Loss of circadian behavioural rhythms in rats kept in constant darkness

Ten male Sprague-Dawley rats were exposed to 1-h light and 1-h dark (LD 1 : 1) cycles for 50 days. They were then released into constant darkness (DD) for 104 days. Exposure to LD 1 : 1 caused gradual disruption of circadian rhythms in their ambulatory and drinking activities until, finally, all the animals lost their circadian behavioural rhythms. After their release into DD, eight rats showed free-running circadian behavioural rhythms, whereas the remaining two rats showed circadian arrhythmicity for approximately 50 days in DD before they restored their free-running rhythms spontaneously.     

An essential role for peptidergic signalling in the control of circadian rhythms in the suprachiasmatic nuclei

Two structurally related neuropeptides, pituitary adenylate cyclase-activating polypeptide (PACAP), colocalized with glutamate in neurones of the retinohypothalamic tract, and vasoactive intestinal peptide (VIP), present in light-responsive cells of the suprachiasmatic nuclei (SCN), appear to play distinct and important roles in the control of mammalian circadian rhythms. Mice deficient in the PACAP-selective PAC1 receptor exhibit altered responsiveness of the SCN clock to light-induced phase-shifts, but display robust circadian patterns of wheel-running behaviour. By contrast, our studies of mice lacking the VPAC2 receptor, which responds to both PACAP and VIP, indicate that this receptor plays a critical role in rhythm generation in the SCN. The predominant factor determining wheel-running activity in VPAC2 receptor null (Vipr2-/-) mice is "masking" by light. Mutant animals re-entrain immediately to advances or delays in the light/dark cycle and do not exhibit robust circadian rhythms of behaviour when in constant darkness. The mice do not exhibit circadian expression of core clock genes (mPer1, mPer2, mCry1), or of the clock-controlled gene arginine vasopressin (AVP), in the SCN. We propose that VIP signalling between SCN neurones provides a paracrine reinforcing signal that is essential for sustained rhythm generation. The presence of VIP signalling in the SCN may explain why SCN neurones are capable of generating long-lasting self-sustained oscillations, whereas rhythmic clock gene expression in other tissues is dependent on periodic reinforcement by neural or hormonal signals.     

Clorgyline delays the phase-position of circadian neurotransmitter receptor rhythms

The number of alpha- and beta-adrenergic, muscarinic cholinergic, opiate, and benzodiazepine receptors in rat forebrain, and dopamine and benzodiazepine receptors in striatum, change throughout the day. The diurnal rhythms of these receptors were altered by treatment with the monoamine-oxidase inhibitor clorgyline: following treatment some or all rhythm characteristics of wave form, amplitude, 24-h mean, and phase, were affected. One common effect of treatment was a delay in phase-position of binding to alpha- and beta-adrenergic, opiate and benzodiazepine receptors. Additionally, the nocturnal elevation in pineal melatonin which normally returns to baseline at light onset, persisted 3 h into the light period after clorgyline administration. These biochemical observations extend behavioural findings that clorgyline can delay the phase-position of rodent nocturnal activity onset, and does so by slowing the central circadian pacemaker.     

Clorgycline delays the phase-position of circadian neurotransmitter receptor rhythms

The number of α- and β-adrenergic, muscarinic cholinergic, opiate, and benzodiazepine receptors in rat forebrain, and dopamine and benzodiazepine receptors in striatum, change throughout the day. The diurnal rhythms of these receptors were altered by treatment with the monoamine-oxidase inhibitor clorgyline: following treatment some or all rhythm characteristics of wave form, amplitude, 24-h mean, and phase, were affected. One common effect of treatment was a delay in phase-position of binding to α- and β-adrenergic, opiate and benzodiazepine receptors. Additionally, the nocturnal elevation in pineal melatonin which normally returns to baseline at light onset, persisted 3 h into the light period after clorgyline administratio. These biochemical observations extend bevavioural findings that clorgyline can delay the phase-position of rodent nocturnal activity onset, and does so by slowing the central circadian pacemaker.     

Methamphetamine-induced,suprachiasmatic nucleus-independent circadian rhythms of activity and mPer gene expression in the striatum of the mouse

While the suprachiasmatic nucleus (SCN) coordinates the majority of daily rhythms, some circadian patterns of expression are controlled from outside of the SCN. These include responses to daily methamphetamine (MAP) injection, or daily restricted feeding. The mechanisms underlying these SCN-independent circadian rhythms are unknown. A circadian oscillation in the expression of mPer1 and/or mPer2, mouse period, in the SCN is considered necessary to generate an SCN-dependent circadian rhythm. Therefore, in this experiment, we examined the association between mPer gene expression and the MAP-induced, SCN-independent circadian rhythm. Acute injection of MAP caused an elevation of mPer1, mBmal1, and mNpas2 gene expression in the striatum and mPer1 in the liver. Daily MAP injection at a fixed time for 6 days shifted the rhythmic mPer1 and mPer2 expression in the striatum from a nocturnal to a diurnal rhythm, but failed to affect that in the SCN. Although lesion of the SCN 'flattened'mPer gene oscillation in the striatum and liver, daily MAP injection caused both behavioural and mPer gene expression rhythms. Daily MAP injection at variable injection intervals (12-36 h) for 6 days, however, failed to produce mPer gene rhythm in the striatum. Daily repeated MAP signals may strengthen the oscillatory force of SCN-independent circadian behavioural and molecular rhythms. The present results suggest that daily oscillation of mPer genes outside the SCN is closely associated with the regulation of SCN-independent rhythms. Thus, the present experiment highlights strongly the important role of clock gene expression, in the brain, that underlies the circadian behavioural rhythm.     

Modification of circadian sleep rhythms by gonadal steroids and the neural mechanisms involved

The effects of gonadectomy and gonadal hormone treatment of castrated rats or ovariectomized (OVX) rats bearing brain lesions on the circadian rhythms of slow wave sleep (SWS) and paradoxical sleep (PS) have been studied under a 14/10 light-dark schedule. Cortical EEGs and dorsal neck EMG were used to monitor SWS, PS and alertness. Intact female rats showed two daytime SWS peaks, one daytime PS peak and a small night PS peak except during proestrus. In intact male rats, the morning SWS peak and night PS peak were variable and SWS and PS peaks in daytime were dissociated. Orchidectomized (ORX) rats showed the morning SWS peak and disrupted the dissociation of SWS and PS peaks. Furthermore, gonadectomy increased the night PS peak. Posterior deafferentation of the hypothalamus (PDM) eliminated the night PS peak. Estradiol (E₂B) injection to long term OVX rats eliminated the night PS peak from the first day of injection. However, E₂B injection into androgenized OVX rats, ORX rats and OVX rats bearing septal lesion or MPO roof cut did not eliminate night PS peak. E₂B injection to short term OVX rats or OVX rats with PDM lesions delayed the E₂B-induced elimination of night PS peak. From these results, it is suggested that: (1) sexual dimorphism exists in the circadian sleep rhythm itself, and this difference partly depends on the hormonal environment produced by sex steroids; (2) the rise and fall of night PS peak reflects the neurohumoral environment in female rats; (3) the appearance of night PS peak involves the abolition of negative feedback of sex steroids and the posterior neural input into hypothalamus; and (4) the elimination of night PS peak on natural proestrus and following E₂B treatment of OVX rats requires the intact positive feedback system of estradiol.     

Correlation between the circadian sleep propensity rhythm and hormonal rhythms under ultra-short sleep–wake cycle

The aim of this study was to clarify effects of hormonal and temperature rhythms on circadian fluctuations of sleep propensity. Ten healthy females underwent 24-h sleep deprivation and entered the circadian sleep propensity assessment setting under the ultra-short sleep-wake schedule. During the experiment, sleep propensity rhythm, rectal temperature, and 24-h serum hormone profiles (melatonin, cortisol and thyroid-stimulating hormone) were investigated. The circadian sleep propensity rhythms had two apparent peaks (afternoon and nocturnal peaks) and a trough (nocturnal sleep gate). The timings of the nocturnal sleep gate and the nocturnal peak were correlated exclusively with temperature and melatonin rhythms (P < 0.05), while that of the afternoon peak was significantly correlated with habitual wake time and melatonin rhythm. These results indicate that the circadian sleep propensity rhythm is influenced not only by the circadian pacemaker, but also by sleep habit.     

Acute intracerebral haemorrhage: circadian and circannual patterns of onset

Hypothesis of the circannual and circadian variation in onset of intracerebral haemorrhage (CH) was verified, by means of single cosinor method and chi-square test for goodness of fit, in 161 consecutive patients (94 men and 67 women) admitted into the Institute of Neurosurgery of Ferrara Hospital, Italy, over 9 years. The majority of CH occurred in the morning between 06.00 AM and 12.00 noon (36.7% of cases, p<0.001); when considering the specific anatomical sites, typical supratentorial haemorrhages showed a similar pattern (37.4%, p= 0.01). A similar morning behavior was found when considering subgroups by sex (men 36.2%, women 37.3%), age ≥60 years (42.5%), no presence of hypertension (39.7%), no presence of diabetes mellitus (33.3%) and non-smokers (30.4%). The results by cosinor analysis yielded a circadian rhythmicity both for total sample and, for the men's subgroup, with a morning peak at 11.44 and 11.25, respectively. For women, however, spectral analysis found a significant ultradian cycle, having a period of 12 h (p = 0.01). A circannual periodicity, with a prevalent peak in February, was found for total sample and males subgroups, too. The results of this study confirm that intracerebral haemorrhages present a characteristic circadian and circannual pattern in onset.     

Discrimination of circadian phase in intact and suprachiasmatic nuclei-ablated rats

This study examined whether the circadian system of rats can serve as a consulted clock for discriminating time of day. Food restricted rats housed in activity wheels were trained to lever press for food in a two-lever T-maze in which the left arm was correct in a morning feeding session, and the right arm in an afternoon session (7 h interval). All six rats learned the task (discrimination ratios>chance on 85–95% of sessions) and exhibited anticipatory wheel running prior to most sessions. Performance was not disrupted by inverting the LD cycle or by omitting 1–3 sessions, indicating that learning was not dependent on light-dark cues, alternation strategies, or physiological states associated with intermeal interval. Five of six additional rats with ablations of the suprachiasmatic nucleus light-entrainable pacemaker acquired the discrimination, indicating that time-of-day cues can be provided by another circadian pacemaker (likely food-entrainable). The results provide the first clear evidence that the circadian system in a mammal can function as a consulted clock that provides discriminative time cues for cognitive processes subserving behavioral plasticity.     

Normal sleep-wake and circadian rhythms in a case of Gerstmann-Sträussler-Sheinker (GSS) disease

Abstract. A polysomnographic study showed normal sleep patterns and circadian rhythms in a patient with ataxic type GSS. Our results suggest that sleep and circadian rhythm changes in prion diseases are due to the involvement of specific brain areas.     

Effect of lithium on the circadian rhythms of locomotor activity and glycogen synthase kinase-3 protein expression in the mouse suprachiasmatic nuclei

A circadian clock located in the suprachiasmatic nucleus (SCN) regulates the period of physiological and behavioural rhythms to approximately 24 h. Lithium can lengthen the period of circadian rhythms in most organisms although little is known about the underlying mechanism. In the present study, we examined Drosophila shaggy ortholog glycogen synthase kinase-3 (GSK-3) protein expression in the SCN after lithium treatment. When locomotor activity was assessed, we found an association between the effect of lithium and the period of circadian oscillation as well as the level of GSK-3 protein expression. The decreased expression of GSK-3 and increased expression of phosphorylated GSK-3 (pGSK-3) resulted in an antiphasic circadian rhythm between the two in the SCN of lithium-treated mice housed under both light-dark and constant dark conditions. The enzyme activity of GSK-3 in the SCN was low when the level of pGSK-3 protein was high, as examined by immunoblotting analysis. Thus, GSK-3 enzyme activity has a correlation with the expression of GSK-3 protein in the SCN. Although both GSK-3 and pGSK-3 proteins are also expressed in the arcuate nucleus, lithium did not affect their expression. Based on the association that we found between lengthened circadian period and GSK-3 protein and GSK-3 activity in the SCN, we suggest that GSK-3 plays a role in regulating the period of the mammalian circadian pacemaker.     

Opiate analgesia: Evidence for circadian rhythms in mice

The effects of a 12-12 h light-dark (L-D) cycle and of constant light (L-L) on nociceptive thresholds and morphine-induced analgesia were studied in two strains of mice: C57BL/6 and SEC/1Re.Under the L-D condition, a diurnal rhythm was observed in the responsivity of mice to nociceptive stimuli, and in the analgesic effects of morphine. Under the L-L schedule clear patterns of daily rhythmicity were evident in both strains for both nociceptive thresholds and responsivity to morphine. Finally, under the L-L schedule, the overall responsivity to pain and the antinociceptive effects of morphine were clearly increased in comparison to the patterns evident in the L-D condition. The administration of naloxone decreased the nociceptive thresholds, thus indicating an involvement of the endogenous opioid peptides.These phenomena are discussed in terms of the different pacemaker mechanisms involved in the expression of various patterns of daily behavioral and neurochemical rhythmicity in these strains, and of the effects of light in controlling the release of endogenous opioids.     

Ontogenesis of circadian sleep-wakefulness rhythms and developmental changes of sleep in the altricial rat and in the precocial guinea pig

Ontogenesis of circadian sleep-wakefulness rhythms and other developmental changes of sleep were examined in the altricial rat and in the precocial guinea pig. Circadian sleep-wakefulness rhythms began to emerge on the 4th–4.5th week after birth in rats born and raised not only under light-dark (LD) 12:12 h lighting schedule but also under a constant light condition. Only ultradian rhythms were recognized in the rat up to 2.5–3.5 weeks after birth. Unlike the rat, the guinea pig did not develop significant diurnal and circadian sleep rhythms during the course of its growth. Sleep-wakefulness rhythms of the 1-week-old infant guinea pig were very similar to those of the adult. During the development of the altricial rat the periods of slow wave sleep (SWS) and paradoxical sleep (PS) cycles, and the amounts of SWS per day increased. On the other hand, the amounts of PS per day and the length of the PS episodes decreased. For the guinea pig, however, these parameters of sleep were apparently constant throughout development.     

Sympathetic regulation of chicken pineal rhythms

Adult hens were chronically cannulated and held in light-dark (LD) 12:12 h lighthing regimes or in constant darkness (DD). Periodic blood sampling for 5–9 days revealed circadian rhythms in plasma melatonin titres. Superior cervical ganglionectomy (SCG-X) performed 1 week after hatching had little or no effect on these rhythms in LD, but unlike normals. SCG-X birds did not sustain persistent rhythms in DD. In SCG-X birds, norepinephrine (NE) infusion for 12 h of each 24 h in DD significantly reduced plasma melatonin titres during the infusion and re-established a rhythm.After each experiment, hens were killed, their pineals were removed and assayed by HPLC-EC for NE. dopamine (DA). serotonin (5-HT) and 5-hydroxy-3-indole-acetic acid (5-HIAA). SCG-X resulted in a 90% depletion of pineal NE: DA content was reduced to undetectable levels. Pineal 5-HT and 5-HIAA were also reduced by SCG-X.The chicken pineal contains circadian oscillators which persist in vitro^8.19.29. The results reported here suggest that noradrenergic fibres from the SCG regulate the pineal's inherent rhythmicity. NE normally released from sympathetic terminals during the bird's day may synchronize oscillators within the pineal by inhibiting melatonin synthesis.     

The lower entrainable limit of rat circadian rhythm to sinusoidal light intensity cycles: A preliminary study

The lower entrainable limit of the circadian behavioral rhythm was examined in rats exposed to sinusoidal light intensity cycles with maximum illuminance of 20 lux and the minimum of 0.01 lux. The period (T) of the light intensity cycle was initially kept at 23.5 h for 20 cycles, and then shortened to 23 h for 33 cycles. Thereafter the rats were released into constant darkness. Five out of 10 rats entrained their circadian rhythms to T = 23.5-h cycle, and they also entrained to the T = 23-h cycle. The phase angle of entrainment was almost unchanged when T was shortened from 23.5 h to 23 h. These results suggest that the T = 23-h cycle was close to the lower limit of entrainment.     

A retinohypothalamic pathway in man: Light mediation of circadian rhythms

It has been proposed that, in animals, a retinohypothalamic pathway exists which mediates the synchronization of the diurnal light-dark cycle with the central neural components regulating endogenous rhythms. There have been numerous anatomic, physiologic and behavioral investigations to substantiate this proposed connection in experimental animals. Morphologic investigation of a retinohypothalamic tract in man has awaited the development of a technique capable of axonal tracing in the human brain. The paraphenylenediamine method was applied to 7 post-mortem human brains. Degenerated axons were found in the suprachiasmatic nuclei of the hypothalamus in each of the 4 patients who had incurred prior optic nerve damage. The retinosuprachiasmatic pathway may be the anatomical substrate for the integration of retinal light information with endogenous rhythms in man.