CCK-A receptor activation induces fos expression in myenteric neurons of rat small intestine

Cholecystokinin (CCK), a hormone secreted from endocrine cells of the small intestine, participates in the control of motility and secretion in the gastrointestinal tract, and in the control of food intake. At least some of the effects of CCK on intestinal function appear to be mediated via activation of intrinsic neurons in the myenteric plexus. However, the distribution of CCK-responsive enteric neurons within the rat small intestine is not known. Neither has the role of CCK-A receptors in the activation of rat myenteric neurons been investigated. Therefore, to determine the distribution of CCK-responsive neurons in the small intestinal myenteric plexus we utilized immunohistochemical detection of Fos, the protein product of the immediate early gene c-fos, to identify neurons that were activated by exogenous CCK. We also monitored Fos expression in the dorsal hindbrain, and examined CCK-induced Fos expression in the presence or absence of a receptor antagonist for the type-A CCK receptor. We found that CCK significantly increased Fos expression in the hindbrain and in myenteric neurons of the duodenum and jejunum, but not the ileum. Neuronal Fos responsiveness in both brain and myenteric neurons was mediated by CCK-A receptors, as CCK-induced Fos expression was eliminated in rats pretreated with a CCK-A receptor antagonist. We conclude that CCK activates small intestinal myenteric neurons, via CCK-A receptors. Activation of these intrinsic intestinal neurons may participate in reflexes and behaviors that are mediated by CCK.     

Dopaminergic activation of estrogen receptors induces fos expression within restricted regions of the neonatal female rat brain

Steroid receptor activation in the developing brain influences a variety of cellular processes that endure into adulthood, altering both behavior and physiology. Recent data suggests that dopamine can regulate expression of progestin receptors within restricted regions of the developing rat brain by activating estrogen receptors in a ligand-independent manner. It is unclear whether changes in neuronal activity induced by dopaminergic activation of estrogen receptors are also region specific. To investigate this question, we examined where the dopamine D1-like receptor agonist, SKF 38393, altered Fos expression via estrogen receptor activation. We report that dopamine D1-like receptor agonist treatment increased Fos protein expression within many regions of the developing female rat brain. More importantly, prior treatment with an estrogen receptor antagonist partially reduced D1-like receptor agonist-induced Fos expression only within the bed nucleus of the stria terminalis and the central amygdala. These data suggest that dopaminergic activation of estrogen receptors alters neuronal activity within restricted regions of the developing rat brain. This implies that ligand-independent activation of estrogen receptors by dopamine might organize a unique set of behaviors during brain development in contrast to the more wide spread ligand activation of estrogen receptors by estrogen.     

Neonatal maternal separation induces sex-specific augmentation of the hypercapnic ventilatory response in awake rat.

Neonatal maternal separation (NMS) is a form of stress that exerts persistent, sex-specific effects on the hypoxic ventilatory response. Adult male rats previously subjected to NMS show a 25% increase in the response, whereas NMS females show a response 30% lower than controls (8). To assess the extent to which NMS affects ventilatory control development, we tested the hypothesis that NMS alters the ventilatory response to hypercapnia in awake, unrestrained rats. Pups subjected to NMS were placed in a temperature- and humidity-controlled incubator 3 h/day for 10 consecutive days (P3 to P12). Control pups were undisturbed. At adulthood (8 to 10 wk old), rats were placed in a plethysmography chamber for measurement of ventilatory parameters under baseline and hypercapnic conditions (inspired CO(2) fraction = 0.05). After 20 min of hypercapnia, the minute ventilation response measured in NMS males was 47% less than controls, owing to a lower tidal volume response (22%). Conversely, females previously subjected to NMS showed minute ventilation and tidal volume responses 63 and 18% larger than controls respectively. Although a lower baseline minute ventilation contributes to this effect, the higher minute ventilation/CO(2) production response observed in NMS females suggests a greater responsiveness to CO(2)/H(+) in this group. We conclude that NMS exerts sex-specific effects on the hypercapnic ventilatory response and that the neural mechanisms affected by NMS likely differ from those involved in the hypoxic chemoreflex.     

Anxiogenic effects of developmental bisphenol a exposure are associated with gene expression changes in the juvenile rat amygdala and mitigated by soy

Early life exposure to Bisphenol A (BPA), a component of polycarbonate plastics and epoxy resins, alters sociosexual behavior in numerous species including humans. The present study focused on the ontogeny of these behavioral effects beginning in adolescence and assessed the underlying molecular changes in the amygdala. We also explored the mitigating potential of a soy-rich diet on these endpoints. Wistar rats were exposed to BPA via drinking water (1 mg/L) from gestation through puberty, and reared on a soy-based or soy-free diet. A group exposed to ethinyl estradiol (50 μg/L) and a soy-free diet was used as a positive estrogenic control. Animals were tested as juveniles or adults for anxiety-like and exploratory behavior. Assessment of serum BPA and genistein (GEN), a soy phytoestrogen, confirmed that internal dose was within a human-relevant range. BPA induced anxiogenic behavior in juveniles and loss of sexual dimorphisms in adult exploratory behavior, but only in the animals reared on the soy-free diet. Expression analysis revealed a suite of genes, including a subset known to mediate sociosexual behavior, associated with BPA-induced juvenile anxiety. Notably, expression of estrogen receptor beta (Esr2) and two melanocortin receptors (Mc3r, Mc4r) were downregulated. Collectively, these results show that behavioral impacts of BPA can manifest during adolescence, but wane in adulthood, and may be mitigated by diet. These data also reveal that, because ERβ and melanocortin receptors are crucial to their function, oxytocin/vasopressin signaling pathways, which have previously been linked to human affective disorders, may underlie these behavioral outcomes.     

Gender difference in BK channel expression in amygdala complex of rat brain

The expression of large-conductance Ca²⁺-activated K⁺ (BK) channel protein in amygdala complex was higher in adult (8-10 weeks old) male rats than in female. Castration at 4-6 weeks old significantly reduced BK channel expression in amygdala to the level similar to that in female. Immunocytochemical analyses of pyramidal-like neurons isolated from amygdala revealed that somas with relatively large size were highly immunoreactive to both anti-androgen receptor (AR) and anti-BK channel antibodies, while those with smaller size were not. The double-immunopositive neurons were dominant (60%) among pyramidal-like neurons isolated from amygdala of male rats but rare among those from female. The membrane current sensitive to penitrem A, a BK channel blocker, was the major K⁺ current component in large neurons and showed higher current-density than that in smaller ones. These results suggest the gender-dependent cell population expressing BK channels in amygdala complex and its up-regulation by AR stimulation.     

Estrogen induces insulin-like growth factor-I expression in the rat uterus

The inability to convincingly demonstrate a mitogenic effect of estrogen on isolated uterine cells in culture suggests that autocrine or paracrine growth factors may be important in the estrogen-induced uterine proliferative response. Here we report that uterine expression of insulin-like growth factor-I (IGF-I), an important mediator of GH action, is increased after 17 beta-estradiol (5 micrograms/100 g bw, ip) administration to ovariectomized prepubertal rats. An increase in uterine IGF-I mRNA abundance, approximately 14-fold above untreated controls, was apparent 6 h after estrogen administration and the level achieved exceeded that seen in the uterus from intact mature rats during diestrus. In contrast to the increase in IGF-I expression in the uterus, no significant change in serum IGF-I concentration or hepatic or renal IGF-I mRNA abundance was demonstrable after 17 beta-estradiol injection of ovariectomized prepubertal rats. The increase in uterine IGF-I expression, was similar in both pituitary-intact and hypophysectomized, ovariectomized rats. We believe this is the first report of induction of IGF-I expression by estrogen in vivo. As such, the finding expands the role and significance of IGF-I as a mediator of growth beyond that related to GH.     

慢性低氧对大鼠肺内几种原癌基因表达的影响

本文用原位杂交技术动态观察了慢性低氧大鼠肺内原癌基因ｊｕｎ、ｆｏｓ和ｍｙｂ的表达,结果发现：（１）正常大鼠肺内有一定量的ｊｕｎｍＲＮＡ表达,少见到ｍｙｂ及ｆｏｓ的表达;（２）低氧１周时,ｊｕｎ的表达较正常下降,２周后有所增加。低氧３周后,ｊｕｎ的ｍＲＮＡ表达较正常明显上升;（３）低氧１、２周后,ｍｙｂ的表达明显增加,３周时基本恢复到正常水平;（４）ｆｏｓ原癌基因在低氧１、２周时有一定量的表达,低氧３周时达最大值。提示低氧可刺激大鼠肺内原癌基因ｊｕｎ、ｍｙｂ和ｆｏｓ的表达。     

Antipsychotic drug treatment induces differential gene expression in the rat cortex

Antipsychotic drug treatment is known to modulate gene expression in experimental animals. In this study, candidate target genes for antipsychotic drug action were searched using microarrays after acute clozapine treatment (1, 6 and 24 h) in the rat prefrontal cortex. Microarray data clustering with a self-organizing map algorithm revealed differential expression of genes involved in presynaptic function following acute clozapine treatment. The differential expression of 35 genes most profoundly regulated in expression arrays was further examined using in situ hybridization following acute clozapine, and chronic clozapine and haloperidol treatments. Acute administration of clozapine regulated the expression of chromogranin A, synaptotagmin V and calcineurin A mRNAs in the cortex. Chronic clozapine treatment induced differential cortical expression of chromogranin A, son of sevenless (SoS) and Sec-1. Chronic treatment with haloperidol regulated the mRNA expression of inhibitor of DNA-binding 2 (ID-2) and Rab-12. Furthermore, the expression of visinin-like proteins-1, -2 and -3 was regulated by chronic drug treatments in various brain regions. Our data suggest that acute and chronic treatments with haloperidol and clozapine modulate the expression of genes involved in synaptic function and in regulation of intracellular Ca2+ in cortex.     

Ischemia induces metallothionein III expression in neurons of rat brain

Metallothionein III (MT-III) is a brain-specific member of the metallothionein family and binds zinc in vivo. In order to confirm the precise localization of MT-III in normal rat brain and the change of MT-III expression after transient whole brain ischemia, we raised a high affinity phagemid-antibody specific for rat MT-III. Immunohistochemical analysis revealed that MT-III in normal brain is localized abundantly in neuronal cell bodies in CA1-3 regions of hippocampus, dentate gyrus, cerebral cortex, olfactory bulb and Purkinje cells in cerebellum. This expression pattern of MT-III was similar to that of MT-III mRNA observed by in situ hybridization studies. ELISA and Northern blot analysis revealed that MT-III protein as well as mRNA levels were up-regulated in cerebrum soon after ischemic stress. Immunohistochemical analysis also demonstrated intense staining in neurons in injured brain after ischemia, which distributed in the same regions as in normal brain. These results suggest that MT-III plays an important role in protecting neurons from ischemic insult by reducing neurotoxic zinc levels and inhibits uncontrolled growth of neurites after ischemia.     

Neonatal caffeine induces sex-specific developmental plasticity of the hypoxic respiratory chemoreflex in adult rats

Caffeine is widely used to treat apneas of prematurity during the neonatal period; however, the potential consequences of administering a neonatal caffeine treatment (NCT) during a critical period for respiratory control development are unknown. The present study therefore determined whether NCT in rats alters the hypoxic respiratory chemoreflex measured at adulthood. Newborn rats received either caffeine (15 mg/kg) or water (control) each day from postnatal day 3 to 12. The ventilatory response to a hypoxic challenge (inspired O(2) fraction = 0.12) was first evaluated in awake adult female and male rats using whole body plethysmography. Results showed that NCT increased the initial phase of the breathing frequency response to hypoxia in males only. This result was confirmed in anesthetized and artificially ventilated adult male rats where NCT also increased the phrenic burst frequency response to hypoxia. RT-PCR assessment of mRNA encoding for adenosine A(1A) and A(2A) receptors, dopamine D(2) receptors, and tyrosine hydroxylase in the rat carotid bodies showed that NCT enhanced mRNA expression levels of adenosine A(2A), dopamine D(2) receptors, and tyrosine hydroxylase of males but not females. Subsequent experiments on awake male rats showed that injection of the adenosine A(2A) receptor antagonist ZM2413855 (1 mg/kg ip) before ventilatory measurements abolished, in NCT rats, the enhanced respiratory frequency response observed during the early phase of hypoxia. We propose that NCT elicits a sex-specific increase in the hypoxic respiratory chemoreflex, which is related, at least partially, to an enhancement in adenosine A(2A) receptors in the rat carotid body.     

Opioid receptor blockade in rat nucleus tractus solitarius alters amygdala dynorphin gene expression

It has been suggested that an opioidergic feeding pathway exists between the nucleus of the solitary tract (NTS) and the central nucleus of the amygdala. We studied the following three groups of rats: 1) artificial cerebrospinal fluid (CSF) infused in the NTS, 2) naltrexone (100 microg/day) infused for 13 days in the NTS, and 3) artificial CSF infused in the NTS of rats pair fed to the naltrexone-infused group. Naltrexone administration resulted in a decrease in body weight and food intake. Also, naltrexone infusion increased dynorphin, but not enkephalin, gene expression in the amygdala, independent of the naltrexone-induced reduction in food intake. Gene expression of neuropeptide Y in the arcuate nucleus and neuropeptide Y peptide levels in the paraventricular nucleus did not change because of naltrexone infusion. However, naltrexone induced an increase in serum leptin compared with pair-fed controls. Thus chronic administration of naltrexone in the NTS increased dynorphin gene expression in the amygdala, further supporting an opioidergic feeding pathway between these two brain sites.     

Blood supply of the rat amygdala.

The cerebral vessels of the rat were filled with inks of different colours. The topography of the vessels of the amygdala were reconstructed from serial sections. The circulation of the individual amygdaloid nuclei was studied in detail. The arteries of the amygdala arise from the deep and cortical branches of the internal carotid and middle cerebral arteries. Eight major arteries were found to supply blood to the amygdala. All amygdaloid nuclei receive branches from both arterial trunks. The vast majority of the veins are collected by the middle cerebral and basal veins. Only a small fraction drains into the hippocampal vein. Of particular importance are the veins ending in the basal vein and those cortical ones that run in the rhinal sulcus. All amygdaloid nuclei have a multidirectional drainage.     

The fos proto-oncogene protein:regulation by morphine in the rat hypothalamus

Mechanisms by which opiates alter neuronal functions, including neuroendocrine functions, are not well defined. We have previously demonstrated that morphine rapidly and transiently increases expression of the proto-oncogene c-fos in the rat caudate-putamen. This regulation of the c-fos gene by morphine may represent a portion of the intracellular cascade coupling activation of opiate receptors on the cell surface to subsequent alterations in neuropeptide gene expression. In the present study, we have focussed on effects of morphine on c-fos expression in the ventromedial hypothalamus, which contains estrogen-concentrating neurons and a large number of neurons expressing the opioid proenkephalin and Proopiomelanocortin. The hypothalamus has been identified as a "final common pathway" between the remainder of the central nervous system and the pituitary gland. As a marker for c-fos expression, we have detected pp50 c-fos (FOS) protein immunocytochemically, using a polyclonal antibody to the M peptide of FOS, and revealed an intense nuclear stain in many neurons. Labeled nuclei were drawn by camera lucida from 12 matched sections (one side only) covering the rostral and middle levels of the ventromedial nucleus of six rats given morphine and six given phosphate buffered saline. Morphine treatment significantly increased the number and density of immuno-labeled nuclei in the ventromedial nucleus, but not in the arcuate nucleus. These results suggest effects of morphine (directly or indirectly) on neurons in the ventromedial hypothalamic nucleus, despite the relative absence of morphine receptors in this nucleus. These results may also provide an anatomical basis for neuroendocrine alterations following morphine treatment.