Fos Induction Research Articles

Activation of neurokinin 3 receptors in the median preoptic nucleus decreases core temperature in the rat.

Estrogens have pronounced effects on thermoregulation, as illustrated by the occurrence of hot flushes secondary to estrogen withdrawal in menopausal women. Because neurokinin B (NKB) gene expression is markedly increased in the infundibular (arcuate) nucleus of postmenopausal women, and is modulated by estrogen withdrawal and replacement in multiple species, we have hypothesized that NKB neurons could play a role in the generation of flushes. There is no information, however, on whether the primary NKB receptor [neurokinin 3 receptor (NK(3)R)] modulates body temperature in any species. Here, we determine the effects of microinfusion of a selective NK(3)R agonist (senktide) into the rat median preoptic nucleus (MnPO), an important site in the heat-defense pathway. Senktide microinfusion into the rat MnPO decreased core temperature in a dose-dependent manner. The hypothermia induced by senktide was similar in ovariectomized rats with and without 17β-estradiol replacement. The hypothermic effect of senktide was prolonged in rats exposed to an ambient temperature of 29.0 C, compared with 21.5 C. Senktide microinfusion also altered tail skin vasomotion in rats exposed to an ambient temperature of 29.0 but not 21.5 C. Comparisons of the effects of senktide at different ambient temperatures indicated that the hypothermia was not secondary to thermoregulatory failure or a reduction in cold-induced thermogenesis. Other than a very mild increase in drinking, senktide microinfusion did not affect behavior. Terminal fluorescent dextran microinfusion showed targeting of the MnPO and adjacent septum, and immunohistochemical studies revealed that senktide induced a marked increase in Fos-activation in the MnPO. Because MnPO neurons expressed NK(3)R-immunoreactivity, the induction of MnPO Fos by senktide is likely a direct effect. By demonstrating that NK(3)R activation in the MnPO modulates body temperature, these studies support the hypothesis that hypothalamic NKB neurons could be involved in the generation of menopausal flushes.

Differential Changes in the Hypothalamic‐Pituitary‐Adrenal Axis and Prolactin Responses to Stress in Early Pregnant Mice

Stress can cause pregnancy failure but it is unclear how the mother's neuroendocrine system responds to stress to impair mechanisms establishing implantation. We analysed stress-evoked hypothalamic-pituitary-adrenal (HPA) axis responses in early pregnant mice. HPA axis secretory responses to immune stress in early-mid pregnancy were strong and similar to that in virgins, although activation of hypothalamic vasopressin neurones, rather than corticotrophin-releasing hormone neurones, may be more important in the stress response in pregnancy. The site and mode of detrimental glucocorticoid action in pregnancy is not established. Because circulating prolactin is important for progesterone secretion and pregnancy establishment, we also hypothesised that stress negatively impacts on prolactin and its neuroendocrine control systems in early pregnant mice. Basal prolactin secretion was profoundly inhibited by either immune or fasting stress in early pregnancy. Prolactin release is inhibited by tonic dopamine release from tuberoinfundibular (TIDA) neurones. However, immune stress did not increase TIDA neurone activity in the median eminence in pregnant mice [measured by 3,4-dihydroxyphenylacetic acid (DOPAC) content and the DOPAC:dopamine ratio]. By contrast, both immune stress and fasting caused weak induction of Fos in TIDA neurones. However, Fos induction does not always reflect dopamine secretion. Taken together, the data suggest that the stress-evoked profound reduction in prolactin secretion does not involve substantially increased dopamine activity as anticipated. In pregnancy, there was also attenuated recruitment of parvocellular paraventricular nucleus neurones and increased activation of brainstem noradrenergic nuclei after immune stress, indicating that other mechanisms may be involved in the suppression of prolactin secretion. In summary, low prolactin and increased circulating glucocorticoids together may partly explain how a mother's endocrine system mediates stress-induced pregnancy failure.

Two-Step Binding of Transcription Factors Causes Sequential Chromatin Structural Changes at the Activated IL-2 Promoter

Most gene promoters have multiple binding sequences for many transcription factors, but the contribution of each of these factors to chromatin remodeling is still unclear. Although we previously found a dynamic change in the arrangement of nucleosome arrays at the Il2 promoter during T cell activation, its timing preceded that of a decrease in nucleosome occupancy at the promoter. In this article, we show that the initial nucleosome rearrangement was temporally correlated with the binding of NFAT1 and AP-1 (Fos/Jun), whereas the second step occurred in parallel with the recruitment of other transcription factors and RNA polymerase II. Pharmacologic inhibitors for activation of NFAT1 or induction of Fos blocked the initial phase in the sequential changes. This step was not affected, however, by inhibition of c-Jun phosphorylation, which instead blocked the binding of the late transcription factors, the recruitment of CREB-binding protein, and the acetylation of histone H3 at lysine 27. Thus, the sequential recruitment of transcription factors appears to facilitate two separate steps in chromatin remodeling at the Il2 locus.

Induction of Fos expression in the rat forebrain after intragastric administration of monosodium l-glutamate, glucose and NaCl

l-glutamate, an umami taste substance, is a key molecule coupled to a food intake signaling pathway. Furthermore, recent studies have unveiled new roles for dietary glutamate on gut-brain axis communication via activation of gut glutamate receptors and subsequent vagus nerve. In the present study, we mapped activation sites of the rat forebrain after intragastric load of 60 mM monosodium l-glutamate (MSG) by measurement of Fos protein, a functional marker of neuronal activation. The same concentration of d-glucose (sweet) and NaCl (salty) was used as controls. MSG administration exclusively produced enhanced Fos expression in four hypothalamic regions (the medial preoptic area, lateral hypothalamic area, dorsomedial nucleus, and arcuate nucleus). On the other hand, glucose administration exclusively enhanced Fos induction in the nucleus accumbens. Both MSG and glucose enhanced Fos induction in three brain regions (the habenular nucleus, paraventricular nucleus, and central nucleus of the amygdala). However, MSG induced Fos inductions were more potent than those of glucose in the habenular nucleus and paraventricular nucleus. Importantly, the present study identified for the first time two brain areas (the paraventricular and arcuate hypothalamic nuclei) that are more potently activated by intragastric MSG loads compared with glucose and NaCl. Overall, our results suggest significant activation of a neural network comprising the habenular nucleus, amygdala, and the hypothalamic subnuclei following intragastric load with glutamate.

Comparison of CRF-immunoreactive neurons distribution in mouse and rat brains and selective induction of Fos in rat hypothalamic CRF neurons by abdominal surgery

Mice and rats are widely used in stress-related behavioral studies while little is known about the distribution of the stress hormone, corticotropin-releasing factor (CRF) in the mouse brain. We developed and characterized a novel rat/mouse CRF polyclonal antibody (CURE ab 200101) that was used to detect and compare the brain distributions of CRF immunoreactivity in naïve and colchicine-treated rats and mice. We also assessed whether the visceral stressor of abdominal surgery activated brain CRF neurons using double labeling of Fos/CRF in naïve rats. CRF-ir neurons were visualized in the cortex, bed nucleus of the stria terminalis, central amygdala, hypothalamic paraventricular nucleus (PVN), Barrington's nucleus and dorsolateral tegmental area in naïve rats. CRF-immunoreactive (ir) neurons in the mouse brain were detected only after colchicine. The pattern shows fundamental similarity compared to the colchicine-treated rat brain, however, there were differences with a lesser distribution in both areas and density except in the lateral septum and external subnucleus of the lateral parabrachial nucleus which contained more CRF-ir neurons in mice, and CRF-ir neurons in the dorsal motor nucleus of the vagus were found only in mice. Abdominal surgery in naïve rats induced Fos-ir in 30% of total CRF-ir neurons in the PVN compared with control (anesthesia alone) while Fos was not co-localized with CRF in other brain nuclei. These data indicate that CRF-ir distribution in the brain displays similarity as well as distinct features in mice compared to rats that may underlie some differential stress responses. Abdominal surgery activates CRF-ir neurons selectively in the PVN of rats without colchicine treatment.

The main and accessory olfactory systems of female mice are activated differentially by dominant versus subordinate male urinary odors

Previous studies have shown that female preferences for male pheromones depend on the female's reproductive condition and the dominance status of the male. However, it is unknown which olfactory system detects the odors that result in a preference for a dominant male. Therefore, in the present study, we asked whether dominant versus subordinate male urinary odors differentially activate the main and accessory olfactory systems in female ( C57Bl/6j) mice by monitoring the induction of the immediate early gene, c-fos. A more robust induction of Fos was observed in female mice which had direct nasal contact with dominant male urinary odors in four specific segments of the accessory olfactory system, i.e., the posteroventral part of the medial amygdala, the bed nucleus of the stria terminalis, the medial part of the preoptic nucleus and the ventrolateral part of the ventromedial hypothalamus, compared to females that were exposed to subordinate male urine. This greater activation of the accessory olfactory pathway by dominant male urine suggests that there are differences in the nonvolatile components of dominant versus subordinate male urine that are detected by the vomeronasal organ. By contrast, subordinate male urinary odors induced a greater activation in the piriform cortex which is part of the main olfactory system, suggesting that female mice discriminate between dominant and subordinate male urine using their main olfactory system as well.

Interactions between amygdala central nucleus and the ventral tegmental area in the acquisition of conditioned cue-directed behavior in rats

Rats orient to and approach localizable visual cues paired with food delivery. Previous studies from this laboratory show that the acquisition and expression of these learned cue-directed responses depend on integrity of a system including the central nucleus of the amygdala (CeA), the substantia nigra pars compacta (SNc) and the dorsolateral striatum (DLS). Other investigators have suggested that cue-directed behaviors may also depend on interaction between CeA and the ventral striatum, perhaps via CeA projections to the ventral tegmental area (VTA). In Experiment 1, we examined the effects of unilateral lesions of CeA and/or VTA on rats' acquisition of conditioned responses to visual cues paired with food. Contrary to the results of previous studies that examined interactions of CeA with either SNc or DLS, rats with contralateral disconnection lesions of CeA and VTA were unimpaired in their acquisition of cue-directed responses. By contrast, rats with lesions of both structures in the same hemisphere failed to learn cue-directed responses, but were normal in their acquisition of conditioned responses directed to the food cup. In Experiment 2, we attempted to characterize the influence of VTA on CeA by examining FOS induction in CeA by a visual cue for food in rats with unilateral lesions of VTA. The results suggested an excitatory influence of VTA on CeA in the presence of food cues. Implications of these results for brain circuits involved in learned orienting and incentive motivation are discussed.

Androgen Receptors in the Posterior Bed Nucleus of the Stria Terminalis Increase Neuropeptide Expression and the Stress-Induced Activation of the Paraventricular Nucleus of the Hypothalamus

The posterior bed nuclei of the stria terminalis (BST) are important neural substrate for relaying limbic influences to the paraventricular nucleus (PVN) of the hypothalamus to inhibit hypothalamic-pituitary-adrenal (HPA) axis responses to emotional stress. Androgen receptor-expressing cells within the posterior BST have been identified as projecting to the PVN region. To test a role for androgen receptors in the posterior BST to inhibit PVN motor neurons, we compared the effects of the non-aromatizable androgen dihydrotestosterone (DHT), the androgen receptor antagonist hydroxyflutamide (HF), or a combination of both drugs implanted unilaterally within the posterior BST. Rats bearing unilateral implants were analyzed for PVN Fos induction in response to acute-restraint stress and relative levels of corticotrophin-releasing hormone and arginine vasopressin (AVP) mRNA. Glutamic acid decarboxylase (GAD) 65 and GAD 67 mRNA were analyzed in the posterior BST to test a local involvement of GABA. There were no changes in GAD expression to support a GABA-related mechanism in the BST. For PVN neuropeptide expression and Fos responses, basic effects were lateralized to the sides of the PVN ipsilateral to the implants. However, opposite to our expectations of an inhibitory influence of androgen receptors in the posterior BST, PVN AVP mRNA and stress-induced Fos were augmented in response to DHT and attenuated in response to HF. These results suggest that a subset of androgen receptor-expressing cells within the posterior BST region may be responsible for increasing the biosynthetic capacity and stress-induced drive of PVN motor neurons.

Norepinephrine and ß 1-adrenergic signaling facilitate activation of hippocampal CA1 pyramidal neurons during contextual memory retrieval

We previously described a role for adrenergic signaling in the hippocampus to promote contextual and spatial memory retrieval. A subsequent study performing expression analysis of the immediate-early gene (IEG) Arc suggested that activation of CA1 but not CA3 pyramidal neurons during memory retrieval is impaired in the absence of NE. The current study sought to confirm and extend those observations by performing expression analysis of a second IEG product, Fos, following a much greater variety of testing conditions. In mutant mice lacking NE, induction of Fos was normal in all regions of the hippocampus and amygdala shortly after fear conditioning. In contrast, when testing contextual fear 1 day after training, induction of Fos in CA1 and the central nucleus of the amygdala (CeA), but not CA3, the dentate gyrus or other amygdaloid nuclei, was impaired in the mutant mice. This pattern corresponded to the memory retrieval deficit exhibited by these mice. On the other hand, induction was normal in CA1 and CeA when testing cued fear 1 day after training, or contextual fear 1 week or 1 month after training, conditions in which retrieval are normal in the absence of NE. Acute restoration of NE in the mutant mice before testing but not before training rescued retrieval of contextual fear and restored Fos induction in CA1 and CeA. Because NE facilitates retrieval through the activation of β 1-adrenergic receptors, β 1 knockout mice were also examined and found to exhibit reduced induction of Fos in CA1 and CeA following retrieval. Based on these and previous results, we hypothesize that adrenergic signaling is critical for the full activation of CA1 pyramidal neurons in response to excitatory input from CA3 pyramidal neurons conveying retrieved contextual information.

NEURAL ACTIVITY ASSOCIATED WITH SOCIAL BEHAVIORS IN THE BROWN ANOLE, ANOLIS SAGREI

Event Abstract Back to Event NEURAL ACTIVITY ASSOCIATED WITH SOCIAL BEHAVIORS IN THE BROWN ANOLE, ANOLIS SAGREI David Kabelik1*, Emily Burford1, Leah Singh1, Veronica Alix1, Aaron Kala1, William Hawes1, Salar Rafieetary1 and Jason Ballard1 1 Rhodes College, Biology, United States The purpose of this research is to define the social behavior neural network within a lizard model system. Due to the relatively small size and low complexity of the Brown Anole (Anolis sagrei) brain in relation to mammalian species, analyses of neural activity encompassing the entire brain are readily possible. Using immunofluorescent multilabeling, we are examining induction of the immediate early gene product Fos, a marker of neural activity, and its colocalization with vasotocin and the dopaminergic enzyme tyrosine hydroxylase, in relation to participation in social behaviors. We are comparing Fos induction and colocalization rates between control males, males involved in an aggressive same-sex confrontation, and in males involved in the courtship of a female. Areas of the brain that show differences from the control group in a limited sample (n=5 per group) will be examined across a larger sample per group of interest (n=22), in order to correlate qualitative and quantitative behavioral differences with Fos expression. In the future, areas of interest will be examined across species and different hormonal conditions in order to examine the plasticity of neural mechanisms underlying hormonally induced and species-specific differences in social behaviors. Keywords: Aggression, Anole, Courtship, Dopamine, Fos, Vasotocin Conference: ISAREN 2011: 7th International Symposium on Amphibian and Reptilian Endocrinology and Neurobiology, Ann Arbor, United States, 11 Jul - 13 Jul, 2011. Presentation Type: Poster Topic: Brain and behavior Citation: Kabelik D, Burford E, Singh L, Alix V, Kala A, Hawes W, Rafieetary S and Ballard J (2011). NEURAL ACTIVITY ASSOCIATED WITH SOCIAL BEHAVIORS IN THE BROWN ANOLE, ANOLIS SAGREI. Front. Endocrinol. Conference Abstract: ISAREN 2011: 7th International Symposium on Amphibian and Reptilian Endocrinology and Neurobiology. doi: 10.3389/conf.fendo.2011.03.00023 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 22 Jul 2011; Published Online: 09 Aug 2011. * Correspondence: Prof. David Kabelik, Rhodes College, Biology, Memphis, TN, 38112, United States, kabelik@gmail.com Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers David Kabelik Emily Burford Leah Singh Veronica Alix Aaron Kala William Hawes Salar Rafieetary Jason Ballard Google David Kabelik Emily Burford Leah Singh Veronica Alix Aaron Kala William Hawes Salar Rafieetary Jason Ballard Google Scholar David Kabelik Emily Burford Leah Singh Veronica Alix Aaron Kala William Hawes Salar Rafieetary Jason Ballard PubMed David Kabelik Emily Burford Leah Singh Veronica Alix Aaron Kala William Hawes Salar Rafieetary Jason Ballard Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

Partial agonists in schizophrenia – why some work and others do not: insights from preclinical animal models

While dopamine D2 receptor partial agonists (PAs) have been long considered for treating schizophrenia, only one, aripiprazole, is clinically available for therapeutic use. This raises critically important questions as to what is unique about aripiprazole and to what extent animal models can predict therapeutic success. A number of PAs whose clinical fate is known: aripiprazole, preclamol, terguride, OPC-4392 and bifeprunox were compared to haloperidol (a reference antipsychotic) in several convergent preclinical animal models; i.e. amphetamine-induced locomotion (AIL) and conditioned avoidance response (CAR), predictive of antipsychotic effects; unilateral nigrostriatal lesioned rats, a model of hypo-dopaminergia; striatal Fos induction, a molecular marker for antipsychotic activity; and side-effects common to this class of drugs: catalepsy (motor side-effects) and prolactaemia. The results were compared across drugs with reference to their measured striatal D2 receptor occupancy. All the PAs occupied striatal D2 receptors in a dose dependent manner, inhibited AIL and CAR, and lacked motor side-effects or prolactinaemia despite D2 receptor occupancy exceeding 80%. At comparative doses, aripiprazole distinguished itself from the other PAs by causing the least rotation in the hypo-dopaminergic model (indicating the least intrinsic activity) and showed the highest Fos expression in the nucleus accumbens (indicating functional D2 antagonism). Although a number of PAs are active in antipsychotic animal models, not all of them succeed. Given that only aripiprazole is clinically available, it can be inferred that low functional intrinsic activity coupled with sufficient functional antagonism as reflected in the animal models may be a marker of success.

Role of dorsal medial prefrontal cortex dopamine D1-family receptors in relapse to high-fat food seeking induced by the anxiogenic drug yohimbine.

In humans, relapse to maladaptive eating habits during dieting is often provoked by stress. In rats, the anxiogenic drug yohimbine, which causes stress-like responses in both humans and nonhumans, reinstates food seeking in a relapse model. In this study, we examined the role of medial prefrontal cortex (mPFC) dopamine D1-family receptors, previously implicated in stress-induced reinstatement of drug seeking, in yohimbine-induced reinstatement of food seeking. We trained food-restricted rats to lever press for 35% high-fat pellets every other day (9-15 sessions, 3 h each); pellet delivery was accompanied by a discrete tone-light cue. We then extinguished operant responding for 10-16 days by removing the pellets. Subsequently, we examined the effect of yohimbine (2 mg/kg, i.p.) on reinstatement of food seeking and Fos (a neuronal activity marker) induction in mPFC. We then examined the effect of systemic injections of the D1-family receptor antagonist SCH23390 (10 μg/kg, s.c.) on yohimbine-induced reinstatement and Fos induction, and that of mPFC SCH23390 (0.5 and 1.0 μg/side) injections on this reinstatement. Yohimbine-induced reinstatement was associated with strong Fos induction in the dorsal mPFC and with weaker Fos induction in the ventral mPFC. Systemic SCH23390 injections blocked both yohimbine-induced reinstatement and mPFC Fos induction. Dorsal, but not ventral, mPFC injections of SCH23390 decreased yohimbine-induced reinstatement of food seeking. In addition, dorsal mPFC SCH23390 injections decreased pellet-priming-induced reinstatement, but had no effect on ongoing high-fat pellet self-administration or discrete-cue-induced reinstatement. Results indicate a critical role of dorsal mPFC dopamine D1-family receptors in stress-induced relapse to palatable food seeking, as well as relapse induced by acute re-exposure to food taste, texture, and smell.

The central amygdala projection to the substantia nigra reflects prediction error information in appetitive conditioning

The central amygdala nucleus (CeA) plays a critical role in cognitive processes beyond fear conditioning. For example, intact CeA function is essential for enhancing attention to conditioned stimuli (CSs). Furthermore, this enhanced attention depends on the CeA's connections to the nigrostriatal system. In the current study, we examined the role of the CeA's connections to two midbrain dopamine regions, the substantia nigra pars compacta (SNc) and the ventral tegmental area (VTA), in processing CS information when predictions of reward or nonreward were confirmed or disconfirmed. Initially, two different retrograde tracers were injected into the SNc and the VTA of rats, to label CeA cells. Different groups of rats then received a visual CS either paired or unpaired with food. Finally, Fos induction was assessed after a test session in which rats were exposed to the visual CS alone or paired with food. Colabeling of Fos and the retrograde tracer(s) showed that CeA neurons projecting to the SNc, but not to the VTA, were engaged in processing CS information when the training and testing conditions differed. These results suggest that the CeA-nigral pathway represents prediction error information during appetitive conditioning.

PHOTIC INDUCTION OF Fos IN THE SUPRACHIASMATIC NUCLEUS OF AFRICAN MOLE-RATS: RESPONSES TO INCREASING IRRADIANCE

African mole-rats (family Bathyergidae) are strictly subterranean rodent species that are rarely exposed to environmental light. Morphological and physiological adaptations to the underground environment include a severely reduced eye size and regressed visual system. Responses of the circadian system to light, however, appear to be intact, since mole-rats are able to entrain their circadian activity rhythms to the light-dark cycle and light induces Fos expression in the suprachiasmatic nucleus (SCN). Social organization varies from solitary species to highly elaborated eusocial structures, characterized by a distinct division of labor and in which one reproductive female regulates the behavior and reproductive physiology of other individuals in the colony. The authors studied light-induced Fos expression in the SCN to increasing light intensities in four mole-rat species, ranging from strictly solitary to highly social. In the solitary Cape mole-rat, light induces significant Fos expression in the SCN, and the number of Fos-immunopositive cells increases with increasing light intensity. In contrast, Fos induction in the SCN of social species was slightly greater than, but not statistically different from, the dark-control animals as is typical of most rodents. One species showed a trend for an increase in expression with increased light, whereas a second species showed no trend in expression. In the naked mole-rat, Fos expression appeared higher in the dark-controls than in the animals exposed to light, although the differences in Fos expression were not significant. These results suggest a gradient in the sensitivity of the circadian system to light in mole-rats, with a higher percentage of individuals that are unresponsive to light in correlation with the degree of sociality. In highly social species, such as the naked mole-rat that live in a relatively stable subterranean milieu in terms of food availability, temperature, constant darkness, and devoid of 24-h cyclic environmental cues, the temporal coordination of rest-wake activities may be dependent on social interactions and social status rather than on photic regulation of the circadian timing system. (Author correspondence: moosthuizen@zoology.up.ac.za)

Alarin stimulates food intake in male rats and LH secretion in castrated male rats

Alarin is a newly identified member of the galanin family of neuropeptides that includes galanin-like peptide (GALP) and galanin. Alarin was discovered as an alternate transcript of the GALP gene in neuroblastoma cells, and subsequently alarin mRNA was detected in the brain of rodents. GALP and galanin are important central regulators of both feeding and reproductive behavior. We hypothesized, that, as a member of the galanin family of peptides, alarin would also have central effects on feeding and reproduction. To test this hypothesis, we treated male rats with alarin intracerebroventricularly (i.c.v.) and measured its effects on food intake and energy homeostasis as well as sexual behavior and luteinizing hormone (LH) secretion. We observed that i.c.v. injection of 1.0 nmol alarin significantly increased food intake (p<0.01) and body weight (p<0.05). Alarin did not affect sexual behavior in male rats; however, alarin did significantly (p<0.01) increase LH levels in castrated, but not intact, male rats. Alarin immunoreactive cell bodies were detected within the locus coeruleus and locus subcoeruleus of the midbrain, which is a brainstem nucleus involved in coordinating many physiological activities, including food intake and reproduction. Lastly, alarin stimulated Fos induction in hypothalamic nuclei, such as the paraventricular nucleus and the nucleus of the tractus solitarious. Our studies demonstrate that alarin, like other members of the galanin family, is a neuromediator of food intake and body weight.

Impact of age and retinal degeneration on the light input to circadian brain structures

Aging causes anatomical and functional changes in visual and circadian systems. In wild type mice rods, cones, and photosensitive retinal ganglion cells (pRGCs) decline with age. In rd/rd cl mice, the early loss of rods and cones is followed by protracted transneuronal loss of inner retinal neurons as well as the pRGCs. Here we use Fos induction to study the light input pathway to the suprachiasmatic nuclei (SCN), the intergeniculate leaflets (IGL) and ventral lateral geniculate nuclei (vLGN) of old (∼700 days) and young (∼150 days) wild type and rd/rd cl mice. Cholera toxin tracing was used in parallel to study the anatomy of this pathway. We find that aging rather than retinal degeneration is a more important factor in reducing light input to the SCN, causing both a reduction in Fos expression and retinal afferents. Furthermore, we show light-induced Fos within the vLGN and IGL is predominantly subserved by rods and cones, and once again aging reduces the amplitude of this response.

Abstract 2041: PDGF inducible microRNAs regulate gene expression in cancer cells

Abstract Background: Platelet derived growth factor (PDGF)-regulated signaling plays a critical role in oncogenesis. By binding to the PDGF receptors, the Akt, Src and MAPK pathways are activated to regulate genes expression and cellular physiological processes. There are subtle functional differences between the effects of specific PDGF ligands (e.g. AA, BB), which are incompletely understood. MicroRNA(miR)s are single-stranded, 21-23 nucleotide regulatory RNAs that repress target gene expression either by inhibiting protein translation or by promoting target mRNA degradation. The hypothesis tested in this study was that PDGFs regulate genes expression by modulating microRNAs level in cancer cells. Methods: MicroRNA microarrays were performed to identify miR alterations after treatment of U118 glioblastoma cells with PDGF-AA and BB. Several time points were tested: 2, 6, 12, and 24 hour. Significance Analysis of Microarrays (SAM) was performed to compare expression level between different time points. Q-PCR was conducted to confirm the microarrays data. Pre-miR probe/miRCury LNA probe were transfected into cancer cells to over-express/knock down selected miRs. Western blots were used to detect miRs’ target proteins. Results: Candidate PDGF-inducible/repressible miRs were identified based on significant change in expression level after AA/BB treatment (p-value &amp;lt;0.01). Expression levels of 7 miRs were altered by PDGF AA and expression levels of 14 miRs were altered after PDGF BB treatment. By Q-PCR we showed that let 7d was repressed by PDGF-AA, but not by PDGF-BB in cancer cells (U118, C272 and U87) and in fibroblasts (WI38). Cyclin D1 is a known target of let7d. By overexpression and knock-down strategies we show that PDGF-AA induced cyclin D1 expression is mediated via repression of let7d. Q-PCR confirmed that miR-146b and miR-106b are inducible by PDGF-BB, but not by PDGF-AA in cancer cells (U118, C272 and U87) and in fibroblasts (WI38). MiR-146b induction by PDGF-BB is modulated via MAPK-dependent induction of Fos. A Fos binding element was identified in the promoter region of miR146b and Fos binding to this region in the presence of PDGF-BB was confirmed by chromatin immunoprecipitation. Several targets of miR146b are altered by PDGF in cancer cells. Conclusions: Our data demonstrate for the first time that PDGF signaling regulates expression of several critical oncogenic miRs which in turn alter target gene expression and function. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2041.

Hypertensive background modifies central nervous system stress responses in male rats

Hypertension is a risk factor for stroke, and has been correlated with HPA axis sensitization and the initiation of immune/inflammatory processes during chronic stress. Our study examined whether hypertension affects the stress‐induced activation of brain regions involved in the generation of stress responses and cardiovascular control. Spontaneously hypertensive (SHR) and normotensive Wistar‐Kyoto (WKY) male rats were acutely (30min x 1d) or repeatedly (30min x 14d) restrained or were not stressed. Perfused brain sections were immunohistochemically stained for Fos, which was quantified in the paraventricular (PVH) and dorsomedial (DMH) nuclei and the central (CeA) and medial (MeA) amygdala. Acute stress increased Fos in the PVH and DMH of WKY rats, compared to controls, and this response habituated with repeated stress exposure. Similar results were seen in control and acute SHRs, but the habituation of Fos induction was not seen in the PVH and only partially occurred in the DMH of these animals after repeated stress. MeA and CeA responses varied by stress treatment but were largely similar in the two strains. The data suggest that hypertension leads to enhanced responses to chronic stress at the level of neuronal activation. Understanding stress‐hypertension interactions in the central nervous system will aid the development of treatments for these and related conditions. Support: NIH 5G12 RR008124

Effect of serotonergic anorectics on food intake and induction of Fos in brain of mice with disruption of melanocortin 3 and/or 4 receptors

Previous studies have indicated that type 3 or 4 melanocortin receptors (MCR) are downstream of the critical anorectic action of drugs that stimulate 5-HT 2C receptors. To characterize further the receptor types involved, we have studied the effect of serotonergic anorectics in mice with genomic disruption of either MC3R or MC4R, or their combined knockout. In a first experiment, we showed that wild type (WT) and MC4R−/− mice showed comparable inhibition of food intake following acute treatment with dexnorfenfluramine. In a second experiment using WAY-161503, a 5-HT receptor full agonist with selectivity for 2B and 2C subtypes, we found that MC4R−/− responded comparably to WT, while MC3R−/− had reduced sensitivity. Double receptor knockout (DKO) mice responded comparably to WT and MC4R−/−. Surprisingly, brain Fos-ir was not strongly induced in any brain region by WAY-16103 with the exception of the paraventricular nucleus of DKO. These data suggest that MC3Rs may be involved in the response to serotonergic anorectic agents, and more generally in control of food intake.

Light induces Fos expression via extracellular signal‐regulated kinases 1/2 in melanopsin‐expressing PC12 cells

The photopigment melanopsin is expressed in a subtype of mammalian ganglion cells in the retina that project to the circadian clock in the hypothalamic suprachiasmatic nucleus to mediate non-visual light information. Melanopsin renders these retinal ganglion cells intrinsically photosensitive and the cells respond to light by a membrane depolarization and induction of the immediate early response gene Fos. Previous studies showed that the light activated melanopsin-induced signaling, the phototransduction, leading to depolarization of the membrane resembles the invertebrate opsins, which involves a Galpha(q/11) coupled phospholipase C activation. However, the signaling proteins mediating melanopsin-induced Fos expression are unresolved. In this study, we examined the phototransduction leading to Fos expression in melanopsin-transfected PC12 cells. A pivotal role of the extracellular signal-regulated protein kinase 1/2 (ERK1/2) was found as pharmacological blockage of this kinase suppressed the light-induced Fos expression. Illumination increased the inositol phosphate turnover and induced phosphorylation of ERK1/2 and p38 but not the c-Jun N-terminal kinase. The Galpha(q/11) protein inhibitor YM254890 attenuated these intracellular light responses. Our data strongly indicate that Galpha(q/11)-mediated ERK1/2 activation is essential for expression of Fos upon illumination of melanopsin-expressing PC12 cells.