficient to enhance drug reward sensitivity. Experiment 4 tested whether food restriction in the absence of body weight loss alters drug reward sensitivity. Rats received chronic i.c.v. infusion of the orexigenic melanocortin receptor antagonist, SHU9119 (0.02 microg/0.5 microl/hr for 12 days), and a subset were pair-fed to vehicle-infused controls. Although these subjects ingested approximately 50% of the amount of food ingested by free-feeding SHU9119-infused rats, they displayed no weight loss and no change in sensitivity to D-amphetamine. Together, results of this study support the importance of weight loss, but not leptin, in the enhancement of drug reward sensitivity

The present communication reports on DA uptake in rat striatum in a model of chronic food restriction. The K(m) for DA uptake was unaltered, but the V(max) was reduced by 32%, not supporting the idea that the enhanced behavioral sensitivity to cocaine or d-amphetamine upon chronic food restriction is due to a greater density of DAT at the plasma membrane for drug interaction. Chronic food restriction did not alter the potency of cocaine or D-amphetamine in inhibiting DA uptake in the striatum, suggesting that the enhanced behavioral sensitivity to these drugs upon chronic food restriction is not due to their enhanced affinity for DAT. These results point to factors other than DAT density or affinity underlying the sensitized response to psychostimulants in food restriction

Chronic food restriction increases exploratory behavior, cognitive function, and the rewarding effects of abused drugs. Recently, striatal neuroadaptations that may be involved in these effects were observed. Specifically, D-1 dopamine (DA) receptor agonist challenge produced stronger activation of extracellular signal-regulated kinase (ERK), calcium-calmodulin-dependent kinase II (CaMKII), and the nuclear transcription factor cAMP response element binding protein (CREB) in nucleus accumbens (NAc) of food-restricted (FR) relative to ad libitum fed (AL) rats. Further, when FR rats were injected intracerebroventricularly (i.c.v.) with vehicle (saline) they displayed stronger activation of c-Jun N-terminal protein kinase (JNK), ERK and CaMKII than did AL rats. It is not known to what extent the latter effects represent the basal state of FR rats or an amplified response to the brief handling involved in the i.c.v. injection procedure. Using Western blotting it was found that basal phospho-JNK is higher in caudate-putamen (CPu) and NAc of FR relative to AL rats. Interestingly, brief handling decreased phospho-JNK levels in FR subjects. Basal phospho-ERK1/2 also tended to be elevated in CPu and NAc of FR rats but the elevation was not significant. However, phospho-MEK--the activated kinase upstream of ERK1/2--was significantly elevated in NAc of FR rats. Neither ERK1/2 nor MEK were activated by brief handling. CaMKII was selectively activated by handling in NAc of FR rats, suggesting a state-dependent response to a salient event. Given the established involvement of mitogen-activated protein kinase (MAPK) and CaMKII in synaptic plasticity, learning and memory, the increase in basal phospho-MEK and hyperresponsiveness of CaMKII in NAc may represent adaptive cellular responses to persistent negative energy balance that facilitate associative learning in connection with food-seeking

Behavioral studies have demonstrated that chronic food restriction augments the rewarding and motor-activating effects of centrally injected psychostimulants and direct dopamine (DA) receptor agonists. Recently, it has been shown that intracerebroventricular (i.c.v.) injection of the D-1 DA receptor agonist, SKF-82958, produces an enhanced locomotor-activating effect as well as increased activation of striatal ERK 1/2 MAP kinase, CaM kinase II, CREB, and c-fos in food-restricted (FR) relative to ad libitum fed (AL) rats. Striatal neurons that express the D-1 DA receptor coexpress dynorphin and substance P, and CREB is known to couple D-1 DA receptor stimulation to preprodynorphin (ppD) gene expression. The purpose of the present study was to examine possible genomic consequences of FR using real-time quantitative RT-PCR to measure striatal neuropeptide gene expression 3 h after i.c.v. injection of SKF-82958 (20 microg). Results indicate that, in nucleus accumbens (NAc), basal levels of ppD and preprotachykinin (ppT) mRNA are lower in FR than AL rats. This may reflect a decrease in tonic DA transmission during FR which precedes the compensatory upregulation of postsynaptic D-1 DA receptor-mediated cell signaling. In response to SKF-82958 challenge, however, FR subjects displayed greater levels of ppD and ppT mRNA in NAc than did AL subjects. A similar trend was seen in caudate-putamen (CPu). SKF-82958 also increased preproenkephalin (ppE) mRNA in Nac, but not CPu, with no difference between feeding groups. The present findings regarding ppD and ppT are consistent with prior findings of increased behavioral and cellular responses to acute D-1 DA agonist challenge in FR rats. The functional consequences of increased neuropeptide gene expression in response to acute drug challenge remain to be investigated but may include modulation of behavioral effects that emerge with repeated drug exposure, including sensitization, tolerance, and addiction

The brain melanocortin system mediates downstream effects of hypothalamic leptin and insulin signaling. Yet, there have been few studies of chronic intracerebroventricular (i.c.v.) melanocortin receptor (MCR) agonist or antagonist infusion. Although there is evidence of interaction between melanocortin and dopamine (DA) systems, effects of chronic MCR ligand infusion on behavioral sensitivity to non-ingestive reward stimuli have not been investigated. The objective of this study was to investigate effects of chronic i.c.v. infusion of the MCR agonist, MTII, and the MCR antagonist, SHU9119, on food intake, body weight, and sensitivity to rewarding lateral hypothalamic electrical stimulation (LHSS) and the reward-potentiating (i.e., threshold-lowering) effect of D-amphetamine. The MCR antagonist, SHU9119 (0.02 microg/h) produced sustained hyperphagia and weight gain during the 12-day infusion period, followed by compensatory hypophagia and an arrest of body weight gain during the 24-day post-infusion period. At no point during the experiment was sensitivity to LHSS or D-amphetamine (0.25mg/kg, i.p.) altered. The MCR agonist, MTII (0.02 microg/h) produced a brief hypophagia (3 days) followed by a return to control levels of daily intake, but with body weight remaining at a reduced level throughout the 12-day infusion period. This was followed by compensatory hyperphagia and weight gain during the 24-day post-infusion period. There was no change in sensitivity to non-ingestive reward stimuli during the infusion of MTII. However, sensitivity to D-amphetamine was increased during the 24-day post-infusion period. It therefore seems that changes in ingestive behavior that occur during chronic MCR ligand infusion may not affect the response to non-ingestive reward stimuli. However, it is possible that the drive to re-feed and restore body weight following MCR agonist treatment includes neuroadaptations that enhance the incentive effects of drug stimuli