The effects of ziprasidone on regional c-Fos expression in the rat forebrain.

RATIONALE: Typical and atypical antipsychotic drugs produce characteristic patterns of immediate early gene expression in rat forebrain that are considered to reflect their effects in schizophrenia subjects. OBJECTIVE: To use c-Fos immunohistochemistry to investigate the functional neuroanatomical profile of the newly introduced atypical agent ziprasidone. MATERIALS AND METHODS: c-Fos immunohistochemistry was performed on paraformaldehyde-fixed cryosections of rat brains obtained, initially, from animals 2, 4, or 6 h after oral administration of 10 mg/kg ziprasidone or vehicle and, subsequently, from animals 2 h after oral administration of 1, 3, or 10 mg/kg ziprasidone or vehicle. The density of immunoreactive nuclei was assessed in pre-determined forebrain regions. RESULTS: Ziprasidone induced a time-dependent increase in the density of c-Fos-positive nuclei that was maximal at 2 h. At the 2 h time-point, c-Fos expression was significantly (p<0.05) elevated in the shell and core of the nucleus accumbens, lateral and medial caudate putamen, and lateral septum. At 4 h post-dose, c-Fos expression was also significantly increased in the cingulate gyrus. Ziprasidone-induced c-Fos expression was dose-dependent with significant (p<0.05) c-Fos expression observed in the nucleus accumbens (shell and core) and caudate putamen (lateral and medial) at 3 and 10 mg/kg and in the lateral septum at 10 mg/kg. CONCLUSIONS: Increased c-Fos expression in the nucleus accumbens and lateral septum is considered to be predictive of activity against positive symptoms, in the caudate putamen of motor side effect liability, and in the cingulate gyrus of efficacy against negative symptoms. Thus, the observed pattern of c-Fos expression induced in rat brain by ziprasidone is consistent with its reported clinical effects, namely, efficacy against positive symptoms with a therapeutic window over motor side effects and with some activity against negative symptoms.

LPA1 receptor-deficient mice have phenotypic changes observed in psychiatric disease.

Several psychiatric diseases, including schizophrenia, are thought to have a developmental aetiology, but to date no clear link has been made between psychiatric disease and a specific developmental process. LPA(1) is a G(i)-coupled seven transmembrane receptor with high affinity for lysophosphatidic acid. Although LPA(1) is expressed in several peripheral tissues, in the nervous system it shows relatively restricted temporal expression to neuroepithelia during CNS development and to myelinating glia in the adult. We report the detailed neurological and behavioural analysis of mice homozygous for a targeted deletion at the lpa(1) locus. Our observations reveal a marked deficit in prepulse inhibition, widespread changes in the levels and turnover of the neurotransmitter 5-HT, a brain region-specific alteration in levels of amino acids, and a craniofacial dysmorphism in these mice. We suggest that the loss of LPA(1) receptor generates defects resembling those found in psychiatric disease.

Protein distribution of the orexin-2 receptor in the rat central nervous system.

Orexin-A and -B are neuropeptides mainly expressed in the lateral hypothalamic area (LHA). A role for orexins was first demonstrated in the regulation of feeding behaviour. Subsequently, the peptides have been implicated in the control of arousal. To date, two receptors for orexins have been characterised: orexin-1 and -2 receptors (OX-R1 and OX-R2). Both receptor genes are widely expressed within the rat brain. Particularly high expression of both receptor genes in certain hypothalamic and pons nuclei could be responsible for the orexigenic and arousal properties of the peptides. It is, however, presently unclear if one given receptor subtype or both subtypes may mediate a specific biological effect of orexins such as an increase in food intake. We have recently reported the distribution of the OX-R1 protein in the rat nervous system. In this study, we report the distribution of the OX-R2 protein in the rat brain and spinal cord using specific anti-peptide antisera raised against the OX-R2 protein. We also assess the expression profile of the OX-R2 gene in different brain regions. Immunolabelling for the OX-R2 protein was observed in brain regions that exhibited OX-R1-like immunoreactivity (cerebral neocortex, basal ganglia, hippocampal formation, and many other regions in the hypothalamus, thalamus, midbrain and reticular formation). Differences in the OX-R1 and OX-R2 distribution were, however, noticed in the hippocampus, hypothalamus and dorso-lateral pons.

Distribution and expression of TREK-1, a two-pore-domain potassium channel, in the adult rat CNS.

TREK-1 is a member of the two-pore-domain potassium channel family which is expressed predominantly in the CNS. Using an anti-peptide polyclonal antiserum, we have determined the distribution of TREK-1 in the brain and spinal cord of adult rats. Specificity of the antiserum was tested using a TREK-1-transfected cell line and confirmed with c-myc-tagged TREK-1. In thin tissue sections, immunoreactivity was widespread throughout the rat brain and spinal cord. TREK-1-like signals were observed in the cerebral cortex, basal ganglia, hippocampus, and various other subcortical nuclei in the hypothalamus, thalamus, mesencephalon and rhombencephalon. TREK-1 labelling appeared to be over the entire cell membrane, including the cell body and processes. Cells that morphologically resembled projection neurones and interneurones but not glial cells were labelled. As interneurones and known GABAergic projection neurones were the predominant population labelled, we investigated the possibility that TREK-1 is expressed in GABA-containing neurones using a specific anti-GABA antiserum. Expression of TREK-1 in GABA-containing neurones was observed in a number of areas, including the isocortex, hippocampus and thalamus. Thus, TREK-1 expression defines a unique and specific subset of interneurones and principal cells. These studies indicate a widespread distribution of TREK-1 potassium channels throughout the rat brain and spinal cord, with expression in a number of areas being demonstrated to be present on GABA-containing neurones.

Gene expression and protein distribution of the orexin-1 receptor in the rat brain and spinal cord.

Orexins-A and -B are neuropeptides derived from a single precursor prepro-orexin. The mature peptides are mainly expressed in the lateral hypothalamic and perifornical areas. The orexins have been implicated in the control of arousal and appear to be important messengers in the regulation of food intake. Two receptors for orexins have been characterised so far: orexin-1 and -2 receptors. To gain a further understanding of the biology of orexins, we studied the distribution of the orexin-1 receptor messenger RNA and protein in the rat nervous system. We first assessed the expression profile of the orexin-1 receptor gene (ox-r1) in different regions by using quantitative reverse transcription followed by polymerase chain reaction. Using immunohistochemical techniques, we investigated the distribution of orexin-1 receptor protein in the rat brain using a rabbit affinity-purified polyclonal antiserum raised against an N-terminal peptide. The orexin-1 receptor was widely and strongly expressed in the brain. Thus, immunosignals were observed in the cerebral cortex, basal ganglia, hippocampal formation, and various other subcortical nuclei in the hypothalamus, thalamus, midbrain and reticular formation. In particular, robust immunosignals were present in many hypothalamic and thalamic nuclei, as well as in the locus coeruleus. The distribution of the receptor protein was generally in agreement with the distribution of the receptor messenger RNA in the brain as reported previously by others and confirmed in the present study. In addition, we present in situ hybridisation and immunohistochemical data showing the presence of orexin-1 receptor messenger RNA and protein in the spinal cord and the dorsal root ganglia. Finally, due to the shared anatomical and functional similarities between orexins and melanin-concentrating hormone, we present a comparison between the neuroanatomical distribution of the orexin-1 receptor and melanin-concentrating hormone receptor protein-like immunoreactivities in the rat central nervous system, and discuss some functional implications. In conclusion, our neuroanatomical data are consistent with the biological effects of orexins on food intake and regulation of arousal. In addition, the data suggest other physiological roles for orexins mediated through the orexin-1 receptor.

Expression of melanin-concentrating hormone receptors in insulin-producing cells: MCH stimulates insulin release in RINm5F and CRI-G1 cell-lines.

Melanin-concentrating hormone (MCH) is a hypothalamic orexigenic peptide. Recently, an orphan G-protein-coupled receptor (SLC-1) was identified that binds MCH with high affinity. Here, we demonstrate the mRNA expression of this receptor in insulin-producing cells including CRI-G1 and RINm5F cells, and in rat islets of Langerhans. Immunofluorescence studies in CRI-G1 and RINm5F cell-lines demonstrated cell-surface expression of the receptor. Rat MCH significantly stimulated insulin secretion in both cell-lines. The potency and the efficacy of MCH were significantly increased in the simultaneous presence of forskolin, suggesting that MCH may amplify the insulinotropic effect of cyclic AMP elevating stimuli. Salmon MCH, which differs from rat/human MCH by six amino acids, was less efficacious than rat/human MCH in stimulating insulin release. The data provide evidence for the expression of MCH receptors in insulin producing cells. The insulinotropic effect of MCH may contribute to the regulation of metabolism and energy balance by this peptide.

The distribution of the mRNA and protein products of the melanin-concentrating hormone (MCH) receptor gene, slc-1, in the central nervous system of the rat.

Melanin-concentrating hormone (MCH), a 19 amino acid cyclic peptide, is largely expressed in the hypothalamus. It is implicated in the control of general arousal and goal-orientated behaviours in mammals, and appears to be a key messenger in the regulation of food intake. An understanding of the biological actions of MCH has been so far hampered by the lack of information about its receptor(s) and their location in the brain. We recently identified the orphan G-protein-coupled receptor SLC-1 as a receptor for the neuropeptide MCH. We used in situ hybridization histochemistry and immunohistochemistry to determine the distribution of SLC-1 mRNA and its protein product in the rat brain and spinal cord. SLC-1 mRNA and protein were found to be widely and strongly expressed throughout the brain. Immunoreactivity was observed in areas that largely overlapped with regions mapping positive for mRNA. SLC-1 signals were observed in the cerebral cortex, caudate-putamen, hippocampal formation, amygdala, hypothalamus and thalamus, as well as in various nuclei of the mesencephalon and rhombencephalon. The distribution of the receptor mRNA and immunolabelling was in good general agreement with the previously reported distribution of MCH itself. Our data are consistent with the known biological effects of MCH in the brain, e.g. modulation of the stress response, sexual behaviour, anxiety, learning, seizure production, grooming and sensory gating, and with a role for SLC-1 in mediating these physiological actions.

Melanin-concentrating hormone is the cognate ligand for the orphan G-protein-coupled receptor SLC-1.

The underlying causes of obesity are poorly understood but probably involve complex interactions between many neurotransmitter and neuropeptide systems involved in the regulation of food intake and energy balance. Three pieces of evidence indicate that the neuropeptide melanin-concentrating hormone (MCH) is an important component of this system. First, MCH stimulates feeding when injected directly into rat brains; second, the messenger RNA for the MCH precursor is upregulated in the hypothalamus of genetically obese mice and in fasted animals; and third, mice lacking MCH eat less and are lean. MCH antagonists might, therefore, provide a treatment for obesity. However, the development of such molecules has been hampered because the identity of the MCH receptor has been unknown until now. Here we show that the 353-amino-acid human orphan G-protein-coupled receptor SLC-1 expressed in HEK293 cells binds MCH with sub-nanomolar affinity, and is stimulated by MCH to mobilize intracellular Ca2+ and reduce forskolin-elevated cyclic AMP levels. We also show that SLC-1 messenger RNA and protein is expressed in the ventromedial and dorsomedial nuclei of the hypothalamus, consistent with a role for SLC-1 in mediating the effects of MCH on feeding.