Induction of Fos-like immunoreactivity in the rat brain following Pasteurella multocida endotoxin administration.

The functional neuroanatomy of the immune system link to the CNS was investigated by assessing neuronal activity with Fos immunohistochemistry following systemic lipopolysaccharide (LPS) administration. Two hours after LPS robust Fos-like immunoreactivity (Fos-IR) was observed in several nuclear groups in the brain including the paraventricular and supraoptic nuclei of the hypothalamus, central nucleus of the amygdala, and nucleus of the solitary tract. A similar but diminished pattern of Fos-IR was present at 6 hours and was absent 24 hours after LPS administration. Investigation of the functional neuroanatomy of the acute phase reaction could prove to be critical in enhancing the ability of individuals to combat insults such as tissue damage and inflammation. The central nervous system (CNS), particularly the hypothalamus, is intimately involved in the coordination of the various aspects of the acute phase reaction (reviewed in 1). Understanding the functional neuroanatomy by which the brain responds to immune system challenges would greatly augment the ability to control the deleterious and enhance the beneficial aspects of the acute phase reaction. In this study we have used lipopolysaccharide (LPS or endotoxin) administration as an experimental model to study immune system activation. LPS is a complex glycolipid and a component of the outer membrane of most Gram-negative bacteria (2). Administration of LPS has been demonstrated to induce the secretion of several proteins including interleukin-1 (IL-1), tumor necrosis factor (TNF), and interleukin-6 (IL-6; reviewed in 3). Further, it has been hypothesized that LPS induction of IL-1 and TNF is the key event in the pathogenesis of Gram-negative bacterial septic shock syndrome (2). Many recent studies have utilized immunohistochemistry for Fos, the product of the immediate early gene c-fos, as a marker of neuronal activation. Fos is a nuclear-binding protein that is expressed at increased levels in activated neurons (4). Although the exact function of Fos in the CNS is still unknown, it is thought that Fos is transcribed after cellular stimulation as a means to convert a stimulus into long-term genetic action (for reviews see 5,6). This study investigated the activation of the CNS by peripherally administered LPS isolated from the bacterium Pasteurella multocida. As a marker of neuronal activation, immunohistochemistry for the Fos protein was performed and image analysis was utilized to quantify the Fos induction in the CNS.

Localization of cholecystokinin binding sites in the adult and developing Brazilian opossum brain.

Cholecystokinin (CCK) is now recognized as one of the most abundant peptides in the mammalian central nervous system. We have previously used immunohistochemistry to localize CCK in the adult and developing Brazilian opossum brain. However, little is known about the distribution of CCK binding sites in the developing mammalian brain. Therefore, to further our knowledge of the sites of action for CCK during development, we initiated a series of studies to localize CCK binding sites in the adult and developing Brazilian opossum. This species was chosen because pups are born in a fetus-like state. Receptor autoradiography was performed on coronally sectioned brains of 1 to 60 day postnatal (PN) animals and adults with 125I-Bolton Hunter-CCK-8 as the radioligand. Binding is evident in the 1PN opossum brainstem and is observed in the developing forebrain by 5PN. Region-specific binding increases during development, and binding in the 35PN brain resembles the adult pattern. Binding is evident prior to the detection of CCK-like immunoreactivity in many areas. The facial motor nucleus is identifiable and exhibits high levels of binding in Brazilian opossum pups of 10 to 35 days of age. However, binding is undetectable in the facial motor nucleus of 45 and 60PN pups. In general, the binding patterns for CCK in the adult opossum resemble those of other mammals and likely mediate similar physiological functions. However, some cholecystokininergic pathways appear to be unique to neonatal mammals.

Developmental distribution of GFAP and vimentin in the Brazilian opossum brain.

Cells of glial origin are involved in the morphogenesis of the mammalian central nervous system (CNS). Characterization of glial-associated proteins during neurogenesis and differentiation may aid in understanding the complexity of CNS development. We have utilized immunoblotting and immunohistochemistry to characterize the developmental profiles of glial fibrillary acidic protein (GFAP) and vimentin (VIM) in the brain of the Brazilian opossum, Monodelphis domestica. Typical of marsupials, CNS morphogenesis and neurogenesis in the opossum extend well into the postnatal period. Opossum GFAP and VIM were found as single bands at molecular weights consistent with those reported for other species, thus indicating conservation of the VIM and GFAP proteins through mammalian evolution. Differential developmental trends were observed for both proteins with relative VIM levels decreasing and GFAP levels increasing with age. Vimentin-like immunoreactivity (VIM-IR) was present at day 1 of postnatal life throughout the brain. The density of VIM-IR was maximal at 10 and 15 days postnatal (especially in radial glial elements) and decreased slightly by 25 days postnatal. In the adult brain, VIM-IR was markedly reduced compared to that of younger ages. In contrast, GFAP-like immunoreactivity (GFAP-IR) in the brain of Monodelphis increased dramatically with age. No GFAP-IR was observed in the 1 and 5 day postnatal brains. By 25 days postnatal, the pattern of GFAP-IR in the brainstem resembled that of the adult. In the forebrain, more GFAP-IR was present than at younger ages. The adult distribution of GFAP-IR was very similar to that reported for other mammalian species. These results indicate that GFAP and VIM are reciprocally related during periods of morphogenesis and differentiation of the opossum brain.

Differential down-regulation of estrogen receptor-like immunoreactivity by estradiol in the female Brazilian opossum brain.

The effects of estrus and prolonged estradiol benzoate (EB) treatment on estrogen receptor-like immunoreactivity (ER-LI) were studied in the female Brazilian opossum (monodelphis domestica) brain. In general, both estrus animals and EB treated animals had reduced ER-LI in specific brain regions when compared to ovariectomized controls. These results support a down-regulatory role for estrogen in relation to its receptor in the brain, and indicate that ER-LI is differentially regulated in the Monodelphis forebrain.

Localization of cells containing estrogen receptor-like immunoreactivity in the Brazilian opossum brain.

The Brazilian opossum (Monodelphis domestica) is a small, pouchless marsupial whose young are born in an immature, sexually undifferentiated state. Etgen and Fadem, and Handa and coworkers have biochemically detected and characterized estrogen receptors in the forebrain of the Brazilian opossum. In this study, we have examined the distribution of estrogen receptor-like immunoreactive (ER-LI) cells in the brains of gonadectomized male and female Brazilian opossums using Abbott H222 rat monoclonal estrogen receptor antibody (H222 is a gift of Abbott Labs). An indirect immunohistochemical procedure employing the Vectastain Elite system and a nickel-enhanced DAB chromogen was used. A large number of ER-LI cell nuclei were observed in the medial preoptic area, ventral septal nucleus, medial division of the bed nucleus of the stria terminalis, lateral part of the ventromedial hypothalamus, premammillary nucleus, arcuate nucleus, posterior amygdaloid nucleus, and the midbrain central grey. Lower numbers of ER-LI cell nuclei were observed in the intermediate subdivision of the lateral septal nucleus, and in the anterior, medial, and posterior cortical amygdaloid nuclei. The anatomical distribution of ER-LI in the Brazilian opossum brain is similar to that which has been reported for estrogen binding sites following biochemical analysis. Based on these findings, we believe specific regions of the Brazilian opossum brain may serve as substrata for the action of estrogen in the adult. In addition, these results are supportive of the use of this animal model to investigate the organizational effects of estrogen on the developing central nervous system.

Ontogeny of cells containing estrogen receptor-like immunoreactivity in the Brazilian opossum brain.

In this study, we have used the Brazilian short-tailed opossum (Monodelphis domestica) as a model to study the ontogeny of estrogen receptors in the mammalian brain. Monodelphis is a small, pouchless marsupial which breeds well under laboratory conditions and whose young are born in an immature sexually undifferentiated state. The Abbott H222 monoclonal rat estrogen receptor antibody (gift of Abbott Laboratories) was utilized in an indirect immunohistochemical procedure to detect estrogen receptors in developing opossum brains. Estrogen receptors were first expressed in the dorsomedial and ventromedial hypothalamus of the opossum 10 days after birth (10PN). Most regions that contained estrogen receptor-like immunoreactivity (ER LI) in the adult opossum contained ER LI at 15 PN. These areas include the lateral septum, medial preoptic area, bed nucleus of the stria terminalis, periventricular preoptic area and hypothalamus, amygdala, dorsomedial and ventromedial hypothalamic nuclei, arcuate nucleus, ventral premammillary nucleus, and the midbrain central grey. The number of cells that contain ER LI increased through 60PN in all regions that will contain ER LI in the adult opossum. These results indicate that estrogen receptors are present in early development of the Monodelphis brain and may mark the beginning of a critical period for sexual differentiation of the opossum brain.

Ontogeny of cholecystokinin-like immunoreactivity in the Brazilian opossum brain.

We have studied the anatomical distribution of cholecystokinin-like immunoreactive (CCK-IR) somata and fibers in the brain of the adult and developing Brazilian short-tailed opossum, Monodelphis domestica. Animals ranged in age from the day of birth (1PN) to young adulthood (180PN). A nickel enhanced, avidin-biotin, indirect immunohistochemical technique was used to identify CCK-IR structures. Somata containing CCK immunoreactivity were observed in the cerebral cortex, hippocampus, hypothalamus, thalamus, midbrain, and brainstem in the adult. Cholecystokinin immunoreactive fibers had a wide distribution in the adult Monodelphis brain. The only major region of the brain that did not contain CCK-IR fibers was the cerebellum. The earliest expression of CCK immunoreactivity was found in fibers in the dorsal brainstem of 5-day-old opossum pups. It is possible that the CCK-IR fibers in the brainstem at 5PN are of vagal origin. Cholecystokinin immunoreactive somata were observed in the brainstem on 10PN. The CCK-IR cell bodies observed in the brainstem at 10PN may mark the first expression of CCK-IR elements intrinsic to the brain. A broad spectrum of patterns of onset of CCK expression was observed in the opossum brain. The early occurrence and varied ontogenesis of CCK-IR structures indicates CCK may be involved in the function of a variety of circuits from the brainstem to the cerebral cortex. The early expression of CCK-IR structures in the dorsal brainstem suggests that CCK may modulate feeding behavior in the Monodelphis neonate. Cholecystokinin immunoreactivity in forebrain structures such as the suprachiasmatic nucleus, medial preoptic area, thalamus and cortical structures indicates that CCK may also be involved in circadian rhythmicity, reproductive functions, as well as the state of arousal of the Brazilian opossum. The ontogenic timing of CCK immunoreactivity in specific circuitry also indicates that CCK expression does not occur simultaneously throughout the brain. This pattern of CCK onset may relate to the temporal need for CCK in specific circuits of the central nervous system (CNS) during development.

Galanin-like immunoreactivity in the adult and developing Brazilian opossum brain.

The distribution of galanin-like immunoreactivity has been characterized in the brain of the adult and developing Brazilian opossum (Monodelphis domestica). Two commercially available antisera were used to examine the distribution of galanin-like immunoreactive (GAL-IR) cells and fibers. Nuclear groups containing GAL-IR cell bodies and fibers were seen throughout the adult opossum brain. The distribution of GAL-IR elements seen is similar to that reported for other mammals. Based on these findings, we believe that galanin may have similar physiological functions in the adult Brazilian opossum as has been reported for other mammals. In the developing brain, GAL-IR structures were seen as early as 1 day postnatal (PN) in the developing hypothalamus and brainstem. By days 5 and 10 PN, there was a robust expression of galanin-like immunoreactivity in specific regions of the brain. Since neurogenesis and brain morphogenesis are actively occuring postnatally in the opossum, galanin may be playing a role in the differentation of specific regions of the brain.

Postnatal neurogenesis of the hypothalamic paraventricular and supraoptic nuclei in the Brazilian opossum brain.

We have used bromodeoxyuridine (BrdU) single and BrdU-arginine vasopressin-oxytocin (BrdU-AVP-OT) double and triple label immunohistochemistry to characterize postnatal neurogenesis of the supraoptic and paraventricular nuclei in the Brazilian opossum. Developing pups received a single injection of BrdU between days 1 and 11 postnatally. All brains were collected on day 60 of postnatal life (60 PN). Single label BrdU immunohistochemistry revealed that an injection at 1 PN resulted in heavy labelling in the hypothalamus including the area of the paraventricular nucleus, whereas only approximately one third of the cells in the supraoptic nucleus were labelled. Analysis of data indicated that neurogenesis of the supraoptic and paraventricular nuclei is completed by days 5 and 7 PN, respectively. Double and triple label immunohistochemistry demonstrated that following BrdU injection on day 1 or 2 PN, few of the AVP and OT secreting cells in the supraoptic nucleus were double labelled with either peptide and BrdU, and no double labelled cells were seen following BrdU injection on day 5 PN. Similarly, in the paraventricular nucleus most of the AVP and OT secreting magnocellular cells were not double labelled with either peptide and BrdU. Whereas several double labelled cells were observed in the parvicellular part following BrdU injection on day 1 or 2 PN. No double labelled cells were present in any component of the paraventricular nucleus following injection on day 7 PN or later. These results indicate that the majority of the AVP and OT secreting magnocellular neurons are born prenatally and the OT and AVP parvicellular group of neurons are born during postnatal life. Our results also demonstrate that in contrast to that of eutherian rodents such as the rat and mouse, neurogenesis in the opossum hypothalamus continues into the postnatal period and provides a unique opportunity to study the neuroanatomical development of diverse regions such as the paraventricular nucleus.

The effects of medical evidence and pain intensity on medical student judgments of chronic pain patients.

This study examined symptom judgments made by medical students of hypothetical chronic low back pain patients. Eight vignettes were varied as to the pain intensity reported by the hypothetical patient (low vs. moderate vs. high vs. very high) and the availability of medical evidence supportive of the pain report (present vs. absent). Ninety-five subjects read vignettes and made judgments of patient emotional distress, pain intensity, and pain-related disability. Subjects significantly discounted pain level when intensity was high but slightly augmented pain level when intensity was low. Judgments of pain and disability were higher for patients for whom medical evidence was present compared to those for whom it was absent. The results support and extend previous research on the effects of situational and patient variables on observer pain judgments. Future research should examine the influence of these biasing variables on the assessment and treatment of chronic pain patients.

Cholecystokinin like immunoreactivity in the brains of young Meishan and Duroc pigs(4).

SUMMARY: Cholecystokinin (CCK), a peptide found in both the gastrointestinal tract and brain, has been shown to be involved in the control of feed intake in a variety of animals including the pig. Chinese breeds of pigs such as the Meishan are noted for slow growth and heavy adipose deposition. In this study we have described the regional cholecystokinin-like immunoreactivity (CCK-IR) concentrations in the brain of young Duroc and Meishan pigs utilizing radioimmunoassay. Brains of days 1, 10, and 20 postnatal pigs from each breed were examined. The CCK-IR increased with age in all three areas examined (cortex, medulla, and hypothalamus). The cortical concentrations rose significantly from days 1 to 10 and from days 10 to 20. The levels in the hypothalamus and medulla increased significantly between days 1 and 20. There were no statistically significant differences in CCK-IR between the breeds at any of the three ages examined. Our results indicate that a rise in CCK-IR in the regions of the brain involved in the control of feed intake may parallel the ability of the young pigs to assimilate nutrients from a solid diet. ZUSAMMENFASSUNG: Cholecystokinin-ähnliche Immunreaktivität in den Gehirnen junger Meishan- und Durocschweine Das Peptid Cholecystokinin (CCK) wird im Gastrointestinaltrakt und im Gehirn gefunden und beeinflußt Futteraufnahme in einer Reihe von Tieren einschließlich Schwein. Chinesische Rassen wie Meishan sind wegen ihres langsamen Wachstums und der starken Fettablagerung bekannt. In dieser Studie beschreiben wir regionale Cholecystokinin-ähnliche Immunreaktivitäts-(CCK-IR)Konzentrationen im Gehirn junger Duroc- und Meishantiere, mittels Radioimmunassay bestimmt. Gehirne von 1, 10 und 20 Tage alten Ferkeln jeder Rasse wurden untersucht. CCK-IR nahm mit dem Alter in allen drei untersuchten Organen zu (Kortex, Medulla und Hypothalamus). Die kortikalen Spiegel stiegen vom Tag 1 bis 10 und vom Tag 10 bis 20 signifikant, die des Hypothalamus und der Medulla zwischen Tag 1 und 20. Zwischen Rassen waren keine statistisch signifikanten Unterschiede in CCK-IR. Unsere Ergebnisse zeigen einen Anstieg in CCK-IR in Gehirnregionen, die bei Futteraufnahme involviert sind, und könnten der Fähigkeit junger Schweine zur Assimilation von Nährstoffen aus festen Rationen parallel sein. RÉSUMÉ: Immunoréactivité de type cholécyctokinine dans le cerveau de jeunes porcs Meishan et Duroc La cholécystokinine (CCK) est peptide présent à la fois dans le tractus gastro-intestinal et le cerveau. Son influence sur le contrôle de la consommation alimentaire a été mise en évidence dans plusieurs espèces d'animaux dont le porc. Les races porcines chinoises comme la Meishan présentent une faible croissance et un dépôt de gras important. Cette étude décrit les concentrations régionales des composés présentant une réactivité immunologique similaire à la cholécystokinine (CCK-IR), déterminés par dosage radio-immunologique, dans le cerveau de porcelets Duroc et Meishan. Des cerveaux de porcelets de chacune des deux races ont été examinés à 1, 10 et 20 jours après la naissance. La CCK-IR augmente avec l'âge dans les trois zones du cerveau examinées (cortex, moelle et hypothalamus). Les concentrations corticales augmentent significativement etre 1 et 10 jours d'â ainsi qu'entre 10 et 20 jours d'âge. Les concentrations dans l'hypothalamus et la moelle croissent significativement entre 1 et 20 jours d'âge. Les différences de CCK-IR entre races ne sont statistiquement significatives pour aucun des trois âges étudiés. Nos résultats montrent qu'un accroissement de CCK-IR dans les régions du cerveau impliquées dans le contrôle de la consommation alimentaire peutètre mis en parallèle avec la capacité de porcelets à assimiler les éléments nutritifs d'un régime alimentaire solide. RESUMEN: Immunoreactividad similar a la de la colecistoquinina en los cerebros de lechones jovenes Meishan y Duroc La colecistoquinina (CCK), un péptido que se encuentra en el sistema digestivo y el cerebro, esta rela-cionado con el control del apetito en varias especies, incluido el cerdo. Las razas chinas como la Meishan se distinguen por su crecimiento lento y deposición de tejido adiposo. En este estudio hemos descrito las concentraciones de sustancias immunoló-gicamente similares a la colecistoquinina (CCK-IR) en tres regiones del cerebro de lechones Duroc y Meishan utilizando radioimmunoensayo. Los cerebros de cerdos de 1, 10 y 20 días de edad de cada raza fueron examinados. La CCK-IR aumentó con la edad en las tres áreas examinadas (córtex, médula e hipotálamo). Las concentraciones en el córtex aumentaron significativamente de 1 a 10 y de 10 a 20 dias de edad. Las concentraciones en el hipotálamo y médula aumentaron significativamente entre los días 1 y 20. No hubo diferencias significativas en CCK-IR entre las razas en ninguna de las edades estudiadas. Nuestros resultados indican que un aumento en CCK-IR en las regiones del cerebro que participan en el control del consumo de alimente podrían ir paralelas a la habilidad de los lechones para asimilar nutrientes de una dieta sólida.

Relaxin-induced expression of Fos in the forebrain of the late pregnant rat.

Relaxin, administered parenterally, has been shown to increase the release of oxytocin (OT) into the circulation and increase the firing rate of OTergic neurons. The objective of the present study was to determine if relaxin administration can result in the expression of a transcription factor, suggesting that it alters transcriptional activity within OTergic neurons at the level of the hypothalamus. Primigravid rats were ovariectomized and a jugular cannula was inserted on day 11 of gestation (g11). Pregnancy was maintained by implanting 17 beta-estradiol and progesterone caplets subcutaneously at the time of ovariectomy. At gl9, rats were challenged with intravenous relaxin or isotonic saline and the brains were removed for study. Immunohistochemistry was performed on coronal brain sections, utilizing Fos as a marker of cellular activation. In the group receiving relaxin, Fos-like immunoreactivity (Fos-IR) was abundant only in the supraoptic (SON) and paraventricular nuclei (PVN) of the hypothalamus as well as in the subfornical organ (SFO). In contrast, Fos-IR in the group given isotonic saline was lacking in these three brain regions. A double label study using antibodies against Fos and OT demonstrated that a majority of the Fos-labeled cells in the hypothalamus were OTergic. Because Fos can act as a transcription factor, we interpret these data to indicate that transcription within OTergic cells is altered following relaxin administration, with abundant Fos-IR being limited to the SON and PVN of the hypothalamus and the SFO during late pregnancy in the rat.

Medical education change: a detailed study of six medical schools.

This article reports a comparative case study of six selected USA medical schools, undertaken to identify factors that facilitate or obstruct innovation in medical education. The findings suggest that the culture of each medical school results from a combination of intra-institutional and external factors. Together these forces influence substantially the fate of educational innovations. The institutional culture influences critical elements such as educational philosophy, leadership and resources provided in support of innovation. Equally important, the culture shapes the level and type of change a school considers and implements. The findings also suggest that the availability of resources and the creative impetus present in schools giving priority to research can benefit the educational goals and facilitate educational change.