VTA-NAc内orexin A在吗啡和性奖赏相关行为中作用的研究

There are a lot of differences in the neural mechanisms underlying between drug reward and natural reward despite the common neual basis. Undoubtedly, revealing the common and the different mechanisms underlying drug reward and natural reward will promote the development of research on drug addiction. Among diversified natural rewards, sex is often compared to drug because sexual reward has more similarities to drug. The mesolimbic dopamine system (VTA-NAc pathway) is a common pathway activated by natural reinforcers and addictive drugs, mediating reward, emotion and motivation under physiological conditions. The neuroadaptations taking place in the central nervous system including the mesolimbic dopamine system after repeatedly drug taking leads to persistent drug craving, Orexin, a neuropeptide produced in the lateral hypothalamus, plays an important role in reward-associated, motivated behaviors. Orexin neurons have extensive projections to the mesolimbic dopamine system. In order to further investigate the roles of orexin A in drug reward, this study examined the regulatory roles of orexin A in the VTA and NAcSh on drug reinforcement (acqusition of morphine CPP) and drug-seeking behavior (expression of morphine CPP). Moreover, the roles of orexin A on drug reward were compared with sexual reward. The main results are as follows: 1. The expression of morphine CPP was inhibited by intracerebroventricularly (i.c.v.) administered OX1R antagonist SB334867; 2. The male unconditioned sexual motivation was not affected by i.c.v. administered SB334867. However, i.c.v. given orexin A inhibited unconditioned sexual motivation in sexually high-motivated rats but did not affect sexual motivation in low-motivated rats; 3. The acquisition and expression of morphine CPP was inhibited by SB334867 microinjected into the VTA. SB334867 or orexin A injected into the NAcSh did not influence the acquisition of morphine CPP, but orexin A increased the locomotor activity in rats treated with morphine (3mg/kg); 4. SB334867 microinjected into the VTA did not affect male copulatory behavior, neither affect the acqusition of copulatory CPP; 5. The expression of copulatory CPP was associated with increased Fos protein expression in hypothalamic orexin A neurons, and SB334867 microinjected into the VTA inhibited expression of copulatory CPP. These results suggest that, (1) endogenous orexin A is not involved in male unconditioned sexual motivation, but involved in drug craving; (2) orexin A in the VTA instead of in the NAc is involved in drug reinforcement; (3) orexin A in the VTA is critical for drug-seeking behavior, but it is still unclear for the role of orexin A in the NAcSh; (4) in contrast to drug reinforcement, orexin A in the VTA is not involved in reinforcing effect of sexual reward. Orexin A plays a role both in drug-seeking behavior and in sexual reward-seeking behavior, but the different orexin A neuron populations may be responsible for the roles of orexin A in two types of reward. In a word, the differential roles of orexin A in drug and sexual reward are found in the present study, which provides some evidence for further research on the mechanisms of drug addiction.

Low-level laser therapy and light-emitting diode effects in the secretion of neuropeptides SP and CGRP in rat skin

The phototherapy effects in the skin are related to biomodulation, usually to accelerate wound healing. However, there is no direct proof of the interrelation between the effects of low-level laser therapy (LLLT) and light-emitting diode (LED) in neuropeptide secretion, these substances being prematurely involved in the neurogenic inflammation phase of wound healing. This study therefore focused on investigating LLLT and LED in Calcitonin gene-related peptide (CGRP) and substance P (SP) secretion in healthy rat skin. Forty rats were randomly distributed into five groups with eight rats each: Control Group, Blue LED Group (470 nm, 350 mW power), Red LED Group (660 nm, 350 mW power), Red Laser Group (660 nm, 100 mW power), and Infrared Laser Group (808 nm, 100 mW power) (DMCA (R) Equipamentos Ltda., So Carlos, So Paulo, Brazil). the skin of the animals in the experimental groups was irradiated using the punctual contact technique, with a total energy of 40 J, single dose, standardized at one point in the dorsal region. After 14 min of irradiation, the skin samples were collected for CGRP and SP quantification using western blot analysis. SP was released in Infrared Laser Group (p = 0.01); there was no difference in the CGRP secretion among groups. Infrared (808 nm) LLLT enhances neuropeptide SP secretion in healthy rat skin.

A Local Circuit Model of Learned Straitial and Dopamine Cell Responses under Probabilistic Schedules of Reward

Before choosing, it helps to know both the expected value signaled by a predictive cue and the associated uncertainty that the reward will be forthcoming. Recently, Fiorillo et al. (2003) found the dopamine (DA) neurons of the SNc exhibit sustained responses related to the uncertainty that a cure will be followed by reward, in addition to phasic responses related to reward prediction errors (RPEs). This suggests that cue-dependent anticipations of the timing, magnitude, and uncertainty of rewards are learned and reflected in components of the DA signals broadcast by SNc neurons. What is the minimal local circuit model that can explain such multifaceted reward-related learning? A new computational model shows how learned uncertainty responses emerge robustly on single trial along with phasic RPE responses, such that both types of DA responses exhibit the empirically observed dependence on conditional probability, expected value of reward, and time since onset of the reward-predicting cue. The model includes three major pathways for computing: immediate expected values of cures, timed predictions of reward magnitudes (and RPEs), and the uncertainty associated with these predictions. The first two model pathways refine those previously modeled by Brown et al. (1999). A third, newly modeled, pathway is formed by medium spiny projection neurons (MSPNs) of the matrix compartment of the striatum, whose axons co-release GABA and a neuropeptide, substance P, both at synapses with GABAergic neurons in the SNr and with the dendrites (in SNr) of DA neurons whose somas are in ventral SNc. Co-release enables efficient computation of sustained DA uncertainty responses that are a non-monotonic function of the conditonal probability that a reward will follow the cue. The new model's incorporation of a striatal microcircuit allowed it to reveals that variability in striatal cholinergic transmission can explain observed difference, between monkeys, in the amplitutude of the non-monotonic uncertainty function. Involvement of matriceal MSPNs and striatal cholinergic transmission implpies a relation between uncertainty in the cue-reward contigency and action-selection functions of the basal ganglia. The model synthesizes anatomical, electrophysiological and behavioral data regarding the midbrain DA system in a novel way, by relating the ability to compute uncertainty, in parallel with other aspects of reward contingencies, to the unique distribution of SP inputs in ventral SN.

Investigations into the cellular and molecular changes in the amygdala in models of temporal lobe epilepsy and maternal immune activation

The amygdala is a limbic structure that is involved in many of our emotions and processing of these emotions such as fear, anger and pleasure. Conditions such as anxiety, autism, and also epilepsy, have been linked to abnormal functioning of the amygdala, owing to improper neurodevelopment or damage. This thesis investigated the cellular and molecular changes in the amygdala in models of temporal lobe epilepsy (TLE) and maternal immune activation (MIA). The kainic acid (KA) model of temporal lobe epilepsy (TLE) was used to induce Ammon’s-horn sclerosis (AHS) and to investigate behavioural and cytoarchitectural changes that occur in the amygdala related to Neuropeptide Y1 receptor expression. Results showed that KA-injected animals showed increased anxiety-like behaviours and displayed histopathological hallmarks of AHS including CA1 ablation, granule cell dispersion, volume reduction and astrogliosis. Amygdalar volume and neuronal loss was observed in the ipsilateral nuclei which was accompanied by astrogliosis. In addition, a decrease in Y1 receptor expressing cells in the ipsilateral CA1 and CA3 sectors of the hippocampus, ipsi- and contralateral granule cell layer of the dentate gyrus and ipsilateral central nucleus of the amygdala was found, consistent with a reduction in Y1 receptor protein levels. The results suggest that plastic changes in hippocampal and/or amygdalar Y1 receptor expression may negatively impact anxiety levels. Gamma-aminobutyric acid (GABA) is the main inhibitory neurotransmitter in the brain and tight regulation and appropriate control of GABA is vital for neurochemical homeostasis. GABA transporter-1 (GAT-1) is abundantly expressed by neurones and astrocytes and plays a key role in GABA reuptake and regulation. Imbalance in GABA homeostasis has been implicated in epilepsy with GAT-1 being an attractive pharmacological target. Electron microscopy was used to examine the distribution, expression and morphology of GAT-1 expressing structures in the amygdala of the TLE model. Results suggest that GAT-1 was preferentially expressed on putative axon terminals over astrocytic processes in this TLE model. Myelin integrity was examined and results suggested that in the TLE model myelinated fibres were damaged in comparison to controls. Synaptic morphology was studied and results suggested that asymmetric (excitatory) synapses occurred more frequently than symmetric (inhibitory) synapses in the TLE model in comparison to controls. This study illustrated that the amygdala undergoes ultrastructural alterations in this TLE model. Maternal immune activation (MIA) is a risk factor for neurodevelopmental disorders such as autism, schizophrenia and also epilepsy. MIA was induced at a critical window of amygdalar development at E12 using bacterial mimetic lipopolysaccharide (LPS). Results showed that MIA activates cytokine, toll-like receptor and chemokine expression in the fetal brain that is prolonged in the postnatal amygdala. Inflammation elicited by MIA may prime the fetal brain for alterations seen in the glial environment and this in turn have deleterious effects on neuronal populations as seen in the amygdala at P14. These findings may suggest that MIA induced during amygdalar development may predispose offspring to amygdalar related disorders such as heightened anxiety, fear impairment and also neurodevelopmental disorders.

A functionally atypical amidating enzyme from the human parasite Schistosoma mansoni

Many neuropeptide transmitters require the presence of a carboxy-terminal alpha-amide group for biological activity. Amidation requires conversion of a glycine-extended peptide intermediate into a C-terminally amidated product. This post-translational modification depends on the sequential action of two enzymes (peptidylglycine alpha-hydroxylating monooxygenase or PHM, and peptidyl-alpha-hydroxyglycine alpha-amidating lyase or PAL) that in most eukaryotes are expressed as separate domains of a single protein (peptidylglycine alpha-amidating monooxygenase or PAM). We identified a cDNA encoding PHM in the human parasite Schistosoma mansoni. Transient expression of schistosome PHM (smPHM) revealed functional properties that are different from other PHM proteins; smPHM displays a lower pH-optimum and, when expressed in mammalian cells, is heavily N-glycosylated. In adult worms, PHM is found in the trans-Golgi network and secretory vesicles of both central and peripheral nerves. The widespread occurrence of PHM in the nervous system confirms the important role of amidated neuropeptides in these parasitic flatworms. The differences between schistosome and mammalian PHM suggest that it could be a target for new chemotherapeutics.

Modulation of contractile function through NPY receptors during development of cardiomyocyte hypertrophy.

Severity of left ventricular hypertrophy (LVH) correlates with elevated plasma levels of neuropeptide Y (NPY) in hypertension. NPY elicits positive and negative contractile effects in cardiomyocytes through Y(1) and Y(2) receptors, respectively. This study tested the hypothesis that NPY receptor-mediated contraction is altered during progression of LVH. Ventricular cardiomyocytes were isolated from spontaneously hypertensive rats (SHRs) pre-LVH (12 weeks), during development (16 weeks), and at established LVH (20 weeks) and age-matched normotensive Wistar Kyoto (WKY) rats. Electrically stimulated (60 V, 0.5 Hz) cell shortening was measured using edge detection and receptor expression determined at mRNA and protein level. The NPY and Y(1) receptor-selective agonist, Leu(31)Pro(34)NPY, stimulated increases in contractile amplitude, which were abolished by the Y(1) receptor-selective antagonist, BIBP3226 [R-N(2)-(diphenyl-acetyl)-N-(4-hydroxyphenyl)methyl-argininamide)], confirming Y(1) receptor involvement. Potencies of both agonists were enhanced in SHR cardiomyocytes at 20 weeks (2300- and 380-fold versus controls). Maximal responses were not attenuated. BIBP3226 unmasked a negative contraction effect of NPY, elicited over the concentration range (10(-12) to 3 x 10(-9) M) in which NPY and PYY(3-36) attenuated the positive contraction effects of isoproterenol, the potencies of which were increased in cardiomyocytes from SHRs at 20 weeks (175- and 145-fold versus controls); maximal responses were not altered. Expression of NPY-Y(1) and NPY-Y(2) receptor mRNAs was decreased (55 and 69%) in left ventricular cardiomyocytes from 20-week-old SHRs versus age-matched WKY rats; parallel decreases (32 and 80%) were observed at protein level. Enhancement of NPY potency, producing (opposing) contractile effects on cardiomyocytes together with unchanged maximal response despite reduced receptor number, enables NPY to contribute to regulating cardiac performance during compensatory LVH.

Carboxypeptidase-mediated metabolism of calcitonin gene-related peptide in human gingival crevicular fluid - a role in periodontal inflammation?

Background: Metabolism by peptidases plays an important role in modulating the levels of biologically-active neuropeptides. The metabolism of the anti-inflammatory neuropeptide calcitonin gene-related peptide (GCRP), but not the pro-inflammatory neuropeptides substance P (SP) and neurokinin A (NKA) by components of the gingival crevicular fluid (GCF), could potentiate the inflammatory process in periodontitis.

Gross anatomy of the muscle systems and associated innervation of Apatemon cobitidis proterorhini metacercaria (Trematoda: Strigeidea), as visualized by confocal microscopy.:Strigeidea), as visualized by confocal microscopy

The major muscle systems of the metacercaria of the strigeid trematode, Apatemon cobitidis proterorhini have been examined using phalloidin as a site-specific probe for filamentous actin. Regional differences were evident in the organization of the body wall musculature of the forebody and hindbody, the former comprising outer circular, intermediate longitudinal and inner diagonal fibres, the latter having the inner diagonal fibres replaced with an extra layer of more widely spaced circular muscle. Three orientations of muscle fibres (equatorial, meridional, radial) were discernible in the oral sucker, acetabulum and paired lappets. Large longitudinal extensor and flexor muscles project into the hindbody where they connect to the body wall or end blindly. Innervation to the muscle systems of Apatemon was examined by immunocytochemistry, using antibodies to known myoactive substances: the flatworm FMRFamide-related neuropeptide (FaRP), GYIRFamide, and the biogenic amine, 5-hydroxytryptamine (5-HT). Strong immunostaining for both peptidergic and serotoninergic components was found in the central nervous system and confocal microscopic mapping of the distribution of these neuroactive substances revealed they occupied separate neuronal pathways. In the peripheral nervous system, GYIRFamide-immunoreactivity was extensive and, in particular, associated with the innervation of all attachment structures; serotoninergic fibres, on the other hand, were localized to the oral sucker and pharynx and to regions along the anterior margins of the forebody.

Neurogenic inflammation and asthma

Over the past number of decades there has been considerable interest in the role of neurogenic inflammation in asthma with the identification of many biologically active neuropeptides in the lung. Whilst there is convincing evidence of neurogenic inflammation in various animal models of asthma, the evidence in humans is less clear and replicating the experimental approaches in humans has proven difficult with different studies producing conflicting results. In terms of human studies, research has focused on whether pro-inflammatory neuropeptides are elevated in the asthmatic airway, and if so, what their functional effects are. There have also been studies to assess the efficacy of tachykinin receptor antagonists in improving indices of asthma control. Information to date would suggest that neuropeptides are present in human airways and are possibly upregulated in asthma, but this effect does not appear to be specific and may occur in other inflammatory airways conditions (chronic obstructive pulmonary disease (COPD) and smoking). At present there is insufficient evidence to suggest that tachykinin receptor antagonists confer any additional benefit over inhaled corticosteroid regimes for asthmatic patients. © 2007 Bentham Science Publishers Ltd.

Inter-phyla studies on neuropeptides: the potential for broad-spectrum anthelmintic and/or endectocide discovery

Flatworm, nematode and arthropod parasites have proven their ability to develop resistance to currently available chemotherapeutics. The heavy reliance on chemotherapy and the ability of target species to develop resistance has prompted the search for novel drug targets. In view of its importance to parasite/pest survival, the neuromusculature of parasitic helminths and pest arthropod species remains an attractive target for the discovery Of novel endectocide targets. Exploitation of the neuropeptidergic system in helminths and arthropods has been hampered by a limited Understanding of the functional roles of individual peptides and the structure of endogenous targets, such as receptors. Basic research into these systems has the potential to facilitate target characterization and its offshoots (screen development and drug identification). Of particular interest to parasitologists is the fact that selected neuropeptide families are common to metazoan pest species (nematodes, platyhelminths and arthropods) and fulfil specific roles in the modulation of muscle function in each of the three phyla. This article reviews the inter-phyla activity of two peptide families, the FMRFamide-like peptides and allatostatins, on motor function in helminths and arthropods and discusses the potential of neuropeptide signalling as a target system that could uncover novel endectocidal agents.

Flatworm nerve-muscle: structural and functional analysis

Platyhelminthes occupy a unique position in nerve-muscle evolution, being the most primitive of metazoan phyla. Essentially, their nervous system consists of an archaic brain and associated pairs of longitudinal nerve cords cross-linked as an orthogon by transverse commissures. Confocal imaging reveals that these central nervous system elements are in continuity with an array of peripheral nerve plexuses which innervate a well-differentiated grid work of somatic muscle as well as a complexity of myofibres associated with organs of attachment, feeding, and reproduction. Electrophysiological studies of flatworm muscles have exposed a diversity of voltage-activated ion channels that influence muscle contractile events. Neuronal cell types are mainly multi- and bi-polar and highly secretory in nature, producing a heterogeneity of vesicular inclusions whose contents have been identified cytochemically to include all three major types of cholinergic, aminergic, and peptidergic messenger molecules. A landmark discovery in flatworm neurobiology was the biochemical isolation and amino acid sequencing of two groups of native neuropeptides: neuropeptide F and FMRFamide-related peptides (FaRPs). Both families of neuropeptide are abundant and broadly distributed in platyhelminths, occurring in neuronal vesicles in representatives of all major flatworm taxa. Dual localization studies have revealed that peptidergic and cholinergic substances occupy neuronal sets separate from those of serotoninergic components. The physiological actions of neuronal messengers in flatworms are beginning to be established, and where examined, FaRPs and 5-HT are myoexcitatory, while cholinomimetic substances are generally inhibitory. There is immunocytochemical evidence that FaRPs and 5-HT have a regulatory role in the mechanism of egg assembly. Use of muscle strips and (or) muscle fibres from free-living and parasitic flatworms has provided baseline information to indicate that muscle responses to FaRPs are mediated by a G-protein-coupled receptor, and that the signal transduction pathway for contraction involves the second messengers cAMP and protein kinase C.

Organisation of the nervous system in the Acoela: an immunocytochemical study

In order to broaden the information about the organisation of the nervous system in taxon Acoela, an immunocytochemical study of an undetermined Acoela from Cape Kartesh, Faerlea glomerata, Avagina incola and Paraphanostoma crassum has been performed. Antibodies to 5-HT and the native flatworm neuropeptide GYIRFamide were used. As in earlier studies, the pattern of 5-HT immunoreactivity revealed an anterior structure composed mainly of commissures, a so-called commissural brain. Three types of brain shapes were observed. No regular orthogon was visualised. GYIRFamide immunoreactive cell clusters were observed peripherally to the 5-HT immunoreactive commissural brain. Staining with anti-GYIRFamide revealed more nerve processes than did staining with anti-FMRFamide. As no synapomorphies were found in the organisation of the nervous system of the Acoela and that of the Platyhelminthes, the results support the view that the Acoela is not a member of the Platyhelminthes. (C) 2001 Harcourt Publishers Ltd.

Basiepidermal nervous system in Nemertoderma westbladi (Nemertodermatida): GYIRFamide immunoreactivity

The Nemertodermatida are a small group of microscopic marine worms. Recent molecular studies have demonstrated that they are likely to be the earliest extant bilaterian animals. What was the nervous system (NS) of a bilaterian ancestor like? In order to answer that question, the NS of Nemertoderma westbladi was investigated by means of indirect immunofluorescence technique and confocal scanning laser microscopy. The antibodies to a flatworm neuropeptide GYIRFamide were used in combination with anti-serotonin antibodies and phalloidin-TRITC staining. The immunostaining revealed an entirely basiepidermal NS. A ring lying outside the body wall musculature at the level of the statocyst forms the only centralisation, the

Evolution of the nervous system in Paraphanostoma (Acoela)

According to recent molecular studies, the Acoela are the earliest extant bilaterian group. Their nervous system displays a striking variety of patterns. The aim of the present investigation was to study the variability of the nervous system in a monophyletic group of the Acoela. Six species of Paraphanostoma were chosen for the study. Using immunocytochemical methods and confocal scanning laser microscopy, the immunoreactive patterns of serotonin (5-HT) and the neuropeptide GYIRFamide were described in detail. The study has demonstrated that the brains in Paraphanostoma species, although diverse in detail, still follow the same general pattern. 18S rDNA sequences were used to generate a hypothesis of the phylogeny within the group. Characters of the nervous system revealed in this study were coded and analysed together with 18S rDNA data. Several synapomorphies in the nervous system characters were identified. However, numerous parallelisms in the nervous system evolution have occurred. Data obtained demonstrate that the genus Paraphanostoma is closely related to Childia and should belong to the same family, Childiidae.

FMRFamide-related peptides (FaRPs): A new family of peptides from amphibian defensive skin secretions

Amphibian defensive skin secretions are known to contain a plethora of biologically-active peptides that are often structural and functional analogues of vertebrate neuropeptides. Here we report the structures of two invertebrate neuropeptide analogues, IPPQFMRF amide (IF-8 amide) and EGDEDEFLRF amide (EF-10 amide), from the defensive skin secretions of two different species of African hyperoliid frogs, Kassina maculata and Phylictimantis verrucosus, respectively. These represent the first canonical FMRF amide-related peptides (FaRPs) from a vertebrate source. The cDNA encoding IF-8 amide was cloned from a skin secretion library and found to contain a single copy of the peptide located at the C-terminus of a 58 amino acid residue open-reading frame. These data extend the potential targets of the defensive arsenal of amphibian tegumental secretions to parasitic/predatory invertebrates and the novel peptides described may represent the first vertebrate peptidic endectocides.