[The possible contribution of community pharmacist upstream of drug prescription in France: An experiment of practice groups between general practitioners and community pharmacists]. / La possible contribution du pharmacien d'officine en amont de la prescription médicamenteuse en France : une expérimentation de groupes de pratique médecins généralistes ­ pharmaciens d'officine.

OBJECTIVES: The implementation of practice groups between general practitioners and community pharmacists in several European countries (Belgium, the Netherlands, Switzerland) emphasizes the possible and relevant role of the community pharmacist upstream of drug prescription. In these groups, the pharmacist provides knowledge and faciltates pluriprofessional exchanges on prescription practices. This research assesses the potential of implementing these practice groups in France. METHODS: An experiment was set up in France at 9 sites. Its evaluation was based on the use of questionnaires and semi-structured interviews. The operational feasibility and the adherence of pharmacists to these practice groups were assessed. RESULTS: Our results emphasize that the integration of this practice into the pharmacist's activity is possible but encounters limits, particularly in terms of time investment by the meeting leader pharmacists, regardless of their professional status. The satisfaction of the participating pharmacists with this practice, as well as the previous characteristics of their activity, such as an interprofessional practice already established, are positive factors for the adherence of these professionals to this practice. CONCLUSIONS: If these practice groups can represent a potential new activity for the community pharmacist, several conditions could facilitate their implementation: the possibility of animation in pairs, pharmacist-physician, in order to accentuate the interprofessional collaboration while mobilizing the skills of community pharmacists, an evolution of the initial training of community pharmacists in order to strengthen the skills required to facilitate these groups, as well as a financial or professional recognition in terms of continuing education.

Neuromyelitis optica study model based on chronic infusion of autoantibodies in rat cerebrospinal fluid.

BACKGROUND: Devic's neuromyelitis optica (NMO) is an autoimmune astrocytopathy, associated with central nervous system inflammation, demyelination, and neuronal injury. Several studies confirmed that autoantibodies directed against aquaporin-4 (AQP4-IgG) are relevant in the pathogenesis of NMO, mainly through complement-dependent toxicity leading to astrocyte death. However, the effect of the autoantibody per se and the exact role of intrathecal AQP4-IgG are still controversial. METHODS: To explore the intrinsic effect of intrathecal AQP4-IgG, independent from additional inflammatory effector mechanisms, and to evaluate its clinical impact, we developed a new animal model, based on a prolonged infusion of purified immunoglobulins from NMO patient (IgG(AQP4+), NMO-rat) and healthy individual as control (Control-rat) in the cerebrospinal fluid (CSF) of live rats. RESULTS: We showed that CSF infusion of purified immunoglobulins led to diffusion in the brain, spinal cord, and optic nerves, the targeted structures in NMO. This was associated with astrocyte alteration in NMO-rats characterized by loss of aquaporin-4 expression in the spinal cord and the optic nerves compared to the Control-rats (p = 0.001 and p = 0.02, respectively). In addition, glutamate uptake tested on vigil rats was dramatically reduced in NMO-rats (p = 0.001) suggesting that astrocytopathy occurred in response to AQP4-IgG diffusion. In parallel, myelin was altered, as shown by the decrease of myelin basic protein staining by up to 46 and 22 % in the gray and white matter of the NMO-rats spinal cord, respectively (p = 0.03). Loss of neurofilament positive axons in NMO-rats (p = 0.003) revealed alteration of axonal integrity. Then, we investigated the clinical consequences of such alterations on the motor behavior of the NMO-rats. In a rotarod test, NMO-rats performance was lower compared to the controls (p = 0.0182). AQP4 expression, and myelin and axonal integrity were preserved in AQP4-IgG-depleted condition. We did not find a major immune cell infiltration and microglial activation nor complement deposition in the central nervous system, in our model. CONCLUSIONS: We establish a link between motor-deficit, NMO-like lesions and astrocytopathy mediated by intrathecal AQP4-IgG. Our study validates the concept of the intrinsic effect of autoantibody against surface antigens and offers a model for testing antibody and astrocyte-targeted therapies in NMO.

Animal models of REM dysfunctions: what they tell us about the cause of narcolepsy and RBD?

Rapid eye movement sleep behavior disorder (RBD) is a parasomnia characterized by the loss of muscle atonia during paradoxical (REM) sleep (PS). Conversely, cataplexy, one of the key symptoms of narcolepsy, is a striking sudden episode of muscle weakness triggered by emotions during wakefulness, and comparable to REM sleep atonia. The neuronal dysfunctions responsible for RBD and cataplexy are not known. In the present review, we present the most recent results on the neuronal network responsible for PS. Based on these results, we propose an updated integrated model of the mechanisms responsible for PS and explore different hypotheses explaining RBD and cataplexy. We propose that RBD is due to a specific degeneration of a subpopulation of PS-on glutamatergic neurons specifically responsible of muscle atonia, localized in the caudal pontine sublaterodorsal tegmental nucleus (SLD). Another possibility is the occurrence in RBD patients of a specific lesion of the glycinergic/GABAergic premotor-neurons localized in the medullary ventral gigantocellular reticular nucleus. Conversely, cataplexy in narcoleptics would be due to the activation during waking of the caudal PS-on SLD neurons responsible for muscle atonia. A direct or indirect pathway activated during positive emotion from the central amygdala to the SLD PS-on neurons would induce such activation. In normal conditions, the activation of SLD neurons would be blocked by the simultaneous excitation by the hypocretins of the PS-off GABAergic neurons localized in the ventrolateral periaqueductal gray and the adjacent deep mesencephalic reticular nucleus gating the activation of the PS-on SLD neurons.

A mutation in a case of early onset narcolepsy and a generalized absence of hypocretin peptides in human narcoleptic brains.

We explored the role of hypocretins in human narcolepsy through histopathology of six narcolepsy brains and mutation screening of Hcrt, Hcrtr1 and Hcrtr2 in 74 patients of various human leukocyte antigen and family history status. One Hcrt mutation, impairing peptide trafficking and processing, was found in a single case with early onset narcolepsy. In situ hybridization of the perifornical area and peptide radioimmunoassays indicated global loss of hypocretins, without gliosis or signs of inflammation in all human cases examined. Although hypocretin loci do not contribute significantly to genetic predisposition, most cases of human narcolepsy are associated with a deficient hypocretin system.

A circuit perspective on narcolepsy.

The sleep disorder narcolepsy is associated with symptoms related to either boundary state control that include excessive daytime sleepiness and sleep fragmentation, or rapid eye movement (REM) sleep features including cataplexy, sleep paralysis, hallucinations, and sleep-onset REM sleep events (SOREMs). Although the loss of Hypocretin/Orexin (Hcrt/Ox) peptides or their receptors have been associated with the disease, here we propose a circuit perspective of the pathophysiological mechanisms of these narcolepsy symptoms that encompasses brain regions, neuronal circuits, cell types, and transmitters beyond the Hcrt/Ox system. We further discuss future experimental strategies to investigate brain-wide mechanisms of narcolepsy that will be essential for a better understanding and treatment of the disease.

Secondary findings from whole-exome/genome sequencing evaluating stakeholder perspectives. A review of the literature.

With the development of next generation sequencing, beyond identifying the cause of manifestations that justified prescription of the test, other information with potential interest for patients and their families, defined as secondary findings (SF), can be provided once patients have given informed consent, in particular when therapeutic and preventive options are available. The disclosure of such findings has caused much debate. The aim of this work was to summarize all opinion-based studies focusing on SF, so as to shed light on the concerns that this question generate. A review of the literature was performed, focusing on all PubMed articles reporting qualitative, quantitative or mixed studies that interviewed healthcare providers, participants, or society regarding this subject. The methodology was carefully analysed, in particular whether or not studies made the distinction between actionable and non-actionable SF, in a clinical or research context. From 2010 to 2016, 39 articles were compiled. A total of 14,868 people were interviewed (1259 participants, 6104 healthcare providers, 7505 representatives of society). When actionable and non-actionable SF were distinguished (20 articles), 92% of respondents were keen to have results regarding actionable SF (participants: 88%, healthcare providers: 86%, society: 97%), against 70% (participants: 83%, healthcare providers: 62%, society: 73%) for non-actionable SF. These percentages were slightly lower in the specific situation of children probands. For respondents, the notion of the «patient's choice¼ is crucial. For healthcare providers, the importance of defining policies for SF among diagnostic lab, learning societies and/or countries is outlined, in particular regarding the content and extension of the list of actionable genes to propose, the modalities of information, and the access to information about adult-onset diseases in minors. However, the existing literature should be taken with caution, since most articles lack a clear definition of SF and actionability, and referred to hypothetical scenarios with limited information to respondents. Studies conducted by multidisciplinary teams involving patients with access to results are sadly lacking, in particular in the medium term after the results have been given. Such studies would feed the debate and make it possible to measure the impact of such findings and their benefit-risk ratio.

Excess of infections due to a multi-drug sensitive Salmonella enterica serotype Typhimurium in France in June 2008.

An unusually high number of cases of Salmonella Typhimurium was reported in France in June 2008. In the course of epidemiological investigations 112 cases were ascertained, of whom 75 were interviewed. Subtyping by PFGE and MLVA identified a strain named "majority profile". Subtyping results were available for 45 interviewed cases, 30 of whom (majority below 15 years of age) were found to be infected with the majority profile strain. Evidence suggested the occurrence of an outbreak due to a monoclonal S. Typhimurium strain with the single PFGE profile XTYM-50. Cases with identical PFGE profile were also detected in Switzerland but no link with outbreaks occurring in the same period in Denmark and in the Netherlands was found. Contamination of a product distributed nationally was suggested as the cause of the outbreak but investigations did not reveal any specific food source.

Endoscopic and surgical removal of oesophageal and gastric fishhook foreign bodies in 33 animals.

OBJECTIVES: To describe the location, retrieval, frequency of surgery and complications associated with fishhook foreign bodies. MATERIALS AND METHODS: Retrospective evaluation of the medical records of cats and dogs admitted between 2010 and 2016 after fishhook ingestion. RESULTS: A total of 33 cases (2 cats and 31 dogs) were included. The most common locations were the proximal oesophagus [12/33 (36%)] and stomach [11/33 (33%)]. Endoscopic retrieval was successful in 27 of 33 cases (82%); oesophageal perforation was the only recorded complication, occurring in six of 33 (18%) cases. Surgery was performed in six cases (18%), and no early complications were recorded. The survival rate was 100%. CLINICAL SIGNIFICANCE: The endoscopic removal of ingested fishhooks is highly successful. In the present study, survival to discharge was 100%, even in cases of oesophageal perforation or in cases requiring surgery.

[Changes in clinical practice related to the arrival of next-generation sequencing in the genetic diagnosis of developmental diseases]. / Les modifications de pratique clinique liées à l'arrivée du séquençage haut débit dans le diagnostic génétique des maladies du développement.

INTRODUCTION: The arrival of high-throughput sequencing (HTS) has led to a sweeping change in the diagnosis of developmental abnormalities (DA) with or without intellectual deficiency (ID). With the prospect of deploying these new technologies, two questions have been raised: the representations of HTS among geneticists and the costs incurred due to these analyses. METHODS: Geneticists attending a clinical genetics seminar were invited to complete a questionnaire. The statistical analysis was essentially descriptive and an analysis of costs was undertaken. RESULTS: Of those responding to the questionnaire, 48% had already prescribed exome analysis and 25% had already had the occasion to disclose the results of such analyses. Ninety-six percent were aware that whole-exome sequencing (WES) had certain limits and 74% expressed misgivings concerning its use in medical practice. In parallel, the evaluation of costs showed that WES was less expensive than conventional procedures. DISCUSSION: The survey revealed that geneticists had already come to terms with HTS as early as 2015. Among the major concerns expressed were the complexity of interpreting these tests and the many ethical implications. Geneticists seemed to be aware of the advantages but also the limits of these new technologies. The cost analysis raises questions about the place of HTS and in particular WES in the diagnostic work-up: should it be used early to obtain an etiological diagnosis rather than as the last resort? CONCLUSION: It is essential for future generations of doctors and for the families concerned to learn about the concepts of HTS, which is set to become a major feature of new genomic medicine.

The hypocretin/orexin ligand-receptor system: implications for sleep and sleep disorders.

The molecules originally described as the hypocretins and subsequently as the orexins were initially implicated in the control of food intake. Recent observations implicate this newly-described neurotransmitter system in the sleep disorder narcolepsy and, potentially, in the regulation of normal sleep processes. This article reviews the research that led to the isolation of the hypocretin/orexin peptides, their receptors and the activity of these molecules as we currently understand them. A model is proposed in which the cells that make these peptides might be involved in arousal state control.

Gene expression in the rat brain during sleep deprivation and recovery sleep: an Affymetrix GeneChip study.

Previous studies have demonstrated that macromolecular synthesis in the brain is modulated in association with the occurrence of sleep and wakefulness. Similarly, the spectral composition of electroencephalographic activity that occurs during sleep is dependent on the duration of prior wakefulness. Since this homeostatic relationship between wake and sleep is highly conserved across mammalian species, genes that are truly involved in the electroencephalographic response to sleep deprivation might be expected to be conserved across mammalian species. Therefore, in the rat cerebral cortex, we have studied the effects of sleep deprivation on the expression of immediate early gene and heat shock protein mRNAs previously shown to be upregulated in the mouse brain in sleep deprivation and in recovery sleep after sleep deprivation. We find that the molecular response to sleep deprivation and recovery sleep in the brain is highly conserved between these two mammalian species, at least in terms of expression of immediate early gene and heat shock protein family members. Using Affymetrix Neurobiology U34 GeneChips , we also screened the rat cerebral cortex, basal forebrain, and hypothalamus for other genes whose expression may be modulated by sleep deprivation or recovery sleep. We find that the response of the basal forebrain to sleep deprivation is more similar to that of the cerebral cortex than to the hypothalamus. Together, these results suggest that sleep-dependent changes in gene expression in the cerebral cortex are similar across rodent species and therefore may underlie sleep history-dependent changes in sleep electroencephalographic activity.

Sacroiliac joints imaging in axial spondyloarthritis.

Imaging of the sacroiliac joints is the key point in diagnosing and classifying spondyloarthritis. Since the integration of MRI criteria to the Assessment of Spondyloarhtitis Society (ASAS) classification in 2009, the attention was focused on the presence of bone marrow edema to characterize sacroiliitis. However, returning to basics and analysing structural signs is of utmost importance to avoid overdiagnosis of spondyloarthritis.

Role and origin of the GABAergic innervation of dorsal raphe serotonergic neurons.

Extracellular electrophysiological recordings in freely moving cats have shown that serotonergic neurons from the dorsal raphe nucleus (DRN) fire tonically during wakefulness, decrease their activity during slow wave sleep (SWS), and are nearly quiescent during paradoxical sleep (PS). The mechanisms at the origin of the modulation of activity of these neurons are still unknown. Here, we show in the unanesthetized rat that the iontophoretic application of the GABA(A) antagonist bicuculline on dorsal raphe serotonergic neurons induces a tonic discharge during SWS and PS and an increase of discharge rate during quiet waking. These data strongly suggest that an increase of a GABAergic inhibitory tone present during wakefulness is responsible for the decrease of activity of the dorsal raphe serotonergic cells during slow wave and paradoxical sleep. In addition, by combining retrograde tracing with cholera toxin B subunit and glutamic acid decarboxylase immunohistochemistry, we demonstrate that the GABAergic innervation of the dorsal raphe nucleus arises from multiple distant sources and not only from interneurons as classically accepted. Among these afferents, GABAergic neurons located in the lateral preoptic area and the pontine ventral periaqueductal gray including the DRN itself could be responsible for the reduction of activity of the serotonergic neurons of the dorsal raphe nucleus during slow wave and paradoxical sleep, respectively.

Lower brainstem catecholamine afferents to the rat dorsal raphe nucleus.

A large body of data suggests that the activation of alpha 1 receptors by a tonic noradrenergic input might be responsible for the tonic discharge of the serotonergic neurons of the dorsal raphe nucleus (DRN). To test this hypothesis, it was necessary to determine the origin of the noradrenergic and adrenergic innervation of these neurons. For this purpose, we combined small iontophoretic injections of the sensitive retrograde tracer cholera toxin b subunit (CTb) in the different subdivisions of the DRN with tyrosine hydroxylase immunohistochemistry. After CTb injections in the ventral or dorsal parts of the central DRN, a small number of double-labeled cells was observed in the locus coeruleus (A6 noradrenergic cell group), the A5 noradrenergic group, the dorsomedial medulla (C3 adrenergic cell group), and the lateral paragigantocellular nucleus (C1 adrenergic cell group). After CTb injections in the lateral wings or the dorsal part of the rostral DRN, a similar number of double-labeled cells was seen in C3. Slightly more double-labeled cells were seen in A6 and A5. In addition, a substantial to large number of double-labeled cells appeared in C1, the commissural part of the nucleus of the solitary tract (A2 noradrenergic cell group) and the caudoventrolateral medulla (A1 noradrenergic cell group). These results indicate that the noradrenergic and adrenergic inputs to the DRN arise from all the catecholaminergic cell groups of the lower brainstem except the A7 noradrenergic group. They further reveal the existence of a topographical organization of these afferents to the different subdivisions of the DRN.

Specific potentialities of embryonic rat serotonergic neurons to innervate different periventricular targets in the adult brain.

During the development of the central nervous system, neurons are directed by both genetic and environmental factors to differentiate and form connections with their targets. We took advantage of the abundant homogeneous serotonergic innervations of the ependyma forming the supra- and subependymal plexuses to investigate possible commitment of embryonic neurons to innervate specific targets during axogenesis in the rat. The origin of the supraependymal innervation was determined by retrograde transport of cholera toxin (CT) from the ventricles. The supraependymal plexuses of the fourth ventricle mainly originated from neurons in the dorsocaudal region of the raphe dorsalis (DRN), while the rostral DRN and raphe centralis (CRN) contained perikarya projecting into the third ventricle. This suggested the existence, along the rostrocaudal axis of the raphe, of different neuronal subsets able to form distinct supraependymal plexuses in the third or fourth ventricle. To determine whether serotonergic neurons were committed to innervate specific areas of the ependyma, different embryonic metencephalic segments (rostral, median, or caudal) from 14-day-old rat embryos were independently grafted into the third or fourth ventricle of an adult brain in which the serotonergic neurons had been previously destroyed. The distinctive patterns of re-innervation specific to each of grafted segments indicate that subsets of embryonic serotonergic neurons are indeed committed to innervate certain restricted ependymal areas of the adult brain, presumably in response to different neurotropic and/or neurotrophic cues.

Hypocretin (orexin) activation and synaptic innervation of the locus coeruleus noradrenergic system.

Hypocretin has been identified as a regulator of metabolic and endocrine systems. Several brain regions involved in the central regulation of autonomic and endocrine processes or attention are targets of extensive hypocretin projections. The most dense arborization of hypocretin axons in the brainstem was detected in the locus coeruleus (LC). Multiple labeling immunocytochemistry revealed a massive synaptic innervation of catecholaminergic LC cells by hypocretin axon terminals in rats and monkeys. In both species, all tyrosine hydroxylase-immunopositive cells in the LC examined by electron microscopy were found to receive asymmetrical (excitatory) synaptic contacts from multiple axons containing hypocretin. In parallel electrophysiological studies with slices of rat brain, all LC cells showed excitatory responses to the hypocretin-2 peptide. Hypocretin-2 uniformly increased the frequency of action potentials in these cells, even in the presence of tetrodotoxin, indicating that receptors responding to hypocretin were expressed in LC neurons. Two mechanisms for the increased firing rate appeared to be a reduction in the slow component of the afterhyperpolarization (AHP) and a modest depolarization. Catecholamine systems in other parts of the brain, including those found in the medulla, zona incerta, substantia nigra or olfactory bulb, received significantly less hypocretin input. Comparative analysis of lateral hypothalamic input to the LC revealed that hypocretin-containing axon terminals were substantially more abundant than those containing melanin-concentrating hormone. The present results provide evidence for direct action of hypothalamic hypocretin cells on the LC noradrenergic system in rats and monkeys. Our observations suggest a signaling pathway via which signals acting on the lateral hypothalamus may influence the activity of the LC and thereby a variety of CNSfunctions related to noradrenergic innervation, including vigilance, attention, learning, and memory. Thus, the hypocretin innervation of the LC may serve to focus cognitive processes to compliment hypocretin-mediated activation of autonomic centers already described.

Distribution of glycine-immunoreactive cell bodies and fibers in the rat brain.

To localize glycinergic cell bodies and fibers in the rat brain, we developed a sensitive immunohistochemical method combining the use of specific glycine antibodies (Campistron G. et al. (1986) Brain Res. 376, 400-405; Wenthold R. J. et al. (1987) Neuroscience 22, 897-912) with the streptavidin-horseradish peroxidase technique and 3,3'-diaminobenzidine.4HCl-nickel intensification. We confirmed the presence of numerous glycine-immunoreactive cell bodies and fibers in the cochlear nuclei, superior olivary complex, nucleus of the trapezoid body, cerebellar cortex, deep cerebellar nuclei and area postrema. For the first time in rats, we described a large to very large number of cell bodies in the medial vestibular ventral part, prepositus hypoglossal, gracile, raphe magnus and sensory trigeminal nuclei. A large number of cells was also observed in the oral and caudal pontine, parvocellular, parvocellular pars alpha, gigantocellular and gigantocellular pars alpha reticular nuclei. In addition, glycine-immunoreactive cells were seen in the ambiguous and subtrigeminal nuclei, the lateral habenula and the subfornical organ. We also provide the first evidence in rats for a very large number of fibers in the trigeminal, facial, ambiguous and hypoglossal motor nuclei, all nuclei of the medullary and pontine reticular formation, and the raphe and trigeminal sensory nuclei. We further revealed the presence of a substantial number of fibers in regions where glycine was not considered as a main inhibitory neurotransmitter, such as the pontine nuclei, the periaqueductal gray, the mesencephalic reticular formation, the anterior pretectal nucleus, the intralaminar thalamic nuclei, the zona incerta, the fields of Forel, the parvocellular parts of the paraventricular nucleus, the posterior hypothalamic areas, the anterior hypothalamic area, and the lateral and medial preoptic areas. These results indicate that, in contrast to previous statements, glycine may be an essential inhibitory neurotransmitter not only in the lower brainstem and spinal cord, but also in the upper brainstem and the forebrain.

Forebrain afferents to the rat dorsal raphe nucleus demonstrated by retrograde and anterograde tracing methods.

The dorsal raphe nucleus through its extensive efferents has been implicated in a great variety of physiological and behavioural functions. However, little is know about its afferents. Therefore, to identify the systems likely to influence the activity of serotonergic neurons of the dorsal raphe nucleus, we re-examined the forebrain afferents to the dorsal raphe nucleus using cholera toxin b subunit and Phaseolus vulgaris-leucoagglutinin as retrograde or anterograde tracers. With small cholera toxin b subunit injection sites, we further determined the specific afferents to the ventral and dorsal parts of the central dorsal raphe nucleus, the rostral dorsal raphe nucleus and the lateral wings. In agreement with previous studies, we observed a large number of retrogradely-labelled cells in the lateral habenula following injections in all subdivisions of the dorsal raphe nucleus. In addition, depending on the subdivision of the dorsal raphe nucleus injected, we observed a small to large number of retrogradely-labelled cells in the orbital, cingulate, infralimbic, dorsal peduncular, and insular cortice, a moderate or substantial number in the ventral pallidum and a small to substantial number in the claustrum. In addition, we observed a substantial to large number of cells in the medial and lateral preoptic areas and the medial preoptic nucleus after cholera toxin b subunit injections in the dorsal raphe nucleus excepting for those located in the ventral part of the central dorsal raphe nucleus, after which we found a moderate number of retrogradely-labelled cells. Following cholera toxin b subunit injections in the dorsal part of the central dorsal raphe nucleus, a large number of retrogradely-labelled cells was seen in the lateral, ventral and medial parts of the bed nucleus of the stria terminalis whereas only a small to moderate number was visualized after injections in the other dorsal raphe nucleus subdivisions. In addition, respectively, a substantial and a moderate number of retrogradely-labelled cells was distributed in the zona incerta and the subincertal nucleus following all tracer injections in the dorsal raphe nucleus. A large number of retrogradely-labelled cells was also visualized in the lateral, dorsal and posterior hypothalamic areas and the perifornical nucleus after cholera toxin b subunit injections in the dorsal part of the central raphe nucleus and to a lesser extent following injections in the other subdivisions. We further observed a substantial to large number of retrogradely-labelled cells in the tuber cinereum and the medial tuberal nucleus following cholera toxin b subunit injections in the dorsal part of the central dorsal raphe nucleus or the lateral wings and a small to moderate number after injections in the two other dorsal raphe nucleus subdivisions. A moderate or substantial number of labelled cells was also seen in the ventromedial hypothalamic area and the arcuate nucleus following cholera toxin injections in the dorsal part of the central dorsal raphe nucleus and the lateral wings and an occasional or small number with injection sites located in the other subdivisions. Finally, we observed, respectively, a moderate and a substantial number of retrogradely-labelled cells in the central nucleus of the amygdala following tracer injections in the ventral or dorsal parts of the central dorsal raphe nucleus and a small number after injections in the other subnuclei. In agreement with these retrograde data, we visualized anterogradely-labelled fibres heterogeneously distributed in the dorsal raphe nucleus following Phaseolus vulgaris-leucoagglutinin injections in the lateral orbital or infralimbic cortice, the lateral preoptic area, the perifornical nucleus, the lateral or posterior hypothalamic areas, the zona incerta, the subincertal nucleus or the medial tuberal nucleus. (ABSTRACT TRUNCATED)

Prepro-hypocretin (prepro-orexin) expression is unaffected by short-term sleep deprivation in rats and mice.

The hypocretin/orexin ligand-receptor system has recently been implicated in the sleep disorder narcolepsy. During the dark (active) period, null mutants of the prepro-orexin (prepro-hypocretin) gene have cataplectic attacks and increased levels of both rapid eye movement (REM) and non-REM (NREM) sleep. Intracerebroventricular injection of one of the encoded neuropeptides, orexin-A, early in the light period increases wakefulness and reduces REM sleep in the rat, suggesting that this system may be involved in the normal regulation of sleep and wakefulness. To further test this hypothesis, we measured hypocretin (hcrt) mRNA levels by both Northern hybridization and Taqman analysis in mouse and rat hypothalamus after short-term (6 h) sleep deprivation (SD) and 2-4 hours after recovery from SD. Although our SD procedures effectively induced a sleep debt and increased c-fos mRNA expression in the cortex and hypothalamus as described by other investigators, we found that hcrt mRNA levels were not significantly changed in either species either after SD or after recovery from SD. If the hcrt system is involved in normal regulation of sleep and wakefulness, longer periods of SD may be necessary to affect hcrt mRNA levels or changes may occur at the protein rather than mRNA level. Alternatively, this system may also be involved in another function that counterbalances any SD-induced changes in hcrt mRNA levels.