High corticosterone levels in prenatally stressed rats predict persistent paradoxical sleep alterations.

Prenatal stress predisposes rats to long-lasting disturbances that persist throughout adulthood (e.g., high anxiety, dysfunction of the hypothalamo-pituitary-adrenal axis, and abnormal circadian timing). These disturbances parallel to a large extent those found in depressed patients, in which hypercortisolemia and sleep alterations may be related to stress-inducing events. We studied sleep-wake parameters in control and prenatally stressed adult rats (3-4 months old) and examined possible relationships with their corticosterone levels (determined at 2 months of age). Under baseline conditions, prenatally stressed rats showed increased amounts of paradoxical sleep, positively correlated to plasma corticosterone levels. Other changes include increased sleep fragmentation, total light slow-wave sleep time, and a slight decrease in the percentage of deep slow-wave sleep relative to total sleep time. During recovery sleep from acute restraint stress, all sleep changes persisted and were correlated with stress-induced corticosterone secretion. High corticosterone levels under baseline conditions as well as an acute stress challenge may thus predict long-term sleep-wake alterations in rats. Taken together with other behavioral and hormonal abnormalities in prenatally stressed animals, the pronounced changes in sleep-wake parameters that are similar to those found in depressed patients suggest that prenatal stress may be a useful animal model of depression.

Changes in the phase response curve of the circadian clock to a phase-shifting stimulus.

Experiments were conducted in hamsters to determine whether the phase response curve (PRC) to injections of the short-acting benzodiazepine triazolam is a fixed or a labile property of the circadian clock. The results indicated that (1) both the shape and the amplitude of the PRC to triazolam generated on the first day of transfer from a light-dark cycle (LD 14:10) to constant darkness (DD) (i.e., PRCLD) were different from those of the PRC generated after many days in DD (PRCDD); and (2) the phase-shifting effects of triazolam on the activity rhythms of hamsters transferred from LD 14:10 or 12:12 to DD changed dramatically within the first 8-9 days spent in DD. In an attempt to accelerate the resynchronization of the circadian clock of hamsters subjected to an 8-hr advance in the LD cycle, triazolam was given to the animals at a time selected on the basis of the characteristics of PRCLD. The activity rhythms of five of eight triazolam-treated animals were resynchronized to the new LD cycle within 2-4 days after the shift, whereas those of most of the control animals were resynchronized 21-29 days after the shift. These findings suggest that attempts to use pharmacological or nonpharmacological tools to phase-shift circadian clocks under entrained conditions should take into account information derived from PRCs generated at the time of transition from entrained to free-running conditions.

Circadian clock functioning is linked to acute stress reactivity in rats.

At least two major physiological systems are involved in the adaptation of the organism to environmental challenges: the circadian system and the stress reaction. This study addressed the possibility that interindividual differences in stress sensitivity and in the functioning of the circadian system are related. At 2 months of age, corticosterone secretion in response to a 20-min restraint stress was assessed in 9 Sprague-Dawley rats for which running wheel activity was recorded as a rhythmic behavioral marker of the circadian clock. Two weeks later, the adaptive response of the circadian system to an abrupt shift in the light:dark (LD) cycle was assessed in those rats using a jet-lag paradigm. Finally, after resynchronization to the new LD cycle, rats were transferred to constant darkness to assess the free-running period of their circadian rhythm of running-wheel activity. Results indicate that stress-induced corticosterone secretion was (1) positively correlated with the number of days to resynchronize the circadian activity rhythm to the new LD cycle, and with the value of its free-running period, and (2) negatively correlated with the intensity of daily locomotor activity. Those data, emphasizing the interactions between the stress response of an organism and the functioning of its circadian system, could explain interindividual differences in humans' susceptibility to shift work or other circadian-related disorders.

Sleepiness, performance, and neuroendocrine function during sleep deprivation: effects of exposure to bright light or exercise.

The temporal profiles of subjective fatigue (as assessed by the Stanford Sleepiness Scale), of cognitive performance (on a digit symbol substitution test and a symbol copying task), of body temperature, and of the peripheral concentrations of melatonin, thyroid-stimulating hormone (TSH), and cortisol were obtained simultaneously at frequent intervals in 17 normal young subjects submitted to a 43-h period of constant routine conditions involving continuous wakefulness at bed rest in dim indoor light. The subjects had knowledge of time of day. Caloric intake was exclusively in the form of an intravenous glucose infusion, and plasma glucose levels were monitored continuously in 8 of the 17 subjects. Under these conditions, fluctuations in plasma glucose reflect primarily changes in glucose use because endogenous glucose production is suppressed by the exogenous infusion. Following the completion of a baseline constant routine study, the volunteers participated in two subsequent studies using the same protocol to determine the immediate psychophysiological effects of exposure to a 3-h pulse of bright light or to a 3-h pulse of physical exercise. Sleepiness and performance varied in a mirror image, with significant negative correlations. Sleepiness scores were minimal around noon and then increased at a modest rate throughout the rest of the normal waking period. Staying awake during usual bedtime hours was associated with an acceleration in the rate of increase in sleepiness, which coincided with decreasing body temperature, rapidly rising cortisol concentrations, and maximal levels of melatonin and TSH. When body temperature reached its nadir, a further major increase in sleepiness occurred in parallel with a pronounced decrease in plasma glucose (reflecting increased glucose use). Recovery from maximal sleepiness started when blood glucose levels stopped falling and when significant decreases in cortisol and melatonin concentrations were initiated. Lower levels of subjective sleepiness resumed when glucose concentrations and body temperature had returned to levels similar to those observed prior to sleep deprivation and when melatonin and TSH concentrations had returned to daytime levels. The synchrony of behavioral, neuroendocrine, and metabolic changes suggests that circulating hormonal levels could exert modulatory influences on sleepiness and that metabolic alterations may underlie the sudden increase in fatigue consistently occurring at the end of a night of sleep deprivation. Effects of bright light or exercise exposure on subjective sleepiness appeared to be critically dependent on the timing of exposure.

Preliminary studies on the immediate phase-shifting effects of light and exercise on the human circadian clock.

The aim of the present research was to determine the magnitude and direction of immediate phase shifts of human rhythms following a single exposure to a 3-hr pulse of bright light or physical activity. The pulse of light or activity was presented under "constant-routine" conditions, and measurements of the resultant phase shifts were performed under the same constant-routine conditions on the first day following pulse presentation. Four overt rhythms that are strongly dependent on circadian timing--namely, the rhythms of plasma cortisol, plasma thyroid-stimulating hormone (TSH), plasma melatonin, and body temperature--were monitored. The analysis of the TSH profiles indicated that exposure to light at about the time of the minimum of body temperature resulted in phase advances averaging less than 1 hr in magnitude. Exposure to light approximately 3 hr before the time of the minimum of body temperature resulted in phase delays of 1-2 hr. Preliminary analyses of the melatonin profiles have confirmed these observations. Our findings regarding the effects of exercise are still inconclusive.

Effects of age on the circadian system.

While aging has been associated with changes in the period and amplitude of circadian rhythms, little is known about how aging influences the response of the circadian clock to environmental stimuli. In this paper, we report on recent studies designed to determine the effects of advanced age on the response of the circadian clock to both photic and nonphotic stimuli in old hamsters (e.g., over 16 mo of age). Among the most pronounced age-related changes in the circadian rhythm of locomotor activity are: (a) alterations in the phase-angle of entrainment to the light-dark cycle; (b) an increase in the magnitude of phase shifts induced by pulses of light presented at specific circadian times; and (c) a loss of responsiveness to the phase shifting or entraining effects of stimuli which induce an acute increase of activity. Depletion of brain monoamine levels in young animals can induce changes in the responsiveness of the circadian clock to environmental stimuli which are similar to those which occur spontaneously in old animals, suggesting that aging alters monoaminergic inputs to the clock. Some of the age-related changes in the response of the clock to an activity-inducing stimulus can be reversed by implanting old animals with fetal SCN tissue. Determining the physiological basis for age related changes in the responsiveness of the clock to both internal and external stimuli, and the mechanisms by which normal circadian function can be restored, should lead to new insight into the functioning of the circadian clock and may lead to new approaches for normalizing disturbed circadian rhythms.

Prenatal stress and long-term consequences: implications of glucocorticoid hormones.

We have shown that prenatal restraint stress (PNRS) induces higher levels of anxiety, greater vulnerability to drugs, a phase advance in the circadian rhythm of locomotor activity and an increase in the paradoxical sleep in adult rats. These behavioral effects result from permanent modifications to the functioning of the brain, particularly in the feedback mechanisms of the hypothalamic-pituitary-adrenal (HPA) axis: the secretion of corticosterone is prolonged after stress and the number of the central glucocorticoid receptors is reduced. These abnormalities are associated with modifications in the synthesis and/or release of certain neurotransmitters. Dysfunction of the HPA axis is due, in part, to stress-induced maternal increase of glucocorticoids, which influences fetal brain development. Some biological abnormalities in depression can be related to those found in PNRS rats reinforcing the idea of the usefulness of PNRS rats as an appropriate animal model to study new pharmacological approaches.

Changes in neurohypophysial cholecystokinin content during oestrous cycle in the rat.

Cholecystokinin content in the neurointermediate lobe of the rat pituitary was measured by radioimmunoassay during the different stages of the oestrous cycle. Higher levels were observed in pro-oestrus and oestrus than in metoestrus and dioestrus rats. This difference is similar to the variation observed in the same circumstance concerning oxytocin in the neurohypophysis and neurosecretory activity in magnocellular neurons. These results are discussed in relation to the coexistence of oxytocin and cholecystokinin in neurons of the hypothalamoneurohypophysial system.

Autoradiographic localization of cholecystokinin binding sites in human cerebellar system using a [(125)I]CCK8 probe.

Carboxyl-terminal cholecystokinin octapeptide (CCK8) binding sites were studied in the human cerebellar system by autoradiography. High affinity CCK8 binding sites were demonstrated in the main cerebellar afferent nuclei, namely the inferior olivary complex and the pontine nuclei. This localization of CCK8 binding sites was partly correlated with already described CCK containing terminals. In the cerebellar cortex, high affinity CCK8 binding sites were detected with a laminar distribution. Levels were higher in the granular layer (mostly in the superficial part) and lower in the white matter and the Purkinje cell layer. The non-specific binding was homogenous and particularly low (9%) in the cerebellar cortex but a non-specific binding was selectively localized in the deep cerebellar nuclei. Those results illustrate the species variability of CCK binding sites in the cerebellum and are briefly discussed in relation with the low level of CCK immunoreactivity in this structure. The presence of CCK8 binding sites in cerebellar afferent nuclei and cortex suggests a role of CCK in human cerebellar physiology and particularly in the modulation of afferent inputs to the cerebellum.

Co-existence of cholecystokinin- or gastrin-like peptides with other peptides in the hypophysis and the hypothalamus.

The presence of cholecystokinin and gastrin has been reported in the hypothalamohypophyseal system. These peptides present a peculiar distribution in the hypothalamic nuclei, the median eminence, and the neurohypophysis. CCK and gastrin have close relationships with other peptides like oxytocin, CRF, vasopressin, and the enkephalins; these relationships vary in different projecting areas and in different types of hypothalamic neurons. The functional role of G-CCK in neurosecretion seems to be linked to the role of these closely associated peptides and certainly deserves further investigation.

Sleep in the Wistar-Kyoto rat, a putative genetic animal model for depression.

The Wistar-Kyoto (WKY) rat exhibits several behavioral and hormonal abnormalities often associated with depression. One of the hallmarks of depression consists of alterations in the sleep-wake cycle, particularly in rapid eye movement (REM) sleep. If the WKY rat is indeed an animal model for depression, we hypothesized that it should also show sleep abnormalities relative to the control strain, the Wistar (WIS) rat Under baseline conditions, WKY rats showed a 50% increase in total REM sleep time during the 12 h light phase and an increase in sleep fragmentation during both the light and dark phase. The WKY rats also exhibited lower EEG power densities over the entire frequency range (0.2-25.0 Hz) during REM sleep. After a 6 h sleep deprivation, the REM sleep rebound was more pronounced during the dark but not the light phase in the WKY rats. Since the WKY rat represents a genetic model for depression with altered EEG sleep patterns, this strain may be particularly useful for investigating the relationship between depression and sleep abnormalities.

Resynchronisation of a diurnal rodent circadian clock accelerated by a melatonin agonist.

Using 'jet lag' paradigms involving phase shifts in the light-dark (LD) cycle, we studied the effects of S-20098 on the circadian clock of a diurnal rodent. Arvicanthis mordax, entrained to a regular LD cycle, were subjected to advance shifts (i.e. 4, 6 or 8 h) in the LD cycle and injected with vehicle or the melatonin agonist S-20098 (20 mg/kg) the day of the shift (and also on subsequent days in the 6 h or 8 h shift paradigms). In each condition, S-20098 accelerated by about 30% resynchronization to the new LD cycle. These data, which are the first to demonstrate the chronobiotic effects of a melatonin agonist in a diurnal rodent, provide new insights for the design of human chronopharmacological protocols.

Distribution of neuropeptide Y immunoreactivity in human visual cortex and underlying white matter.

Immunocytochemical techniques have been used to study neuropeptide Y (NPY) distribution in the human visual cortex (Brodman's areas 17, 18 and 19) NPY cell bodies belong mostly to inhibitory (multipolar and bitufted) but also to excitatory (bipolar and some pyramidal) neuronal types. Their distribution is similar in the three cortical areas studied: 20 to 40% of the NPY perikarya are located in the cortical gray matter, mostly in the deep layers, while the remaining 60 to 80% are located in the underlying white matter. Immunoreactive NPY processes form a rich network of intersecting fibers throughout the entire visual cortex. A superficial plexus (layers I and II) and a deep plexus (deep layer V and layer VI) of NPY fibers are present in areas 17, 18 and 19. In area 17, an additional well developed plexus is present in layers IVb and IVc. These plexuses receive branches from long parallel fibers arising from deep cortical layers or underlying white matter and terminating in superficial layers. Local or extrinsic NPY terminals wind around vessels in the cortex as well as in the white matter, and either penetrate them or form clusters of club endings on their walls. Our results suggest a role for NPY in human visual circuitry and in cortical blood flow regulation.

A melatonin agonist facilitates circadian resynchronization in old hamsters after abrupt shifts in the light-dark cycle.

Age-related changes in the mammalian circadian system may be associated with a decline in circulating melatonin levels. Using 'jet lag' paradigms involving abrupt shifts in the light-dark cycle, we showed that a melatonin agonist, S-20098, accelerated by approximately 25% resynchronization of the circadian activity rhythm in old hamsters to the new light-dark cycle. It suggests the usefulness of melatonin-related compounds to treat circadian disorders associated with aging.

The effects of short periods of immobilization on the hamster circadian clock.

Recent findings indicate that stimuli which induce an acute increase in locomotor activity can induce phase shifts in the circadian clock of hamsters. Support for the actual role of the acute increase in activity in the mediation of these phase shifts is provided by the observation that immobilization can totally block phase shifts in the activity rhythm that are normally induced in response to exposure to two of these stimuli, either a pulse of darkness or an injection of a benzodiazepine. In order to further examine the effects of immobilization on the circadian system of hamsters, 3 studies were carried out. In a first study, the effects of a 3-h period of immobilization procedure on the phase of the free running circadian rhythm of locomotor activity were tested at 8 different circadian times. Immobilization during the highly active part of the animal's activity cycle resulted in phase delays in the activity rhythm, while immobilization at other circadian times had little or no effect on the circadian time-keeping system. In two other studies, we reported that immobilization had no effect on phase shifts normally induced by 3-h pulses of light or injections of the protein synthesis inhibitor, cycloheximide, two stimuli that are clearly not associated with an increase in locomotor activity in hamsters. Thus, the ability of immobilization to block stimulus-induced phase shifts in the circadian clock appears to be specific to those stimuli that induce an acute increase in locomotor activity.(ABSTRACT TRUNCATED AT 250 WORDS)

Grafting fetal suprachiasmatic nuclei in the hypothalamus of old hamsters restores responsiveness of the circadian clock to a phase shifting stimulus.

In the present study, 18-25-month-old hamsters free-running in constant dim light were injected, both before and after receiving fetal grafts containing either cerebellar tissue or the suprachiasmatic nuclei (SCN), with a dose of triazolam given at a time known to reliably phase shift the rhythm of locomotor activity in young hamsters. SCN-grafted animals, but not control animals implanted with fetal cerebellar tissue, showed a significantly greater response to the phase shifting effects of triazolam, demonstrating that at least some age-related changes in the circadian system can be reversed by neuronal transplantation. These results raise the possibility that neuropharmacological interventions that can simulate the effects of fetal SCN grafts might be useful in the treatment of age-related disorders in circadian function.

A benzodiazepine antagonist, Ro 15-1788, can block the phase-shifting effects of triazolam on the mammalian circadian clock.

A single injection of the short acting benzodiazepine, triazolam, can induce permanent phase advances as well as phase delays in the onset of the circadian rhythm of wheel running behavior in hamsters free-running under constant environmental conditions. If the phase shifting effects of triazolam on the circadian system are mediated through the benzodiazepine-GABA receptor complex, then it should be possible to block these effects with RO 15-1788, a selective benzodiazepine antagonist, which acts at the benzodiazepine-GABA receptor level. To test this hypothesis, hamsters free running in constant light received an intraperitoneal injection of various doses of Ro 15-1788 15 min before a single i.p. injection of 0.5 mg of triazolam. This dose of triazolam is known to induce maximal phase shifts in the circadian rhythm of wheel running behavior in hamster. Treatment with Ro 15-1788 totally blocked both the phase advancing and phase delaying effects of triazolam, while the administration of Ro 15-1788 alone did not phase shift the activity rhythm. These results support the hypothesis that the phase shifting effects of triazolam are mediated through the benzodiazepine-GABA receptor complex. The absence of any phase shifting effects of Ro 15-1788 when delivered alone suggests that Ro 15-1788 has no partial agonist properties in this experimental paradigm.

Sleep deprivation decreases phase-shift responses of circadian rhythms to light in the mouse: role of serotonergic and metabolic signals.

The circadian pacemaker in the suprachiasmatic nuclei is primarily synchronized to the daily light-dark cycle. The phase-shifting and synchronizing effects of light can be modulated by non-photic factors, such as behavioral, metabolic or serotonergic cues. The present experiments examine the effects of sleep deprivation on the response of the circadian pacemaker to light and test the possible involvement of serotonergic and/or metabolic cues in mediating the effects of sleep deprivation. Photic phase-shifting of the locomotor activity rhythm was analyzed in mice transferred from a light-dark cycle to constant darkness, and sleep-deprived for 8 h from Zeitgeber Time 6 to Zeitgeber Time 14. Phase-delays in response to a 10-min light pulse at Zeitgeber Time 14 were reduced by 30% in sleep-deprived mice compared to control mice, while sleep deprivation without light exposure induced no significant phase-shifts. Stimulation of serotonin neurotransmission by fluoxetine (10 mg/kg), a serotonin reuptake inhibitor that decreases light-induced phase-delays in non-deprived mice, did not further reduce light-induced phase-delays in sleep-deprived mice. Impairment of serotonin neurotransmission with p-chloroamphetamine (three injections of 10 mg/kg), which did not increase light-induced phase-delays in non-deprived mice significantly, partially normalized light-induced phase-delays in sleep-deprived mice. Injections of glucose increased light-induced phase-delays in control and sleep-deprived mice. Chemical damage of the ventromedial hypothalamus by gold-thioglucose (600 mg/kg) prevented the reduction of light-induced phase-delays in sleep-deprived mice, without altering phase-delays in control mice. Taken together, the present results indicate that sleep deprivation can reduce the light-induced phase-shifts of the mouse suprachiasmatic pacemaker, due to serotonergic and metabolic changes associated with the loss of sleep.

Prenatal stress in rats predicts immobility behavior in the forced swim test. Effects of a chronic treatment with tianeptine.

Prenatally-stressed (PS) rats are characterized by a general impairment of the hypothalamo-pituitary-adrenal (HPA) axis and sleep disturbances indicating that this model has face validity with some clinical features observed in a subpopulation of depressed patients. The prolonged corticosterone secretion shown by PS rats in response to stress was positively correlated with an increased immobility behavior in the forced swim test. To investigate the predictive validity of this model, a separate group of animals was chronically treated with the antidepressant tianeptine (10 mg/kg i.p. for 21 days). Such chronic treatment reduced in PS rats immobility time in the forced swim test. These findings suggest that the PS rat is an interesting animal model for the evaluation of antidepressant treatment.

Aging alters the entraining effects of an activity-inducing stimulus on the circadian clock.

In young hamsters, a single injection of the short-acting benzodiazepine, triazolam, can induce permanent phase shifts in the circadian clock, while repeated injections of triazolam entrain the circadian clock to the period of the injections. Triazolam appears to act on the circadian clock by inducing an acute increase in the activity of the animals, which in turn phase-shifts the circadian clock. Surprisingly, single injections of benzodiazepines do not phase-shift the activity rhythm of old hamsters, despite the fact that such treatment induces similar acute changes in the activity state of young and old animals. We compared the entraining effects of repeated injections of triazolam on the circadian clock of young and old hamsters; while six out of seven young hamsters were entrained to the triazolam injections, only one out of seven old animals was entrained by this treatment. Three of the remaining six old hamsters showed a lengthening of the activity rhythm, while no consistent effect on the period of the activity rhythm was observed in the remaining three old animals. These results indicate that the circadian system of old hamsters becomes selectively unresponsive to synchronizing signals mediated by the activity-rest state, and suggest that aging is associated with a weakened coupling between the activity-rest cycle and the circadian clock.