Selected Brain Areas Research Articles

Aluminum l-glutamate complex in rat brain cortex: in vivo prevention of aluminum deposit by magnesium d-aspartate

Our previous experiments in the rat showed that aluminum l-glutamate complex (Al l-Glu) crosses the blood–brain barrier and accumulates in selective brain areas and that Al salts may increase d-aspartic acid forms in living brain proteins, probably by inducing more thermodynamically stable structures than L isomers. As magnesium blocks NMDA receptors, d-aspartic acid was used in the present study in the form of magnesium salt to prevent the excitotoxicity of dicarboxylic amino acids. Effects on brain amino acids and Al cortex levels in mature rats were studied after chronic treatment with Al l-Glu or Na l-Glu alone or in association with magnesium d-aspartate (Mg d-Asp). Results demonstrate that treatment with Mg d-Asp induces a decrease in the Al concentration in brain cortex of Al l-Glu-treated rats. In aluminum-free treated controls, treatment with Mg d-Asp in association with Na l-Glu also induces a decrease in Al concentration in brain cortex. These data indicate that Mg d-Asp administration protects rat brain cortex from Al accumulation and suggest that this treatment may be useful in preventing brain Al intoxication.

Antidepressant treatments induce the expression of basic fibroblast growth factor in cortical and hippocampal neurons.

New experimental evidence suggests that the mechanism of action of antidepressants includes the induction of neurotrophic factor synthesis in selected brain areas. The present study is aimed at establishing whether prolonged antidepressant treatments increase the expression of basic fibroblast growth factor (FGF2), a polypeptide growth factor that has a broad neurotrophic activity in the adult central nervous system. Rats received a single dose or long-term (3 weeks) administration of desipramine (DMI), fluoxetine (FLU), and mianserin (MIA), then were sacrificed at 5 and 24 h after the last injection. RNase protection assay and Western blot analysis revealed that all antidepressant drugs elicited an anatomically specific increase in FGF2 mRNA and protein. The increase in FGF2 mRNA after a single injection was seen only at 5 h after the injection and was restricted to the entorhinal cortex, whereas the effect of the long-term treatments lasted up to 24 h and occurred in the entire cortex and hippocampus. Immunohistochemical analysis of FGF2 immunoreactivity was carried out to investigate which cell types responded to the antidepressant treatments. DMI and MIA increased FGF2 proteins predominantly in neurons of layer V throughout the cerebral cortex and in some neurofilament-positive cells of the hippocampus. FLU increased FGF2 immunoreactivity mainly in neurofilament-positive cells of the hippocampus. These findings may explain the therapeutic efficacy of antidepressants in affective disorders.

Pharmacological characterization of vanilloid receptor located in the brain

Specific [ 3H]resiniferatoxin (RTX) binding detects the vanilloid receptor type I (VR1). In the present study we demonstrate specific, high-affinity, saturable [ 3H]RTX binding in various areas of monkey brain not known to be innervated by primary afferent neurons as well as in spinal cord and dorsal root ganglion neurons of the same origin. Detailed pharmacological characterization and comparison revealed no major difference in binding affinities between the peripheral and the central sites as measured by K d/ K i values. In general, lower receptor density was measured in selected brain areas than in the periphery. Areas with higher receptor density were detected in the locus ceruleus, preoptic area, and medial basal hypothalamus of the brain. Both capsaicin and the competitive antagonist capsazepine inhibited the specific binding of [ 3H]RTX to membrane preparations of the dorsal horn of the spinal cord and dorsal root ganglia with K i values of 4.3±0.32 μM and 2.7±0.33 μM, respectively. Inhibition was observed in the central areas (hypothalamus) with K i values of 0.95±0.1 μM for capsaicin and 0.86±0.11 μM for capsazepine. Previous biological and pharmacological evidence suggested that vanilloid receptors were present in the brain. Our results demonstrate that the pharmacological properties of both the peripheral and central receptor sites display appropriate pharmacological similarity to represent the same receptor class. The modest differences in ligand affinities for the vanilloid receptor expressed in the brain nuclei and the dorsal root ganglion neurons may correspond to differences in sequence, modification or associated proteins.

Selective age-related changes in the PKC-sensitive, calmodulin-binding protein, neurogranin, in the mouse brain.

Brain ageing is associated with a dysregulation of intracellular calcium (Ca(2+)) homeostasis which leads to deficits in Ca(2+)-dependent signalling pathways and altered neuronal functions. Given the crucial role of neurogranin/RC3 (Ng) in the post-synaptic regulation of Ca(2+) and calmodulin levels, age-dependent changes in the levels of Ng mRNA and protein expression were analysed in 3, 12, 24 and 31-month-old mouse brains. Ageing produced significant decreases in Ng mRNA expression in the dorsal hippocampal subfields, retrosplenial and primary motor cortices, whereas no reliable changes were seen in any other cortical regions examined. Western blot indicated that Ng protein expression was also down-regulated in the ageing mouse brain. Analysis of Ng immunoreactivity in both hippocampal CA1 and retrosplenial areas indicated that Ng protein in aged mice decreased predominantly in the dendritic segments of pyramidal neurones. These data suggest that age-related changes of post-synaptic Ng in selected brain areas, and particularly in hippocampus, may contribute to altered Ca(2+)/calmodulin-signalling pathways and to region-specific impairments of synaptic plasticity and cognitive decline.

Induction of the immediate early genes egr-1 and c-fos by methamphetamine in mouse brain

Methamphetamine (METH) is one of the most commonly abused psychostimulant, and is known to induce dopaminergic neurotoxicity by generating oxidative stress and free radicals. In the present study we investigated the effects of METH on egr-1 and c-fos immediate early gene induction in different regions of mouse brain, at different doses and different time courses. We also measured the tissue levels of monoamines in order to correlate their changes with gene expression. A single injection of METH (40 mg/kg) significantly increased egr-1 and c-fos mRNA expression within 30 min in frontal cortex, nucleus accumbens, caudate putamen, septum and CA1 region of hippocampus. Time course studies showed that in most cases, both genes were expressed within 30 min and decreased after 60 min. METH produced a significant decrease in striatal dopamine level, reaching a very low level after 24 h. Striatal serotonin level significantly increased and returned to control levels after 2 h. These data show that METH induced egr-1 and c-fos mRNA expression in selective brain areas, which correlated with an alteration in monoamines.

Sexual pheromone or conventional odors increase extracellular lactate without changing glucose utilization in specific brain areas of the rat.

Brain extracellular lactate levels increase following physiological stimuli. Monitoring lactate levels might be a tool for detecting dynamic changes in brain activity. In this study we compared changes of extracellular lactate in selected brain areas with rates of glucose utilization as measured by the [14C]2-deoxyglucose method, following olfactory stimulation. Conventional (green pepper essence, heptanal, exanal, octanal) and, above all, non-conventional (sexual pheromone) odors increased lactate in the rhinencephalum, but not in the striatum. Glucose utilization did not change in any area. This discrepancy may result from the different temporal resolution of the two methods employed and/or from the clearance of lactate, whose tissue content increases transiently following neuronal activation as a reflection of the initial oxygen debt.

Estradiol Modulation of Central Monoamine Activity in Female Mountain Spiny Lizards

Although estradiol (E2) mediates many behaviors in females, relatively little is known about its role in female aggression. Previous studies in female mountain spiny lizards indicated that female aggression is modulated by ovariectomy and sex steroid hormone replacement and that expression of aggressive behavior is accompanied by changes in serotonin activity. This study examines if E2 modulates the activity of serotonin and other central monoamines. Free-living females were caught and housed in the laboratory and received one of 3 treatments: sham surgery (SHAM), ovariectomy plus empty implant (OVEX), or sham surgery plus a long lasting E2 implant (E2-IMP). After 3 weeks of treatment, selected brain areas were examined for levels of monoamines and their metabolites. Changes in monoamine activity were most pronounced in the septum where levels of serotonin (5-HT), 5-hydroxyindoleacetic acid (5-HIAA), and norepinephrine (NE) were higher in E2-IMP females relative to SHAM, and levels of 5-HIAA were higher in OVEX females relative to SHAM. Changes in dopamine (DA) activity were also found, with increased DA concentration and decreased ratio of forebrain:brainstem HVA concentrations in E2-IMP relative to SHAM females. These results suggest that the actions of E2 on aggression might be mediated, in part, by dose-dependent effects on 5-HT activity in the septum.

Intersubject Variability in Cortical Activations during a Complex Language Task

Intersubject variability in the functional organization of the human brain has theoretical and practical importance for basic and clinical neuroscience. In the present study, positron emission tomography (PET) and anatomical magnetic resonance imaging (MRI) were used to study the functional anatomy of language processes. Intersubject variability in task-induced activations in six brain regions was assessed in 20 normal subjects (10 men and 10 women) for frequency of occurrence, location, intensity, and extent. A complex, but well-studied task (overt verb generation) was compared to a simple baseline (visual fixation) to induce activations in brain areas serving perceptual, motoric, and cognitive functions. The frequency of occurrence was high for all selected brain areas (80–95%). The variability in response location in Talairach space, expressed as the standard deviation along each axis (x, y, z), ranged from 5.2 to 9.9 mm. This variability appears to be uniformly distributed across the brain, uninfluenced by regional differences in the complexity of gyral anatomy or mediated behavior. The variability in response location, expressed as the average Euclidean distances (averaged across subjects) about mean locations of activations, varied from 9.40 to 13.36 mm and had no significant differences by region (P > 0.05, β = 0.20). Intensity variability was also relatively small and homogenous across brain regions. In contrast, response extent was much more variable both across subjects and across brain regions (0.79 to 1.77, coefficient of variation). These findings are in good agreement with previous PET studies of intersubject variability and bode well for the possibility of using functional neuroimaging to study neural plasticity subsequent to congenital and acquired brain lesions.

Age, strain, and semi-chronic hydergine treatment effects on motor activity and neuronal nucleic acid-protein metabolism in male mice

This study was designed to examine the effects of Hydergine (DHET), co-dergocrine mesylate, treatment on motor activity and neuronal nucleoprotein metabolism in several motor areas of the aging rodent brain, specifically the caudate-putamen (CP), the substantia nigra (SN), and the cerebral cortex layer V (CX). Three age groups of two different strains of mice were used which represented two different aging rates: DBA/2 male mice (short lived) and C57BL/6 male mice (long lived). A representative sample of each age group was injected (IP) daily with a single dose of either DHET (1 mg/kg) or vehicle (0.9% saline) solution for one month. Total spontaneous motor activity was measured using a File apparatus to assess the functional ability of the selected brain areas. Histochemical parameters measured included the relative RNA and protein contents from a homogeneous population of neurons within each nuclei. The RNA and protein contents were assessed with a scanning microdensitometer using azure B and Coomassie staining protocols, respectively. The results of this study provide evidence that DHET does have significant effects on neuronal functioning in the motor compartments studied at the behavioral as well as the histochemical level for DBA/2 male mice. The C57BL/6 strain showed parallel, but less significant, changes in the histochemical parameters and no statistical differences in motor activity. In addition, DHET treatment produced no sign of neurotoxicity within any of the brain nuclei in either strain.

Calculation of maximal hybridization capacity (Hmax) for quantitative in situ hybridization: a case study for multiple calmodulin mRNAs.

In estimations of mRNA copy numbers, quantitative in situ hybridization (ISH) is expected to be performed at saturating probe concentration. In practice, however, this condition can rarely be fulfilled when medium to high amounts of transcripts exist and/or in large-scale experiments. To resolve this problem, we developed and tested a double-step procedure involving the use of calmodulin (CaM) I, II, and III [(35)S]-cRNA probes on adult rat brain sections; the hybridization signals were detected with a phosphorimager. By means of hybridization at increasing probe concentrations for a time sufficient for saturation, saturation curves were created for and maximal hybridization capacity (Hmax) values were assigned to selected brain areas. The Kd values of these various brain areas did not differ significantly, which allowed the creation and use of one calibration graph of Hmax vs hybridized [(35)S]-cRNA values for all brain areas for a given probe concentration. Large-scale ISH experiments involving a subsaturating probe concentration were performed to estimate Hmax values for multiple CaM mRNAs. A calibration graph corresponding to this probe concentration was created and Hmax values (expressed in ISH copy no/mm(2) units) were calculated for several brain regions via the calibration. The value of the method was demonstrated by simultaneous quantification of the total accessible multiple CaM mRNA contents of several brain areas in a precise and economical way.

Region-specific regulation of cytochrome P450 aromatase messenger ribonucleic acid by androgen in brains of male rhesus monkeys.

We demonstrated previously that testosterone regulates aromatase activity in the anterior/dorsolateral hypothalamus of male rhesus macaques. To determine the level of the androgen effect, we developed a ribonuclease protection assay to study the effects of testosterone or dihydrotestosterone (DHT) on aromatase (P450(AROM)) mRNA in selected brain areas. Adult male rhesus monkeys were treated with testosterone or DHT. Steroids in serum were quantified by RIA. Fourteen brain regions were analyzed for P450(AROM) mRNA. Significant elevations of its message over controls (P<0.05) were found in the medial preoptic area/anterior hypothalamus of both androgen treatment groups and the medial basal hypothalamus of the testosterone-treated males. Other brain areas were not affected by androgen treatment. We conclude that testosterone and DHT regulate P450(AROM) mRNA in brain regions that mediate reproductive behaviors and gonadotropin release. The P450(AROM) mRNA of other brain areas is not androgen dependent. Brain-derived estrogens may also be important for maintaining neural circuitry in brain areas not related to reproduction. The control of P450(AROM) mRNA in these areas may differ from what we report here, but it is equally important to understand the function of in situ estrogen formation in these areas.

Differential regulation of calmodulin content and calmodulin messenger RNA levels by acute and repeated, intermittent amphetamine in dopaminergic terminal and midbrain areas

Repeated doses of psychoactive drugs often produce adaptive responses that differ from the initial drug application and additional adaptive processes occur following cessation of the drug. The relationship between alterations in calmodulin protein and messenger RNA produced by an initial versus a repeated dose of amphetamine was examined, as well as changes following drug cessation. Calmodulin protein and messenger RNA of the three individual calmodulin genes were measured in rat dopaminergic cell body and terminal areas following acute or repeated amphetamine. Rats were either injected once with 2.5 mg/kg amphetamine or saline and decapitated after 3 h, or given 10 injections of amphetamine three to four days apart and decapitated 3 h after the final injection. Calmodulin messenger RNA and protein were also measured three and seven days after ceasing drug treatment. Acute amphetamine increased calmodulin 1.7-fold in the striatum and threefold in the ventral mesencephalon, with corresponding elevations in calmodulin messenger RNAs. In response to the 10th dose of amphetamine, however, the degree of increase in calmodulin was diminished in the striatum and ablated in the ventral mesencephalon. Correspondingly, select species of calmodulin messenger RNA were decreased from control levels. In the frontal cortex or nucleus accumbens, calmodulin levels were basically unaltered by the first or 10th doses of amphetamine, but both calmodulin and its messenger RNA were altered with time upon cessation of the drug. Three days later, both calmodulin protein and messenger RNA were decreased in select brain areas. By seven days after the 10th injection, calmodulin content was altered compared to saline controls in all areas, but the change in messenger RNA no longer paralleled the change in protein. Our findings demonstrate that both calmodulin protein and select species of calmodulin messenger RNA are altered by acute amphetamine, but this effect is attenuated after repeated, intermittent amphetamine. There are further time-dependent changes after cessation of repeated amphetamine, which may reflect compensatory neuronal responses. The alterations in calmodulin content and synthesis could contribute to changes in patterns or duration of behaviors that occur upon cessation of repeated amphetamine.

Localization of a glutathione-dependent dehydroascorbate reductase within the central nervous system of the rat

In this study, we describe for the first time the occurrence, within the central nervous system of the rat, of a dehydroascorbate reductase analogous to the one we recently described in the liver. Dehydroascorbate reductase plays a pivotal role in regenerating ascorbic acid from its oxidation product, dehydroascorbate. In a first set of experiments, we showed that a dehydroascorbate reductase activity is present in brain cytosol; immunoblotting analysis confirmed the presence of an immunoreactive cytosolic protein in selected brain areas. Immunotitration showed that approximately 65% of dehydroascorbate reductase activity of brain cytosol which was recovered in the ammonium sulphate fraction can be attributed to this enzyme. Using immunohistochemistry, we found that a variety of brain areas expresses the enzyme. Immunoreactivity was confined to the gray matter. Amongst the several brain regions, the cerebellum appears to be the most densely stained. The enzyme was also abundant in the hippocampus and the olfactory cortex. The lesion of norepinephrine terminals following systemic administration of DSP-4 markedly decreased immunoreactivity in the cerebellum. Apart from the possible co-localization of the enzyme with norepinephrine, the relative content of dehydroascorbate reductase in different brain regions might be crucial in conditioning regional sensitivity to free radical-induced brain damage. Given the scarcity of protective mechanisms demonstrated in the brain, the discovery of a new enzyme with antioxidant properties might represent a starting-point to increase our knowledge about the antioxidant mechanisms operating in several central nervous system disorders.

Differential distribution of G-protein β-subunits in brain: An immunocytochemical analysis

Heterotrimeric G proteins play central roles in signal transduction of neurons and other cells. The variety of their alpha-, beta-, and gamma-subunits allows numerous combinations thereby confering specificity to receptor-G-protein-effector interactions. Using antisera against individual G-protein beta-subunits we here present a regional and subcellular distribution of Gbeta1, Gbeta2, and Gbeta5 in rat brain. Immunocytochemical specificity of the subtype-specific antisera is revealed in Sf9 cells infected with various G-protein beta-subunits. Since Gbeta-subunits together with a G-protein gamma-subunit affect signal cascades we include a distribution of the neuron-specific Ggamma2- and Ggamma3-subunits in selected brain areas. Gbeta1, Gbeta2, and Gbeta5 are preferentially distributed in the neuropil of hippocampus, cerebellum and spinal cord. Gbeta2 is highly concentrated in the mossy fibres of dentate gyrus neurons ending in the stratum lucidum of hippocampal CA3-area. High amounts of Gbeta2 also occur in interneurons innervating spinal cord alpha-motoneurons. Gbeta5 is differentially distributed in all brain areas studied. It is found in the pyramidal cells of hippocampal CA1-CA3 as well as in the granule cell layer of dentate gyrus and in some interneurons. In the spinal cord Gbeta5 in contrast to Gbeta2 concentrates around alpha-motoneurons. In cultivated mouse hippocampal and hypothalamic neurons Gbeta2 and Gbeta5 are found in different subcellular compartments. Whereas Gbeta5 is restricted to the perikarya, Gbeta2 is also found in processes and synaptic contacts where it partially colocalizes with the synaptic vesicle protein synaptobrevin. An antiserum recognizing Ggamma2 and Ggamma3 reveals that these subunits are less expressed in hippocampus and cerebellum. Presumably this antiserum specifically recognizes Ggamma2 and Ggamma3 in combinations with certain G alphas and/or Gbetas. The widespread but regionally and cellularly rather different distribution of Gbeta- and Ggamma2/3-subunits suggests that region-specific combinations of G-protein subunits mediate signal transduction in the central nervous system. The different subcellular distribution of Gbeta-subunits in cultivated neurons reflects that observed in tissue where Gbeta5 and Gbeta2 associate preferentially with the perikarya and the neuropil, respectively, and suggests an additional association of Gbeta2 with secretory vesicles.

Selective early induction of synaptosomal-associated protein (molecular weight 25,000) following systemic administration of kainate at convulsant doses in the rat

SNAP-25 (synaptosomal-associated protein of mol. wt 25,000) is an essential component for neurotransmitter release, and its expression has been related to the plastic responses that follow CNS injury. In the present study, transient induction of SNAP-25 in selected brain areas is shown by immunohistochemistry at short times after a single intraperitoneal injection of kainate at convulsant doses. Six hours after kainate injection, SNAP-25 immunoreactivity was noticed in the perikarya of certain neurons of the perirhinal and lateral cortices, polymorphic layer of the dentate gyrus, CA3 pyramidal area of the hippocampus, and thalamus. In the same areas, a strong increase in SNAP-25 immunorectivity was detected at 12 and 24 h after kainate injection in cell bodies and fibers. Four days after kainate administration, the immunostaining pattern was similar to that observed in control animals. Intraperitoneal injection of cycloheximide blocked the expression of SNAP-25, thus suggesting de novo SNAP-25 protein synthesis following kainate administration. Kainate-dependent induction of SNAP-25a messenger RNA synthesis was observed by in situ hybridization in the mentioned brain areas. Heat shock protein of mol. wt 72,000 (HSP70/72) is a chaperone whose expression is induced early under stress conditions. Its expression and distribution were compared to that of SNAP-25 after the excitotoxic insult. Brain areas overexpressing SNAP-25 and HSP70/72 overlapped. In addition, partial co-localization of both antigens was observed by double-labeling immunohistochemistry. These results provide evidence of an involvement of SNAP-25 in the reactive response that follows kainate administration, and support the role of this protein in the plastic events that take place after kainate excitotoxicity.

Chapter 2.1.2 Neurophysiology of magnocellular neuroendocrine cells: Recent advances

Magnocellular neuroendocrine cells of the hypothalamic paraventricular and supraoptic nuclei are responsible for most of the vasopressin and oxytocin in the peripheral blood as well as for central release of these peptides in selected brain areas. As the principal component of the hypothalamo-neurohypophysial system, these neurons have been a subject of continual study for half a century. The wealth of solid information from decades of in vivo studies has provided a firm basis for in vitro, brain slice and explant investigations of neural mechanisms involved in the control and regulation of vasopressin and oxytocin neurons. In vitro methods have revealed the presence and permitted the study of monosynaptic projections to supraoptic neurons from the olfactory bulbs, the tuberomammillary nuclei of the posterior hypothalamus and from the organum vasculosum of the lamina terminalis. Such methods have also facilitated the elucidation of the various ionic currents controlling neurosecretory cell activity as well as the roles of calcium binding proteins and release of calcium from internal stores. This review summarizes recent advances in our understanding of the afferent inputs that impinge upon these two cell types, and the cellular and molecular mechanisms intrinsic to these neurons that determine their activity patterns and, in part, their responses to incoming stimuli.

Decrease of the Number of Opioid Receptors and of the Responsiveness to Morphineduring Neuronal Differentiation Induced by 17β-Estradiol in Estrogen Receptor-Transfected Neuroblastoma Cells (SK-ER3)

Estrogens modulate the density of opioid receptors in selected brain areas; however, it is not clear whether they exert such an effect directly on the cells which express the opioid receptors. Therefore, we analyzed the binding of [³H]-diprenorphine in human neuroblastoma cells stably transfected with the estrogen receptor cDNA (SK-ER3 cell line). A 16-hour exposure of these cells with 1 nM 17β-estradiol induces a progressive morphological differentiation which appears clearly established 6 days after the suspension of the treatment. The binding of [³H]-diprenorphine was then measured immediately after the exposure to 17β-estradiol (16 h) as well as 6 days later. The results shows that the number of opioid receptors in SK-ER3 cells is unaffected at 16 h but appears significantly reduced at 6 days. This effect is blocked by the estrogen antagonist ICI-182780, and is coincident to a decrease of the inhibitory effect of morphine on cyclic AMP accumulation. Binding experiments performed using selective ligands suggest that the µ subclass of opioid receptors is down-regulated by estradiol in SK-ER3 cells.

Chronic administration of aluminium l-glutamate in young mature rats: effects on iron levels and lipid peroxidation in selected brain areas

Clinical and experimental studies have demonstrated the neurotoxicity of aluminium (Al), notably as a result of lipid peroxidation in vitro. We previously showed that Al is able to cross the blood–brain barrier as an l-glutamate complex and be deposited in rat brain. The present work in young mature rats investigated the in vivo effects of chronic Al– l-glutamate treatment on Al and iron movement in plasma and selected brain regions. Brain lipid peroxidation was determined by evaluating the production of thiobarbituric acid reactive substances (TBARS) and analysing polyunsaturated fatty acids (PUFAs) such as C20:4 n-6 and C22:6 n-3. Our results indicate that iron concentration was decreased in plasma and that Al accumulated especially in striatum where iron levels were decreased and in the hippocampus where TBARS were increased without PUFA modifications. These data show that Al administered chronically as an l-glutamate complex is neurotoxic in vivo and thus provides a good model for studying Al toxic mechanisms.

Dexamethasone Induces Hypertrophy of Developing Medial Septum Cholinergic Neurons: Potential Role of Nerve Growth Factor

Glucocorticoid hormones influence neuronal plasticity during development; however little is known about the mechanisms of this trophic activity. Because glucocorticoids increase nerve growth factor (NGF) synthesis in selected brain areas and NGF plays a role in the development of basal forebrain cholinergic neurons, we tested the hypothesis that glucocorticoids may foster maturation of the cholinergic phenotype during postnatal development via the induction of NGF biosynthesis. The synthetic glucocorticoid dexamethasone (DEX) was injected systemically (0.5 mg/kg, s.c.) once a day for 1 week in 7-d-old (P7) rats. DEX elicited an increase in NGF mRNA and protein levels in the cerebral cortex and hippocampus as well as specific NGF responses, such as TrkA tyrosine phosphorylation in the septum, choline acetyltransferase (ChAT) and p75 neurotrophin receptor (p75NTR) immunoreactivity, and a relative number of cholinergic neurons in the medial septum. To examine whether the effect of DEX is age-related, we treated 1- and 14-d-old rats with DEX for 1 week. DEX increased NGF expression in rats treated from P1 to P8 but not in those treated from P14 to P21. The age-related increased expression of NGF correlated with the induction of ChAT immunoreactivity in the medial septum. Moreover, in the spinal cord, neither NGF nor ChAT levels were increased by DEX, suggesting that the glucocorticoid-mediated changes seen in the basal forebrain are associated with specific NGF responses. Our data suggest that by increasing NGF levels, glucocorticoids may play a role in the maturation of postnatal cholinergic neurons.

Evidence That Hypothalamic Neurotensin Signals Leptin Effects on Feeding Behavior in Normal and Fat-Preferring Rats

Leptin inhibits food intake when it is injected in the periphery or in the central nervous system. It is likely that its action is not only mediated by the inhibition of orexigenic peptides such as neuropeptide Y. Therefore, we characterized the pharmacological and physiological relationships of leptin with neurotensin (NT), a central feeding inhibitor. Firstly, we investigated the central interactions of leptin and NT. Intracerebro-ventricular (ICV) injection were done in normal Long-Evans rats. NT had a short lasting (30 minutes; p<0.01) inhibitory effect on spontaneous food intake measured at the beginning of the dark phase whereas the effect of ICV leptin was observed after 24 hours (p<0.001). Co-injection with leptin potentiated NT effect at 30 minutes (p<0.001) and prolonged it for 30 additional minutes (p<0.01). In addition, NT potentiated the effect of leptin at 30 and 60 minutes (p<0.02 and p<0.001 respectively) but not at 24 hours. Secondly, we observed that NT concentrations were augmented in selective brain areas in fat-preferring rats (+ 34% for hypothalamic NT; p<0.03). This increase was observed in the parvocellular part of the paraventricular nucleus (PVNp) only and was associated with an increase in circulating leptin levels (+ 75%; p<0.003). Interestingly, plasma leptin and NT in the PVNp were strongly correlated (r=0.57; p<0.003), suggesting changes of NT processing or release in this nucleus. These results strongly suggest that the short-term anorexigenic effects of leptin in normal rats are at least partly mediated by changes in NT processing or release. They also suggest that these processes take place in the hypothalamus, most probably in the PVNp and that they might be sensitive to fat ingestion. Therefore, the neurotensin increase observed in fat-preferring rats would limit the overconsumption of energy, a physiological mechanism translated by leptin.