Florence Cayetanot

Age-related effects on the biological clock and its behavioral output in a primate.

In humans, activity rhythms become fragmented and attenuated in the elderly. This suggests an alteration of the circadian system per se that could in turn affect the expression of biological rhythms. In primates, very few studies have analyzed the effect of aging on the circadian system. The mouse lemur provides a unique model of aging in non‐human primates. To assess the effect of aging on the circadian system of this primate, we recorded the circadian and daily rhythms of locomotor activity of mouse lemurs of various ages. We also examined age‐related changes in the daily rhythm of immunoreactivities for vasoactive intestinal polypeptide (VIP) and arginine‐vasopressin (AVP) in suprachiasmatic nucleus neurons (SCN), two major peptides of the biological clock. Compared to adult animals, aged mouse lemurs showed a significant increase in daytime activity and an advanced activity onset. Moreover, when maintained in constant dim red light, aged animals exhibited a shortening of the free‐running period compared to adult animals. In adults, AVP immunoreactivity (ir) peaked during the second part of the day, and VIP ir peaked during the night. In aged mouse lemurs, the peaks of AVP ir and VIP ir were significantly shifted with no change in amplitude. AVP ir was most intense at the beginning of the night; whereas, VIP ir peaked at the beginning of the daytime. A weakened oscillator could account for the rhythmic disorders often observed in the elderly. Changes in the daily rhythms of AVP ir and VIP ir may affect the ability of the SCN to transmit rhythmic information to other neural target sites, and thereby modify the expression of some biological rhythms.

Shortened seasonal photoperiodic cycles accelerate aging of the diurnal and circadian locomotor activity rhythms in a primate

The gray mouse lemur (Microcebus murinus), a prosimian primate, exhibits seasonal rhythms strictly controlled by photoperiodic variations. Previous studies indicated that longevity can be altered by long-term acceleration of seasonal rhythms, providing a model for assessing various aspects of aging. To assess the effect of aging and accelerated aging on the circadian system of this primate, we compared the circadian rhythm of the locomotor activity in adult mouse lemurs (2-4.5 years, n = 9), aged mouse lemurs (5-9 years, n = 10), and adult mouse lemurs that had been exposed from birth to a shortened seasonal photoperiodic cycle (2-4.5 years, n = 7). Compared to adult animals, aged mouse lemurs showed a significant increase in intradaily variability and an advanced activity onset. Aging was characterized by a decrease in amplitude, with both a decrease in nocturnal activity and an increase in daytime activity. When maintained in constant dim red light, aged animals exhibited a shortening of the free-running period (22.8 ± 0.1 h) compared to adult animals (23.5 ± 0.1 h). A 3- to 5-year exposure to an accelerated seasonal photoperiodic rhythm (“annual” duration of 5 months) in accelerated mouse lemurs produced disturbances of the locomotor activity rhythm that resembled those of aged mouse lemurs, whether animals were studied in entrained or in free-running conditions. The present study demonstrated a weakened and fragmented locomotor activity rhythm during normal aging in this primate. Increasing the number of expressed seasonal cycles accelerated aging of parameters related to circadian rhythmicity in adult animals.

Arginine-vasopressin and vasointestinal polypeptide rhythms in the suprachiasmatic nucleus of the mouse lemur reveal aging-related alterations of circadian pacemaker neurons in a non-human primate

The suprachiasmatic nucleus (SCN) of the hypothalamus, the mammalian circadian pacemaker, is entrained by external cues and especially by photic information. Light is transmitted primarily via the retinohypothalamic tract, which terminates in the ventral part (or core) of the SCN, where vasoactive intestinal polypeptide (VIP)‐containing neurons are located. VIP cells are mainly intrinsic and project to the dorsal part (or shell) of the SCN, where neurons containing arginine‐vasopressin (AVP) reside. As aging leads to marked changes in the expression of circadian rhythms, we examined in primates whether age‐related decay in biological rhythmicity is associated with changes in the oscillation of peptide expression in SCN neurons. We used double immunohistochemistry and quantitative analysis in the SCN of mouse lemurs, which provide a unique model of aging in non‐human primates. In adult animals, VIP‐positive and AVP‐positive SCN neurons exhibited daily rhythms of their number and immunostaining intensity: AVP immunoreactivity peaked during the second part of the day, and VIP peaked during the night. In aged mouse lemurs, the peaks of AVP and VIP immunopositivity were significantly shifted, so that AVP was most intense at the beginning of the night, whereas VIP peaked at the beginning of daytime. The results show that the circadian rhythm of neuropeptides in the SCN is modified by aging in primates, with a differential regulation of the two main peptidergic cell populations. These changes may affect the ability of the SCN to transmit rhythmic information to other neural target sites, and thereby to modify the expression of some biological rhythms.

Redistribution of CB1 cannabinoid receptors during evolution of cholinergic basal forebrain territories and their cortical projection areas: A comparison between the gray mouse lemur (Microcebus murinus, primates) and rat

Endocannabinoid signaling, mediated by presynaptic CB1 cannabinoid receptors on neurons, is fundamental for the maintenance of synaptic plasticity by modulating neurotransmitter release from axon terminals. In the rodent basal forebrain, CB1 cannabinoid receptor-like immunoreactivity is only harbored by a subpopulation of cholinergic projection neurons. However, endocannabinoid control of cholinergic output from the substantia innominata, coincident target innervation of cholinergic and CB1 cannabinoid receptor-containing afferents, and cholinergic regulation of endocannabinoid synthesis in the hippocampus suggest a significant cholinergic-endocannabinergic interplay. Given the functional importance of the cholinergic modulation of endocannabinoid signaling, here we studied CB1 cannabinoid receptor distribution in cholinergic basal forebrain territories and their cortical projection areas in a prosimian primate, the gray mouse lemur. Perisomatic CB1 cannabinoid receptor immunoreactivity was unequivocally present in non-cholinergic neurons of the olfactory tubercule, and in cholecystokinin-containing interneurons in layers 2/3 of the neocortex. Significantly, CB1 cannabinoid receptor-like immunoreactivity was localized to cholinergic perikarya in the magnocellular basal nucleus. However, cortical cholinergic terminals lacked detectable CB1 cannabinoid receptor levels. A dichotomy of CB1 cannabinoid receptor distribution in frontal (suprasylvian) and parietotemporal (subsylvian) cortices was apparent. In the frontal cortex, CB1 cannabinoid receptor-containing axons concentrated in layers 2/3 and layer 6, while layer 4 and layer 5 were essentially devoid of CB1 cannabinoid receptor immunoreactivity. In contrast, CB1 cannabinoid receptors decorated axons in all layers of the parietotemporal cortex with peak densities in layer 2 and layer 4. In the hippocampus, CB1 cannabinoid receptor-containing terminals concentrated around pyramidal cell somata and proximal dendrites in the CA1-CA3 areas, and granule cell dendrites in the molecular layer of the dentate gyrus. CB1 cannabinoid receptors frequently localized to inhibitory GABAergic terminals while leaving glutamatergic boutons unlabeled. Aging did not affect either the density or layer-specific distribution of CB1 cannabinoid receptor-immunoreactive processes. We concluded that organizing principles of CB1 cannabinoid receptor-containing neurons and their terminal fields within the basal forebrain are evolutionarily conserved between rodents and prosimian primates. In contrast, the areal expansion and cytoarchitectonic differentiation of neocortical subfields in primates is associated with differential cortical patterning of CB1 cannabinoid receptor-containing subcortical and intracortical afferents.

Consequences of in utero caffeine exposure on respiratory output in normoxic and hypoxic conditions and related changes of Fos expression: a study on brainstem-spinal cord preparations isolated from newborn rats.

Several aspects of the central regulation of respiratory control have been investigated on brainstem-spinal cord preparations isolated from newborn rats whose dam was given 0.02% caffeine in water as drinking fluid during the whole period of pregnancy. Analysis of the central respiratory drive estimated by the recording of C4 ventral root activity was correlated to Fos pontomedullary expression. Under normoxic conditions, preparations obtained from the caffeine-treated group of animals displayed a higher respiratory frequency than observed in the control group (9.2 ± 0.5 versus 7.2 ± 0.6 burst/min). A parallel Fos detection tends to indicate that the changes of the respiratory rhythm may be due to a decrease in neuronal activity of medullary structures such as the ventrolateral subdivision of the solitary tract, the area postrema, and the nucleus raphe obscurus. Under hypoxic conditions, the preparations displayed a typical hypoxic respiratory depression associated with changes in the medullary Fos expression pattern. In addition, the hypoxic respiratory depression is clearly emphasized after in utero exposure to caffeine and coincides with an increased Fos expression in the area postrema and nucleus raphe obscurus, two structures in which it is not increased in the absence of caffeine. Taken together, these results support the idea that in utero caffeine exposure could affect central respiratory control.

Possible role of retrotrapezoid nucleus and parapyramidal area in the respiratory response to anoxia: an in vitro study in neonatal rat

The brainstem-spinal cord preparation from neonatal rat has been used in several reports to evaluate the central effect of low oxygen level on the respiratory network. We demonstrate that bilateral lesion of retrotrapezoid nucleus and parapyramidal area unmasks an early reinforcement of the respiratory output in response to anoxia. This suggests that neurons in both areas might trigger or relay a central depressive influence of hypoxia on the respiratory network.

Attenuated effect of increased daylength on activity rhythm in the old mouse lemur, a non-human primate

Adaptation of physiological and behavioral functions to seasonal changes in daylength is of major relevance for optimal fitness and survival. Because aging is characterized by changes in biological rhythms, it may be hypothesized that old animals fall short of showing a full adaptation to prolonged changes in the duration of daily light exposure, as naturally occurring in relation to season in younger individuals. To test this hypothesis, we analyzed changes in the patterns of daily locomotor activity and body temperature rhythms of young and old mouse lemurs (Microcebus murinus, Primates) exposed to short and long daylengths. The effect of an increase in the duration of daily light exposure was attenuated in old animals, as compared to younger lemurs. Although some age-related differences in the locomotor activity rhythm could be seen under exposure to short daylength, they were predominant under long daylength. Some mechanisms allowing adaptation to changing daylength thus seem to be impaired at old age. Changes in coupling of circadian oscillators to the light-dark cycle and disturbances in the physiological responses to change in light duration should be further investigated.

Calbindin D28K protein cells in a primate suprachiasmatic nucleus: localization, daily rhythm and age-related changes.

In mammals, the suprachiasmatic nucleus (SCN) of the hypothalamus is the master circadian pacemaker. The SCN controls daily rhythms and synchronizes the organism to its environment and especially to photic signals. Photic signals via the retinohypothalamic tract reach the ventral part of the SCN, where the majority of calbindin‐containing neurons are located. Calbindin cells seem important for the control of circadian rhythmicity. As ageing leads to marked changes in the expression of circadian rhythms, we investigated in the mouse lemur, a nocturnal primate, age‐related changes in the oscillation of calbindin protein expression in SCN neurons. We used immunohistochemistry and quantitative analysis of calbindin expression in the SCN of adult and aged mouse lemurs. In this primate, a dense cluster of calbindin‐positive neurons was found in the ventral part of the SCN. In adult animals, calbindin‐positive SCN neurons did not exhibit daily rhythms in their number or intensity, but exhibited significant daily variations in the percentage of cells with a calbindin‐positive nucleus, characterized by high values during the daytime and low values during the night. Immunoreactive intensity peaked in the middle of the daytime. Calbindin expression in the nuclei of calbindin cells in the SCN tends to be modified by ageing. The amplitude of daily variation in calbindin expression was damped, with a lower immunointensity during the daytime and a delayed decrease during the night. These changes may affect the ability of the SCN to transmit rhythmic information to other SCN cells and thereby modify the synchronization of the different cell populations in the SCN.

5-HT acting on 5-HT1/2 receptors does not participate in the in vitro hypoxic respiratory depression

The involvement of serotoninergic mechanisms in the central respiratory depression produced by hypoxia was studied in the newborn rat brainstem-spinal cord preparation. The respiratory frequency measured by the C4 ventral root activity was recorded. 5-HT (30 microM) superfusion elicited a rapid increase in respiratory frequency, prevented by a treatment with methysergide (a 5-HT(1/2) receptor antagonist) (40 microM). To investigate the possible participation of 5-HT in hypoxic respiratory depression, this concentration of methysergide was added to the bathing medium during hypoxia. Methysergide did not modify the decrease in respiratory frequency produced by hypoxia. In order to ensure that other 5-HT subtype receptors were not involved in hypoxic respiratory depression, 5-HT was added to the bath during hypoxic-methysergide tests; no effect on respiratory frequency was observed. These results suggest that in the newborn rat brainstem-spinal cord preparation, serotoninergic mechanisms are not involved in the elaboration of the in vitro respiratory response to hypoxia.

Fos study of ponto-medullary areas involved in the in vitro hypoxic respiratory depression

In this study, the brainstem–spinal cord preparation isolated from newborn rats, an established model for the study of the hypoxic respiratory depression (HRD), has been used. The comparison of Fos expression in ponto-medullary areas in these preparations placed either in normoxic or hypoxic conditions suggests that only the retrotrapezoid nucleus (RTN) and the ventrolateral medulla (VLM) are involved in the in vitro HRD. Hypoxic preparations exhibit a Fos expression enhanced in the RTN, suggesting that the RTN might play a crucial role in the HRD. As well as this, VLM neurons presented a decrease in Fos expression that could be related to the decline of the respiratory output induced by hypoxia.