Biomedical subjects

P J Shiromani

Publications and source records attributed to P J Shiromani.

At least 19 recordsLinked to original sources

PubMed2005

Beneficial effects of regular exercise on sleep in old F344 rats.

With aging there is a significant decline in the normal architecture of sleep and a reduction in the diurnal amplitude of core body temperature. Regular moderate exercise has been shown to have a positive impact in the elderly and here we investigate whether sleep-wake patterning can also be improved. Young (3 months) and old (22 months) F344 rats were exercised once a day for 50min at night onset over an 8-week period. Thereafter, polysomnographic recordings were obtained immediately after exercise. To determine the lasting consequences of exercise, sleep was also recorded 2 days and 2 weeks after exercise had ended. Old rats that were exercised had a significant weight loss, were awake more during the last third of their active period, had less sleep fragmentation and the amplitude of the diurnal rhythm of core body temperature was significantly increased. Old exercised rats also had an overall increase in the amplitude of EEG power (0.5-16Hz) during wake and theta EEG power during REM sleep. In young rats regular exercise increased EEG delta power (0.5-4Hz) during NREM sleep. Our data indicate regular exercise in old rats improves sleep architecture, EEG power and diurnal rhythm of temperature.

Aging↗

PubMed2005

Insomnia following hypocretin2-saporin lesions of the substantia nigra.

The neuropeptide hypocretin, also known as orexin, has been implicated in waking since its deletion leads to the sleep disorder narcolepsy. Hypocretin neurons project to major arousal areas, and in an effort to determine which region is responsible for the changes in sleep-wake architecture we have developed the neurotoxin hypocretin2-saporin, which lesions hypocretin receptor bearing neurons. Here, in rats, we investigate the effects of hypocretin2-saporin lesions of the substantia nigra and ventral tegmental area in the regulation of sleep and wakefulness. Bilateral injection of hypocretin2-sap into both the ventral tegmental area and substantia nigra (92 and 184 ng/microl, 0.25 microl in the ventral tegmental area and 0.5 microl in the substantia nigra) or into the substantia nigra alone (184 ng/microl, 0.5 microl) produced insomnia. The insomnia seemed to be associated with a large increase in locomotion on days 4 and 6 postinjection, as hyperactivity and stereotypic movements were consistently observed on the video recordings in all lesioned rats. In these rats, a nearly complete loss of both tyrosine hydroxylase and neuron-specific nuclear protein (neuronal nuclei) immunoreactive cells in the substantia nigra as well as diminution of tyrosine hydroxylase-immunoreactive fibers in the caudate putamen was found. Following bilateral injection of hypocretin2-sap at a lower concentration (46 ng/microl, 0.25 microl in the ventral tegmental area and 0.5 microl in the substantia nigra), very little reduction in the number of tyrosine hydroxylase- and neuronal nuclei-immunoreactive neurons and only a temporary increase in wakefulness (17.4% increase during light-off period on day 6 postinjection) were observed. Ventral tegmental area lesions (184 ng/mul of hypocretin2-sap, 0.25 microl, bilateral injections) did not produce significant changes in sleep, although most of the tyrosine hydroxylase- and neuronal nuclei-immunoreactive neurons in the ventral tegmental area were destroyed. Insomnia following hypocretin2-sap lesions of the substantia nigra could be secondary to increased motor activity resulting from reduction of tonic inhibitory control by the substantia nigra.

Animals↗

PubMed2004

The diurnal rhythm of adenosine levels in the basal forebrain of young and old rats.

There are significant decrements in sleep with age. These include fragmentation of sleep, increased wake time, decrease in the length of sleep bouts, decrease in the amplitude of the diurnal rhythm of sleep, decrease in rapid eye movement sleep and a profound decrease in electroencephalogram Delta power (0.3-4 Hz). Old rats also have less sleep in response to 12 h-prolonged wakefulness (W) indicating a reduction in sleep drive with age. The mechanism contributing to the decline in sleep with aging is not known but cannot be attributed to loss of neurons implicated in sleep since the numbers of neurons in the ventral lateral preoptic area, a region implicated in generating sleep, is similar between young (3.5 months) and old (21.5 months) rats. One possibility for the reduced sleep drive with age is that sleep-wake active neurons may be stimulated less as a result of a decline in endogenous sleep factors. Here, we test this hypothesis by focusing on the purine, adenosine (AD), one such sleep factor that increases after prolonged W. In experiment 1, microdialysis measurements of AD in the basal forebrain at 1 h intervals reveal that old (21.5 months) rats have more extracellular levels of AD compared with young rats across the 24 h diurnal cycle. In experiment 2, old rats kept awake for 6 h (first half of lights-on period) accumulated more AD compared with young rats. If old rats have more AD then why do they sleep less? To investigate whether changes in sensitivity of the AD receptor contribute to the decline in sleep, experiments 3 and 4 determined that for the same concentration of AD or the AD receptor 1 agonist, cyclohexyladenosine, old rats have less sleep compared with young rats. We conclude that even though old rats have more AD, a reduction in the sensitivity of the AD receptor to the ligand does not transduce the AD signal at the same strength as in young rats and may be a contributing factor to the decline in sleep drive in the elderly.

Adenosine↗

PubMed2003

Effects of lateral hypothalamic lesion with the neurotoxin hypocretin-2-saporin on sleep in Long-Evans rats.

Narcolepsy, a disabling neurological disorder characterized by excessive daytime sleepiness, sleep attacks, sleep fragmentation, cataplexy, sleep-onset rapid eye movement sleep periods and hypnagogic hallucinations was recently linked to a loss of neurons containing the neuropeptide hypocretin. There is considerable variability in the severity of symptoms between narcoleptic patients, which could be related to the extent of neuronal loss in the lateral hypothalamus. To investigate this possibility, we administered two concentrations (90 ng or 490 ng in a volume of 0.5 microl) of the neurotoxin hypocretin-2-saporin, unconjugated saporin or saline directly to the lateral hypothalamus and monitored sleep, the entrained and free-running rhythm of core body temperature and activity. Neurons stained for hypocretin or for the neuronal specific marker were counted in the perifornical area, dorsomedial and ventromedial nucleus of the hypothalamus. More neuronal nuclei (NeuN) cells were destroyed by the higher concentration of hypocretin-2-saporin (-55%) compared with the lower concentration (-34%) in the perifornical area, although both concentrations lesioned the hypocretin neurons almost equally well (high concentration=91%; low concentration=88%). The high concentration of hypocretin-2-saporin also lesioned neurons in the dorsomedial nucleus of the hypothalamus and ventromedial nucleus of the hypothalamus. Narcoleptic-like sleep behavior was produced by both concentrations of the hypocretin-2-saporin. The high concentration produced a larger increase in non-rapid eye movement sleep amounts during the normally active night cycle than low concentration. Neither concentration of hypocretin-2-saporin disrupted the phase or period of the core temperature or activity rhythms. The low concentration of unconjugated saporin did not significantly lesion hypocretin or neurons and did not alter sleep. The high concentration of unconjugated saporin produced some loss of neuronal nuclei-immunoreactive (NeuN-ir) neurons and hypocretin immunoreactive neurons, but only a transient increase in non-rapid eye movement sleep. These results led us to conclude that the extent of hypocretin neuronal loss together with an accompanying loss of cells in the lateral hypothalamus may explain the differences in severity of symptoms seen in human narcolepsy.

Animals↗

PubMed2003

A secretin i.v. infusion activates gene expression in the central amygdala of rats.

For the last 100 years secretin has been extensively studied for its hormonal effects on digestion. Recent observations that the deficits in social reciprocity skills seen in young (3-4-year-old) autistic children are improved after secretin infusions suggest an additional influence on neuronal activity. We show here that i.v. administration of secretin in rats induces Fos protein expression in the neurons of the central amygdala as well as the area postrema, bed nucleus of the stria terminalis, external lateral parabrachial nucleus and supraoptic nucleus. However, secretin infusion did not induce Fos expression in the solitary tract nucleus or paraventricular nucleus, regions normally activated by related peptides such as cholecystokinin. The peak blood levels of secretin that induce Fos protein expression in rat brain are similar to the peak blood levels observed during i.v. treatment with secretin in humans. The amygdala is known to be critical for developing reciprocal social interaction skills and abnormalities in this brain region have been demonstrated in autistic children.

Amygdala↗

PubMed2001

Hypocretin-2-saporin lesions of the lateral hypothalamus produce narcoleptic-like sleep behavior in the rat.

Hypocretins (Hcrts) are recently discovered peptides linked to the human sleep disorder narcolepsy. Humans with narcolepsy have decreased numbers of Hcrt neurons and Hcrt-null mice also have narcoleptic symptoms. Hcrt neurons are located only in the lateral hypothalamus (LH) but neither electrolytic nor pharmacological lesions of this or any other brain region have produced narcoleptic-like sleep, suggesting that specific neurons need to be destroyed. Hcrt neurons express the Hcrt receptor, and to facilitate lesioning these neurons, the endogenous ligand hypocretin-2/orexin B (Hcrt2) was conjugated to the ribosome-inactivating protein saporin (SAP). In vitro binding studies indicated specificity of the Hcrt2-SAP because it preferentially bound to Chinese hamster ovary cells containing the Hcrt/orexin receptor 2 (HcrtR2/OX(2)R) or the Hcrt/orexin receptor 1 (HcrtR1/OX(1)R) but not to Kirsten murine sarcoma virus transformed rat kidney epithelial (KNRK) cells stably transfected with the substance P (neurokinin-1) receptor. Administration of the toxin to the LH, in which the receptor is known to be present, eliminated some neurons (Hcrt, melanin-concentrating hormone, and adenosine deaminase-containing neurons) but not others (a-melanocyte-stimulating hormone), indicating specificity of the toxin in vivo. When the toxin was administered to the LH, rats had increased slow-wave sleep, rapid-eye movement (REM) sleep, and sleep-onset REM sleep periods. These behavioral changes were negatively correlated with the loss of Hcrt-containing neurons but not with the loss of adenosine deaminase-immunoreactive neurons. These findings indicate that damage to the LH that also causes a substantial loss of Hcrt neurons is likely to produce the multiple sleep disturbances that occur in narcolepsy.

Adenosine Deaminase↗

PubMed2001

Effects of hypocretin-saporin injections into the medial septum on sleep and hippocampal theta.

Neurons containing the peptide hypocretin, also known as orexin, were recently implicated in the human sleep disorder narcolepsy. Hypocretin neurons are located only in the lateral hypothalamus from where they innervate virtually the entire brain and spinal cord. This peptide is believed to be involved in regulating feeding and wakefulness. However, to fully understand what other behaviors are regulated by this peptide it is necessary to investigate each hypocretin target site. In the present study, we focus on one hypocretin target site, the medial septum, where there is a dense collection of hypocretin-2 receptor-containing cells, and degenerating axons are present here in canines with narcolepsy [J. Neurosci. 19 (1999) 248]. We utilize a saporin toxin conjugated to the hypocretin receptor binding ligand, hypocretin-2, and find that when this toxin is injected into the medial septum, it lesions the parvalbumin and cholinergic neurons. We contrast the effects of the hypocretin-saporin with another saporin conjugated toxin, 192 IgG-saporin, that lesions only the cholinergic neurons in the basal forebrain. 192 IgG-saporin reduced theta activity, a finding consistent with previous reports [J. Neurophysiol. 79 (1998) 1633; Neurodegeneration 4 (1995) 61; Neuroscience 62 (1994) 1033]. However, hypocretin-saporin completely eliminated hippocampal theta activity by day 12, indicating that parvalbumin-containing cells in the medial septum generate theta. The daily amount of sleep and wakefulness were not different between hypocretin-saporin, 192 IgG-saporin, or saline-treated rats. The homeostatic response to 12 h prolonged wakefulness was also not affected in hypocretin-saporin lesioned rats. These findings suggest that hypocretin neurons could facilitate theta generation during episodes of purposeful behavior by activating GABAergic neurons in the MS/VDB. In this way, hypocretin, which is implicated in feeding, energy metabolism and wakefulness, serves to influence cognitive processes critical for the animal's survival.

Acetylcholine↗

PubMed2001

Effects of prolonged wakefulness on c-fos and AP1 activity in young and old rats.

Recent studies have demonstrated that the immediate-early gene c-fos is induced in neuronal populations responsible for specific sleep-wake states. The induction of this gene may be functionally relevant to sleep homeostasis since without the gene mice (c-fos null) take longer to fall asleep and have a selective reduction in slow-wave sleep. This suggests that a build-up of c-fos during wakefulness increases the drive to sleep and lack of c-fos is associated with reduced sleep. Sleep also has an effect on c-Fos serving to eliminate the protein rapidly. Waxing and waning of transcription factors such as c-Fos may influence slow, oscillating events such as sleep and wakefulness. To further examine what role c-Fos may play in regulating sleep, the present study examined the effects of prolonged wakefulness on c-Fos and AP-1 activity in young (3.5 months old) and old (21.5 months old) Sprague--Dawley rats. Previously we found that old rats slept less even after prolonged wakefulness, and other investigators have found that aging is also associated with a decline in c-Fos. In the present study, we reasoned that prolonged wakefulness would also fail to increase c-Fos in old versus young rats. The baseline levels of c-Fos and AP-1 activity were not different between young and old rats. However, in response to 6 or 12 h of prolonged wakefulness, old rats demonstrated significantly less c-Fos and AP-1 activity compared to young rats. These findings suggest that in old rats the mechanism responsible for c-Fos induction in response to wakefulness is deficient. Such a decline at the molecular level could contribute to the decline in sleep that typically occurs with age.

Aging↗

PubMed2001

Hypocretin receptor protein and mRNA expression in the dorsolateral pons of rats.

The hypocretins (also known as orexins) are hypothalamic peptides that have been implicated in feeding and sleep regulation. Previous reports have described the distribution of the mRNAs encoding two hypocretin receptors (HCRT-R), but the pattern of protein expression has not been investigated. Here we examine the distribution of the mRNA and protein for the HCRT receptor 1 (HCRT-R1) and HCRT receptor 2 (HCRT-R2) in the pontine brainstem and demonstrate that they are present in many pontine nuclei including those associated with REM sleep. Immunohistochemistry indicates that one or both of the receptor subtypes are expressed in the dorsal raphe, the lateral dorsal tegmental (LDT), the pedunculo pontine (PPT), the locus coeruleus (LC), the locus subcoeruleus, pontis oralis, Barrington's, the trigeminal complex (mesencephalic trigeminal and motor nucleus of the trigeminal nerve), the dorsal tegmental nucleus of Gudden (DTG), the ventral cochlear nucleus (VCA), trapezoid nucleus (TZ), pontine raphe nucleus and the pontine reticular formation. These regions have been shown to be involved in mastication, bladder control, gastrointestinal function and in arousal. Given these projection sites and the functions associated with these sites, we suggest that HCRT may play a role in maintaining alertness and vigilance while the animal is engaged in consummatory behavior.

Animals↗

PubMed2001

Hypothalamic regulation of sleep.

The recent discovery linking narcolepsy, a sleep disorder characterized by very short REM sleep latency, with a neuropeptide that regulates feeding and energy metabolism, provides a way to understand how several behaviors may be disrupted as a result of a defect in this peptide. In this chapter we review the evidence linking hypocretin and sleep, including our own studies, and propose that a defect in the lateral hypothalamus that also involves the hypocretin neurons is likely to produce a disturbance in sleep, mood, appetite, and rhythms.

Animals↗

PubMed2000

Sleep and wakefulness in c-fos and fos B gene knockout mice.

G-protein coupled receptor (GPCR) stimulation has been implicated in the regulation of sleep. Upon stimulation of a GPCR an intracellular cascade involving second and third messengers is initiated. The latter include the fos-family of immediate early genes (IEGs). Although there is considerable evidence indicating that IEGs are expressed in response to sleep, the effects of their deletion on sleep is not known. The present study examined sleep-wakefulness in mice lacking the c-fos or fos B genes. Null c-fos mice compared to their wildtype (WT) and heterozygote (het) siblings had more wakefulness and less slow wave sleep (SWS); REM sleep was not affected. The null c-fos mice also had increased delta activity (0.3-4 Hz). In contrast, the null and heterozygote fos B mice had less REM sleep, but the time spent in SWS or wakefulness was not different from their wild-type (WT) siblings. In the null c-fos mice, the increased wakefulness and the reduction in SWS could not be due to a systemic alteration in temperature since the core temperature was similar in all mice. By demonstrating that these IEGs are involved in sleep, we suggest that the deletion of specific genes, even within a family of genes, can have a specific effect on sleep.

Animals↗

PubMed2000

c-Fos expression in the cholinergic basal forebrain after enforced wakefulness and recovery sleep.

The present study used c-Fos expression to examine cellular activity in cholinergic regions in the basal forebrain (BF) following enforced waking and recovery sleep. Cholinergic cells within the vertical and horizontal limbs of the diagonal band of Broca (VDB and HDB, respectively) showed significantly higher c-Fos immunoreactivity after prolonged waking than after recovery sleep. Cholinergic cells within the medial septal nucleus (MS), however, showed no change in c-Fos expression under these conditions. Consistent with our previous findings, c-Fos immunoreactivity in the ventral lateral preoptic area (VLPO) was increased after 1-2h of recovery sleep compared with enforced waking. These results indicate state-specific effects on transcription and subsequent protein expression in cell populations associated with behavioral state and further show that the HDB, VDB and VLPO are good candidates for the further study of intracellular events associated with sleep and wakefulness.

Animals↗

PubMed2000

Compensatory sleep response to 12 h wakefulness in young and old rats.

There is a pronounced decline in sleep with age. Diminished output from the circadian oscillator, the suprachiasmatic nucleus, might play a role, because there is a decrease in the amplitude of the day-night sleep rhythm in the elderly. However, sleep is also regulated by homeostatic mechanisms that build sleep drive during wakefulness, and a decline in these mechanisms could also decrease sleep. Because this question has never been addressed in old animals, the present study examined the effects of 12 h wakefulness on compensatory sleep response in young (3.5 mo) and old (21.5 mo) Sprague-Dawley and F344 rats. Old rats in both strains had a diminished compensatory increase in slow-wave sleep (SWS) after 12 h of wakefulness (0700-1900, light-on period) compared with the young rats. In contrast, compensatory REM sleep rebound was unaffected by age. To assess whether the reduced SWS rebound in old rats might result from loss of neurons implicated in sleep generation, we counted the number of c-Fos immunoreactive (c-Fos-ir) cells in the ventral lateral preoptic (VLPO) area and found no differences between young and old rats. These findings indicate that old rats, similar to elderly humans, demonstrate less sleep after prolonged wakefulness. The findings also indicate that although old rats have a decline in sleep, this cannot be attributed to loss of VLPO neurons implicated in sleep.

Adaptation, Physiological↗

PubMed2000

Distribution of hypocretin-containing neurons in the lateral hypothalamus and C-fos-immunoreactive neurons in the VLPO.

The present study investigated the distribution of neurons implicated in the regulation of sleep in three species generally used in sleep research, i.e., mice, rats and cats. We focused on sleep active neurons in the ventral lateral preoptic (VLPO) area and the hypocretin/orexin-containing neurons in the lateral hypothalamus. The latter groups of neurons were found recently to play an important role in the regulation of REM sleep. The expression of the transcription factor, c-Fos, was used to identify the VLPO. In mice and rats, in response to sleep, a discrete cluster of c-Fos positive cells was found in the VLPO. In mice, this cluster was located more medially compared to the rat, and as in the rat, galanin immunostained neurons were found in the VLPO. In the cat, c-Fos positive cells did not segregate to a specific location but were more diffusely represented in the preoptic area. In all three species, orexin/hypocretin-containing neurons were located only in the lateral hypothalamus with the distribution being more diffuse in the cat. The grouping of sleep-active cells in rodents makes it feasible to extract these cells for tissue culture and molecular analysis. Moreover, given that rodents have a distinct circadian distribution of sleep-wakefulness, the connectivity with the suprachiasmatic nucleus can also be determined.

Animals↗

PubMed1999

Choline acetyltransferase expression during periods of behavioral activity and across natural sleep-wake states in the basal forebrain.

The present study examined whether the expression of the messenger RNA encoding the protein responsible for acetylcholine synthesis is associated with sleep-wakefulness. Choline acetyltransferase messenger RNA levels were analysed using a semi-quantitative assay in which reverse transcription was coupled to complementary DNA amplification using the polymerase chain reaction. To examine the relationship between steady-state messenger RNA and behavioral activity, rats were killed during the day (4.00 p.m.) or night (4.00 a.m.), and tissue from the vertical and horizontal limbs of the diagonal bands of Broca was analysed. Choline acetyltransferase messenger RNA levels were higher during the day than during the night. The second study examined more closely the association between choline acetyltransferase messenger RNA levels and individual bouts of wakefulness, slow-wave sleep or rapid eye movement sleep. Choline acetyltransferase messenger RNA levels were low during wakefulness, intermediate in slow-wave sleep and high during rapid eye movement sleep. In contrast, protein activity, measured at a projection site of cholinergic neurons of the basal forebrain, was higher during wakefulness than during sleep. These findings suggest that choline acetyltransferase protein and messenger RNA levels exhibit an inverse relationship during sleep and wakefulness. The increased messenger RNA expression during sleep is consistent with a restorative function of sleep.

Animals↗

PubMed1998

REM sleep deprivation increases the levels of tyrosine hydroxylase and norepinephrine transporter mRNA in the locus coeruleus.

The present study was conducted to determine the effects of REM sleep deprivation on the levels of tyrosine hydroxylase (TH) and norepinephrine transporter (NET) mRNA in the locus coeruleus (LC) of rats. The animals were deprived of REM sleep for 1, 3 or 5 days, then killed and changes in the mRNA levels were determined using in situ hybridization. The levels of both TH and NET mRNA increased in animals deprived of REM sleep for 3 days or longer whereas no change in these messages were observed in the LC of control animals. REM sleep deprivation has been used as a mode of treatment for major depression. Others have shown that treatment with tricyclic antidepressants also results in increased levels of TH and NET mRNA in LC. Our results suggest that the antidepressant effect of REM sleep deprivation and tricyclic antidepressants may share similar molecular changes in the norepinephrine system.

Animals↗

PubMed1998

Expression of cholinergic markers in the pons of Flinders rats.

The Flinders sensitive line of rats (FSL rats) have an altered REM sleep pattern which includes a shorter REM sleep latency and an increased percentage of REM sleep [R.M. Benca, D.H. Overstreet, M.A. Gilliland, D. Russell, B.M. Bergmann, W.H. Obermeyer, Increased basal REM sleep but no difference in dark induction or light suppression of REM sleep in Flinders rats with cholinergic supersensitivity, Neuropsychopharmacology 15 (1996) 45-51; P.J. Shiromani, D.H. Overstreet, D. Levy, C.A. Goodrich, S.A. Campbell, J. C. Gillin, Increased REM sleep in rats selectively bred for cholinergic hyperactivity, Neuropsychopharmacology 1 (1988) 127-133]. Cholinergic mechanisms have been implicated in REM sleep generation [reviewed in P.J. Shiromani, J.C. Gillin, S.J. Henriksen, Acetylcholine and the regulation of REM sleep: basic mechanisms and clinical implication for affective illness and narcolepsy, Annu. Rev. Pharmacol. Toxicol. 27 (1987) 137-156]. In the present study, specific aspects of the cholinergic system were examined in the pontine region of the FSL rats. The number of cholinergic neurons in the LDT and PPT were not different in FSL and control rats. Analysis of steady state levels of mRNAs encoding the acetylcholine synthesizing protein, choline acetyltransferase (ChAT) or the m2, m3 and m5 muscarinic receptor subtypes were also comparable in FSL and control rats. These data raise the possibility that the cellular events underlying the altered REM sleep pattern in FSL rats may include mechanisms that effect the muscarinic or nicotinic receptor in the pons.

Acetylcholine↗

PubMed1997

Effects of sleep on wake-induced c-fos expression.

We investigated the effects of sleep on wake-induced c-fos expression in the cerebral cortex of rats and c-fos-lacZ transgenic mice. In the cortex of rats, the levels of c-Fos, detected both by immunocytochemistry and Western blot, remained high during 6 or 12 hr of enforced wakefulness but declined rapidly (within 1 hr) with increasing time of recovery sleep. Similarly, in the transgenic mice in which lacZ expression is driven from the c-fos promoter, beta-galactosidase activity was high after enforced wakefulness and declined with increasing amounts of sleep. These results suggest that the decrease in c-Fos protein in cortical neurons during sleep may be attributable to cessation of c-fos expression, activation of a process that degrades the wake-induced c-Fos, or both.

Animals↗