Glenda C. Harris

A role for lateral hypothalamic orexin neurons in reward seeking

The lateral hypothalamus is a brain region historically implicated in reward and motivation, but the identity of the neurotransmitters involved are unknown. The orexins (or hypocretins) are neuropeptides recently identified as neurotransmitters in lateral hypothalamus neurons. Although knockout and transgenic overexpression studies have implicated orexin neurons in arousal and sleep, these cells also project to reward-associated brain regions, including the nucleus accumbens and ventral tegmental area. This indicates a possible role for these neurons in reward function and motivation, consistent with previous studies implicating these neurons in feeding. Here we show that activation of lateral hypothalamus orexin neurons is strongly linked to preferences for cues associated with drug and food reward. In addition, we show that chemical activation of lateral hypothalamus orexin neurons reinstates an extinguished drug-seeking behaviour. This reinstatement effect was completely blocked by prior administration of an orexin A antagonist. Moreover, administration of the orexin A peptide directly into the ventral tegmental area also reinstated drug-seeking. These data reveal a new role for lateral hypothalamus orexin neurons in reward-seeking, drug relapse and addiction.

Arousal and reward: a dichotomy in orexin function

The orexins (or hypocretins) are neuropeptide transmitters made exclusively in hypothalamic neurons that have extensive CNS projections. Previous studies reported that this system is most strongly associated with feeding, arousal and the maintenance of waking. We review here recent studies that reveal a novel and important role for the orexin/hypocretin neuronal system in reward processing and addiction. We propose that the current evidence indicates a dichotomy in orexin function, such that orexin neurons in the lateral hypothalamus regulate reward processing for both food and abused drugs, whereas those in the perifornical and dorsomedial hypothalamus regulate arousal and response to stress. Evidence also indicates roles for lateral hypothalamus orexin neurons and ventral tegmental orexin receptors in reward-based learning and memory.

Brain substrates for increased drug seeking during protracted withdrawal.

Studies are reviewed indicating that both increased anxiety and altered hedonic processing accompany protracted withdrawal from opiates. Increased anxiety may be most apparent in response to stress, whereas decreased motivation for natural rewards but increased interest in drugs reveals substantial alterations in hedonic values. Our recent work indicates that increased norepinephrine (NE) release in the bed nucleus of the stria terminalis (BNST) may underlie anxiety associated with protracted withdrawal. Altered plasticity in afferents to the ventral tegmental area (VTA; accumbens, amygdala and lateral hypothalamus), or in the VTA itself, may be involved in the altered hedonic processing that occurs during protracted withdrawal. We hypothesize that conditioned release of NE in the BNST in response to stressors (including drug-associated stimuli) may elevate anxiety which then augments the reward value of drugs by a negative reinforcement mechanism. We also propose that plasticity in VTA neurons and their afferents during chronic drug exposure and protracted withdrawal decreases the valence of natural rewards whereas sensitization occurs to the motivational effects of drugs that increases their motivational valence. The combination of anxiety, decreased valence of natural rewards, and sensitized incentive for drugs make a potent formula for relapse and drug seeking during protracted withdrawal.

Lateral hypothalamic orexin neurons are critically involved in learning to associate an environment with morphine reward

Previously, we reported that lateral hypothalamic (LH) orexin neurons are stimulated in proportion to the preference shown for reward-associated cues during conditioned place preference (CPP) testing. Here, we examine for the first time the role of these neurons in the acquisition of morphine CPP. Results show that LH orexin neurons, but not those in the perifornical area (PFA), are stimulated during conditioning when morphine is given in a novel drug-paired environment (CPP compartment) but not when given in the home cage, nor when saline was given in the CPP environment. Furthermore, bilateral excitotoxic lesions of the LH orexin area completely blocked the acquisition of morphine CPP. Lesions that spared LH orexin neurons had no effect. Orexin neurons in the LH project to the ventral tegmental area (VTA), an area important in the acquisition of morphine CPP. Therefore, we investigated the importance of the LH orexin connection to the VTA in the acquisition of a morphine CPP using a disconnection technique involving a unilateral excitotoxic lesion of LH orexin neurons and contralateral blockade of VTA orexin receptors. Results indicated that a unilateral LH orexin lesion together with a microinjection of the orexin A antagonist (SB 334867) into the contralateral VTA prior to each morphine-pairing session was sufficient to block the development of a morphine CPP. Either of these treatments by themselves was not sufficient to block CPP development. These results demonstrate the importance of LH orexin neurons and their projections to the VTA in the formation of associations between environmental cues and drug reward.

Critical Role for Ventral Tegmental Glutamate in Preference for a Cocaine-Conditioned Environment

Cocaine administration induces glutamatergic activation within the mesolimbic-accumbens system. This activation has been linked to the behavioral effects of cocaine and recently to the induction of long-term potentiation in dopamine neurons within the ventral tegmental area (VTA). We sought to determine if glutamate receptor activation is also crucial to the development of a conditioned place preference (CPP) to cocaines rewarding effects. Two groups of rats were given intra-VTA injections of either vehicle or a combination of NMDA (N-methyl-D-aspartate) and AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid) receptor antagonists (AP5 0.24 nmol plus CNQX 0.12 nmol per side) prior to each cocaine place-conditioning trial. Microinjections of the glutamate antagonists completely blocked the development of a cocaine CPP when given within, but not when given outside of, the VTA. These data indicate that glutamatergic activity in the VTA may be crucial for learning to associate environmental stimuli with cocaine exposure.

Glutamate-associated plasticity in the ventral tegmental area is necessary for conditioning environmental stimuli with morphine

We sought to determine if plasticity in the ventral tegmental area (VTA) of the midbrain is involved in learning to associate morphine exposure with a specific environment. For this, we tested whether activation of glutamate receptors and protein kinase A is needed for the acquisition and expression of a morphine-conditioned place preference (CPP). Rats received bilateral microinjections of either the NMDA antagonist AP5 (0.48 nmol/0.3 microl), the AMPA antagonist CNQX (0.21 nmol/0.3 microl), or vehicle into the VTA prior to each of three morphine-conditioning sessions. Both the AMPA and NMDA receptor antagonists blocked the development of morphine CPP when given into the VTA but not when given outside the VTA. In similar studies the protein kinase A (PKA) inhibitor, Rp-cAMPS (13 nmol/0.3 microl), blocked the acquisition of morphine CPP when given into the VTA immediately after morphine conditioning. In separate experiments, glutamate antagonists, or Rp-cAMPS, immediately prior to the preference test blocked the expression of morphine CPP when microinjected into the VTA. These data indicate that the VTA is an important site for synaptic modifications involved in the learning and memory of environmental cues predicting reward, and that glutamate input and PKA activation are crucial to this process.

Enhanced Morphine Preference Following Prolonged Abstinence: Association with Increased Fos Expression in the Extended Amygdala

We previously found that chronically morphine-pretreated, abstinent rats show stronger preferences for morphine-associated environments than placebo-pretreated rats. Here we show that this increased preference persisted for at least 5 weeks after withdrawal of chronic morphine. To determine brain regions involved in this behavior, we examined neural activation (as indexed by Fos-like proteins) induced by a morphine-conditioned place preference test. Placebo-pretreated (P) morphine-conditioned rats showed significantly elevated Fos in the anterior cingulate cortex (Cg), nucleus accumbens core (Ac-C) and shell (Ac-S), ventral lateral and dorsal lateral bed nucleus of the stria terminialis (BNST-VL and -DL), and central and basolateral amygdala nuclei (ACE, ABL) when compared to nonconditioned P rats. Chronically morphine-pretreated (M) rats that exhibited enhanced morphine preference 5 weeks after morphine withdrawal showed significantly greater Fos in all the same areas except the BNST-DL relative to conditioned P or nonconditioned M rats. Place preference measures and Fos expression were positively correlated in the Cg and ABL, for conditioned P animals, and in the Cg, ABL and BNST-VL for conditioned M animals. These results indicate a relationship between place preference behavior and neural indices of activation in the forebrain in response to morphine-conditioned cues that may be chronically modulated by prior morphine exposure.

Activation in extended amygdala corresponds to altered hedonic processing during protracted morphine withdrawal

Previously we reported that during protracted morphine abstinence rats show reduced conditioned place preferences (CPP) for food-associated environments, compared to non-dependent subjects. To determine the brain regions involved in this altered reward behavior, we examined neural activation (as indexed by Fos-like proteins) induced by a preference test for a food-associated environment in 5-week morphine-abstinent versus non-dependent animals. The results indicate that elevated Fos expression in the anterior cingulate cortex (Cg) and basolateral amygdala (BLA) correlated positively with preference behavior in all groups. In contrast, Fos expression in stress-associated brain areas, including the ventral lateral bed nucleus of the stria terminalis (VL-BNST), central nucleus of the amygdala (CE), and noradrenergic (A2) neurons in the nucleus tractus solitarius (NTS) was significantly elevated only in morphine-abstinent animals. Furthermore, the number of Fos positive neurons in these areas was found to correlate negatively with food preference in abstinent animals. These results indicate that the altered hedonic processing during protracted morphine withdrawal leading to decreased preference for cues associated with natural rewards may involve heightened activity in stress-related brain areas of the extended amygdala and their medullary noradrenergic inputs.

Altered Motivation and Learning Following Opiate Withdrawal: Evidence for Prolonged Dysregulation of Reward Processing

Opiate abuse has been associated with cognitive deficits in human addicts. To determine if prior opiate exposure alters the ability to learn, we trained animals in an instrumental learning task for a food reward. During a 2-week period after withdrawal, morphine-abstinent rats were significantly slower at learning an escalating fixed-ratio response for food reward compared to placebo-treated animals. When these same animals were trained in a conditioned suppression paradigm (two tone–shock pairings given in the operant box), the morphine-withdrawn animals showed greater retention by taking significantly longer to resume responding for food reward when the tone was presented. In a third experiment, morphine-abstinent rats withdrawn 2 or 5 weeks were tested for their ability to associate a highly palatable food reward with a specific environment using a place-conditioning paradigm. At 2 weeks postwithdrawal, morphine-abstinent rats did not show any significant place preference for a food they readily consumed, while placebo-treated rats readily learned to prefer the food-paired environment. At 5 weeks postwithdrawal, rats developed significantly less preference for food-associated cues, but more preference for morphine-associated cues, compared to placebo-treated animals. These data suggest that prior morphine exposure may have prolonged effects on the motivation for natural rewards, which in turn may compromise the ability of former addicts to overcome their addictions.

Augmented accumbal serotonin levels decrease the preference for a morphine associated environment during withdrawal.

Recent studies have found that acute morphine administration increases serotonin (5-HT) transmission within the nucleus accumbens and other forebrain regions. In contrast, 5-HT transmission is depressed during withdrawal from chronic morphine. We show that pharmacological agents that increase brain 5-HT levels (fluoxetine or 5-hydoxytryptophan, 5-HTP) abolish the preference of chronically morphine-treated, withdrawn rats for a morphine-associated environment. Similar results were seen when fluoxetine was microinjected into the nucleus accumbens. Conversely, rats given morphine acutely showed an enhanced preference for a morphine-associated environment when pretreated with these agents. Fluoxetine also decreased the heightened anxiety found in morphine withdrawn rats. The results of our study indicate that drugs that augment 5-HT levels may reduce the desire for morphine during withdrawal.