James Buggy

Circadian rhythmicity after neural transplant to hamster third ventricle: specificity of suprachiasmatic nuclei.

Neural transplants into the third ventricle utilized to quantitatively assess the effectiveness of fetal tissue from selected brain sites in restoring circadian locomotor rhythmicity of adult hamsters rendered arrhythmic by lesions of the suprachiasmatic nuclei (SCN). Circadian function was continuously monitored in recording wheel cages under controlled environmental conditions. Animals which remained arrhythmic for 3-4 weeks after SCN lesions received transplants of neural tissue from 13-14-day-old fetuses: either SCN tissue or non-SCN tissue (cerebral cortex or hypothalamus excluding SCN). Quantitative evaluation of the data indicated partial restoration of circadian rhythmicity in 37% of 19 animals with SCN transplants, but in 0% of the 9 animals with non-SCN neural transplants. The mean time for reappearance of rhythmicity was 20 days after SCN transplantation. Animals were sacrificed 8-10 weeks after transplantation for histological analysis in order to visualize lesion placement and to characterize transplants. The cytoarchitecture and neuropeptide organization of the transplants were consistent with the brain region. Only SCN transplants were characterized by aggregates of small neurons with codistributed immunoreactivity for SCN-characteristic neuropeptides.

Proto-oncogene c-fos and the regulation of vasopressin gene expression during dehydration

Secretion of the antidiuretic hormone (ADH) vasopressin is increased when body fluid homeostasis is disturbed by dehydration. Associated with this increased secretion is an elevation of vasopressin mRNA in magnocellular hypothalamic neurons projecting to the posterior pituitary. The proto-oncogene c-fos codes for a nuclear phospho-protein Fos which binds to specific DNA elements and acts as a transcriptional regulator coupling short-term extracellular stimuli to long-term responses by altering secondary target gene expression. This study in rats examined the time courses of dehydration induced c-fos expression and the change of vasopressin gene expression in the magnocellular neurons of the hypothalamus. Immunocytochemical and in situ hybridization study demonstrated that c-fos was induced by acute intracellular dehydration in the hypothalamic magnocellular nuclei of paraventricular (PVN), supraoptic (SON), and accessory groups such as nucleus circularis. Double-label immunocytochemical study co-localized Fos and vasopressin-neurophysin immunoreactivity in the same magnocellular neurons in the SON and PVN. In situ hybridization analysis after acute dehydration revealed a rapid and transient c-fos induction followed by a persistent increase in vasopressin mRNA for up to 2 days even after rehydration. Furthermore, prevention of c-fos translation by pretreatment with protein synthesis inhibitor cycloheximide attenuated this dehydration induced increase in vasopressin mRNA. This study demonstrated that an increase in vasopressin transcription after acute dehydration is dependent on an early phase of protein synthesis.

Sodium appetite decreased by central angiotensin blockade.

Disturbances in body water and electrolytes that trigger sodium appetite, such as sodium depletion or hypovolemia, are potent activators of the renin-angiotensin system. In the absence of an actual deficit in body fluids, angiotensin injections are adequate to stimulate increased sodium ingestion. To assess whether angiotensin is a significant mediator of sodium appetite induced by acute alterations in body fluids, sodium intake was examined in rats during central or peripheral angiotensin blockade. Central blockade of angiotensin receptors by intracerebroventricular (ICVT) injection of the analogue antagonist saralasin decreased (but did not eliminate) sodium intake after polethylene glycol-induced hypovolemia or sodium depletion resulting from dialysis against glucose. Conversely, peripheral blockade of angiotensin converting enzyme with orally active captopril potentiated rather than decreased sodium appetite and stimulated water intake after sodium depletion. This increased water and salt intake after peripheral inhibition of converting enzyme was reversed, however, by concurrent central blockade of angiotensin receptors. These data support the hypothesis that angiotensin participates in sodium appetite associated with acute alteration in body fluids. Furthermore, the brain is the site at which angiotensin exerts its influence on sodium appetite. While the involvement of angiotensin of brain origin is not ruled out, the change in sodium appetite after peripheral blockade of converting enzyme suggests that circulating angiotensin derived from renal renin may interact with central angiotensin receptors regulating sodium appetite.

Angiotensin-estrogen central interaction: localization and mechanism.

Intracerebroventricular (ICVT) administration of estradiol benzoate (EB) to ovariectomized female rats decreased drinking and pressor responses to central injections of angiotensin II (AII). Estrogen treatment does not have this effect in male rats. As EB given ICVT reaches many brain areas, the site of action of EB was localized using crystalline implants of EB in the medial preoptic area or the ventromedial nucleus of the hypothalamus. These areas were chosen as they have a high density of estrogen receptors. Only medial preoptic area application of estrogen decreased angiotensin II-induced drinking. Angiotensin receptor binding was examined in homogenates from different brain regions to determine if the mechanism through which estrogen decreases central responses to AII involves altered receptor function. Systemic EB did not affect AII receptor binding in several brain regions but binding was decreased in homogenates from the preoptic area and septum-thalamus blocks which encompassed structures (median preoptic nucleus, organum vasculosum, and subfornical organ) implicated in central actions of AII. The sex specificity of the effect of estrogen was dependent on sexual differentiation of the brain. Manipulation of the neonatal hormone environment, which alters this brain differentiation, also altered the characteristic responses of the two sexes to estrogen. Neonatal androgenization of females, which causes masculinization and defeminization, resulted in animals which as adults no longer responded to EB with decreased drinking. On the other hand, preventing the development of a male brain by neonatal castration produced animals which as adults tended to decrease their drinking following estrogen. In summary, this study found that EB acts in the preoptic area to depress AII-induced responses by a site specific modulation of central AII receptors. Alteration of early brain development changed the responses of the two sexes to estrogen, perhaps by altering sexual differentiation of the preoptic area.

Depression of ad lib and angiotensin-induced sodium intake at oestrus.

During the rat oestrous cycle, cholinergic-stimulated water intake is constant across days; while both daily ad lib and angiotensin II-stimulated water intakes are least on the day of oestrus. If permitted to self select, however, rats will elect to drink sodium chloride solutions as well as water under ad lib or angiotensin-stimulated conditions. Therefore, adult female rats with continuous access to both water and 1.8% NaCl were studied to determine if saline intake also varied with the oestrous cycle. Ad lib drinking and drinking stimulated by intracerebroventricular injections of angiotensin II or carbachol were monitored daily for the duration of an entire oestrous cycle. Both ad lib and angiotensin-induced sodium intakes were depressed at oestrus, while the minimal sodium intake seen after carbachol was not changed. The depression in sodium intake so mirrored that observed for water intake that the ratio of saline to water intake did not vary over the oestrous cycle.

AP-1 DNA binding activity induced by hyperosmolality in the rat hypothalamic supraoptic and paraventricular nuclei

Immediate early gene products (c-fos, c-jun and their cognates) act as transcription factors coupling physiologically relevant stimuli to long-term responses by binding to the AP-1 site in the promoter region of target genes. The induction of c-fos has been identified in the paraventricular (PVN) and supraoptic (SON) hypothalamic magnocellular nuclei after hyperosmotic stimulation by using in situ hybridization and immunocytochemistry. In this study, AP-1 DNA binding activity, an indicator of the functional form of the c-fos transcription factor, was examined in nuclear extracts prepared from these brain regions using an electrophoretic mobility shift assay and a labeled oligonucleotide containing the AP-1 consensus sequence. Two hours after hypertonic saline injection (i.p.), rats were killed and nuclear proteins were extracted from tissue punches of brain regions to assess AP-1 binding activity. Hyperosmolality induced an increase of AP-1 binding activity in nuclear protein from SON and PVN, but not striatum. This binding was competitively displaced by excess unlabeled AP-1 oligonucleotide whereas addition of increasing amounts of unlabeled SP-1 oligonucleotide (promoter site on housekeeping genes for the ubiquitous SP-1 transcription factor) did not decrease the binding. The binding protein was shown to contain c-Fos/Fra and c-Jun since addition of c-Fos/Fra antiserum formed a supershift of the DNA, protein and antibody complex, and c-Jun antibody blocked the protein DNA binding. These data suggest that hyperosmolality leads to a selective and specific increase in AP-1 DNA binding activity which may be responsible for regulating secondary target gene expression in the hypothalamic SON and PVN.

Brain functional activity during PAG stimulation-produced analgesia: A 2-DG study

The autoradiographic 2-deoxyglucose method for regional cerebral metabolic activity was modified for use with tritium label to determine which brain stem and spinal cord nuclei changed their functional neural activity during periaqueductal gray stimulation-produced analgesia. The greatest changes in activity during electrical stimulation of the periaqueductal gray occurred in nucleus paragigantocellularis, the ventral portion of the nucleus reticularis gigantocellularis, and the nucleus cuneiformis. Substantial increases in metabolic activity were also evident in the spinal trigeminal nucleus and the substantia gelatinosa. Many of the regions which displayed increased functional activity in the present study have been shown to possess substantial enkephalin immunoreactivity. While several of these structures have previously been implicated in modulation of nociceptive transmission, this study raises the possibility that other brain stem nuclei may also participate in analgesic mechanisms.

3H-2-deoxyglucose uptake after electrical stimulation of cardioactive sites in anterior medial cortex in rabbits.

The anterior medial cortex is an important integrative area for cardiovascular adjustments occurring during learning and conditioning. The autoradiographic 3H-2-deoxyglucose (3H-2DG) method for regional cerebral metabolic activity was used to identify other forebrain regions associated with cardiovascular adjustments elicited by electrical stimulation of anterior medial cortex. Rabbits that received anterior midline stimulation that produced bradycardia and depressor responses showed increased metabolic activity in the ipsilateral mediodorsal (MD) nucleus of the thalamus and the dorsal aspect of the claustrum. Two additional animals with anterior cortical placements in the infralimbic and anterior limbic areas showed ipsilateral increased activity in perirhinal cortex, but more dorsolateral placements in the precentral agranular area did not produce increased perirhinal activity. Control animals did not show this pattern of activity. These data suggest that MD and claustrum participate in a neural circuit which mediates cardiovascular adjustments similar to those elicited by Pavlovian conditioning contingencies.

Muscarinic cholinergic receptors in the rat deep cerebellar nuclei: a quantitative autoradiographic study

The distribution of muscarinic cholinergic receptors within the rat deep cerebellar nuclei was analyzed using in vitro receptor binding of [3H]quinuclidinylbenzilate (QNB) in conjunction with autoradiography. The highest density of QNB binding sites occurred in the lateral cerebellar (dentate) nucleus. Interpositus nuclei displayed an intermediate density of muscarinic cholinergic binding sites with the posterior interpositus nucleus demonstrating higher binding than the anterior nucleus. The fastigial (medial) cerebellar nucleus exhibited the lowest levels of QNB binding among the four cerebellar nuclei. These results indicate that muscarinic cholinergic receptors are present in the deep cerebellar nuclei and that differences in receptor density occur among the four nuclear groups.

Motivating physical activity in animal models

Abstract A variety of physical activity models have been developed for animal studies to simulate the range of activities engaged by humans, from intense athletic training and competition to normal activities of daily life. These animal models of exercise cover a spectrum from high‐intensity to low‐intensity exercise with brief to long durations, and either forced or voluntary in nature. The varieties of animal models of exercise adapted for use in rats and mice, the most frequently studied animals, reviewed here include resistance or weight‐lifting exercise, treadmill running, voluntary wheel running, enriched environment, and skills training. The motivational factors used to induce exercise in these various animal models are considered, and the use of rewarding brain stimulation to motivate exercise in animals is described as well as recent findings that suggest that exercise in itself has rewarding aspects. Challenges of future animal studies of exercise are to better understand quantitative and qualitative aspects of the dimension of exercise and physical activity, and to move beyond descriptive reports of whether or not exercise has an effect with research that elucidates the mechanisms of exercise effects in both the central nervous system and periphery