Altered neurotransmission in brains of autoimmune mice: pharmacological and neurochemical evidence

Depressive-like behavior is the most profound manifestation of autoimmunity-associated behavioral syndrome in lupus-prone MRL-lpr mice. This led to the hypothesis that chronic autoimmunity and inflammation alter the activity of central serotonergic and dopaminergic systems. Three drugs with a selective mode of action were used to probe the functional status of these two systems in vivo. The behavioral effects of single and repeated intraperitoneal (i.p.) injections of sertraline, quinpirole (QNP) and risperidone were measured in the forced swim and brief sucrose preference tests. In comparison to MRL +/+ controls, autoimmune MRL-lpr mice did not show a reduction in sucrose intake after the administration of sertraline. Acute injection of quinpirole increased floating more in the MRL-lpr than in the control group, while intermittent administration induced self-injurious behavior in both groups. Acute injection of risperidone significantly increased floating in MRL-lpr mice, while repeated administration abolished the difference between the substrains in sucrose intake. These discrepancies in responsiveness implied that the central neurotransmitter activity is dissimilar in the two MRL substrains. This notion was confirmed in a cohort of untreated MRL-lpr and MRL +/+ mice by comparing their neurotransmitter/metabolite levels in several brain regions. In particular, MRL-lpr brains showed increased dopamine (DA) levels in the paraventricular nucleus (PVN) and median eminence (ME), decreased concentrations of serotonin in the PVN and enhanced levels in the hippocampus, as well as decreased norepinephrine (NE) levels in the prefrontal cortex. Behavioral deficits correlated with the changes in PVN and median eminence. These results are consistent with the hypothesis that imbalanced neurotransmitter regulation of the hypothalamus-pituitary axis plays an important role in the etiology of behavioral dysfunction induced by systemic autoimmune disease.     

Taste responsiveness and diet preference in autoimmune MRL mice

One of the most profound behavioural deficits in lupus-prone MRL-lpr mice is blunted responsiveness to sweet solutions. Given the systemic nature of autoimmune/inflammatory disease, it was not clear whether impaired taste sensitivity or motivated response to palatable food underlie this deficit. The present study compares response rates of MRL-lpr mice (which develop disease early), congenic MRL +/+ mice (which develop disease later in life) and non-autoimmune Swiss Webster (SW) mice to different tastes and diets. Healthy SW mice showed the highest responsiveness to palatable stimulation throughout the study. Conversely, the preference for palatable solutions progressively declined in MRL-lpr mice as the disease developed. No differences between the two MRL substrains were seen in responsiveness to quinine or saline, suggesting that blunted responsiveness to palatable solutions cannot be accounted for by reduced taste sensory function (hypogeusia). In addition, changes in response rates to palatable solutions were associated with systemic upregulation of pro-inflammatory cytokines. With a new cohort of mice fed on carbohydrate-rich and fat-rich diets, we also examined whether reduced sucrose intake in MRL-lpr mice can be accounted for by a reduced craving for carbohydrates. Contrary to this expectation, diseased MRL-lpr mice preferred carbohydrate-rich food while consuming a food mass comparable to controls. These results further support the hypothesis that the onset of lupus-like disease alters motivated behaviour, independent of changes in neurologic function and food metabolism.     

Impaired response to amphetamine and neuronal degeneration in the nucleus accumbens of autoimmune MRL-lpr mice

Spontaneous development of lupus-like disease in MRL-lpr mice is accompanied by a constellation of behavioral deficits, including blunted responsiveness to sucrose. Although autoimmunity-induced damage of limbic areas is proposed to underlie this deficit, the systemic nature of the disease precludes inference of a causal relationship between CNS damage and functional loss. Based on the stimulatory effects of d-amphetamine sulfate (AMPH) on sucrose intake, the present study pharmacologically probes the functional status of central dopaminergic circuits involved in control of behavioral reward. The response rates were compared between diseased MRL-lpr mice and congenic MRL +/+ controls tested in the sucrose preference paradigm. Neuronal loss was assessed by Fluoro Jade B (FJB) staining of nucleus accumbens and the CA2/CA3 region. While control mice significantly increased intake of sucrose solutions 60 min after administration of AMPH (i.p., 0.5 mg/kg), the intake in drugged MRL-lpr mice was comparable to those given saline injection. Increased FJB staining was detected in the nucleus accumbens and hippocampus of diseased mice, and AMPH treatment neither altered this nor other measures of organ pathology. The results obtained are consistent with previously observed changes in the mesolimbic dopamine system of MRL-lpr mice and suggest that the lesion in the nucleus accumbens and deficits in dopamine release underlie impaired responsiveness to palatable stimulation during the progress of systemic autoimmune disease. As such, they point to a neurotransmitter-specific regional brain damage which may account for depressive behaviors in neuropsychiatric lupus erythematosus.     

Neurotoxic properties of cerebrospinal fluid from behaviorally impaired autoimmune mice

The chronic, lupus-like autoimmune disease in MRL-lpr mice is associated with leucocyte infiltration into the choroid plexus, brain cell death, and deficits in motivated behavior. The presence of lymphoid cells in the ventricular lumen and the increased number of TUNEL-positive cells in periventricular areas led to the hypothesis that immune cells enter into the cerebrospinal fluid (CSF) and induce primary neuronal damage in regions bordering the cerebral ventricles. Using an in vitro approach, we presently examine the possibility that CSF from autoimmune mice is neurotoxic and/or gliotoxic. The CSF and serum from diseased MRL-lpr mice, less symptomatic MRL +/+ controls, and healthy Swiss/Webster mice (non-autoimmune controls) were frozen until their effects on the viability of pyramidal neurons and astrocytes were assessed in a two-color fluorescence assay. Significant reduction in neuronal viability (in some cases as low as 67%) was observed in the co-cultures of hippocampal neurons and astrocytes incubated for 24 h with CSF from autoimmune MRL-lpr mice. The viability of astrocytes did not differ among the groups, and the CSF from autoimmune mice appeared more toxic than the serum. The behavior of MRL-lpr mice differed significantly from the control groups, as indicated by impaired exploration, reduced intake of palatable food, and excessive immobility in the forced swim test. The present results suggest that CSF from the behaviorally impaired lupus-prone mice is neurotoxic and are consistent with the hypothesis that neuroactive metabolites are produced intrathecally in neuropsychiatric lupus erythematosus.     

Behavioral effects of infection with IL-6 adenovector

The onset of autoimmunity in lupus-prone mice is accompanied by a constellation of behavioral deficits, termed Autoimmunity-Associated Behavioral Syndrome (AABS). In particular, a spontaneous increase in serum interleukin-6 (IL-6) levels in five-week old MRL-lpr mice coincides temporally with blunted responsiveness to sucrose and excessive immobility in the forced swim test. These relationships, along with evidence that sucrose intake drops after systemic IL-6 overexpression is induced in healthy mice, have led to the hypothesis that sustained elevation in serum IL-6 also induces other aspects of AABS. This hypothesis is tested by comparing the behavioral profiles of healthy mice infected with Ad5mIL6 adenovirus (2 x 10(8) pfu of virus/mouse i.p.) with those of animals infected with control Ad5 virus. This methodology was used to achieve high circulating levels of IL-6, to overcome the problem of its short half-life, and to avoid the stressful effects of repeated injections. The Ad5mIL6 infection (known to induce excessive IL-6 levels over five days) transiently reduced food, water, and sucrose intake, as well as rectal temperature in MRL +/+ and AKR/J mice. Although the level of locomotor activity did not decline, Ad5mIL6-infected AKR/J mice demonstrated less novel object exploration. Performance in the step-down, plus-maze, and spontaneous alternation tests were disturbed to various degrees in all infected animals. The present results suggest that prolonged exposure to circulating IL-6 primarily impairs ingestive behavior, likely reflecting enhanced catabolism. The inability of circulating IL-6 to alter other aspects of behavior supports the hypothesis that multiple immuno-neuroendocrine mechanisms contribute to the pathogenesis of AABS.     

A behavioral profile of autoimmune lupus-prone MRL mice.

Manifestations of the human autoimmune disease systemic lupus erythematosus (SLE) include a number of behavioral and cognitive deficits. The present study asks whether neurobehavioral dysfunction is present also in MRL mice that spontaneously develop most of the fundamental immunological aberrations of SLE. There are two congenic substrains of MRL mice that differ in the time of disease onset: MRL-lpr mice develop lupus early and MRL(-)+/+ develop the typical signs of disease relatively late in life. The behavior of these substrains was assessed at 7 to 11 weeks of age, a time that coincides with the onset of disease in MRL-lpr mice and the absence of known lupus symptoms in the MRL(-)+/+ group. When compared to the congenic MRL(-)+/+ control substrain, MRL-lpr mice were spontaneously less active, traversed a crossbeam slower, and ceased responding to the novelty of a new environment sooner. They were also more reluctant to leave their home base or travel far away from it and perseverated in their response bias during extinction and reversal learning. Immunological status was characterized by moderate proteinuria in both substrains and high titers of antinuclear antibodies in MRL-lpr but not MRL(-)+/+ mice. Histological analysis revealed minimal or no signs of joint pathology in MRL-lpr mice. Thus, this study shows the presence of behavioral dysfunction in mice with early stages of autoimmune disease and gives support for the idea that MRL mice may provide a useful model of neurobehavioral dysfunction in SLE. It is suggested that the behavioral profile of MRL-lpr mice may indicate increased "timidity," related to genetics, autoimmunity, or both.     

Immunosuppression prevents neuronal atrophy in lupus-prone mice: evidence for brain damage induced by autoimmune disease?

An early onset of systemic, lupus-like disease in MRL-lpr mice is accompanied by deterioration in their behavioral performance and atrophy of pyramidal neurons in the parietal cortex and the hippocampal CA1 area. Using the immunosuppressive drug cyclophosphamide (CY) to attenuate the disease, we have tested the hypothesis that the autoimmune/inflammatory process is responsible for changes in brain morphology. A modified Golgi impregnation method revealed that, in comparison to saline-treated controls, immunosuppressive treatment with CY (100 mg/kg/week i.p. over 8 weeks) increased dendritic branching and spine numerical density in the CA1 region of MRL-lpr mice and MRL +/+ mice, which develop less severe manifestations of the disease. More interestingly, CY selectively prevented the atrophy and aberrant morphology of pyramidal neurons in the parietal cortex of MRL-lpr mice. The neuropathological measures (in particular reduced dendritic spine density) significantly correlated with increased serum levels of antinuclear antibodies and splenomegaly. The present results support the hypothesis that chronic autoimmune disease induces functionally important changes in neuronal morphology, and provide an empirical basis for understanding the behavioral dysfunction in systemic lupus erythematosus and autoimmune phenomena reported in some forms of mental illness.     

Blunted Sensitivity to Sucrose in Autoimmune MRL-lpr Mice: A Curve-Shift Study

Lupus-prone MRL-lpr mice show an autoimmunity-associated behavioral syndrome that has many features similar to the effects of chronic stress. The present study evaluated whether autoimmune MRL-lpr mice show reduced responsiveness to sucrose, as observed in normal animals exposed to chronic mild stress. Sixteen-week old MRL-lpr mice and their age-matched congenic MRL +/+ controls were given 0%, 0.5%, 1%, 2%, 4%, 8%, or 16% sucrose solution to drink every 48 h in a one-bottle test. The MRL-lpr mice drank less than controls at all concentrations, except at 16%. The amount of sucrose consumed vs. solution concentration followed a saturation curve. Estimates were obtained for the concentration yielding the half-maximum response (X₅₀) and the response at saturating concentration of sucrose (R_max). The X₅₀ was significantly higher in MRL-lpr than in MRL +/+ mice, indicating a shift to the right of the concentration-intake curve. The R_max did not differ significantly between substrains, suggesting that the autoimmune process did not affect performance capacity. Pretreatment with the immunosuppressant cyclophosphamide diminished the substrain difference in X₅₀, suggesting that reduced sensitivity to sucrose is related to autoimmune/inflammatory factors. These results support the similarity between autoimmunity-associated behavioral syndrome and behavioral changes produced by chronic stress, and suggest common neuroendocrine mechanisms. Because reduced sensitivity to palatable stimulus may reflect blunted hedonic responsiveness (“anhedonia”), it is hypothesized that an autoimmune/inflammatory factor(s) produces the depression found in human lupus, and some cases of affective disorder. Copyright © 1996 Elsevier Science Inc.     

Purine receptor antagonist modulates serology and affective behaviors in lupus-prone mice: evidence of autoimmune-induced pain?

Neurologic and psychiatric (NP) manifestations are severe complications of systemic lupus erythematosus (SLE). As commonly seen in patients, spontaneous disease onset in the MRL/MpJ-Fas^lpr/J (MRL-lpr) mouse model of NP–SLE is accompanied by increased autoantibodies, pro-inflammatory cytokines and behavioral dysfunction which precede neuroinflammation and structural brain lesions. The role of purinergic receptors in the regulation of immunity and behavior remains largely unexplored in the field of neuropsychiatry. To examine the possibility that purinoception is involved in the development of affective behaviors, the P2X purinoceptor antagonist, suramin, was administered to lupus-prone mice from 5 to 14 weeks of age. In addition to food and water measures, novel object and sucrose preference tests were performed to assess neophobic anxiety- and anhedonic-like behaviors. Enzyme-linked immunosorbant assays for anti-nuclear antibodies (ANA) and pro-inflammatory cytokines were employed in immunopathological analyses. Changes in dendritic morphology in the hippocampal CA1 region were examined by a Golgi impregnation method. Suramin significantly lowered serum ANA and prevented behavioral deficits, but did not prevent neuronal atrophy in MRL-lpr animals. In a new batch of asymptomatic mice, systemic administration of corticosterone was found to induce aberrations in CA1 dendrites, comparable to the “stress” of chronic disease. The precise mechanism(s) through which purine receptor inhibition exerted beneficial effects is not known. The present data supports the hypothesis that activation of the peripheral immune system induces nociceptive-related behavioral symptomatology which is attenuated by the analgesic effects of suramin. Hypercortisolemia may also initiate neuronal damage, and metabolic perturbations may underlie neuro-immuno-endocrine imbalances in MRL-lpr mice.     

Social isolation stress exacerbates autoimmune disease in MRL/lpr mice

The potential physiological mechanisms explaining an influence of psychosocial stress on autoimmune diseases remain undetermined. Exposure of chronic social isolation stress to MRL/lpr mice significantly enhanced the degree of proteinuria after 20 weeks of age and reduced the survival rate. The serum anti-dsDNA IgG2a levels were increased significantly by stress at 19 weeks of age, which was simultaneously accompanied by inhibition of the serum corticosterone elevation. Furthermore, stress caused increased IFN-gamma production from anti-CD3-stimulated splenic mononuclear cells, whereas IL-4 and IL-10 production decreased. These results indicated that isolation stress exacerbated autoimmune disease in MRL/lpr mice, the possible mechanism for which might be related to stress-induced dysregulation of Th1/Th2 balance and inhibition of the blood corticosterone response to inflammatory stimuli.     

Increased TUNEL staining in brains of autoimmune Fas-deficient mice

Profound changes in brain morphology and behavior coincide with the spontaneous development of systemic autoimmune/inflammatory disease in Fas-deficient MRL-lpr mice. The dendrites atrophy, the density of hippocampal and cortical neurons decreases, and an anxious/depressive-like behavior emerges while lymphoid cells infiltrate into the choroid plexus of MRL-lpr mice. We hypothesized that the inherited lack of the Fas-dependent anti-inflammatory mechanism would lead to unsuppressed immune activity, characterized by reduced apoptosis in the MRL-lpr brain. Using the terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeled (TUNEL) method as an indicator of apoptosis, a surprisingly high incidence of TUNEL-positive cells was observed in the hippocampus, choroid plexus and periventricular regions of MRL-lpr mice, 5-10-fold higher than that found in the MRL +/+ control brain. Immunostaining with anti-CD3, CD4 and CD8 monoclonal antibodies showed limited overlap between CD-positive and TUNEL-positive cells, suggesting that the dying cells are for the most part (approximately 70%) not T-lymphocytes. Although further characterization of the phenotype of the dying cells and the mechanism of cell death are required, the present results suggest the involvement of a Fas-independent apoptotic process in neurodegeneration induced by systemic autoimmune disease.     

Food aversion: A critical balance between allergen-specific IgE levels and taste preference

Animals sensitized to allergens change their feeding behavior and avoid drinking the otherwise preferred sweetened solutions containing the allergens. This phenomenon, known as food aversion, appears to be mediated by allergen-specific IgE antibodies. Here we investigated food aversion in BALB/c and C57BL/6 mice, which differ in their allergic responses to the allergen ovalbumin as well as in their preference for sweet taste. BALB/c mice present higher levels of IgE and a natural lower preference for sweet flavors when compared to C57BL/6 mice. Specifically, we studied a conflicting situation in which animals simultaneously experienced the aversive contact with the allergen and the attractive sweet taste of increasing concentrations of sucrose. We found that BALB/c mice were more prone to develop food aversion than C57BL/6 mice and that this aversive behavior could be abolished in both strains by increasing the palatability of the solution containing the allergen. In both strains food aversion was positively correlated with the levels of allergen-specific IgE antibodies and inversely correlated with their preference for sucrose sweetened solutions.     

Neuroimmunopathology in a murine model of neuropsychiatric lupus

Animal models are extremely useful tools in defining pathogenesis and treatment of human disease. For many years researchers believed that structural damage to the brain of neuropsychiatric (NP) patients lead to abnormal mental function, but this possibility was not extensively explored until recently. Imaging studies of NP-systemic lupus erythematosus (SLE) support the notion that brain cell death accounts for the emergence of neurologic and psychiatric symptoms, and evidence suggests that it is an autoimmunity-induced brain disorder characterized by profound metabolic alterations and progressive neuronal loss. While there are a number of murine models of SLE, this article reviews recent literature on the immunological connections to neurodegeneration and behavioral dysfunction in the Fas-deficient MRL model of NP-SLE. Probable links between spontaneous peripheral immune activation, the subsequent central autoimmune/inflammatory responses in MRL/MpJ-Tnfrsf6(lpr) (MRL-lpr) mice and the sequential mode of events leading to Fas-independent neurodegenerative autoimmune-induced encephalitis will be reviewed. The role of hormones, alternative mechanisms of cell death, the impact of central dopaminergic degeneration on behavior, and germinal layer lesions on developmental/regenerative capacity of MRL-lpr brains will also be explored. This model can provide direction for future therapeutic interventions in patients with this complex neuroimmunological syndrome.     

Taste aversion learning in autoimmune Mrl-lpr/lpr and Mrl +/+ mice

Conditioned taste aversion to a neutral stimulus paired with an immunosuppressive drug (cyclophosphamide) was assessed in lupus-prone MRL-lpr/lpr and congenic control (MRL +/+) mice. The presence of lymphoproliferation in MRL-lpr/lpr mice was associated with poorer taste aversion learning and varied as a function of the dose of cyclophosphamide. There were no differences in learning performance between MRL-lpr/lpr and MRL +/+ mice when the animals were tested at an age prior to the development of lymphadenopathy, or when lithium chloride or electric shock were used as unconditioned stimuli. These results are consistent with the hypothesis that the immune status of an organism has an impact on behavior and the possibility that behavior can serve an in vivo immunoregulatory function.     

Elevated immunoglobulin levels in the cerebrospinal fluid from lupus-prone mice

The systemic autoimmune disease lupus erythematosus (SLE) is frequently accompanied by neuropsychiatric manifestations and brain lesions of unknown etiology. The MRL-lpr mice show behavioral dysfunction concurrent with progression of a lupus-like disease, thus providing a valuable model in understanding the pathogenesis of autoimmunity-induced CNS damage. Profound neurodegeneration in the limbic system of MRL-lpr mice is associated with cytotoxicity of their cerebrospinal fluid (CSF) to mature and immature neurons. We have recently shown that IgG-rich CSF fraction largely accounts for this effect. The present study examines IgG levels in serum and CSF, as well as the permeability of the blood-brain barrier in mice that differ in immune status, age, and brain morphology. In comparison to young MRL-lpr mice and age-matched congenic controls, a significant elevation of IgG and albumin levels were detected in the CSF of aged autoimmune MRL-lpr mice. Two-dimensional gel electrophoresis and MALDI-TOF MS confirmed elevation in IgG heavy and Ig light chain isoforms in the CSF. Increased permeability of the blood-brain barrier correlated with neurodegeneration (as revealed by Fluoro Jade B staining) in periventricular areas. Although the source and specificity of neuropathogenic antibodies remain to be determined, these results support the hypothesis that a breached blood-brain barrier and IgG molecules are involved in the etiology of CNS damage during SLE-like disease.     

Autoimmune-induced damage of the midbrain dopaminergic system in lupus-prone mice

Spontaneous development of lupus-like disease is accompanied by impaired dopamine catabolism and degenerating axon terminals in the mesencephalon of MRL-lpr mice. We presently examine the hypothesis that systemic autoimmunity affects the central dopaminergic system in behaviorally impaired animals. The functional damage of the nigrostriatal pathway was assessed from rotational behavior after a single injection of the D1/D2-receptor agonist apomorphine. Neurodegeneration in the midbrain was estimated by Fluoro Jade B (FJB) staining. The causal role of autoimmunity was tested by comparing asymptomatic and diseased MRL-lpr mice, and by employing the immunosuppressive drug cyclophosphamide. Damage of dopaminergic neurons was assessed by tyrosine-hydroxylase (TH) staining of the midbrain. Apomorphine induced significant asymmetry in limb use, which lead to increased circling in the diseased MRL-lpr group. While FJB-positive somas were not seen in the striatum, increased staining in the substantia nigra (SN) and ventral tegmental area (VTA) were detected in behaviorally impaired MRL-lpr mice, but not in age-matched controls. Reduced brain mass and increased levels of TNF-alpha in their cerebrospinal fluid (CSF) suggested cerebral atrophy and inflammation. In addition, CSF was neurotoxic to a dopaminergic progenitor cell line. Immunosuppression attenuated CSF cytotoxicity, TNF-alpha levels, and midbrain neurodegeneration. Supportive of the notion that dying neurons were dopaminergic, the SN of autoimmune mice showed approximately a 35% reduction in the number of TH-positive cells. A three-fold increase in serum brain-reactive antibodies accompanied this loss. Although the source of toxic mediator(s) remains unknown, present results are consistent with the hypothesis that autoimmunity-induced destruction of mesonigral and mesolimbic dopaminergic pathways contributes to the etiology of aberrant behavior in an animal model of neuropsychiatric lupus.     

Decreased Intake of Sucrose Solutions in Orexin Knockout Mice

Orexins are synthesized by lateral hypothalamic neurons and are suggested to be implicated in feeding behavior. Recent studies have shown that intracerebroventricular administration of orexin-A increases intake of sweet-tasting solution. Effects of suppressing the orexin system on consumption of sweet-tasting solution and sensory processing with sweet taste inputs, however, have yet to be examined. We examined the effects of orexin deficiency on sucrose solution intake, locomotor activity, and preference for sweet solution using male orexin knockout (OxKO) and littermate wild-type (WT) mice. In the dark and over 24-h periods, OxKO mice showed significantly less sucrose intake and lower locomotor activity than WT mice without alteration in food intake whereas preferences for 100?mM sucrose were not different between the genotypes. Moreover, sucrose intake of OxKO mice was significantly less than sucrose intake of a subgroup of WT mice with similar locomotor activity compared to that of OxKO mice. These results suggest that factors other than the lower energy expenditure due to lower locomotor activity are likely responsible for the decreased sucrose intake of OxKO mice. Orexin deficiency may lower the satiety threshold resulting in reduced sucrose intake, without altering food intake.     

Decreased motivation to eat in mu-opioid receptor-deficient mice

Altered motivational processes might participate to the physiopathology of eating-related disorders. The endogenous opioid system is thought to mediate the hedonic properties of food intake. To assess the role for the micro-opioid receptor (MOR) pathway in the motivational properties of food intake, in the present study we tested wild-type and MOR-deficient mice (MOR-/-) in a nose-poke operant paradigm for chow or sucrose pellets. To avoid confounding factors linked to food restriction/deprivation experience, mice were always provided with food ad libitum. Although less MOR-/- than wild-type mice initiated operant behaviour, under a fixed ratio-1 (FR-1) reinforcement schedule the two genotypes showed similar patterns of food-driven nose-poking, indicating preserved cognitive abilities in MOR-deficient mice. However, during FR-3 and progressive ratio (PR) reinforcement experiments, MOR-/- mice showed lower levels of nose-poking for either chow or sucrose pellets than wild-type mice, indicating a crucial role for the MOR pathway in the motivational properties of food intake. Moreover, under the PR reinforcement schedule mice nose-poking for sucrose pellets showed higher genotype-independent breakpoint levels than mice working for chow pellets, indicating that the MOR pathway is not essential for hedonic processing of palatable food intake. Finally, MOR-/- mice did not differ from wild-type mice in the rate of operant responding extinction, further supporting the notion of unaltered cognitive abilities in the MOR-deficient mice. The present findings strongly indicate that the MOR pathway mediates the motivational properties of food intake, but it is not essential for hedonic processing of ingestive behaviour.