Regional Distribution of Iodomelatonin Binding Sites within the Suprachiasmatic Nucleus of the Syrian Hamster and the Siberian Hamster

The pineal hormone melatonin is a potent regulator of seasonal and circadian rhythms in vertebrates. In order to characterize potential target tissues of melatonin, the distribution of iodomelatonin (IMEL)-binding sites was examined within neurochemically and anatomically defined subdivisions of the suprachiasmatic nucleus (SCN), a structure necessary for seasonal and circadian rhythms in mammals. Studies were carried out in both the adult Syrian (Mesocricetus auratus) and Siberian (Phodopus sungorus) hamster. The retinoreceptive zone of the SCN was identified anatomically by immunocytochemical (ICC) visualization of cholera toxin B subunit tracer (ChTB-ir) following its intra-ocular injection. Photically-responsive SCN cells were identified by immunostaining for the protein product of the immediate-early gene c-fos (Fos-ir) following exposure of the animal to light. The non-photoresponsive zone of the SCN was identified using in situ hybridization (ISH) for arginine vasopressin (AVP) mRNA, whilst sites of IMEL-binding in the SCN were identified by in vitro film autoradiography using the specific ligand 2-[¹²⁵l]-iodomelatonin. To compare directly the distribution of IMEL-binding sites and one of the functional zones of the nucleus, alternate serial coronal sections through the SCN were processed for autoradiography for IMEL and one of the following: ICC for ChTB-ir or Fos-ir, or ISH for AVP mRNA. Overall, the regional distribution of the various markers within the SCN was comparable in the two species. The retinorecipient (ChTB-ir) and photically-responsive (Fos-ir) zones of the SCN mapped together to the middle and caudal thirds of the nucleus, predominantly in its ventro-lateral division. IMEL-binding was present throughout the full rostro-caudal extent of the SCN, but by far the most extensive area of IMEL-binding was in the rostral half of the nucleus, leading to a clear dissociation along the rostro-caudal axis of the principal zone of IMEL-binding and the retinorecipient zone of the nucleus. In the Syrian hamster, in coronal sections of the caudal SCN which did contain significant amounts of both IMEL-binding and Fos-ir, IMEL-binding was confined to the medial zone, distinct from the Fos-ir region of the ventro-lateral SCN. The segregation was less clear-cut in the Siberian hamster where the area of IMEL-binding was more extensive. The dissociation of IMEL-binding and photically-responsive cells in the Syrian hamster was confirmed in a series of sagittal sections which were processed alternately for Fos-ir and IMEL-binding. Whereas Fos-ir was confined to the ventro-lateral SCN, IMEL-binding was concentrated in the medial zone of the nucleus. In both species, mRNA for AVP was found throughout the rostro-caudal extent of the SCN, but the peak area was located in the rostral half, and so was segregated from the principal retinorecipient zone. The distribution of mRNA for AVP along the rostro-caudal and medio-lateral axes was in direct register with the IMEL-binding in both species. These studies suggest that melatonin acts upon pathways within the SCN different to those addressed by light, and that it may influence directly the efferent activity of the nucleus, possibly via an effect on vasopressinergic cells.     

Fos-like immunoreactivity in the circadian timing system of calorie-restricted rats fed at dawn: daily rhythms and light pulse-induced changes

Daily rhythms of pineal melatonin, body temperature, and locomotor activity are synchronized to the light–dark cycle (LD) via a circadian clock located in the suprachiasmatic nuclei (SCN). A timed caloric restriction in rats fed at dawn induces phase-advances and further phase-stabilization of these rhythms, suggesting that the circadian clock can integrate conflicting daily photic and non-photic cues. The present study investigated the daily expression of Fos-like immunoreactivity (Fos-ir) and light pulse-induced Fos-ir in the SCN, the intergeniculate leaflet (IGL) and the paraventricular thalamic nucleus (PVT) in calorie-restricted rats fed 2 h after the onset of light and in controls fed ad libitum. A daily rhythm of Fos-ir in the SCN was confirmed in control rats, with a peak approximately 2 h after lights on. At this time point (i.e. just prior to the feeding time), the level of SCN Fos-ir was lowered in calorie-restricted rats. Concomitantly, IGL Fos-ir was higher in calorie-restricted vs. control rats. In response to a light pulse during darkness, Fos-ir induction was found to be specifically (i.e. phase-dependently) lowered in the SCN and IGL of calorie-restricted rats. Observed changes of Fos-ir in the PVT were possibly related to the wake state of the animals. This study shows that repetitive non-photic cues presented in addition to a LD cycle affect the Fos expression in the circadian timing system.     

Entrainment of the Circadian Rhythm in Rat Pineal N-Acetyltransferase Activity Under Extremely Long and Short Photoperiods

Entrainment of a pacemaker driving the circadian rhythm in rat pineal N-acetyltransferase activity was studied under extremely long and short photoperiods. Adult male rats maintained under the light-dark regime (LD) 18:6 or under the regime LD 6:18 were exposed to a 1-min light pulse at different times at night, then they were released into darkness, and the next night phase-shifts of the evening N-acetyltransferase rise and of the morning N-acetyltransferase decline caused by light pulses were determined. The evening rise was phase-delayed by at most 0.5 h under LD 18:6, but by as much as 2.8 h under LD 6:18. The morning decline was phase-advanced by at most 1.9 h under LD 18:6, but by as much as 3.5 h under LD 6:18. Hence, the magnitude of phase-shifts and consequently patterns of phase-response curves, which show possibilities of discrete entrainment, depend on the photoperiods under which animals are maintained. A 1-min light pulse applied within 1 h before the end of the dark period phase-advanced the morning N-acetyltransferase decline under LD 18:6 as well as under LD 6:18, while a pulse applied within 1 h after the beginning of the dark period phase-delayed the evening N-acetyltransferase rise only in rats maintained under LD 18:6, but not in those kept under LD 6:18. It seems that under very long photoperiods, the N-acetyltransferase rhythm may be entrained by evening as well as by the morning light, while under very short photoperiods the rhythm may be synchronized by morning light only.     

Effect of melatonin and diazepam on the dissociated circadian rhythm in rats

The main structures involved in the circadian system in mammals are the suprachiasmatic nuclei (SCN) of the hypothalamus. The SCN contain multiple autonomous single-cell circadian oscillators that are coupled among themselves, generating a single rhythm. However, under determined circumstances, the oscillators may uncouple and generate several rhythmic patterns. Rats exposed to an artificially established 22-h light-dark cycle (T22) express two stable circadian rhythms in their motor activity that reflect the separate activities of two groups of oscillators in the morphologically well-defined ventrolateral and dorsomedial SCN subdivisions. In the experiments described in this paper, we studied the effect of melatonin and diazepam (DZP) administration in drinking water on the dissociated components of rat motor activity exposed to T22, to deduce the possible mechanism of these drugs on the circadian system. In order to suppress the endogenous circadian rhythm of melatonin, in some of the rats the pineal gland or the superior cervical ganglia were removed. The results show that melatonin or DZP treatment increased the manifestation of the light-dependent component to the detriment of the manifestation of the non-light-dependent component and that melatonin, but not DZP, shortens the period of the non-light-dependent component. These findings suggest that both DZP and melatonin favor entrainment to external light, and that melatonin could also act on the SCN, producing changes in the period of the circadian cycle.     

Repetitive transcranial magnetic stimulation interrupts phase synchronization during rhythmic motor entrainment

Rhythmic stimuli delivered through the auditory system can facilitate improved motor control following a motor impairment. The synchronization of movement to rhythmic auditory cues is characterized by quick, stable coupling of motor responses to rhythmic auditory cues. The exact neural sites responsible for this transformation of auditory input into timed rhythmic motor output are not clear. Neuroimaging studies have identified left ventral premotor cortex (vPMC) and left superior temporal-parietal (STP) activation during rhythmic auditory-motor synchronization. To investigate brain areas necessary for different types of rhythmic auditory-motor synchronization, we delivered repetitive transcranial magnetic stimulation (rTMS) to 15 healthy individuals prior to a rhythmic-auditory tapping task. Subthreshold rTMS was administered separately to the left vPMC and STP at a frequency of 0.9Hz for 15 min. Phase synchronization error (difference between auditory stimulus and response onsets) significantly increased after rTMS to STP as compared to baseline. Synchronization error also increased after rTMS to vPMC as compared to baseline, but not significantly. Absolute period error, (absolute difference between metronome interval and response interval) was not affected by rTMS. The significant effect of rTMS at the STP expands upon previous imaging research, suggesting that this area is part of the network responsible for rhythmic auditory-motor synchronization. The effect of rTMS on phase synchronization, but not period synchronization suggests these are separate neural processes controlled by different neural networks.     

Twelve-hour days in the brain and behavior of split hamsters

Hamsters will spontaneously ‘split’ and exhibit two rest–activity cycles each day when housed in constant light (LL). The suprachiasmatic nucleus (SCN) is the locus of a brain clock organizing circadian rhythmicity. In split hamsters, the right and left SCN oscillate 12 h out of phase with each other, and the twice‐daily locomotor bouts alternately correspond to one or the other. This unique configuration of the circadian system is useful for investigation of SCN communication to efferent targets. To track phase and period in the SCN and its targets, we measured wheel‐running and FOS expression in the brains of split and unsplit hamsters housed in LL or light–dark cycles. The amount and duration of activity before splitting were correlated with latency to split, suggesting behavioral feedback to circadian organization. LL induced a robust rhythm in the SCN core, regardless of splitting. The split hamsters’ SCN exhibited 24‐h rhythms of FOS that cycled in antiphase between left and right sides and between core and shell subregions. In contrast, the medial preoptic area, paraventricular nucleus of the hypothalamus, dorsomedial hypothalamus and orexin‐A neurons all exhibited 12‐h rhythms of FOS expression, in‐phase between hemispheres, with some detectable right–left differences in amplitude. Importantly, in all conditions studied, the onset of FOS expression in targets occurred at a common phase reference point of the SCN oscillation, suggesting that each SCN may signal these targets once daily. Finally, the transduction of 24‐h SCN rhythms to 12‐h extra‐SCN rhythms indicates that each SCN signals both ipsilateral and contralateral targets.     

Housing conditions influence the expression of food-anticipatory activity in mice

If food is presented for a limited duration once every 24 h, rats will gradually develop increased activity in the hours preceding mealtime, even if this is during a time of day when they are normally inactive. This food-anticipatory activity (FAA) is mediated by the entrainment of an endogenous self-sustaining circadian pacemaker. Mice are increasingly being used for the molecular analysis of behavior because of the large number of genetically manipulated mouse models available, but little is known about food entrainment in this species. This study assessed the impact of different housing conditions on the expression of FAA in a temporally restricted feeding paradigm. Wheel-running activity rhythms were recorded from mice (Mus musculus) undergoing food restriction while housed under one of two conditions. The results demonstrated that mice housed on open shelves showed robust FAA while those housed in isolation boxes did not. These results indicate that differences in susceptibility to food entrainment among mice (e.g., those with different genotypes) should be interpreted cautiously, since at least one strain is strongly affected by a relatively minor procedural difference that was not anticipated to have a significant impact.     

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.     

隐匿性拖带时起搏后间期与慢径消融成功靶点的关系

评价应用隐匿性拖带方法对准确靶点消融的有效性及探讨常规慢径靶点部位与房室结折返性心动过速(AVNRT)折返环的关系。可反复诱发的持续性典型AVNRT的患者 34例 ,消融导管在后或中间隔标测到A/V≤ 0 .5处 ,然后诱发心动过速 ,在高位右房 (HRA)和冠状窦口 (CSO)超速起搏产生隐匿性拖带 ,并按常规方法进行慢径消融。比较隐匿性拖带时靶点部位起搏后间期与心动过速周长的差值 (PPI-TCL值 )在成功靶点与不成功靶点区别。结果 :HRA超速起搏发生隐匿性拖带时 ,His束记录部位A波均为逆向夺获。而CSO超速起搏拖带时 ,32例His束记录部位A波为顺向夺获 ,另 2例为逆向夺获。在这 32例中共记录 5 4个靶点 ,成功靶点的PPI-TCL值明显小于不成功靶点 (12 .4± 5 .8msvs 32 .1± 18.6ms,P <0 .0 1)。PPI-TCL值≤ 2 0ms对靶点成功消融的敏感性和特异性分别为 84%、81%。结论 :本研究提示常规慢径消融成功部位作为房室结外的后部延伸组织参与组成AVNRT折返环或距其非常近。在可持续发作和诱发的AVNRT患者中 ,CSO部位起搏拖带顺向心房夺获时 ,靶点部位测出的PPI-TCL值≤ 2 0ms,可作为一种新的慢径路电生理定位消融方法