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Brigitte L Kieffer

Publications and source records attributed to Brigitte L Kieffer.

At least 19 recordsLinked to original sources
PubMed2007

Circadian-dependent learning and memory enhancement in nociceptin receptor-deficient mice with a novel KUROBOX apparatus using stress-free positive cue task.

Using the novel apparatus KUROBOX, learning and memory behaviors, as well as various parameters of movement activity, were reevaluated in mice deficient for nociceptin/orphanin FQ receptor (NOP-/- mice) or mu-opioid receptor (MOP-/- mice). This method has the advantages that no handling procedures are required throughout the experiments performed over 3 days, positive cue paradigms are used without water or shock stress, and the method does not disturb the nocturnal habit of mice. NOP-/- mice displayed a significant enhancement of learning and memory under stress-free conditions, but there were no changes in the various physical and psychological parameters of movement activity (nest stay ratio, distance moved, speed and angle in the movement) and biological rhythm that were measured. Enhancement of nocturnal learning was observed during the first 12-h dark cycle, and enhancement of memory was observed at the beginning of the second dark cycle in NOP-/- mice. In contrast, MOP-/- mice showed no significant change in learning and memory behaviors or in physical and psychological parameters of movement activity, except for speed, MOP-/- mice showed a significant decrease in speed of movement. Thus, the KUROBOX apparatus provides a useful alternative method to evaluate learning and memory activity under the more physiological conditions. In addition, this apparatus has an advantage that various physical and psychological parameters of movement activity affecting learning and memory behavior are also evaluated at the same time.

Animals↗
PubMed2007

Knock-in mice reveal nonsense-mediated mRNA decay in the brain.

Nonsense-mediated mRNA decay (NMD) is a process of mRNA surveillance that degrades transcripts harboring a premature termination codon (PTC). Mammalian NMD was mostly studied in cultured cells so far and there was no direct evidence yet that NMD could operate in the brain. We introduced, by homologous recombination in mouse, a PTC in the mu opioid receptor gene (mor). mor transcript was severely downregulated in the brain of these knock-in mice. A systemic cycloheximide treatment significantly increased the level of the mutant mRNA, suggesting NMD involvement. To further corroborate this hypothesis, we generated a second knock-in mouse line where the PTC was placed at 10 instead of 96 nucleotides from the downstream splice junction. As predicted by the "termination codon position rule" established in vitro, mor transcript brain expression was rescued to wild-type level. These knock-in mouse lines will be valuable models to better understand and manipulate NMD in vivo.

Animals↗
PubMed2006

Deletion of mu- and kappa-opioid receptors in mice changes epidermal hypertrophy, density of peripheral nerve endings, and itch behavior.

The mu- (MOR) and kappa- (KOR) opioid receptors have been implicated in the regulation of homeostasis of non-neuronal cells, such as keratinocytes, and sensations like pain and chronic pruritus. Therefore, we have studied the phenotype of skin after deletion of MOR and KOR. In addition, we applied a dry skin model in these knockout mice and compared the different mice before and after induction of the dermatitis in terms of epidermal thickness, epidermal peripheral nerve ending distribution, dermal inflammatory infiltrate (mast cells, CD4 positive lymphocytes), and scratching behavior. MOR knockout mice reveal as phenotype a significantly thinner epidermis and a higher density of epidermal fiber staining by protein gene product 9.5 than the wild-type counterparts. Epidermal hypertrophy, induced by the dry skin dermatitis, was significantly less developed in MOR knockout than in wild-type mice. Neither mast cells nor CD4 T(h)-lymphocytes are involved in the changes of epidermal nerve endings and epidermal homeostasis. Finally, behavior experiments revealed that MOR and KOR knockout mice scratch less after induction of dry skin dermatitis than wild-type mice. These results indicate that MOR and KOR are important in skin homeostasis, epidermal nerve fiber regulation, and pathophysiology of itching.

Animals↗
PubMed2006

Presynaptic delta opioid receptors regulate ethanol actions in central amygdala.

Endogenous opioid systems are implicated in the reinforcing effects of ethanol consumption. For example, delta opioid receptor (DOR) knockout (KO) mice show greater ethanol consumption than wild-type (WT) mice (Roberts et al., 2001). To explore the neurobiological correlates underlying these behaviors, we examined effects of acute ethanol application in brain slices from DOR KO mice using whole-cell patch recording techniques. We examined the central nucleus of amygdala (CeA) because the CeA is implicated in alcohol reinforcement (Koob et al., 1998). We found that the acute ethanol effects on GABA(A) receptor-mediated inhibitory postsynaptic currents (IPSCs) were greater in DOR KO mice than in WT mice. Ethanol increased the frequency of miniature IPSCs (mIPSCs) significantly more in DOR KO mice than in WT mice. In CeA of WT mice, application of ICI 174864 [[allyl]2-Tyr-alpha-amino-isobutyric acid (Aib)-Aib-Phe-Leu-OH], a DOR inverse agonist, augmented ethanol actions on mIPSC frequency comparable with ethanol effects seen in DOR KO mice. Superfusion of the selective DOR agonist D-Pen(2),D-Pen(5)-enkephalin decreased the mean frequency of mIPSCs; this effect was reversed by the DOR antagonist naltrindole. These findings suggest that endogenous opioids may reduce ethanol actions on IPSCs of CeA neurons in WT mice through DOR-mediated inhibition of GABA release and that the increased ethanol effect on IPSCs in CeA of DOR KO mice could be, at least in part, due to absence of DOR-mediated inhibition of GABA release. This result supports the hypothesis that endogenous opioid peptides modulate the ethanol-induced augmentation of GABA(A) receptor-dependent circuitry in CeA (Roberto et al., 2003).

Amygdala↗
PubMed2006

Effects of constitutive deletion of opioid receptors on the basal densities of Fas and Fas-associated protein with death domain (FADD) in the mouse brain: a delta-opioid tone inhibits FADD.

The acute effects of opiate drugs and opiate addiction have been associated with modulation of Fas/FADD (Fas-Associated protein with Death Domain) signaling complex in the rat brain. This study investigated the possible existence of endogenous opioid tones regulating the basal activities of Fas receptor forms and FADD in the brain, using gene-targeted mice lacking mu-, delta- or kappa-opioid peptide receptors (KO mice). In mu-KO mice, but not in delta- or kappa-KO mice, the basal immunodensity of native Fas (35 kDa monomeric form) was decreased in the cerebral cortex (33%) when compared with WT littermates. In delta-KO mice, but not in mu- or kappa-KO mice, the basal content of 120 kDa Fas aggregates (complexes of monomers relevant in Fas signaling) was markedly increased in the cortex (93%). In contrast, no differences between genotypes were observed in the basal expression of glycosylated Fas (51/48/45 kDa forms). Notably, the basal content of FADD (the adaptor protein that couples Fas to caspases and transmits the death signal) was increased in the cerebral cortex of delta-KO mice (48%), but not in mu- or kappa-KO mice. In addition, the basal content of phosphorylated FADD at Ser191 (the relevant species of FADD implicated in nonapoptotic signals) was also upregulated in the cortices of delta-opioid receptor KO mice (6.5-11.0-fold). The results suggest that mu-receptors tonically stimulate (through endogenous opioid peptides) the activation of native Fas, whereas delta-receptors tonically inhibit the expression of Fas aggregates and that of FADD and phosphorylated FADD (Ser191) in the mouse brain. These data are in line with the acute opposite modulation of Fas and FADD induced by mu- and delta-opiate agonists, and strongly support the notion of an anti-apoptotic delta-opioid tone that restrains Fas signaling.

Animals↗
PubMed2006

Knockin mice expressing fluorescent delta-opioid receptors uncover G protein-coupled receptor dynamics in vivo.

The combination of fluorescent genetically encoded proteins with mouse engineering provides a fascinating means to study dynamic biological processes in mammals. At present, green fluorescent protein (GFP) mice were mainly developed to study gene expression patterns or cell morphology and migration. Here we used enhanced GFP (EGFP) to achieve functional imaging of a G protein-coupled receptor (GPCR) in vivo. We created mice where the delta-opioid receptor (DOR) is replaced by an active DOR-EGFP fusion. Confocal imaging revealed detailed receptor neuroanatomy throughout the nervous system of knock-in mice. Real-time imaging in primary neurons allowed dynamic visualization of drug-induced receptor trafficking. In DOR-EGFP animals, drug treatment triggered receptor endocytosis that correlated with the behavioral response. Mice with internalized receptors were insensitive to subsequent agonist administration, providing evidence that receptor sequestration limits drug efficacy in vivo. Direct receptor visualization in mice is a unique approach to receptor biology and drug design.

Animals↗
PubMed2006

Mu-delta opioid receptor functional interaction: Insight using receptor-G protein fusions.

Fusion proteins between a receptor and a pertussis toxin-insensitive G(i)alpha subunit were used to gain insight into the molecular interactions that take place upon mu and delta opioid receptor heterodimerization. When mu opioid receptor-G(i1)alpha fusions were coexpressed with nonfused delta opioid receptors in human embryonic kidney 293 cells, or vice versa, receptor heterodimers were detected by coimmunoprecipitation. In pertussis toxin-treated cells, receptor coexpression decreased the amount of guanosine 5'-O-(3-[35S]thio)triphosphate ([35S]GTPgammaS) incorporated in the fused G alpha protein after the addition of agonists specific for the receptor-G(i1)alpha fusion. In addition, activation of the G alpha protein occurred in heterodimers upon addition of an agonist specific for the nonfused receptor. It remained unaffected by an inverse agonist specific for the receptor-G(i1)alpha fusion. These data suggest that signaling through the receptor-G(i1)alpha fusion protein is impaired in heterodimers and support a mechanism in which activation of the G alpha subunit is promoted by a direct interaction with the nonfused receptor. Alternatively, receptor coexpression did not modify the ligand binding properties for the high-affinity state of the receptor-G(i1)alpha fusion nor the EC50 values for agonist-induced [35S]GTPgammaS incorporation in the G(i1)alpha subunit. In addition, no binding competition was observed between delta and mu ligands. Together, the data point to mu-delta opioid receptor heterodimers formed by contact interactions between monomers that retain their structural integrity.

Analgesics, Opioid↗
PubMed2006

Mu-opioid receptors are involved in the tolerance to nicotine antinociception.

Several studies have shown the participation of the endogenous opioid system on the antinociceptive effects and addictive properties of nicotine. The aim of the present study was to explore the involvement of the mu-opioid receptors in the development of tolerance to nicotine antinociception. Chronic treatment of C57BL/6 mice with nicotine (5 mg/kg s.c., three times daily during 12 days) resulted in tolerance to its antinociceptive responses in the tail-immersion test. We investigated the possible existence of adaptive changes in the expression and/or functional activity of mu-opioid receptors in these tolerant mice by using autoradiography of [(3)H]D-Ala(2)-MePhe(4)-Gly-ol(5) enkephalin ([(3)H]DAMGO) binding and DAMGO-stimulated guanosine [(35)S]5'-(gamma-thio)-triphosphate ([(35)S]GTPgammaS) binding. The density of mu-opioid receptors in the spinal cord was not modified in nicotine-tolerant mice, whereas a decrease was found in the caudate-putamen, as well as in the core and the shell of the nucleus accumbens. However, the functional activity of these receptors was significantly increased in the spinal cord as a consequence of nicotine treatment. To further investigate the role of mu-opioid receptors in the tolerance to nicotine-induced antinociception, we evaluated this response in C57BL/6 mu-opioid receptor knockout mice. Chronic nicotine treatment produced tolerance in both wild-type and knockout animals, but tolerance developed faster in mice lacking mu-opioid receptors. These results indicate that mu-opioid receptors play an important role in the development of tolerance to nicotine antinociceptive effects.

Analgesics, Opioid↗
PubMed2006

Big dynorphin as a putative endogenous ligand for the kappa-opioid receptor.

The diversity of peptide ligands for a particular receptor may provide a greater dynamic range of functional responses, while maintaining selectivity in receptor activation. Dynorphin A (Dyn A), and dynorphin B (Dyn B) are endogenous opioid peptides that activate the kappa-opioid receptor (KOR). Here, we characterized interactions of big dynorphin (Big Dyn), a 32-amino acid prodynorphin-derived peptide consisting of Dyn A and Dyn B, with human KOR, mu- (hMOR) and delta- (hDOR) opioid receptors and opioid receptor-like receptor 1 (hORL1) expressed in cells transfected with respective cDNA. Big Dyn and Dyn A demonstrated roughly similar affinity for binding to hKOR that was higher than that of Dyn B. Dyn A was more selective for hKOR over hMOR, hDOR and hORL1 than Big Dyn, while Dyn B demonstrated low selectivity. In contrast, Big Dyn activated G proteins through KOR with much greater potency, efficacy and selectivity than other dynorphins. There was no correlation between the rank order of the potency for the KOR-mediated activation of G proteins and the binding affinity of dynorphins for KOR. The rank of the selectivity for the activation of G proteins through hKOR and of the binding to this receptor also differed. Immunoreactive Big Dyn was detected using the combination of radioimmunoassay (RIA) and HPLC in the human nucleus accumbens, caudate nucleus, hippocampus and cerebrospinal fluid (CSF) with the ratio of Big Dyn and Dyn B being approximately 1:3. The presence in the brain implies that Big Dyn, along with other dynorphins, is processed from prodynorphin and secreted from neurons. Collectively, the high potency and efficacy and the relative abundance suggest that Big Dyn may play a role in the KOR-mediated activation of G proteins.

Animals↗
PubMed2006

RF9, a potent and selective neuropeptide FF receptor antagonist, prevents opioid-induced tolerance associated with hyperalgesia.

Neuropeptide FF (NPFF) has been proposed to play a role in pain modulation, opioid tolerance, and several other physiological processes. However, pharmacological agents that would help define physiological roles for this peptide are still missing. Here we report the discovery of a potent and selective NPFF receptor antagonist, RF9, that can be administered systemically. This compound does not show any effects by itself but can block efficiently the increase in blood pressure and heart rate evoked by NPFF. When chronically coinjected with heroin, RF9 completely blocks the delayed and long-lasting paradoxical opioid-induced hyperalgesia and prevents the development of associated tolerance. Our data indicate that NPFF receptors are part of a bona fide antiopioid system and that selective antagonists of these receptors could represent useful therapeutic agents for improving the efficacy of opioids in chronic pain treatment.

Adamantane↗
PubMed2006

Neuropathic pain is enhanced in delta-opioid receptor knockout mice.

We have evaluated the possible involvement of delta-opioid receptor (DOR) in the development and expression of neuropathic pain. For this purpose, partial ligation of the sciatic nerve was performed in DOR knockout mice and wild-type littermates. The development of mechanical and thermal allodynia, as well as thermal hyperalgesia was evaluated by using the von Frey filament model, the cold-plate test and the plantar test, respectively. In wild-type and DOR knockout mice, sciatic nerve injury led to a neuropathic pain syndrome revealed in these nociceptive behavioural tests. However, the development of mechanical and thermal allodynia, and thermal hyperalgesia was significantly enhanced in DOR knockout mice. These results reveal the involvement of DOR in the control of neuropathic pain and suggest a new potential therapeutic use of DOR agonists.

Analysis of Variance↗
PubMed2005

Dissociation of analgesic and hormonal responses to forced swim stress using opioid receptor knockout mice.

Exposure to stress triggers hormonal and behavioral responses. It has been shown that the endogenous opioid system plays a role in some physiological reactions to stress. The opioid system was described to mediate analgesia induced by mild stressors and to modulate the activation of the hypothalamic-pituitary-adrenal axis. Our study assessed the contribution of opioid receptors in stress-induced analgesia and adrenocorticotropic hormone (ACTH) and corticosterone release by a genetic approach. We performed a parallel analysis of mice deficient in mu, delta, or kappa opioid receptors, as well as of triple opioid receptor knockout mice, following exposure to a mild stress (3-min swim at 32 degrees C). In wild-type mice, stress elicited an increase in jumping latency on the hot plate, which was influenced by gender and genetic background. This analgesic response was reversed both by naloxone and by the triple mutation, and decreased in mu and delta opioid receptor knockout females. In wild-type females, stress also delayed front- and hindpaw behaviors in the hot plate test and increased tail-flick latency in the tail immersion test. Opioid receptor deletion however did not affect these stress responses. In addition, stress produced an increase in ACTH and corticosterone plasma levels. This endocrine response remained unchanged in all mutant strains. Therefore our data indicate that, under our stress conditions, the endogenous opioid system is recruited to produce some analgesia whereas it does not influence hypothalamic-pituitary-adrenal axis activity. This implies that brain circuits mediating analgesic and hormonal responses to stress can be dissociated.

Adrenocorticotropic Hormone↗
PubMed2005

Induction of delta opioid receptor function by up-regulation of membrane receptors in mouse primary afferent neurons.

It is not clear whether primary afferent neurons express functional cell-surface opioid receptors. We examined delta receptor coupling to Ca2+ channels in mouse dorsal root ganglion neurons under basal conditions and after receptor up-regulation. [D-Ala2,Phe4,Gly5-ol]-enkephalin (DAMGO), [D-Ala2,D-Leu5]-enkephalin (DADLE), trans-(+/-)-3,4-dichloro-N-methyl-N-(2-[1-pyrrolidinyl]cyclohexyl) benzene-acetamide methanesulfonate (U-50,488H; 1 microM), and baclofen (50 microM) inhibited Ca2+ currents, whereas the -selective ligands [D-Pen2,Pen5]-enkephalin (DPDPE) and deltorphin II (1 microM) did not. The effect of DADLE (1 microM) was blocked by the mu-antagonist D-Phe-Cys-Tyr-D-Trp-Arg-Thr-Pen-Thr-NH2 (CTAP; 300 nM) but not by the -antagonist Tyr-1,2,3,4-tetrahydroisoquinoline-Phe-Phe-OH (300 nM), implicating mu receptors. Despite a lack of functional delta receptors, flow cytometry revealed cell-surface receptors. We used this approach to identify conditions that up-regulate receptors, including mu receptor gene deletion in dorsal root ganglion neurons of mu-/- mice and 18-h incubation of mu+/+ neurons with CTAP followed by brief (10-min) DPDPE exposure. Under these conditions, the expression of cell-surface delta receptors was up-regulated to 149 +/- 9 and 139 +/- 5%, respectively; furthermore, DPDPE and deltorphin II (1 microM) inhibited Ca2+ currents in both cases. Viral replacement of mu receptors in mu-/- neurons reduced delta receptor expression to mu+/+ levels, restored the inhibition of Ca2+ currents by DAMGO, and abolished receptor coupling. Our observations suggest that receptor-Ca2+ channel coupling in primary afferent fibers may have little functional significance under basal conditions in which mu receptors predominate. However, up-regulation of cell-surface delta receptors induces their coupling to Ca2+ channels. Pharmacological approaches that increase functional delta receptor expression may reveal a novel target for analgesic therapy.

Animals↗
PubMed2005

Lack of stimulant and anxiolytic-like effects of ethanol and accelerated development of ethanol dependence in mu-opioid receptor knockout mice.

The opioid system is implicated in various aspects of alcoholism. Acute ethanol administration produces anxiolytic-like effects in rodents while alcohol withdrawal induces anxiogenic-like effects. Mice lacking the mu-opioid receptor (MOR) do not self-administer ethanol and display decreased anxiety-like behavior. We hypothesized that MOR might be involved in the development and expression of alcoholism, particularly in relation to anxiety states. In mice lacking MOR (MOR-/- mice), we examined the acute anxiolytic-like and locomotor stimulant effects of ethanol (0, 0.75, 1.25, 1.75 g/kg, i.p.). In a separate experiment, mice were submitted to chronic ethanol-containing liquid diet and we assessed somatic and affective ethanol withdrawal on three consecutive withdrawal episodes by scoring handling-induced convulsions and anxiety-like behavior. Deletion of MOR blocked the acute anxiolytic-like and stimulant effects of ethanol. Furthermore, MOR-/- mice displayed affective and physical signs of ethanol withdrawal in earlier withdrawal tests than wild-type mice. The present results implicate MOR in affective and somatic aspects of ethanol exposure and withdrawal. In addition, our findings support the hypothesis that the clinical efficacy of the opioid receptor antagonist naltrexone against relapse to alcoholism might be related to an action on the acute positive effects of alcohol rather than the negative affect of abstinence.

Alcohol Drinking↗
PubMed2005

Deletion of the mu opioid receptor results in impaired acquisition of Pavlovian context fear.

The mu opioid receptor may constitute a critical component of a negative feedback system that regulates Pavlovian fear conditioning. We investigated context fear conditioning acquisition and expression in mu opioid receptor knockout mice (on an inbred, C57 genetic background). We discovered that the mu receptor knockout results in an unexpected and significant deficit in context fear acquisition. Mice lacking the mu receptor showed normal fear acquisition when subjected to a 1-day fear conditioning protocol but evinced deficient fear learning when acquisition was conducted across 5 days. The knockout mice showed normal reactivity to footshock in both fear conditioning protocols. Finally, we confirmed the effectiveness of the receptor deletion in the C57 strain by subjecting the mice to a test of morphine analgesia in the hot-plate assay. As has been seen with mixed genetic background, the receptor deletion resulted in a complete lack of analgesic response to 10 mg/kg morphine. Surprisingly, mice with a single copy of the mu receptor gene (heterozygous knockouts) showed intact sensitivity to morphine but a significant deficit in Pavlovian fear conditioning. The results indicate that deletion of the mu receptor gene impairs fear conditioning and that the conditioning and analgesia effects of heterozygous deletion are dissociable. The conditioning deficit seen in this line of mice may be related to impairment in hippocampus function.

Analgesics, Opioid↗
PubMed2005

Decreased oral self-administration of alcohol in kappa-opioid receptor knock-out mice.

BACKGROUND: Although a large body of evidence suggests a role for the opioid system in alcoholism, the precise role of mu-, delta-, kappa-, and ORL1-opioid receptors and the physiological significance of their natural genetic variation have not been identified. The method of targeted gene disruption by homologous recombination has been used to knock out (KO) genes coding for opioid receptors, and study their effects on alcohol self-administration. Here we examined the effects of targeted disruption of kappa-opioid receptor (KOR) on oral alcohol self-administration and other behaviors. METHODS: Oral alcohol, saccharin and quinine self-administration was assessed in a two-bottle choice paradigm using escalating concentrations of alcohol, or tastant solutions. In preference tests 12% alcohol, 0.033% and 0.066% saccharin, and 0.03 mM and 0.1 mM quinine solutions were used. Open-field activity was determined in an arena equipped with a computer-controlled activity-detection system. Subjects were tested for three consecutive days. Locomotor activity was assessed on days 1 and 2 (after saline injection, i.p.) and on day 3 (after alcohol injection, i.p.). Alcohol-induced locomotor activity was determined as the difference in activity between day 3 and day 2. RESULTS: Male KOR KO mice in preference tests with 12% alcohol consumed about half as much alcohol as wild-type (WT) or heterozygous (HET) mice, showed lower preference for saccharin (0.033% and 0.066%) and higher preference to quinine (0.1 mM) than WT mice. Female KOR KO mice showed similar reduction in alcohol consumption in comparison to WT and HET mice. Partial deletion of KOR in HET mice did not change alcohol consumption in comparison to WT mice. In all genotype-groups females drank significantly more alcohol than males. MANOVA of locomotor activity among KO, WT, and HET mice indicated that strain and sex effects were not significant for alcohol-induced activation (p > 0.05), while strain x sex interaction effects on alcohol-induced activation could be detected (F(1,55) = 6.07, p < 0.05). CONCLUSION: Our results indicating decreased alcohol consumption, lower saccharin preference, and higher quinine preference in KOR KO mice are in line with previous observations of opioid involvement in maintenance of food intake and raise the possibility that the deficient dynorphin/KOR system affects orosensory reward through central mechanisms which reduce alcohol intake and disrupt tastant responses, either as direct effects of absence of kappa-opioid receptors, or as effects of indirect developmental compensatory changes.

Alcohol Drinking↗
PubMed2005

Generation and characterization of Rgs4 mutant mice.

RGS proteins are negative regulators of signaling through heterotrimeric G protein-coupled receptors and, as such, are in a position to regulate a plethora of biological phenomena. However, those have just begun to be explored in vivo. Here, we describe a mouse line deficient for Rgs4, a gene normally expressed early on in discrete populations of differentiating neurons and later on at multiple sites of the central nervous system, the cortex in particular, where it is one of the most highly transcribed Rgs genes. Rgs4(lacZ/lacZ) mice had normal neural development and were viable and fertile. Behavioral testing on mutant adults revealed subtle sensorimotor deficits but, so far, supported neither the proposed status of Rgs4 as a schizophrenia susceptibility gene (by showing intact prepulse inhibition in the mutants) nor (unlike another member of the Rgs family, Rgs9) a role of Rgs4 in the acute or chronic response to opioids.

Animals↗