Rasgrf2 controls noradrenergic involvement in the acute and subchronic effects of alcohol in the brain

Rationale

Alcohol addiction is a major psychiatric disease, and yet, the underlying molecular adaptations in the brain remain unclear. Recent evidence suggests a functional role for the ras-specific guanine-nucleotide releasing factor 2 (Rasgrf2) in alcoholism. Rasgrf2^?/? mice consume less alcohol and show entirely absent dopamine responses to an alcohol challenge compared to wild types (WT).

Objective

In order to further investigate how Rasgrf2 modifies the acute and subchronic effects of alcohol in the brain, we investigated its effects on the noradrenergic and serotonergic systems.

Methods

We measured noradrenaline and serotonin activity in the brain by in vivo microdialysis and RNA expression by chip analysis and RT-PCR after acute and sub-chronic alcohol exposure in Rasgrf2^?/? and WT mice.

Results

In vivo microdialysis showed a significantly reduced noradrenergic response and an absent serotonergic response in the nucleus accumbens (NAcc) and caudate putamen (CPu) after an alcohol challenge in Rasgrf2^?/? mice. A co-expression analysis showed that there is a high correlation between Rasgrf2 and α2 adrenoceptor RNA expression in the ventral striatum in naïve animals. Accordingly, we further assessed the role of Rasgrf2 in the response of the noradrenergic system to subchronic alcohol exposure. A decrease in β1 adrenoceptor gene expression was seen in Rasgrf2^+/+, but not Rasgrf2^?/? mice following alcohol exposure. Conversely, alcohol resulted in a decrease in both β2 and α2 adrenoceptor gene expression in knockout but not WT Rasgrf2 mice.

Conclusions

These findings suggest that adaptations in the noradrenergic system contribute to the Rasgrf2 enhanced risk of alcoholism.     

CRF1 receptor-deficiency induces anxiety-like vulnerability to cocaine

Rationale

The intake of psychostimulant drugs may induce cognitive dysfunction and negative affective-like states, and is associated with increased activity of stress-responsive systems. The corticotropin-releasing factor (CRF) system mediates neuroendocrine, behavioural and autonomic responses to stressors, and might be implicated in substance-related disorders. CRF signalling is mediated by two receptor types, named CRF₁ and CRF₂.

Objectives

The present study aims to elucidate the role for the CRF₁ receptor in cognitive dysfunction and anxiety-like states induced by cocaine.

Results

The genetic inactivation of the CRF₁ receptor (CRF₁+/? and CRF₁?/?) does not influence recognition memory in drug-naïve mice, as assessed by the novel object recognition (NOR) test. Moreover, the chronic administration of escalating doses of cocaine (5–20 mg/kg, i.p.) induces NOR deficits, which are unaffected by CRF₁ receptor-deficiency. However, the same drug regimen reveals an anxiety-like vulnerability to cocaine in CRF₁?/? but not in wild-type or CRF₁+/? mice, as assessed by the elevated plus maze test.

Conclusions

The present findings indicate dissociation of cognitive dysfunction and anxiety-like states induced by cocaine. Moreover, they unravel a novel mechanism of vulnerability to psychostimulant drugs.     

Evaluation of the emotional phenotype and serotonergic neurotransmission of fatty acid amide hydrolase-deficient mice

Rationale

By enhancing brain anandamide tone, inhibitors of fatty acid amide hydrolase (FAAH) induce anxiolytic-like effects in rodents and enhance brain serotonergic transmission. Mice lacking the faah gene (FAAH^?/?) show higher anandamide levels. However, their emotional phenotype is still debated and their brain serotonergic tone remained unexplored.

Objectives and methods

In this study, we tested FAAH^?/? mice in the social interaction and the open field tests performed under different lighting conditions (dim and bright) since variations of the experimental context were proposed to explain opposite findings. Moreover, by microdialysis performed under dim light, we analyzed their serotonergic transmission in frontal cortex (FC) and ventral hippocampus (vHIPP).

Results

In both light conditions, FAAH^?/? mice showed reduced emotionality, compared to wt controls, as suggested by the increased rearing and reduced thigmotaxis displayed in the open field and by the longer time spent in social interaction. Basal serotonergic tone was higher in the FC of mutant mice as compared to control mice, while no difference was observed in the vHIPP. K⁺-induced depolarization produced similar increases of serotonin in both areas of both genotypes. An acute treatment with the CB1 antagonist rimonabant completely abolished the emotional phenotype of FAAH^?/? mice and prevented the K⁺-stimulated release of serotonin in their FC and vHIPP, without producing any effect in wt mice.

Conclusions

Our results support the role of FAAH in the regulation of emotional reactivity and suggest that anandamide-mediated hyperactivation of CB1 is responsible for the emotional phenotype of FAAH^?/? mice and for their enhanced serotonergic tone.     

Cues predicting drug or food reward restore morphine-induced place conditioning in mice lacking delta opioid receptors

Rationale

The exact role of delta opioid receptors in drug-induced conditioned place preference (CPP) remains debated. Under classical experimental conditions, morphine-induced CPP is decreased in mice lacking delta opioid receptors (Oprd1 ^?/?). Morphine self-administration, however, is maintained, suggesting that drug-context association rather than drug reward is deficient in these animals.

Objectives

This study further examined the role of delta opioid receptors in mediating drug-cue associations, which are necessary for the expression of morphine-induced CPP.

Methods

We first identified experimental conditions under which Oprd1 ^?/? mice are able to express CPP to morphine (5, 10 or 20 mg/kg) in a drug-free state and observed that, in this paradigm, CPP was dependent on circadian time conditions. We then took advantage of this particularity to assess the ability of various cues (internal or discrete), predicting either drug or food reward, to restore CPP induced by morphine (10 mg/kg) in Oprd1 ^?/? mice in conditions under which they normally fail to express CPP.

Results

We found that presentation of circadian, drug or auditory cues, predicting morphine or food reward, restored morphine CPP in Oprd1 ^?/? mice, which then performed as well as control mice.

Conclusions

This study reveals that, in contrast to spatial cues, internal or discrete morphine-predicting stimuli permit full expression of morphine CPP in Oprd1 ^?/? mice. Delta receptors, therefore, appear to play a crucial role in modulating spatial contextual cue-related responses. This activity may be critical when context gains control over behavior, as is the case for context-induced relapse in drug abuse.     

Gene Knockout and Metabolome Analysis of Carnitine/Organic Cation Transporter OCTN1

Purpose

Solute carrier OCTN1 (SLC22A4) is an orphan transporter, the physiologically important substrate of which is still unidentified. The aim of the present study was to examine physiological roles of OCTN1.

Methods

We first constructed octn1 gene knockout (octn1 ^?/? ) mice. Metabolome analysis was then performed to identify substrates in vivo. The possible association of the substrate identified with diseased conditions was further examined.

Results

The metabolome analysis of blood and several organs indicated complete deficiency of a naturally occurring potent antioxidant ergothioneine in octn1 ^?/? mice among 112 metabolites examined. Pharmacokinetic analyses after oral administration revealed the highest distribution to small intestines and extensive renal reabsorption of [³H]ergothioneine, both of which were much reduced in octn1 ^?/? mice. The octn1 ^?/? mice exhibited greater susceptibility to intestinal inflammation under the ischemia and reperfusion model. The blood ergothioneine concentration was also much reduced in Japanese patients with Crohn’s disease, compared with healthy volunteers and patients with another inflammatory bowel disease, ulcerative colitis.

Conclusions

These results indicate that OCTN1 plays a pivotal role for maintenance of systemic and intestinal exposure of ergothioneine, which could be important for protective effects against intestinal tissue injuries, providing a possible diagnostic tool to distinguish the inflammatory bowel diseases.     

Increased amphetamine-induced locomotor activity, sensitization, and accumbal dopamine release in M5 muscarinic receptor knockout mice

Introduction

Muscarinic M₅ receptors are the only muscarinic receptor subtype expressed by dopamine-containing neurons of the ventral tegmental area. These cells play an important role for the reinforcing properties of psychostimulants and M₅ receptors modulate their activity. Previous studies showed that M₅ receptor knockout (M ₅ ^?/? ) mice are less sensitive to the reinforcing properties of addictive drugs.

Materials and methods

Here, we investigate the role of M₅ receptors in the effects of amphetamine and cocaine on locomotor activity, locomotor sensitization, and dopamine release using M ₅ ^?/? mice backcrossed to the C57BL/6NTac strain.

Statistical analyses

Sensitization of the locomotor response is considered a model for chronic adaptations to repeated substance exposure, which might be related to drug craving and relapse. The effects of amphetamine on locomotor activity and locomotor sensitization were enhanced in M ₅ ^?/? mice, while the effects of cocaine were similar in M ₅ ^?/? and wild-type mice.

Results

Consistent with the behavioral results, amphetamine-, but not cocaine, -elicited dopamine release in nucleus accumbens was enhanced in M ₅ ^?/? mice.

Discussion

The different effects of amphetamine and cocaine in M ₅ ^?/? mice may be due to the divergent pharmacological profile of the two drugs, where amphetamine, but not cocaine, is able to release intracellular stores of dopamine. In conclusion, we show here for the first time that amphetamine-induced hyperactivity and dopamine release as well as amphetamine sensitization are enhanced in mice lacking the M₅ receptor. These results support the concept that the M₅ receptor modulates effects of addictive drugs.     

Metformin attenuates renal fibrosis in both AMPKα2‐dependent and independent manners

Metformin is a well‐known AMP ‐activated protein kinase (AMPK ) activator, and it has been shown to inhibit organ fibrosis. Whether AMPK α2 mediates metformin protection against renal fibrosis remains unknown. Here, we aimed to investigate the role of the AMPK α2 isoform in mediating the inhibitory effect of metformin on renal fibrosis. Unilateral ureteral obstruction (UUO ) was used to induce renal fibrosis in wild‐type (WT ) and AMPK α2 knockout (AMPK α2^?/?) mice. Metformin treatment was initiated 3 days before UUO and was continued until 7 days after UUO . In WT mice, metformin significantly inhibited UUO ‐induced renal fibrosis. In AMPK α2^?/? mice, metformin also tended to inhibit UUO ‐induced renal fibrosis. Specifically, metformin significantly reduced UUO ‐induced transforming growth factor β1 (TGF β1) mRNA and protein expression in WT mice but not in AMPK α2^?/? mice. In contrast, metformin reduced UUO ‐induced TGF β1 downstream Smad3 phosphorylation in both WT and AMPK α2^?/? mice, suggesting that this regulation occurs in an AMPK α2‐independent manner. In conclusion, the underlying mechanisms for the protective effects of metformin against renal fibrosis include AMPK α2‐dependent targeting of TGF β1 production and AMPK α2‐independent targeting of TGF β1 downstream signalling. In this regard, metformin has an advantage over other AMPK activators for the treatment of renal fibrosis.     

Basolateral Efflux Mediated by Multidrug Resistance-Associated Protein 3 (Mrp3/Abcc3) Facilitates Intestinal Absorption of Folates in Mouse

Purpose

This study investigated the role of an ABC transporter, Mrp3/Abcc3 in intestinal folate absorption.

Methods

Plasma concentrations of folic acid and leucovorin, given orally, were determined in wild-type and Mrp3 ^?/? mice. Mucosal-to-serosal transport was determined in the everted intestinal sacs. The plasma concentrations of endogenous 5-methyltetrahydrofolic acid, homocysteine and vitamin B₁₂, and mRNA levels of hepatic and intestinal folate metabolizing enzymes were compared between wild-type and Mrp3 ^?/? mice.

Results

C _max and area-under plasma concentration–time curve of folic acid were 3.0- and 2.3-fold lower in Mrp3 ^?/? mice compared with wild-type mice, whereas the total body clearance was unchanged. Absorption of leucovorin was significantly delayed in Mrp3 ^?/? mice. Mucosal-to-serosal transport of folic acid and leucovorin was significantly decreased in the duodenum of Mrp3 ^?/? mice, where their PS _serosal was decreased to 6.3 and 22% of that in wild-type mice, respectively. PS _serosal of 5-methyltetrahydrofolic acid was moderately decreased in Mrp3 ^?/? mice. There was no obvious abnormality in folate homeostasis in Mrp3 ^?/? mice.

Conclusions

Mrp3 accounts for the serosal efflux of folic acid and leucovorin, while it makes a moderate contribution to the serosal efflux of 5-methyltetrahydrofolic acid in mice. Mrp3 dysfunction does not disrupt folate homeostasis in mouse.     

Absence of quasi-morphine withdrawal syndrome in adenosine A2A receptor knockout mice

Rationale

Caffeine and other methylxanthines induce behavioral activation and anxiety responses in mice via antagonist action at A_2A adenosine receptors. When combined with the opioid antagonist naloxone, methylxanthines produce a characteristic quasi-morphine withdrawal syndrome (QMWS) in opiate-naive animals.

Objectives

The aim of this study was to establish the role of A_2A receptors in the quasi-morphine withdrawal syndrome induced by co-administration of caffeine and naloxone and in the behavioral effects of caffeine.

Methods

We have used A_2A receptor knockout (A_2AR^?/?) mice in comparison with their wild-type and heterozygous littermates to measure locomotor activity in the open field and withdrawal symptoms induced by caffeine and naloxone. Naïve wild-type and knockout mice were also examined for enkephalin and dynorphin mRNA expression by in situ hybridization and for μ-opiate receptor by ligand binding autoradiography to check for possible opiate receptor changes induced by A_2A receptor inactivation.

Results

Caffeine increases locomotion and anxiety in wild-type animals, but it has no psychomotor effects in A_2AR^?/? mice. Co-administration of caffeine (20 mg/kg) and naloxone (2 mg/kg) resulted in a severe quasi-morphine withdrawal syndrome in wild-type mice that was almost completely abolished in A_2AR^?/? mice. Heterozygous animals exhibited a 40% reduction in withdrawal symptoms, suggesting that there is no genetic/developmental compensation for the inactivation of one of the A_2AR alleles. A_2AR^?/? and wild-type mice have similar levels of striatal μ-opioid receptors, thus the effect is not due to altered opioid receptor expression.

Conclusions

Our results demonstrate that A_2A receptors are required for the induction of quasi-morphine withdrawal syndrome by co-administration of caffeine and naloxone and implicate striatal A_2A receptors and μ-opiate receptors in tonic inhibition of motor activity in the striatum.     

Altered Brain Uptake of Therapeutics in a Triple Transgenic Mouse Model of Alzheimer’s Disease

Purpose

The purpose of this study was to systematically assess the impact of Alzheimer’s disease (AD)-associated blood–brain barrier (BBB) alterations on the uptake of therapeutics into the brain.

Methods

The brain uptake of probe compounds was measured in 18–20 month old wild type (WT) and triple transgenic (3×TG) AD mice using an in situ transcardiac perfusion technique. These results were mechanistically correlated with immunohistochemical and molecular studies.

Results

The brain uptake of the paracellular marker, [¹⁴C] sucrose, did not differ between WT and 3×TG mice. The brain uptake of passively diffusing markers, [³H] diazepam and [³H] propranolol, decreased 54–60% in 3×TG mice, consistent with a 33.5% increase in the thickness of the cerebrovascular basement membrane in 3×TG mice. Despite a 42.4% reduction in P-gp expression in isolated brain microvessels from a sub-population of 3×TG mice (relative to WT mice), the brain uptake of P-gp substrates ([³H] digoxin, [³H] loperamide and [³H] verapamil) was not different between genotypes, likely due to a compensatory thickening in the cerebrovascular basement membrane counteracting any reduced efflux of these lipophilic substrates.

Conclusion

These studies systematically assessed the impact of AD on BBB drug transport in a relevant animal model, and have demonstrated a reduction in the brain uptake of passively-absorbed molecules in this mouse model of AD.     

Memantine-induced brain activation as a model for the rapid screening of potential novel antipsychotic compounds: exemplified by activity of an mGlu2/3 receptor agonist

Rationale

Schizophrenia is a severe, disabling chronic disorder affecting approximately 1% of the population. Improvements and development of more robust and hopefully predictive screening assays for this disease should enhance the identification and development of novel treatments. The present study describes a rapid and robust method for the testing of potential novel antipsychotics by utilising a simplified [¹⁴C]2-deoxyglucose (2-DG) autoradiography method following memantine-induced brain activation.

Methods

Male C57BL/6JCRL mice were given vehicle, ketamine or memantine (10, 20 and 30?mg/kg, subcutaneously (s.c.)) and sacrificed 45?min post-[^14?C]2-DG administration. In subsequent reversal studies, the memantine challenge was further validated with haloperidol (0.32?mg/kg, s.c.) and clozapine (2.5 and 10?mg/kg, s.c.) in parallel with the ketamine model (Duncan et al. 1998a). Lastly, the effects of an mGlu2/3 receptor agonist, LY404039 (10?mg/kg, s.c.), on both ketamine and memantine-induced brain activation was determined.

Results

Both N-methyl-d-aspartate (NMDA) antagonists dose-dependently induced significant region-specific increases in 2-DG uptake. Interestingly, memantine elicited a considerably greater brain activation signature with a larger dynamic window than ketamine. The ??atypical?? antipsychotic clozapine significantly reversed memantine-induced 2-DG uptake whilst the ??typical?? antipsychotic haloperidol was inactive. Pre-treatment with LY404039 fully reversed both the ketamine- and memantine-induced increase in 2-DG uptake without effects on basal 2-DG uptake.

Conclusion

This novel pre-clinical imaging methodology displays potential for the screening of compounds targeting the NMDA receptor hypofunction hypothesis of schizophrenia and should assist in developing compounds from the bench to clinic.     

Superstitious conditioning as a model of delusion formation following chronic but not acute ketamine in humans

Background

Ketamine has previously been shown to induce delusion-like or referential beliefs, both acutely in healthy volunteers and naturalistically among nonintoxicated users of the drug. Delusions are theoretically underpinned by increased superstitious conditioning or the erroneous reinforcement of random events.

Materials and methods

Using a novel and objectively measured superstitious conditioning task, experiment 1 assessed healthy volunteers before and during placebo (n?=?16), low-dose (n?=?15), and high-dose ketamine (n?=?16) under randomized and double-blind conditions. Experiment 2 used the same task to compare ketamine users (n?=?18), polydrug controls (n?=?19), and nondrug-using controls (n?=?17).

Results

In experiment 1, ketamine produced dose-dependent psychotomimetic effects but did not cause changes in superstitious conditioning. Experiment 2 found increased levels of superstitious conditioning among ketamine users compared to polydrug and nondrug-using controls, respectively, as evidenced by both objective task responses and subjective beliefs following the task.

Conclusions

Results indicate that chronic but not acute exposure to ketamine may increase the propensity to adopt superstitious conditioning. These findings are discussed in terms of acute and chronic ketamine models of delusion-like belief formation in schizophrenia.     

Effects of an epilepsy-causing mutation in the SCN1A sodium channel gene on cocaine-induced seizure susceptibility in mice

Rationale

High doses of cocaine can elicit seizures in humans and in laboratory animals. Several mechanisms have been proposed for the induction of seizures by cocaine, including enhanced monoaminergic signaling, blockade of ion channels, and alterations in GABA and glutamate transmission. Mutations in the SCN1A gene, which encodes the central nervous system (CNS) voltage-gated sodium channel (VGSC) Na_v1.1, are responsible for several human epilepsy disorders including Dravet syndrome and genetic (generalized) epilepsy with febrile seizures plus (GEFS+). Mice heterozygous for the R1648H GEFS+ mutation (RH mice) exhibit reduced interneuron excitability, spontaneous seizures, and lower thresholds to flurothyl- and hyperthermia-induced seizures. However, it is unknown whether impaired CNS VGSC function or a genetic predisposition to epilepsy increases susceptibility to cocaine-induced seizures.

Objectives

Our primary goal was to determine whether Scn1a dysfunction caused by the RH mutation alters sensitivity to cocaine-induced behavioral and electrographic (EEG) seizures. We also tested novelty- and cocaine-induced locomotor activity and assessed the expression of Na_v1.1 in midbrain dopaminergic neurons.

Results

We found that RH mice had a profound increase in cocaine-induced behavioral seizure susceptibility compared to wild-type (WT) controls, which was confirmed with cortical EEG recordings. By contrast, although the RH mice were hyperactive in novel environments, cocaine-induced locomotor activity was comparable between the mutants and WT littermates. Finally, immunofluorescence experiments revealed a lack of Na_v1.1 immunoreactivity in dopaminergic neurons.

Conclusion

These data indicate that a disease-causing CNS VGSC mutation confers susceptibility to the proconvulsant, but not motoric, effects of cocaine.     

Developmental alterations in anxiety and cognitive behavior in serotonin transporter mutant mice

Rationale

A promoter variant of the serotonin transporter (SERT) gene is known to affect emotional and cognitive regulation. In particular, the “short” allelic variant is implicated in the etiology of multiple neuropsychiatric disorders. Heterozygous (SERT^+/?) and homozygous (SERT^?/?) SERT mutant mice are valuable tools for understanding the mechanisms of altered SERT levels. Although these genetic effects are well investigated in adulthood, the developmental trajectory of altered SERT levels for behavior has not been investigated.

Objectives

We assessed anxiety-like and cognitive behaviors in SERT mutant mice in early adolescence and adulthood to examine the developmental consequences of reduced SERT levels. Spine density of pyramidal neurons was also measured in corticolimbic brain regions.

Results

Adult SERT^?/? mice exhibited increased anxiety-like behavior, but these differences were not observed in early adolescent SERT^?/? mice. Conversely, SERT^+/? and SERT^?/? mice did display higher spontaneous alternation during early adolescence and adulthood. SERT^+/? and SERT^?/? also exhibited greater neuronal spine densities in the orbitofrontal but not the medial prefrontal cortices. Adult SERT^?/? mice also showed an increased spine density in the basolateral amygdala.

Conclusions

Developmental alterations of the serotonergic system caused by genetic inactivation of SERT can have different influences on anxiety-like and cognitive behaviors through early adolescence into adulthood, which may be associated with changes of spine density in the prefrontal cortex and amygdala. The altered maturation of serotonergic systems may lead to specific age-related vulnerabilities to psychopathologies that develop during adolescence.     

Treatment with a carbon monoxide-releasing molecule inhibits chronic inflammatory pain in mice: nitric oxide contribution

Rationale

Carbon monoxide synthetized by inducible heme oxygenase (HO-1) exerts potent anti-inflammatory and antinociceptive effects during acute and neuropathic pain, but its role in the modulation of chronic inflammatory pain and the possible involvement of nitric oxide in this action remain unknown.

Objectives and methods

The antiallodynic and antihyperalgesic effects of a carbon monoxide releasing molecule, tricarbonyldichlororuthenium(II) dimer (CORM-2), daily administered from days 4 to 14 after complete Freund's adjuvant (CFA) injection in wild-type (WT), neuronal (NOS1-KO), and inducible (NOS2-KO) nitric oxide synthases knockout mice, were evaluated using von Frey filaments and plantar tests. Effects of CORM-2 treatment on the expression of HO-1, NOS1, and NOS2 at 14 days after inflammation induction were assessed by Western blot.

Results

Main inflammatory pain symptoms induced by CFA in WT, NOS1-KO, and NOS2-KO mice were significantly reduced in a time-dependent manner by CORM-2 treatment. In all genotypes, inflammation increased the dorsal root ganglia and paw expression of HO-1, but CORM-2 treatment only over-expressed this enzyme in the paw of all genotypes. The increased NOS1 expression induced by inflammation in WT mice was abolished by CORM-2 treatment, while there was no effect of the inflammation in neither CORM-2 treatment in the expression of NOS2 in WT and NOS1-KO mice.

Conclusions

CORM-2 treatment inhibits inflammatory pain through enhancing HO-1 paw expression in all genotypes and reducing NOS1 over-expression in WT mice. An interaction between HO-1/carbon monoxide and NOS1/nitric oxide systems was also demonstrated. CORM-2 treatment may represent a new approach for management chronic inflammatory pain.     

Netrin-1 receptor-deficient mice show age-specific impairment in drug-induced locomotor hyperactivity but still self-administer methamphetamine

Rationale

The mesocorticolimbic dopamine system undergoes significant reorganization of neuronal connectivity and functional refinement during adolescence. Deleted in colorectal cancer (DCC), a receptor for the guidance cue netrin-1, is involved in this reorganization. Previous studies have shown that adult mice with a heterozygous (het) loss-of-function mutation in DCC exhibit impairments in sensitization and conditioned place preference (CPP) to psychostimulants. However, the commonly abused psychostimulant methamphetamine (METH) has not been assessed, and the role of DCC in drug self-administration remains to be established.

Objectives

Using dcc het mice and wildtype (WT) littermates, we extended previous findings on dcc haplodeficiency by examining self-administration of METH in adult mice, including cue-induced drug seeking following abstinence. We also examined hyperactivity, sensitization, and CPP to a METH-paired context in adult and adolescent mice.

Results

While adult dcc het mice expressed largely similar METH self-administration and cue-induced drug seeking as WT littermates, they failed to modulate responding according to dose of METH. Compared to WT, both adult and adolescent dcc het mice expressed impaired locomotor hyperactivity to acute METH but nevertheless showed comparable behavioral sensitization. Conditioned hyperactivity increased with age in WT but not in dcc het mice.

Conclusions

Impaired METH-induced hyperactivity and dose-related responding in adult dcc het mice suggest that reduced DCC alters METH-related behaviors. Adolescence is identified as a vulnerable period during which impairment in hyperactivity due to reduced DCC can be overcome with repeated METH injections. Nevertheless, DCC appears to have a somewhat limited role in METH-consumption and seeking following abstinence.     

Cholinergic modulation of auditory processing, sensory gating and novelty detection in human participants

Rationale

Suppression of redundant auditory information and facilitation of deviant, novel, or salient sounds can be assessed with paired-click and oddball tasks, respectively. Electrophysiological correlates of perturbed auditory processing found in these paradigms are likely to be a trait marker or candidate endophenotype for schizophrenia.

Objective

This is the first study to investigate the effects of the muscarinic M1 antagonist biperiden and the cholinesterase inhibitor rivastigmine on auditory-evoked potentials (AEPs), sensory gating, and mismatch negativity (MMN) in young, healthy volunteers.

Results

Biperiden increased P50 amplitude and prolonged N100 and P200 latency in the paired-click task but did not affect sensory gating. Rivastigmine was able to reverse the effects of biperiden on N100 and P200 latency. Biperiden increased P50 latency in the novelty oddball task, which was reversed by concurrent administration of rivastigmine. Rivastigmine shortened N100 latency and enhanced P3a amplitude in the novelty oddball paradigm, both of which were reversed by biperiden.

Conclusion

The muscarinic M1 receptor appears to be involved in preattentive processing of auditory information in the paired-click task. Additional effects of biperiden versus rivastigmine were reversed by a combination treatment, which renders attribution of these findings to muscarinic M1 versus muscarinic M2–M5 or nicotinic receptors much more difficult. It remains to be seen whether the effects of cholinergic drugs on AEPs are specifically related to the abnormalities found in schizophrenia. Alternatively, aberrant auditory processing could also be indicative of a general disturbance in neural functioning shared by several neuropsychiatric disorders and/or neurodegenerative changes seen in aging.     

Tissue Distribution and Gender-Specific Protein Expression of Cytochrome P450 in five Mouse Genotypes with a Background of FVB

Purpose

To systematically investigate tissue distribution and gender-specific protein expression of Cytochrome P450 (Cyps) in five mouse genotypes with a background of Friend virus B (FVB).

Methods

The Cyps were extracted from the tissue S9 fractions of the main metabolic organs and then absolutely quantified by applying the UHPLC-MS/MS method.

Results

The liver had the highest expression of Cyps, followed by the small intestine and kidney. In the liver, Cyp1a2, Cyp2c29, Cyp2c39, Cyp2d22, Cyp2e1, and Cyp3a11 were the main isoforms. Cyp1a2 and Cyp2c29 were male-specific, while Cyp2c39 was female-specific. Compared with the expression in Wild-type (WT) FVB mice, the expression of Cyp1a2, Cyp1b1, Cyp2d22, and Cyp3a25 significantly decreased in female efflux transporter (ET) knockout mice. In the small intestine, Cyp2c29 and Cyp3a11 were the major isoforms. Knockout of ET didn’t alter the expression levels of most Cyps. However, female ET knockout mice presented higher Cyp2c29 expression than WT FVB mice. The Cyp7a1 expression was markedly decreased in ET knockout mice except Bcrp1^?/? mice. In the kidney, Cyp2e1 was the main isoform and exhibited male specificity. Knockout of ET slightly affected the protein expression of Cyps in the brain, heart, lung, spleen and stomach.

Conclusions

A comprehensive understanding of the distribution characteristics and gender-specific expression of Cyps in major metabolic organs of WT and ET knockout FVB mice should contribute to a better understanding of drug efficacy and toxicity, and drug-drug interactions.

     

Cocaine self-administration behaviors in ClockΔ19 mice

Rationale

A key role has been identified for the circadian locomotor output cycles kaput (Clock) gene in the regulation of drug reward. Mice bearing a dominant negative mutation in the Clock gene (ClockΔ19 mice) exhibit increased cocaine-induced conditioned place preference, reduced anxiety- and depression-like behavior, increased sensitivity to intracranial self-stimulation, and increased dopaminergic cell activity in the ventral tegmental area.

Objectives

We sought to determine if this hyperhedonic phenotype extends to cocaine self-administration and measures of motivation.

Methods

Two separate serial testing procedures were carried out (n?=?7–10/genotype/schedule). Testing began with acquisition of sucrose pellet self-administration, implantation of intravenous catheter, acquisition of cocaine self-administration, and dose–response testing (fixed ratio or progressive ratio). To evaluate diurnal variations in acquisition behavior, these sessions occurred at Zeitgeber 2 (ZT2) or ZT14.

Results

WT and ClockΔ19 mice exhibited similar learning and readily acquired food self-administration at both ZT2 and ZT14. However, only ClockΔ19 mice acquired cocaine self-administration at ZT2. A greater percentage of ClockΔ19 mice reached acquisition criteria at ZT2 and ZT14. ClockΔ19 mice self-administered more cocaine than WT mice. Using fixed ratio and progressive ratio schedules of reinforcement dose–response paradigms, we found that cocaine is a more efficacious reinforcer in ClockΔ19 mice than in WT mice.

Conclusion

Our results demonstrate that the Clock gene plays an important role in cocaine reinforcement and that decreased CLOCK function increases vulnerability for cocaine use.     

Nicotine exposure during adolescence: cognitive performance and brain gene expression in adult heterozygous reeler mice

Rationale

We have recently reported nicotine-induced stimulation of reelin and glutamic acid decarboxylase 67 (GAD67) mRNA expression levels in the brain of heterozygous reeler mice (HRM), a putative animal model for the study of symptoms relevant to major behavioral disorders.

Objectives

We aimed to evaluate long-term behavioral effects and brain molecular changes as a result of adaptations to nicotine exposure in the developing HRM males.

Methods

Adolescent mice (pnd 37–42) were exposed to oral nicotine (10 mg/l) in a 6-day free-choice drinking schedule. As expected, no differences in total nicotine intake between WT (wild-type) mice and HRM were found.

Results

Long-term behavioral effects and brain molecular changes, as a consequence of nicotine exposure during adolescence, were only evidenced in HRM. Indeed, HRM perseverative exploratory behavior and poor cognitive performance were modulated to WT levels by subchronic exposure to nicotine during development. Furthermore, the expected reduction in the expression of mRNA of reelin and GAD67 in behaviorally relevant brain areas of HRM appeared persistently restored by nicotine. For brain-derived neurotrophic factor (BDNF) mRNA expression, no genotype-dependent changes appeared. However, expression levels were increased by previous nicotine in brains from both genotypes. The mRNA encoding for nicotine receptor subunits (α7, β2 and α4) did not differ between genotypes and as a result of previous nicotine exposure.

Conclusion

These findings support the hypothesis of pre-existing vulnerability (based on haploinsufficiency of reelin) to brain and behavioral disorders and regulative short- and long-term effects associated with nicotine modulation.