Les publications

IF du Neurocentre

799 publications

* equal contribution
Les IF indiqués ont été collectés par le Web of Sciences en Juin 2020

01/06/2003 | J Neurosci
The glucocorticoid receptor as a potential target to reduce cocaine abuse.
Deroche-Gamonet V, Sillaber I, Aouizerate B, Izawa R, Jaber M, Ghozland S, Kellendonk C, Le Moal M, Spanagel R, Schutz G, Tronche F, Piazza PV

Several findings suggest that glucocorticoid hormones are involved in determining the propensity of an individual to develop cocaine abuse. These hormones activate two related transcription factors, the glucocorticoid receptor (GR) and the mineralocorticoid receptor. In this study, we show that the selective inactivation of the GR gene in the brains of mice profoundly flattened the dose-response function for cocaine intravenous self-administration and suppressed sensitization, two experimental procedures considered relevant models of addiction. Furthermore, administration of a GR antagonist dose-dependently reduced the motivation to self-administer cocaine. Importantly, the absence of GR did not modify the basal behavioral and molecular effects of cocaine but selectively modified the excessive response to the drug spontaneously present in certain vulnerable individuals or induced by repeated drug exposure in others. In conclusion, we provide the first genetic evidence that the GR gene can modulate cocaine abuse. This suggests that targeting GR function in the brain could provide new therapeutic strategies to treat cocaine addiction for which there is no available treatment.

01/06/2003 | J Neurosci
Evidence for enhanced neurobehavioral vulnerability to nicotine during periadolescence in rats.
Adriani W, Spijker S, Deroche-Gamonet V, Laviola G, Le Moal M, Smit AB, Piazza PV

Epidemiological studies indicate that there is an increased likelihood for the development of nicotine addiction when cigarette smoking starts early during adolescence. These observations suggest that adolescence could be a 'critical' ontogenetic period, during which drugs of abuse have distinct effects responsible for the development of dependence later in life. We compared the long-term behavioral and molecular effects of repeated nicotine treatment during either periadolescence or postadolescence in rats. It was found that exposure to nicotine during periadolescence, but not a similar exposure in the postadolescent period, increased the intravenous self-administration of nicotine and the expression of distinct subunits of the ligand-gated acetylcholine receptor in adult animals. Both these changes indicated an increased sensitivity to the addictive properties of nicotine. In conclusion, adolescence seems to be a critical developmental period, characterized by enhanced neurobehavioral vulnerability to nicotine.

06/2003 | Conscious Cogn
Impaired strategic regulation of contents of conscious awareness in
Sonntag P , Gokalsing E , Olivier C , Robert P , Burglen F , Kauffmann-Muller F , Huron C , Salame P , Danion JM

Conscious awareness comprises two distinct states, autonoetic and noetic

06/2003 | Development
Wnt signaling mediates reorientation of outer hair cell stereociliary bundles in the mammalian cochlea.
Dabdoub A , Donohue MJ , Brennan A , Wolf V , Montcouquiol M , Sassoon DA , Hseih JC , Rubin JS , Salinas PC , Kelley MW

In the mammalian cochlea, stereociliary bundles located on mechanosensory hair cells within the sensory epithelium are unidirectionally oriented. Development of this planar polarity is necessary for normal hearing as stereociliary bundles are only sensitive to vibrations in a single plane; however, the mechanisms governing their orientation are unknown. We report that Wnt signaling regulates the development of unidirectional stereociliary bundle orientation. In vitro application of Wnt7a protein or inhibitors of Wnt signaling, secreted Frizzled-related protein 1 or Wnt inhibitory factor 1, disrupts bundle orientation. Moreover, Wnt7a is expressed in a pattern consistent with a role in the polarization of the developing stereociliary bundles. We propose that Wnt signaling across the region of developing outer hair cells gives rise to planar polarity in the mammalian cochlea.

05/2003 | Eur J Neurosci
The effects of ibotenic hippocampal lesions on discriminative fear conditioning
Desmedt A, Marighetto A, Garcia R, Jaffard R

To what extent the hippocampus is required for contextual conditioning remains a

05/2003 | Eur J Neurosci
Modulation of GABAergic transmission by endogenous glutamate in the rat supraoptic nucleus.
Piet R, Bonhomme R, Theodosis DT, Poulain DA, Oliet SH

The presence of group III metabotropic glutamate receptors on GABAergic terminals in the supraoptic nucleus suggests that the level of glutamate in the extracellular space may regulate synaptic strength at inhibitory synapses. To test this hypothesis we examined the consequences of increasing ambient glutamate on GABA-mediated synaptic activity in supraoptic neurons. The concentration of the excitatory amino acid in the extracellular space was increased pharmacologically by blocking glutamate transporters. Inhibition of the astrocyte-specific GLT-1 glutamate transporter led to a reversible decrease in evoked inhibitory postsynaptic current amplitude. This modulation had a presynaptic origin as revealed by analysis of paired-pulse ratio and miniature inhibitory currents. Furthermore, blocking group III metabotropic glutamate receptors with the specific antagonist MAP4 prevented the depression of GABAergic transmission induced by glutamate transporter blockade. Thus, presynaptic metabotropic glutamate receptors located on inhibitory terminals in the supraoptic nucleus appear to sense changes in ambient glutamate and modify GABA release accordingly. However, it seems that such changes need to reach a certain magnitude because the discrete deficit in glutamate clearance which occurs in the supraoptic nucleus of lactating rats is not sufficient to modulate GABA-mediated transmission. These results suggest that ambient glutamate contributes to the modulation of synaptic efficacy not only at glutamatergic synapses but also at inhibitory GABAergic synapses.

29/04/2003 | Philos Trans R Soc Lond B Biol Sci
Active dendrites, potassium channels and synaptic plasticity.
Johnston D, Christie BR, Frick A, Gray R, Hoffman DA, Schexnayder LK, Watanabe S, Yuan LL

The dendrites of CA1 pyramidal neurons in the hippocampus express numerous types of voltage-gated ion channel, but the distributions or densities of many of these channels are very non-uniform. Sodium channels in the dendrites are responsible for action potential (AP) propagation from the axon into the dendrites (back-propagation); calcium channels are responsible for local changes in dendritic calcium concentrations following back-propagating APs and synaptic potentials; and potassium channels help regulate overall dendritic excitability. Several lines of evidence are presented here to suggest that back-propagating APs, when coincident with excitatory synaptic input, can lead to the induction of either long-term depression (LTD) or long-term potentiation (LTP). The induction of LTD or LTP is correlated with the magnitude of the rise in intracellular calcium. When brief bursts of synaptic potentials are paired with postsynaptic APs in a theta-burst pairing paradigm, the induction of LTP is dependent on the invasion of the AP into the dendritic tree. The amplitude of the AP in the dendrites is dependent, in part, on the activity of a transient, A-type potassium channel that is expressed at high density in the dendrites and correlates with the induction of the LTP. Furthermore, during the expression phase of the LTP, there are local changes in dendritic excitability that may result from modulation of the functioning of this transient potassium channel. The results support the view that the active properties of dendrites play important roles in synaptic integration and synaptic plasticity of these neurons.

15/04/2003 | J Neurosci
Presynaptic facilitation of glutamatergic synapses to dopaminergic neurons of the rat substantia nigra by endogenous stimulation of vanilloid receptors.
Marinelli S , Di Marzo V , Berretta N , Matias I , Maccarrone M , Bernardi G , Mercuri NB

Growing evidence regarding the function of vanilloid receptor-1 (VR1) in the brain suggests potential central roles of this receptor, previously described to occur primarily in peripheral sensory neurons. In the present study, we used electrophysiological and biochemical techniques to investigate the function and the endogenous stimulation of VR1 in dopaminergic neurons of the substantia nigra pars compacta (SNc). The VR1 agonist capsaicin increased the frequency of both TTX-sensitive and -insensitive spontaneous EPSCs (sEPSCs) without affecting their amplitude, suggesting a presynaptic site of action. In contrast, no effect was detected with regard to GABAergic transmission. No increase in sEPSC frequency was observed in the presence of cadmium chloride, while the voltage-dependent calcium channel antagonist omega-conotoxin MVIIC did not prevent capsaicin action. The VR1 antagonists capsazepine and iodoresiniferatoxin (IRTX) blocked the effects of capsaicin. Importantly, IRTX per se reduced sEPSC frequency, suggesting a tonic activity of VR1. The endogenous VR1 agonist anandamide (AEA) produced an IRTX-sensitive increase in the frequency of sEPSCs on dopaminergic neurons that was more pronounced when protein kinase A had been activated. Furthermore, mass spectrometric analyses and binding experiments revealed high levels of endogenous AEA and specific binding of AEA to VR1 receptors in the SNc. These data suggest a tonic facilitation of glutamate release exerted by VR1 in the SNc through a stimulation of VR1 by endovanilloids, including anandamide. The increase in sEPSC frequency by VR1 onto midbrain dopaminergic neurons suggests the involvement of these receptors in motor and cognitive functions involving the dopaminergic system.

15/04/2003 | J Neurosci
Normalization of Ca2+ signals by small oblique dendrites of CA1 pyramidal neurons.
Frick A, Magee J, Koester HJ, Migliore M, Johnston D

Oblique dendrites of CA1 pyramidal neurons predominate in stratum radiatum and receive approximately 80% of the synaptic input from Schaffer collaterals. Despite this fact, most of our understanding of dendritic signal processing in these neurons comes from studies of the main apical dendrite. Using a combination of Ca2+ imaging and whole-cell recording techniques in rat hippocampal slices, we found that the properties of the oblique dendrites differ markedly from those of the main dendrites. These different properties tend to equalize the Ca2+ rise from single action potentials as they backpropagate into the oblique dendrites from the main trunk. Evidence suggests that this normalization of Ca2+ signals results from a higher density of a transient, A-type K+ current [I(K(A))] in the oblique versus the main dendrites. The higher density of I(K(A)) may have important implications for our understanding of synaptic integration and plasticity in these structures.

03/2003 | Int J Obes Relat Metab Disord
Endogenous cannabinoid system as a modulator of food intake.
Cota D, Marsicano G, Lutz B, Vicennati V, Stalla GK, Pasquali R, Pagotto U

The ability of Cannabis sativa (marijuana) to increase hunger has been noticed for centuries, although intensive research on its molecular mode of action started only after the characterization of its main psychoactive component Delta(9)-tetrahydrocannabinol in the late 1960s. Despite the public concern related to the abuse of marijuana and its derivatives, scientific studies have pointed to the therapeutic potentials of cannabinoid compounds and have highlighted their ability to stimulate appetite, especially for sweet and palatable food. Later, the discovery of specific receptors and their endogenous ligands (endocannabinoids) suggested the existence of an endogenous cannabinoid system, providing a physiological basis for biological effects induced by marijuana and other cannabinoids. Epidemiological reports describing the appetite-stimulating properties of cannabinoids and the recent insights into the molecular mechanisms underlying cannabinoid action have proposed a central role of the cannabinoid system in obesity. The aim of this review is to provide an extensive overview on the role of this neuromodulatory system in feeding behavior by summarizing the most relevant data obtained from human and animal studies and by elucidating the interactions of the cannabinoid system with the most important neuronal networks and metabolic pathways involved in the control of food intake. Finally, a critical evaluation of future potential therapeutical applications of cannabinoid antagonists in the therapy of obesity and eating disorders will be discussed.

The endogenous cannabinoid system has been shown recently to play a crucial role in the extinction of aversive memories. As the amygdala is presumably involved in this process, we investigated the effects of the cannabinoid receptor agonist WIN 55,212-2 (WIN-2) on synaptic transmission in the lateral amygdala (LA) of wild-type and cannabinoid receptor type 1 (CB1)-deficient mice. Extracellular field potential recordings and patch-clamp experiments were performed in an in vitro slice preparation. We found that WIN-2 reduces basal synaptic transmission and pharmacologically isolated AMPA receptor- and GABA(A) receptor-mediated postsynaptic currents in wild-type, but not in CB1-deficient mice. These results indicate that, in the LA, cannabinoids modulate both excitatory and inhibitory synaptic transmission via CB1. WIN-2-induced changes of paired-pulse ratio and of spontaneous and miniature postsynaptic currents suggest a presynaptic site of action. Inhibition of G(i/o) proteins and blockade of voltage-dependent and G protein-gated inwardly rectifying K(+) channels inhibited WIN-2 action on basal synaptic transmission. In contrast, modulation of the adenylyl cyclase-protein kinase A pathway, and blockade of presynaptic N- and P/Q- or of postsynaptic L- and R/T-type voltage-gated Ca(2+) channels did not affect WIN-2 effects. Our results indicate that the mechanisms underlying cannabinoid action in the LA partly resemble those observed in the nucleus accumbens and differ from those described for the hippocampus.

02/2003 | Eur J Neurosci
Fos imaging reveals ageing-related changes in hippocampal response to radial maze
Touzani K, Marighetto A, Jaffard R

A two-stage radial arm maze (RAM) task has been designed recently to demonstrate

02/2003 | Cell Mol Life Sci
Effect of maternal under-nutrition on pup body weight and hypothalamic endocannabinoid levels.
Matias I , Leonhardt M , Lesage J , De Petrocellis L , Dupouy JP , Vieau D , Di Marzo V

Dietary long-chain polyunsaturated fatty acids are known to influence brain levels of the endocannabinoid anandamide in newborn pigs and mice. Furthermore, endocannabinoids were shown to control pup suckling and body weight in mice, and food intake in adult rodents. Here we determined the effect of maternal under-nutrition during gestation, lactation, or both, on body weight, and on the levels of endocannabinoids and expression of cannabinoid CB1 receptors and fatty acid amide hydrolase in the hypothalamus of rat pups at weaning (21 days old) or adult rats (4 months old). Maternal under-nutrition resulted in a striking decrease in body weight of weaning rats, paralleled by a decrease in the hypothalamic levels of the endocannabinoid anandamide, but not of 2-arachidonoylglycerol. No significant change in the hypothalamic expression of either cannabinoid CB1 receptors or fatty acid amide hydrolase mRNA was detected in any of the three groups of weaned pups. The decrease in pup body weight and hypothalamic anandamide levels was not observable in 4-month-old rats from any of the three groups. These data suggest that maternal under-nutrition causes a decrease in hypothalamic anandamide levels and loss of body weight, and confirm a crucial role for endocannabinoid signalling in neonatal development.

2003 | Nature
Identification of Vangl2 and Scrb1 as planar polarity genes in mammals.
Montcouquiol M*, Rachel RA*, Lanford PJ, Copeland NG, Jenkins NA, Kelley MW

2003 | Eur J Neurosci
Modulation of GABAergic transmission by endogenous glutamate in the rat supraoptic nucleus
Piet R, Bonhomme R, Theodosis D, Poulain D, Oliet S

2003 | Nat Cell Biol
NMDA receptor trafficking through an interaction between PDZ Proteins and the exocyst complex
Sans N, Prybylowski K, Petralia RS, Chang K, Wang YX, Racca C, Vicini S, Wenthold RW

NMDA (N-methyl-D-aspartate) receptors (NMDARs) are targeted to dendrites and anchored at the post-synaptic density (PSD) through interactions with PDZ proteins. However, little is known about how these receptors are sorted from the endoplasmic reticulum and Golgi apparatus to the synapse. Here, we find that synapse-associated protein 102 (SAP102) interacts with the PDZ-binding domain of Sec8, a member of the exocyst complex. Our results show that interactions between SAP102 and Sec8 are involved in the delivery of NMDARs to the cell surface in heterologous cells and neurons. Furthermore, they suggest that an exocyst-SAP102-NMDAR complex is an important component of NMDAR trafficking.

2003 | J Neurosci
Aberrant formation of glutamate receptor complexes in hippocampal neurons of mice lacking the GluR2 AMPA receptor subunit.
Sans N, Vissel B, Petralia RS, Wang YX, Chang K, Royle GA, Wang CY, O'Gorman S, Heinemann SF, Wenthold RW

The number and type of receptors present at the postsynaptic membrane determine the response to the neurotransmitter released from the presynaptic terminal. Because most neurons receive multiple and distinct synaptic inputs and contain several different subtypes of receptors stimulated by the same neurotransmitter, the assembly and trafficking of receptors in neurons is a complex process involving many levels of regulation. To investigate the mechanism that neurons use to regulate the assembly of receptor subunits, we studied a GluR2 knock-out mouse. GluR2 is a critical subunit that controls calcium permeability of AMPA receptors and is present in most native AMPA receptors. Our data indicate that in the absence of GluR2, aberrant receptor complexes composed of GluR1 and GluR3 are formed in the hippocampus, and that there is an increased number of homomeric GluR1 and GluR3 receptors. We also show that these homomeric and heteromeric receptors are less efficiently expressed at the synapse. Our results show that GluR2 plays a critical role in controlling the assembly of AMPA receptors, and that the assembly of subunits may reflect the affinity of one subunit for another or the stability of intermediates in the assembly process. Therefore, GluR1 may have a greater preference for GluR2 than it does for GluR3.

2003 | Annu. Rev. Pharmacol. Toxicol.
Trafficking of NMDA receptors
Wenthold RW, Prybylowski K, Standley S, Sans N, Petralia RS

2003 | Hippocampus
Hippocampal lesions and discrimination performance of mice in the radial maze:
Etchamendy N, Desmedt A, Cortes-Torrea C, Marighetto A, Jaffard R

The effects of ibotenate hippocampal lesions on discrimination performance in an

10/2002 | j gen virol
Lymphoid activation: a confounding factor in AIDS vaccine development?
Richardson J, Broche S, Baud S, Leste-Lasserre T, Femenia F, Levy D, Moraillon A, Pancino G, Sonigo P

In a previous vaccination trial, inoculation of env gene DNA failed to elicit a detectable antibody response, yet accelerated virus dissemination in most immunized cats following challenge with feline immunodeficiency virus. This result raised the possibility that cell-mediated immune responses had given rise to immune-mediated enhancement of infection. Since high-level replication of immunodeficiency viruses in lymphocytes requires cellular activation, antigen-specific responses or non-specific polyclonal activation may have increased the frequency of optimal target cells. In the present DNA vaccination trial, although designed so as to minimize non-specific polyclonal activation, immune-mediated enhancement was nonetheless observed in certain immunized cats. Moreover, rapid virus dissemination in vivo was associated with the presence of T-helper responses prior to challenge, and was linked to increased susceptibility of lymphocytes to ex vivo infection. Immune activation may thus be a confounding factor in vaccination against lentivirus infection, diminishing vaccine efficacy and giving rise to immune-mediated enhancement.

We investigated whether three calcium-binding proteins, calretinin, parvalbumin, and calbindin, could identify specific aspects of the postnatal development of the rat lateral (LVN) and medial (MVN) vestibular nuclei and their vestibular and cerebellar connections. Calretinin levels in the vestibular nuclei, increased significantly between birth and postnatal day (P) 45. In situ hybridization and immunocytochemical staining showed that calretinin-immunoreactive neurons were mostly located in the parvocellular MVN at birth and that somatic and dendritic growth occurred between birth and P14. During the first week, parvalbumin-immunoreactive fibers and endings were confined to specific areas, i.e., the ventral LVN and magnocellular MVN, and identified exclusively the maturation of the vestibular afferents. Calbindin was located within the dorsal LVN and the parvocellular MVN and identified the first arrival of the corticocerebellar afferents. From the second week, in addition to labeling vestibular afferents in their specific target areas, parvalbumin was also found colocalized with calbindin in mature Purkinje cell afferents. Thus, the specific spatiotemporal distribution of parvalbumin and calbindin could correspond to two successive phases of synaptic remodeling involving integration of the vestibular sensory messages and their cerebellar control. On the basis of the sequence of distribution patterns of these proteins during the development of the vestibular nuclei, calretinin is an effective marker for neuronal development of the parvocellular MVN, parvalbumin is a specific marker identifying maturation of the vestibular afferents and endings, and calbindin is a marker of the first appearance and development of Purkinje cell afferents.

01/08/2002 | Nature
The endogenous cannabinoid system controls extinction of aversive memories.
Marsicano G, Wotjak CT, Azad SC, Bisogno T, Rammes G, Cascio MG, Hermann H, Tang J, Hofmann C, Zieglgansberger W, Di Marzo V, Lutz B

Acquisition and storage of aversive memories is one of the basic principles of central nervous systems throughout the animal kingdom. In the absence of reinforcement, the resulting behavioural response will gradually diminish to be finally extinct. Despite the importance of extinction, its cellular mechanisms are largely unknown. The cannabinoid receptor 1 (CB1) and endocannabinoids are present in memory-related brain areas and modulate memory. Here we show that the endogenous cannabinoid system has a central function in extinction of aversive memories. CB1-deficient mice showed strongly impaired short-term and long-term extinction in auditory fear-conditioning tests, with unaffected memory acquisition and consolidation. Treatment of wild-type mice with the CB1 antagonist SR141716A mimicked the phenotype of CB1-deficient mice, revealing that CB1 is required at the moment of memory extinction. Consistently, tone presentation during extinction trials resulted in elevated levels of endocannabinoids in the basolateral amygdala complex, a region known to control extinction of aversive memories. In the basolateral amygdala, endocannabinoids and CB1 were crucially involved in long-term depression of GABA (gamma-aminobutyric acid)-mediated inhibitory currents. We propose that endocannabinoids facilitate extinction of aversive memories through their selective inhibitory effects on local inhibitory networks in the amygdala.

08/2002 | Eur J Biochem
Presence and regulation of the endocannabinoid system in human dendritic cells.
Matias I , Pochard P , Orlando P , Salzet M , Pestel J , Di Marzo V

Cannabinoid receptors and their endogenous ligands, the endocannabinoids, have been detected in several blood immune cells, including monocytes/macrophages, basophils and lymphocytes. However, their presence in dendritic cells, which play a key role in the initiation and development of the immune response, has never been investigated. Here we have analyzed human dendritic cells for the presence of the endocannabinoids, anandamide and 2-arachidonoylglycerol (2-AG), the cannabinoid CB1 and CB2 receptors, and one of the enzymes mostly responsible for endocannabinoid hydrolysis, the fatty acid amide hydrolase (FAAH). By using a very sensitive liquid chromatography-atmospheric pressure chemical ionization-mass spectrometric (LC-APCI-MS) method, lipids extracted from immature dendritic cells were shown to contain 2-AG, anandamide and the anti-inflammatory anandamide congener, N-palmitoylethanolamine (PalEtn) (2.1 +/- 1.0, 0.14 +/- 0.02 and 8.2 +/- 3.9 pmol x 10(-7) cells, respectively). The amounts of 2-AG, but not anandamide or PalEtn, were significantly increased following cell maturation induced by bacterial lipopolysaccharide (LPS) or the allergen Der p 1 (2.8- and 1.9-fold, respectively). By using both RT-PCR and Western immunoblotting, dendritic cells were also found to express measurable amounts of CB1 and CB2 receptors and of FAAH. Cell maturation did not consistently modify the expression of these proteins, although in some cell preparations a decrease of the levels of both CB1 and CB2 mRNA transcripts was observed after LPS stimulation. These findings demonstrate for the first time that the endogenous cannabinoid system is present in human dendritic cells and can be regulated by cell activation.

06/2002 | Psychopharmacology (Berl)
Study of the addictive potential of modafinil in naive and cocaine-experienced rats.
Deroche-Gamonet V, Darnaudery M, Bruins-Slot L, Piat F, Le Moal M, Piazza PV

RATIONALE: Modafinil is a drug that promotes wakefulness and, as such, is used to treat hypersomnia and narcolepsy. Preclinical and clinical studies suggest that modafinil could possess weak reinforcing effects in drug-experienced subjects. However, its abuse potential in drug-naive healthy individuals is still totally uninvestigated, despite the fact that availability of modafinil has recently increased. OBJECTIVES: The purpose of our study was to investigate the potential addictive properties of modafinil by testing its reinforcing effects in naive rats. The interactions of modafinil with the reinforcing effects of cocaine were also tested. METHODS: First, using i.v. self-administration and place conditioning tests, we studied the reinforcing and rewarding effects of a large range of doses of modafinil in naive rats. Second, we tested the influence of modafinil on reinforcing and incentive effects of cocaine in rats trained for cocaine self-administration. The effects of modafinil were compared with those of amphetamine and haloperidol. RESULTS: Modafinil did not produce reinforcing or rewarding effects and did not modify the effects of cocaine. CONCLUSIONS: Our results suggest that modafinil does not possess an addictive potential in naive individuals. Furthermore, it would be behaviorally distinct from classical central nervous system stimulants which are known to alter cocaine-induced effects. However, as shown previously in nonhuman primates and in humans, modafinil could possibly have reinforcing effects in cocaine-experienced individuals.

06/2002 | Neurotox Res
Individual vulnerability to substance abuse and affective disorders: role of early environmental influences.
Koehl M, Lemaire V, Mayo W, Abrous DN, Maccari S, Piazza PV, Le Moal M, Vallee M

One of the most important questions raised by modern psychiatry and experimental psychopathology is the origin of mental diseases. More concisely, clinical and experimental neurosciences are increasingly concerned with the factors that render one individual more vulnerable than another to a given pathological outcome. Animal models are now available to understand the sources of individual differences for specific phenotypes prone to behavioral disadaptations. Over the last 10 years we have explored the consequences of environmental perinatal manipulations in the rat. We have shown that prenatal stress is at the origin of a wide range of physiological and behavioral aberrances such as alterations in the activity of the hormonal stress axis, increased vulnerability to drug of abuse, emotional liability, cognitive impairments and predisposition to pathological aging. Taken together, these abnormalities define a bio-behavioral syndrome. Furthermore, the cognitive disabilities observed in prenatally-stressed rats were recently related to an alteration of neurogenesis in the dentate gyrus, thus confirming the impact of early life events on brain morphology. A second model (handling model) has also been developed in which pups are briefly separated from their mothers during early postnatal life. In contrast with prenatally-stressed animals, handled rats exhibited a reduced emotion response when confronted with novel situations and were protected against age-induced impairments of both the hormonal stress axis and cognitive functions. Taken together, the results of these investigations show that the bio-behavioral phenotype that characterizes each individual is strongly linked to the nature and timing of perinatal experience. Furthermore, data collected in prenatally-stressed animals indicate that this model could be used profitably to understand the etiology and pathophysiology of affective disorders.

01/05/2002 | J Neurosci
Nicotine self-administration impairs hippocampal plasticity.
Abrous DN, Adriani W, Montaron MF, Aurousseau C, Rougon G, Le Moal M, Piazza PV

Nicotine, the neuroactive compound responsible for tobacco addiction, is primarily believed to have beneficial effects on the adult brain. However, in heavy smokers, abstinence from nicotine is accompanied by cognitive impairments that suggest adverse effects of nicotine on brain plasticity. For this reason, we studied changes in plasticity-related processes in the dentate gyrus (DG) of the hippocampal formation of animals trained to self-administer nicotine. The DG was chosen because it undergoes profound plastic rearrangements, many of which have been related to memory and learning performances. In this region, we examined the expression of the polysialylated (PSA) forms of neural cell adhesion molecule (NCAM), PSA-NCAM, neurogenesis, and cell death by measuring the number of pyknotic cells. It was found that nicotine self-administration profoundly decreased, in a dose-dependent manner, the expression of PSA-NCAM in the DG; a significant effect was observed at all the doses tested (0.02, 0.04, and 0.08 mg/kg per infusion). Neurogenesis was also decreased in the DG, but a significant effect was observed only for the two highest doses of nicotine. Finally, the same doses that decreased neurogenesis also increased cell death. These results raise an important additional concern for the health consequences of nicotine abuse and open new insight on the possible neural mechanisms of tobacco addiction.

Conditioning theories propose that, through a Pavlovian associative process, discrete stimuli acquire the ability to elicit neural states involved in the maintenance and relapse of a drug-taking behaviour. Experimental evidence indicates that drug-related cues play a role in relapse, however, their influence on the development and maintenance of drug self-administration has been poorly investigated. In this report, we analysed the effects of a drug-associated cue light on acquisition, maintenance and reinstatement of intravenous cocaine self-administration. The results show that a cocaine-associated cue light can act as an incentive in absence of the drug, but does not directly modify drug-reinforcing effects. Contingent and non-contingent presentations of a cocaine-associated cue light reinstated an extinguished self-administration behaviour. However, regardless of whether or not a cue light was associated with cocaine infusions, rats acquire cocaine intravenous self-administration reaching the same levels of intake. Furthermore, after self-administration has been acquired in presence of the cue light, the omission of the cue light or its non-contingent presentation did not modify rat behaviour. In conclusion, our work shows that cocaine-associated explicit cues do not directly interfere with the reinforcing effects of the drug.

One of the functions of astroglial cells in the central nervous system is to clear synaptically-released glutamate from the extracellular space. This is performed thanks to specific transporters of the excitatory amino acid expressed on their surface. The way by which astrocytic glutamate uptake contributes to synaptic transmission has been investigated via numerous experimental approaches but has never been addressed under conditions where neuroglial interactions are physiologically modified. Recently, we took advantage of the neuroglial plastic properties of the hypothalamo-neurohypophysial system to examine the consequences of a physiological reduction in the astrocytic coverage of neurons on glutamatergic synaptic transmission. This experimental model has brought some insights on the physiological interactions between glial cells and neurons at the level of the synapse. In particular, it has revealed that the degree of glial coverage of neurons influences glutamate concentration at the vicinity of excitatory synapses and, as a consequence, affects the level of activation of presynaptic glutamate receptors. Astrocytes, therefore, appear to contribute to the regulation of neuronal excitability by modulating synaptic efficacy at glutamatergic nerve terminals.

The supraoptic and paraventricular nuclei of the hypothalamus undergo reversible anatomical changes under conditions of intense neurohypophysial hormone secretion, such as lactation, parturition and chronic dehydration. This morphological remodelling includes a reduction in astrocytic coverage of neurones resulting in an increase in the number and extent of directly juxtaposed somatic and dendritic surfaces. There is a growing body of evidence indicating that such anatomical plasticity is of functional significance. Astrocytic-dependent clearance of electrolytes and neurotransmitters from the extracellular space appears to be altered under conditions where glial coverage of magnocellular neurones is reduced. Glutamate, for example, has been found to accumulate in the extracellular space in the supraoptic nucleus of lactating animals and cause a modulation of synaptic efficacy. On the other hand, the range of action of substances released from astrocytes and acting on adjacent magnocellular neurones is expected to be limited during such anatomical remodelling. It thus appears that the structural plasticity of the magnocellular nuclei does affect neuroglial interactions, inducing significant changes in signal transmission and processing.

Neuroprotective effects have been described for many cannabinoids in several neurotoxicity models. However, the exact mechanisms have not been clearly understood yet. In the present study, antioxidant neuroprotective effects of cannabinoids and the involvement of the cannabinoid receptor 1 (CB1) were analysed in detail employing cell-free biochemical assays and cultured cells. As it was reported for oestrogens that the phenolic group is a lead structure for antioxidant neuroprotective effects, eight compounds were classified into three groups. Group A: phenolic compounds that do not bind to CB1. Group B: non-phenolic compounds that bind to CB1. Group C: phenolic compounds that bind to CB1. In the biochemical assays employed, a requirement of the phenolic lead structure for antioxidant activity was shown. The effects paralleled the protective potential of group A and C compounds against oxidative neuronal cell death using the mouse hippocampal HT22 cell line and rat primary cerebellar cell cultures. To elucidate the role of CB1 in neuroprotection, we established stably transfected HT22 cells containing CB1 and compared the protective potential of cannabinoids with that observed in the control transfected HT22 cell line. Furthermore, oxidative stress experiments were performed in cultured cerebellar granule cells, which were derived either from CB1 knock-out mice or from control wild-type littermates. The results strongly suggest that CB1 is not involved in the cellular antioxidant neuroprotective effects of cannabinoids.

Considerable efforts have been made to identify changes of brain synaptic plasticity associated with fear conditioning. However, for both clinical applications and our fundamental understanding of memory processes, it appears also necessary to investigate synaptic plasticity related to extinction. We previously showed that extinction of freezing to a tone conditioned stimulus (CS; previously paired with footshock) in mice results in a sequence of depression and potentiation of synaptic efficacy in the medial prefrontal cortex (mPFC). These data as well as those from lesion studies suggest that the direction of changes in prefrontal synaptic plasticity may modulate extinction of learned fear. To test this, we analyzed the effects of low-frequency stimulation (LFS) and high-frequency stimulation (HFS) of the mediodorsal thalamic nucleus, known to induce prefrontal long-term depression (LTD) and potentiation (LTP), respectively, on extinction. We found that maintenance of the depression phase, using thalamic LFS, was associated with resistance to extinction. Thalamic HFS applied before extinction testing had no effect on the rate of extinction. However, 1 week follow-up tests revealed that the memory of extinction was intact in these mice (with prefrontal LTP) and in control mice displaying prefrontal LTP-like changes, whereas control mice that did not exhibit such changes displayed a return of freezing to the CS. The results suggest that after extinction the lack of depression-LTP-like conversion sequence in the mPFC synaptic efficacy may profoundly alter the process of consolidation.

01/01/2002 | J Neurosci
Anti-S-nitrosocysteine antibodies are a predictive marker for demyelination in experimental autoimmune encephalomyelitis: implications for multiple sclerosis.
Boullerne AI, Rodriguez JJ, Touil T, Brochet B, Schmidt S, Abrous ND, Le Moal M, Pua JR, Jensen MA, Mayo W, Arnason BG, Petry KG

Multiple sclerosis (MS) is characterized by inflammation within the CNS. This inflammatory response is associated with production of nitric oxide (NO) and NO-related species that nitrosylate thiols. We postulated that MS patients would exhibit an antibody (Ab) response directed against proteins containing S-nitrosocysteine (SNO-cysteine) and showed that anti-NO-cysteine Abs of the IgM isotype are in fact present in the sera of some MS patients (Boullerne et al., 1995). We report here the presence of a seemingly identical Ab response directed against SNO-cysteine in an acute model of MS, experimental autoimmune encephalomyelitis (EAE) induced in Lewis rats with the 68-84 peptide of guinea pig myelin basic protein (MBP(68-84)). Serum levels of anti-SNO-cysteine Abs peaked 1 week before the onset of clinical signs and well before the appearance of anti-MBP(68-84) Abs. The anti-SNO-cysteine Ab peak titer correlated with the extent of subsequent CNS demyelination, suggesting a link between Ab level and CNS lesion formation. In relapsing-remitting MS patients, we found elevated anti-SNO-cysteine Ab at times of relapse and normal values in most patients judged to be in remission. Two-thirds of patients with secondary progressive MS had elevated anti-SNO-cysteine Ab levels, including those receiving interferon beta-1b. The data show that a rise in circulating anti-SNO-cysteine Ab levels precedes onset of EAE. Anti-SNO-cysteine Abs are also elevated at times of MS attacks and in progressive disease, suggesting a possible role for these Abs, measurable in blood, as a biological marker for clinical activity.

The cannabinoid receptor type 1 (CB1) displays unusual properties, including the dual capacity to inhibit or stimulate adenylate cyclase and a brain density considerably higher than the majority of G protein-coupled receptors. Together with overlapping expression patterns of dopamine and serotonin receptors this suggests a potential of CB1 to modulate the function of the dopamine and serotonin system. Indeed, pharmacological studies provide evidence for cross-talks between CB1 and receptors of these neurotransmitter systems. In trying to obtain further insights into possible functional and/or structural interactions between CB1 and the dopamine receptors and the serotonin receptors, we performed double-label in situ hybridization at the cellular level on mouse forebrain sections by combining a digoxigenin-labelled riboprobe for CB1 with 35S-labelled riboprobes for dopamine receptors D1 and D2, and for serotonin receptors 5-HT1B and 5-HT3, respectively. As a general rule, we found that CB1 colocalizes with D1, D2 and 5-HT1B only in low-CB1-expressing cells which are principal projecting neurons, whereas CB1 coexpression with 5-HT3 was also observed in high-CB1-expressing cells which are considered to be mostly GABAergic. In striatum and olfactory tubercle, CB1 is coexpressed to a high extent with D1, D2 and 5-HT1B. Throughout the hippocampal formation, CB1 is coexpressed with D2, 5-HT1B and 5-HT3. In the neocortex, coexpression was detected only with 5-HT1B and 5-HT3. In summary a distinct pattern is emerging for the cannabinoid system with regard to its colocalization with dopamine and serotonin receptors and, therefore, it is likely that different mechanisms underlie its cross-talk with these neurotransmitter systems.

01/10/2001 | J Neurosci
Synapse-associated protein 97 selectively associates with a subset of AMPA receptors early in their biosynthetic pathway.
Sans N, Racca C, Petralia RS, Wang YX, McCallum J, Wenthold RJ

The regulation of AMPA receptors at the postsynaptic membrane is a fundamental component of synaptic plasticity. In the hippocampus, the induction of long-term potentiation increases the delivery of GluR1, a major AMPA receptor subunit in hippocampal pyramidal neurons, to the synaptic plasma membrane through a mechanism that requires the PDZ binding domain of GluR1. Synapse-associated protein 97 (SAP97), a member of the membrane-associated guanylate kinase family, is believed to associate with AMPA receptors (AMPARs) containing the GluR1 subunit, but the functional significance of these interactions is unclear. We investigated the interaction of GluR1 with SAP97, the only PDZ protein known to interact with GluR1. We find that interactions involving SAP97 and GluR1 occur early in the secretory pathway, while the receptors are in the endoplasmic reticulum or cis-Golgi. In contrast, few synaptic receptors associate with SAP97, suggesting that SAP97 dissociates from the receptor complex at the plasma membrane. We also show that internalization of GluR1, as triggered by NMDAR activation, does not require SAP97. These results implicate GluR1-SAP97 interactions in mechanisms underlying AMPA receptor targeting.

RATIONALE: The endogenous cannabinoid system is thought to play a role in reinforcement processes. OBJECTIVES: We tested the effects of five doses of the cannabinoid receptor 1 (CB1) antagonist SR141716 [0, 0.3, 1, 3 and 10 mg/kg intraperitoneal (IP)] on intracranial self-stimulation at the level of the median forebrain bundle (MFB). Self-stimulation was assessed 30 min and 210 min after SR141716 administration. We compared the effect of SR141716 with the effect of a decrease in the magnitude of stimulation (-100 microA) and the effects of a cocaine injection (1, 5 and 10 mg/kg IP). METHODS: a protocol of rate-frequency curve for self-stimulation was applied. Two rate-frequency curves were established daily, 3 h apart. The frequency required to produce half-maximal performance (M50) and the maximal performance (RMax) were used as the parameters to characterize the rate-frequency functions. RESULTS: SR141716 decreased the sensitivity to the electrical brain stimulation. SR141716 induced a shift to the right of the rate-frequency curve. This effect depended on the dose administered and the time after injection. Thirty minutes after the injection, 1, 3 and 10 mg/kg SR141716 induced a significant decrease in sensitivity to electrical stimulation, as shown by an elevation in the M50 value. RMax showed a tendency to decrease with increasing doses. At 210 min after administration, 3 and 10 mg/kg SR141716 maintained their decreasing effect on the sensitivity to the stimulation as shown by the significant increase of the M50, however, the maximal response was restored to the basal value. A decrease in self-stimulation intensity produced an effect comparable to the one observed 30 min after either 3 or 10 mg/kg SR141716, while cocaine (5 and 10 mg/kg) produced the opposite effect. Neither condition affected the rate-frequency curve measured 3 h later. CONCLUSIONS: In accordance with recent observations, these experiments suggest that the endogenous cannabinoid system facilitates the perception or the effects of positive reinforcers. They also suggest that this neurochemical system could be a target of interest for treating psychopathologies implicating the reinforcing system.

Neurosteroids are a subclass of steroids that can be synthesized in the central nervous system independently of peripheral sources. Several neurosteroids influence cognitive functions. Indeed, in senescent animals we have previously demonstrated a significant correlation between the cerebral concentration of pregnenolone sulfate (PREG-S) and cognitive performance. Indeed, rats with memory impairments exhibited low PREG-S concentrations compared to animals with correct memory performance. Furthermore, these memory deficits can be reversed by intracerebral infusions of PREG-S. Neurotransmitter systems modulated by this neurosteroid were unknown until our recent report of an enhancement of acetylcholine (ACh) release in basolateral amygdala, cortex, and hippocampus induced by central administrations of PREG-S. Central ACh neurotransmission is involved in the regulation of memory processes and is affected in normal aging and in human neurodegenerative pathologies like Alzheimer's disease. ACh neurotransmission is also involved in the modulation of sleep-wakefulness cycle and relationships between paradoxical sleep and memory are well documented in the literature. PREG-S infused at the level of ACh cell bodies induces a dramatic increase of paradoxical sleep in young animals. Cognitive dysfunctions, particularly those observed in Alzheimer's disease, have also been related to alterations of cerebral plasticity. Among these mechanisms, neurogenesis has been recently studied. Preliminary data suggest that PREG-S central infusions dramatically increase neurogenesis. Taken together these data suggest that PREG-S can influence cognitive processes, particularly in senescent subjects, through a modulation of ACh neurotransmission associated with paradoxical sleep modifications; furthermore our recent data suggest a role for neurosteroids in the modulation of hippocampal neurogenesis.

Balance epithelia in birds closely resemble their mammalian counterparts, but their cells turnover rapidly and they quickly regenerate hair cells, leading to functional recovery from damage that would be permanent for a mammal. We isolated and cultured sheets of the chicken's utricular epithelium in bromo-deoxyuridine and specific inhibitors of different intracellular signalling pathways to identify signals that influence turnover and regeneration. Synthesis (S-phase) entry was effectively blocked by inhibition of PI3-K, TOR or MAPK, and significantly decreased by inhibitors of PKC. Comparisons indicate that activated PI3-K and TOR are required for S-phase entry in both avian and mammalian balance epithelia, but activation of the MAPK pathway appears to have a more significant role in avian utricles than in mammals. The dissimilarities in the requirements for these signalling pathways do not appear sufficient to explain the marked difference in regenerative capacity between the ears of birds and mammals.

Apical dendrites of layer V cortical pyramidal neurons are a major target for glutamatergic synaptic inputs from cortical and subcortical brain regions. Because innervation from these regions is somewhat laminar along the dendrites, knowing the distribution of glutamate receptors on the apical dendrites is of prime importance for understanding the function of neural circuits in the neocortex. To examine this issue, we used infrared-guided laser stimulation combined with whole cell recordings to quantify the spatial distribution of glutamate receptors along the apical dendrites of layer V pyramidal neurons. Focally applied (<10 microm) flash photolysis of caged glutamate on the soma and along the apical dendrite revealed a highly nonuniform distribution of glutamate responsivity. Up to four membrane areas (extent 22 microm) of enhanced glutamate responsivity (hot spots) were detected on the dendrites with the amplitude and integral of glutamate-evoked responses at hot spots being three times larger than responses evoked at neighboring sites. We found no association of these physiological hot spots with dendritic branch points. It appeared that the larger responses evoked at hot spots resulted from an increase in activation of both alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) and N-methyl-D-aspartate (NMDA) receptors and not a recruitment of voltage-activated sodium or calcium conductances. Stimulation of hot spots did, however, facilitate the triggering of both Na+ spikes and Ca(2+) spikes, suggesting that hot spots may serve as dendritic initiation zones for regenerative spikes.

15/08/2001 | J Neurosci
Alleviation of a selective age-related relational memory deficit in mice by
Etchamendy N, Enderlin V, Marighetto A, Vouimba RM, Pallet V, Jaffard R, Higueret P

Vitamin A and its derivatives, the retinoids, have been implicated recently in

15/08/2001 | Nucleic Acids Res
SOX6 binds CtBP2 to repress transcription from the Fgf-3 promoter.
Murakami A, Ishida S, Thurlow J, Revest JM, Dickson C

Fgf-3 is expressed in a complex pattern during mouse development. Previously, an essential regulatory element PS4A was identified in the promoter region, and shown to bind at least three factors. To identify the transcription factor(s), we used a yeast one-hybrid screen and obtained a novel Sox6 cDNA (SOX6D). When introduced into cells it strongly repressed activity from both an Fgf-3 reporter gene as well as an artificial promoter containing three PS4A elements. In situ hybridisation analysis showed that Sox6 and Fgf-3 are co-expressed in the otic vesicle of E9.5 mouse embryos in a mutually exclusive pattern, consistent with a repression of Fgf-3 transcription by SOX6. To characterise additional factor(s) involved in Fgf-3 gene repression, a yeast two-hybrid screen was used with the N-terminal portion of SOX6D. Mouse CtBP2 cDNA clones were isolated and shown to bind SOX6 in yeast and mammalian cells. Furthermore, mutational analysis of SOX6 showed that binding to CtBP2, and its responsiveness to this co-repressor, were dependent on a short amino acid sequence motif PLNLSS. Co-expression studies in NIH3T3 cells showed that SOX6 and CtBP2 co-operate to repress activity from the Fgf-3 promoter through the enhancer element PS4A. These results show that SOX6 can recruit CtBP2 to repress transcription from the Fgf-3 promoter.

15/08/2001 | J Immunol
Development of the thymus requires signaling through the fibroblast growth factor receptor R2-IIIb.
Revest JM, Suniara RK, Kerr K, Owen JJ, Dickson C

Mice deficient for fibroblast growth factor (Fgf)R2-IIIb show a block in thymic growth after embryonic day 12.5, a stage that just precedes its detection in thymic epithelial cells. Fgf7 and Fgf10, the main ligands for FgfR2-IIIb, are expressed in the mesenchyme surrounding the thymic epithelial primordium, and Fgf10-deficient mice also exhibit impaired thymic growth. Hence, Fgf signaling is essential for thymic epithelial proliferation. In addition to the proliferative block, most thymic epithelial cells fail to progress from an immature cytokeratin 5-positive to a cytokeratin 5-negative phenotype. Nevertheless, sufficient epithelial cell differentiation occurs in the severely hypoplastic thymus to allow the development of CD4/CD8-double-positive thymocytes and a very small number of single-positive thymocytes expressing TCRs.

06/2001 | J Clin Endocrinol Metab
Normal human pituitary gland and pituitary adenomas express cannabinoid receptor type 1 and synthesize endogenous cannabinoids: first evidence for a direct role of cannabinoids on hormone modulation at the human pituitary level.
Pagotto U, Marsicano G, Fezza F, Theodoropoulou M, Grubler Y, Stalla J, Arzberger T, Milone A, Losa M, Di Marzo V, Lutz B, Stalla GK

Little is known about the expression and function of cannabinoid receptor type 1 (CB1) in the human pituitary gland. The aim of this study was to investigate CB1 expression in human normal and tumoral pituitaries by in situ hybridization and immunohistochemistry using an antibody against CB1. CB1 was found in corticotrophs, mammotrophs, somatotrophs, and folliculostellate cells in the anterior lobe of normal pituitary. After examination of 42 pituitary adenomas, CB1 was detected in acromegaly-associated pituitary adenomas, Cushing's adenomas, and prolactinomas, whereas faint or no expression was found in nonfunctioning pituitary adenomas. Experiments with cultured pituitary adenoma cells showed that the CB1 agonist WIN 55,212--2 inhibited GH secretion in most of acromegaly-associated pituitary adenomas tested and that the CB1 antagonist SR 141716A was generally able to reverse this effect. Moreover, WIN 55,212--2 was able to suppress GHRH-stimulated GH release, and this effect was not blocked by coincubation with SR 141716A, possibly indicating a non-CB1-mediated effect. In contrast, WIN 55,212--2 was ineffective on GH-releasing peptide-stimulated GH release. In four Cushing's adenomas tested, WIN 55,212--2 was not able to modify basal ACTH secretion. However, simultaneous application of CRF and WIN 55,212--2 resulted in a synergistic effect on ACTH secretion, and this effect could be abolished by SR 141716A, demonstrating a CB1-mediated effect. In the single case of prolactinomas tested, WIN 55,212--2 was able to inhibit basal secretion of PRL. Finally, the presence of endocannabinoids (anandamide and 2-arachidonoylglycerol) was investigated in normal and tumoral pituitaries. All tumoral samples had higher contents of anandamide and 2-arachidonoylglycerol compared with the normal hypophysis. Moreover, endocannabinoid content in the different pituitary adenomas correlated with the presence of CB1, being elevated in the tumoral samples positive for CB1 and lower in the samples in which no or low levels of CB1 were found. The results of this study point to a direct role of cannabinoids in the regulation of human pituitary hormone secretion.

Analysis of excitatory synaptic transmission in the rat hypothalamic supraoptic nucleus revealed that glutamate clearance and, as a consequence, glutamate concentration and diffusion in the extracellular space, is associated with the degree of astrocytic coverage of its neurons. Reduction in glutamate clearance, whether induced pharmacologically or associated with a relative decrease of glial coverage in the vicinity of synapses, affected transmitter release through modulation of presynaptic metabotropic glutamate receptors. Astrocytic wrapping of neurons, therefore, contributes to the regulation of synaptic efficacy in the central nervous system.

05/2001 | Eur J Neurosci
Glutamate receptor targeting in the postsynaptic spine involves mechanisms that are independent of myosin Va.
Petralia RS, Wang YX, Sans N, Worley PF, Hammer JA 3rd, Wenthold RJ

Targeting of glutamate receptors (GluRs) to synapses involves rapid movement of intracellular receptors. This occurs in forms of synaptic upregulation of receptors, such as long-term potentiation. Thus, many GluRs are retained in a cytoplasmic pool in dendrites, and are transported to synapses for upregulation, presumably via motor proteins such as myosins travelling along cytoskeletal elements that extend up into the spine. In this ultrastructural immunogold study of the cerebellar cortex, we compared synapses between normal rats/mice and dilute lethal mutant mice. These mutant mice lack myosin Va, which has been implicated in protein trafficking at synapses. The postsynaptic spine in the cerebellum lacks the inositol trisphosphate receptor (IP3R) -laden reticular tubules that are found in normal mice and rats (Takagishi et al., Neurosci. Lett., 1996, 215, 169). Thus, we tested the hypothesis that myosin Va is necessary for transport of GluRs and associated proteins to spine synapses. We found that these spines retain a normal distribution of (i) GluRs (delta 1/2, GluR2/3 and mGluR1alpha), (ii) at least one associated MAGUK (membrane-associated guanylate kinase) protein, (iii) Homer (which interacts with mGluR1alpha and IP3Rs), (iv) the actin cytoskeleton, (v) the reticulum-associated protein BiP, and (vi) the motor-associated protein, dynein light chain. Thus, while myosin Va may maintain the IP3R-laden reticulum in the spine for proper calcium regulation, other mechanisms must be involved in the delivery of GluRs and associated proteins to synapses. Other possible mechanisms include diffusion along the extrasynaptic membrane and delivery via other motors running along the spine's actin cytoskeleton.

05/03/2001 | Brain Res Mol Brain Res
Evidence for an endocannabinoid system in the central nervous system of the leech Hirudo medicinalis.
Matias I , Bisogno T , Melck D , Vandenbulcke F , Verger-Bocquet M , De Petrocellis L , Sergheraert C , Breton C , Di Marzo V , Salzet M

In invertebrates, like Hydra and sea urchins, evidence for a functional cannabinoid system was described. The partial characterization of a putative CB1 cannabinoid receptor in the leech Hirudo medicinalis led us to investigate the presence of a complete endogenous cannabinoid system in this organism. By using gas chromatography-mass spectrometry, we demonstrate the presence of the endocannabinoids anandamide (N-arachidonoylethanolamine, 21.5+/-0.7 pmol/g) and 2-arachidonoyl-glycerol (147.4+/-42.7 pmol/g), and of the biosynthetic precursor of anandamide, N-arachidonylphosphatidyl-ethanolamine (16.5+/-3.3 pmol/g), in the leech central nervous system (CNS). Anandamide-related molecules such as N-palmitoylethanolamine (32.4+/-1.6 pmol/g) and N-linolenoylethanolamine (5.8 pmol/g) were also detected. We also found an anandamide amidase activity in the leech CNS cytosolic fraction with a maximal activity at pH 7 and little sensitivity to typical fatty acid amide hydrolase (FAAH) inhibitors. Using an antiserum directed against the amidase signature sequence, we focused on the identification and the localization of the leech amidase. Firstly, leech nervous system protein extract was subjected to Western blot analysis, which showed three immunoreactive bands at ca. approximately 42, approximately 46 and approximately 66 kDa. The former and latter bands were very faint and were also detected in whole homogenates from the coelenterate Hydra vulgaris, where the presence of CB1-like receptors, endocannabinoids and a FAAH-like activity was reported previously. Secondly, amidase immunocytochemical detection revealed numerous immunoreactive neurons in the CNS of three species of leeches. In addition, we observed that leech amidase-like immunoreactivity matches to a certain extent with CB1-like immunoreactivity. Finally, we also found that stimulation by anandamide of this receptor leads, as in mammals, to inhibition of cAMP formation, although this effect appeared to be occurring through the previously described anandamide-induced and CB1-mediated activation of nitric oxide release. Taken together, these results suggest the existence of a complete and functional cannabinoid system in leeches.