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Ghrelin, an orexigenic hormone produced by the stomach, increases food intake and enhances the locomotor and rewarding effects of cocaine. Consistent with these behavioral effects, ghrelin increases dopamine cell activity in the mesolimbic system resulting in elevated levels of dopamine release and turnover in target regions such as the ventral striatum. In the current study, we examined the psychostimulant effects of acute and daily cocaine in mice with targeted deletion of the ghrelin gene (ghrelin knockout; KO) and that of their wild-type (WT) littermates. We hypothesized that ghrelin-KO mice would be hyporesponsive to the effects of cocaine as reflected in attenuated locomotor activity following both acute and chronic injections, and that this would be correlated with striatal dopamine and dopamine metabolite concentrations. Results show that the locomotor stimulating effect of cocaine (10 mg/kg) was decreased in ghrelin-KO mice as compared with their WT littermates. In addition, repeated daily injection of cocaine resulted in gradual increases in locomotor activity in WT mice, an effect that was attenuated in ghrelin-KO mice. These behavioral effects were correlated with changes in dopamine utilization in the striatum of WT mice that were not seen in ghrelin-KO mice unless these were pretreated with ghrelin. These data suggest that ghrelin is important for normal function of the mesolimbic dopaminergic system, potentially modulating both dopamine release and reuptake., (Copyright © 2011 IBRO. Published by Elsevier Ltd. All rights reserved.)

Obesity is associated with increased risk of diabetes, cardiovascular disease and several types of cancers. The hypothalamus is a region of the brain critical in the regulation of body weight. One of the critical and best studied hypothalamic circuits is comprised of the melanocortinergic orexigenic agouti-related protein (AgRP) and anorexigenic α-melanocyte stimulating hormone (α-MSH) neurons. These neurons project axons to the same hypothalamic target neurons and balance each other's activity leading to body weight regulation. We previously showed that the brain proteoglycan syndecan-3 regulates feeding behavior and body weight, and syndecan-3 null (SDC-3(-/-)) mice are lean and obesity resistant. Here we show that the melanocortin agonist Melanotan II (MTII) potently suppresses food intake and activates the hypothalamic paraventricular nuclei (PVN) in SDC-3(-/-) mice based on c-fos immunoreactivity. Interestingly, we determined that the AgRP neuropeptide is reduced in the PVN of SDC-3(-/-) mice compared to wild type mice. In contrast, neuropeptide Y, coexpressed in the AgRP neuron, is not differentially expressed nor is the counteracting neuropeptide α-MSH. These findings are unprecedented and indicate that AgRP protein localization can be selectively regulated within the hypothalamus resulting in altered neuropeptide response and tone., (Copyright © 2010 IBRO. Published by Elsevier Ltd. All rights reserved.)

Ghrelin, an orexigenic hormone produced by the stomach, is secreted in anticipation of scheduled meals and in correlation with anticipatory locomotor activity. We hypothesized that ghrelin is directly implicated in stimulating locomotor activity in anticipation of scheduled meals. To test this hypothesis, we observed 24 h patterns of locomotor activity in mice with targeted mutations of the ghrelin receptor gene (GHSR KO) and wild-type littermates, all given access to food for 4 h daily for 14 days. While wild type (WT) and GHSR KO mice produced increases in anticipatory locomotor activity, anticipatory locomotor activity in GHSR KO mice was attenuated (P<0.05). These behavioral measures correlated with attenuated levels of Fos immunoreactivity in a number of hypothalamic nuclei from GHSR KO placed on the same restricted feeding schedule for 7 days and sacrificed at ZT4. Interestingly, seven daily i.p. ghrelin injections mimicked hypothalamic Fos expression patterns to those seen in mice under restricted feeding schedules. These data suggest that ghrelin acts in the hypothalamus to augment locomotor activity in anticipation of scheduled meals.

Uncoupling proteins in the inner mitochondrial membrane uncouples oxidative phosphorylation from ATP synthesis. It has been suggested that these proteins are involved in thermogenesis as well as in the regulation of reactive oxygen species production in the mitochondria. The present work was conducted to investigate the localization of the uncoupling protein 2-like immunoreactivity (uncoupling protein 2/3 immunoreactivity) in the main catecholaminergic projection fields in the rat brain as well as in the areas of the dopaminergic and noradrenergic nerve cell groups. In particular, the relationships of tyrosine hydroxylase, dopamine beta-hydroxylase and uncoupling protein 2/3 immunoreactivity were assessed by double immunolabeling and confocal laser microscopy analysis associated with computer-assisted image analysis. Uncoupling protein 2/3 immunoreactivity was observed in discrete dopaminergic terminals in the nucleus accumbens and in the cerebral cortex whereas it was found in scattered noradrenergic terminals in the caudate putamen and Islands of Calleja Magna. One interesting finding was that uncoupling protein 2/3 immunoreactivity together with tyrosine hydroxylase immunoreactivity in the shell of nucleus accumbens was observed surrounding the previously characterized D1 receptor rich nerve cell column system characterized by a relative lack of tyrosine hydroxylase immunoreactivity. Moreover, in animal models of dopaminergic pathway degeneration, plastic changes in uncoupling protein 2/3 terminals have been shown in the cerebral cortex and striatum as seen from the increased size and intensity of uncoupling protein 2/3 immunoreactivity of their varicosities. Taken together, these findings open up the possibility that uncoupling protein 2/3 could play an important role modulating the dopaminergic and noradrenergic neurotransmission within discrete brain regions.

It has been shown that gonadal steroids have the capacity to induce synaptic plasticity in certain areas of the nervous system. Previously we have demonstrated that due to the effect of estradiol there is a transient decrease in the number of GABAergic axo-somatic synapses in the arcuate nucleus. By using systemic application of the tracer Fluorogold we retrogradely labeled a subpopulation of arcuate neurons that project to the median eminence. We than applied the disector method for synapse quantification and found that these "hypophysiotropic neurons" receive less axo-somatic inputs. We found that 17beta-estradiol induced a decrease in the numerical density of axo-somatic contacts of these retrogradely-labeled neoroendocrine cells. Our data support the hypothesis that the hormonally driven morphological synaptic plasticity is neuron specific within the arcuate nucleus and plays a decisive role in the regulation of anterior pituitary.

Unlike primates who undergo ovarian failure and loss of sex steroids at the end of reproduction, aging rodents undergo constant vaginal estrus followed by constant diestrus and finally anestrus, which indicates the absence of responsive ovarian follicles. The latter state is analogous to menopause in women. The timing of the appearance of constant estrus is determined by many factors including estrogen exposure in the brain during development and the number of times that the animal gets pregnant. The chief site of this reproductive aging in rat brains is the arcuate nucleus of the hypothalamus. The transition from normal cycles to constant estrus parallels the females' gradually decreased ability to respond to administered estradiol with a cycle of inhibition followed by disinhibition of gonadotrophin-releasing hormone. Evidence has accumulated indicating this to be due to a loss of the rat's ability to respond to markedly elevated estradiol with the usual arcuate nucleus neuro-glial plasticity that supports the estrogen-induced gonadotrophin surge (EIGS). Just as male rats are not capable of an EIGS, aged females loose this ability through repeated EIGS. Experiments indicate that in male rats the hypothalamic synaptology that develops as a result of exposure to testicular androgens in the perinatal period (brain sexual differentiation) is a result of conversion of testosterone from the testes to estrogen in the brain and is therefore due to early estrogen exposure. Aging females appear to reach a synaptology similar to males and constant estrus as a result of repeated exposure to ovarian estrogens during their reproductive careers. The relative role of aging and hormonal factors remains unclear. Morphological evidence is presented that indicates the above effects of estrogen involve changes in hypothalamic arcuate nucleus neurons and glia, including changes in the organization of perikaryal membranes as well as arcuate nucleus synaptology and the load of peroxidase in the astroglia. A possible role for free radicals (reactive oxygen species) in hypothalamic reproductive aging has been proposed. Such a mechanism is supported by evidence that the anti-oxidant vitamin E delays the onset of constant estrus and the accumulation of glial peroxidase in aging female rats. However, since the synaptology and peroxidase load in constant estrus females is independent of the age at which the constant estrus occurs, it appears that the role of (repeated) estradiol exposure is more deterministic of hypothalamic failure than is aging, per se.

It is well established that estrogen has positive effects on the density of pyramidal cell spines in the hippocampal CA1 subfield. This study explored whether afferent connections of the hippocampus that come from estrogen-sensitive subcortical structures, including the septal complex, median raphe and supramammillary area, play a role in this estrogen-induced hippocampal synaptic plasticity. These particular subcortical structures have major influences on hippocampal activity, including theta rhythm and long-term potentiation. The latter also promotes the formation of new synapses. All of the rats were ovariectomized; the fimbria/fornix, which contains the majority of subcortical efferents to the hippocampus, was transected unilaterally in each, and half of the animals received estrogen replacement. Using unbiased electron microscopic stereological methods, the CA1 pyramidal cell spine synapse density was calculated. In the estrogen-treated rats, contralateral to the fimbria/fornix transection, the spine density of CA1 pyramidal cells increased dramatically, compared to the spine density values of both the ipsilateral and contralateral hippocampi of non-estrogen-treated animals and to that of the ipsilateral hippocampus of the estrogen replaced rats. These observations indicate that fimbria/fornix transection itself does not considerably influence CA1 area pyramidal cell spine density and, most importantly, that the estrogenic effect on hippocampal morphology, in addition to directly affecting the hippocampus, involves subcortical mediation.

Central injection of galanin elicits feeding in satiated rats. We recently observed galanin-immunoreactive fibers in synaptic connection with a population of beta-endorphin-immunopositive cell bodies and dendrites in the basal hypothalamus. Because beta-endorphin also stimulates food intake, these morphological findings raised the possibility that stimulation of feeding by galanin may, in part, be mediated by beta-endorphin release. First, we observed that ICV injection of galanin (1.5-6.0 nmol) stimulated feeding in a dose-related fashion. Next, the effect on food intake of the opioid receptor antagonist naloxone (20-200 micrograms, ICV) administered immediately preceding galanin (3 nmol, ICV) was evaluated. Galanin-induced feeding was suppressed by naloxone in a dose-dependent manner with a maximal suppression of 76% at the highest naloxone dose. These findings support the existence of a functional link between galanin and beta-endorphin and are in accord with the view that stimulation of food intake by galanin may, in part, be mediated by increased beta-endorphin release.

In the absence of cellular estrogen receptors or proven direct estrogen action in the rat, it is assumed that estrogen indirectly regulates the secretory activity of the preoptic area luteinizing hormone-releasing hormone-producing cells. We have previously shown that pro-opiomelanocortin neurons in the arcuate nucleus of the rat send axons rostrally to connect with luteinizing hormone-releasing hormone neurons of the preoptic area. An experiment combining retrograde tracing and double-immunostaining was used to test the hypothesis that rat GABAergic and/or catecholaminergic neurons can influence luteinizing hormone-releasing hormone-producing cells via mediobasal hypothalamic beta-endorphin neurons. The retrograde tracer horseradish peroxidase was injected into the medial preoptic area; two days later, arcuate nucleus Vibratome sections were double-immunostained for beta-endorphin and glutamate decarboxylase or tyrosine hydroxylase. Light and electron microscopic analysis of these triple-labeled sections demonstrated that a population of beta-endorphin-immunoreactive neurons concentrated in the ventromedial arcuate nucleus contain retrogradely transported horseradish peroxidase granules and form synaptic contacts with glutamate decarboxylase- and tyrosine hydroxylase-immunoreactive axon terminals. The present data suggest that arcuate nucleus GABA and catecholamine fibers may influence luteinizing hormone-releasing hormone-containing neurons via projective pro-opiomelanocortin cells.

All of the progesterone receptor-containing cells of the monkey hypothalamus are GABAergic. The aim of this study was to further characterize these GABAergic progesterone receptor-containing neurons based on their calbindin or parvalbumin content. These calcium-binding proteins are characteristic markers of different populations of GABAergic neurons in the central nervous system. Double-immunolabeling for progesterone receptor and either calbindin or parvalbumin was performed on hypothalamic Vibratome sections of estrogen primed African green monkeys (Cercopithecus aethiops). Progesterone receptor-containing calbindin-immunoreactive neurons were observed in the ventromedial and periventricular areas of the hypothalamus. Forty-one per cent of the progesterone receptor-containing cells in this area were calbindin immunopositive. No double-immunolabeled neurons could be detected in the infundibular (arcuate) nucleus. In tissue double-immunolabeled for progesterone receptor and parvalbumin, none of the progesterone receptor-containing neurons exhibited immunoreactivity for parvalbumin. Electron microscopic double-immunostaining for progesterone receptor and calbindin confirmed the light microscopic results. Furthermore, a large number of asymmetric synaptic contacts were observed on the calbindin-immunoreactive neurons. These observations demonstrate that progesterone receptor-containing cells in the monkey mediobasal hypothalamus consist of at least two different types of GABA neurons, and indicate that progesterone receptor-containing calbindin cells may be postsynaptic targets of excitatory fibers.