Circadian Clock Protein Content and Daily Rhythm of Locomotor Activity Are Altered after Chronic Exposure to Lead in Rat

Lead exposure has been reported to produce many clinical features, including parkinsonism. However, its consequences on the circadian rhythms are still unknown. Here we aimed to examine the circadian rhythms of locomotor activity following lead intoxication and investigate the mechanisms by which lead may induce alterations of circadian rhythms in rats. Male Wistar rats were injected with lead or sodium acetate (10 mg/kg/day, i.p.) during 4 weeks. Both groups were tested in the “open field” to quantify the exploratory activity and in the rotarod to evaluate motor coordination. Then, animals were submitted to continuous 24 h recordings of locomotor activity under 14/10 Light/dark (14/10 LD) cycle and in complete darkness (DD). At the end of experiments, the clock proteins BMAL1, PER1-2, and CRY1-2 were assayed in the suprachiasmatic nucleus (SCN) using immunohistochemistry. We showed that lead significantly reduced the number of crossing in the open field, impaired motor coordination and altered the daily locomotor activity rhythm. When the LD cycle was advanced by 6 h, both groups adjusted their daily locomotor activity to the new LD cycle with high onset variability in lead-intoxicated rats compared to controls. Lead also led to a decrease in the number of immunoreactive cells (ir-) of BMAL1, PER1, and PER2 without affecting the number of ir-CRY1 and ir-CRY2 cells in the SCN. Our data provide strong evidence that lead intoxication disturbs the rhythm of locomotor activity and alters clock proteins expression in the SCN. They contribute to the understanding of the mechanism by which lead induce circadian rhythms disturbances

Mélatonine, rythme de la température corporelle et organisation de l'hypothalamus et des noyaux suprachiasmatiques chez le dromadaire (Camelus dromedarius) (Démonstration de l'entraînement de l'horloge circadienne par la photopériode et par la température ambiante)

Dans les zones désertiques, les animaux sont exposés à un milieu hostile. Dans ce biotope particulier la question de la nature des facteurs environnementaux utilisés pour contrôler la saisonnalité est importante. En étudiant les variations saisonnières du profil de la sécrétion de la mélatonine qui sont parallèles aux changements annuels de la photopériode, nous avons pu montrer que le Dromadaire est capable de mesurer et d intégrer l information photopériodique. La synthèse de la mélatonine dépend d une régulation post-transcriptionelle de l enzyme AANAT avec des niveaux égaux de jour et de nuit des ARNm-Aa-nat. Dans un 2ème temps, nous avons étudié les noyaux suprachiasmatiques (SCN) du Dromadaire, siège de l horloge biologique. Une cartographie de l hypothalamus a été réalisée. Les SCN sont longs et contiennent des neurones à tyrosine hydroxylase, à ocytocine et plusieurs types d innervation (NPY, 5-HT, Met-enk ). Le système hypothalamo-neurohypophysaire est large et comprend une multitude de noyaux. Dans le biotope désertique, nous avons supposé qu a côté de la photopériode, d autres facteurs pouvaient entraîner l horloge biologique. Nous avons étudié la possibilité d un entraînement non photique de l horloge par le cycle de la température ambiante (TA). Après avoir démontré que le rythme de la température corporelle (TCR) est contrôlé par l horloge circadienne et dépendant de la photopériode, nous avons pu établir que ce rythme et celui de la mélatonine pouvait être entraînés par la TA. Chez le Dromadaire, ce cycle de la TA est donc un véritable zeitgeber qui doit désormais être pris en compte dans les études du contrôle des rythmes saisonniers (reproduction ).In desert areas, the dromedary camel is exposed to a hostile environment. In such habitat, the question of the nature of the environmental parameters which are important in the control of seasonality has to be raised. By the observation that the seasonal profile of melatonin secretion parallels the annual changes in photoperiod, we have demonstrated that the dromedary Camel is able to measure and integrate annual photoperiod. The melatonin synthesis depends of a post-transcriptional regulation of the enzyme AA-NAT with equal high levels of mRNA gene Aa-nat during the day and during the night (the mRNA expression of AA-NAT is constitutive). Afterward, we have studied the suprachiasmatic nuclei (SCN) which contains the biological clock. The SCN are long and show numerous tyrosine hydroxylase neurons and some of oxytocin but also several types of fibers. A mapping of the hypothalamus nuclei of the camel has been also performed. The hypothalamic-neurohypophysial system is vast and includes a multitude of nuclei. Within the desert area it is probable that, beside photoperiod, other environmental cues could entrain the biological clock. We examined the possibility of a non-photic synchronization of the clock by the ambient temperature cycle (AT). Having demonstrated the circadian clock control of the rhythm of body temperature (CRT) and its dependence on the photoperiod, we established then that this rhythm and that of melatonin could be driven by the AT. In the Camel, the AT is a strong zeitgeber and should now be taken into account in studies aiming control of seasonal rhythms.STRASBOURG-Sc. et Techniques (674822102) / SudocSudocFranceF

Manganese-induced atypical parkinsonism is associated with altered Basal Ganglia activity and changes in tissue levels of monoamines in the rat.

Manganese neurotoxicity is associated with motor and cognitive disturbances known as Manganism. However, the mechanisms underlying these deficits remain unknown. Here we investigated the effects of manganese intoxication on motor and non-motor parkinsonian-like deficits such as locomotor activity, motor coordination, anxiety and "depressive-like" behaviors. Then, we studied the impact of this intoxication on the neuronal activity, the globus pallidus (GP) and subthalamic nucleus (STN). At the end of experiments, post-mortem tissue level of the three monoamines (dopamine, norepinephrine and serotonin) has been determined. The experiments were carried out in adult Sprague-Dawley rats, daily treated with MnCl2 (10 mg/kg/, i.p.) for 5 weeks. We show that manganese progressively reduced locomotor activity as well as motor coordination in parallel with the manifestation of anxiety and "depressive-like" behaviors. Electrophysiological results show that, while majority of GP and STN neurons discharged regularly in controls, manganese increased the number of GP and STN neurons discharging irregularly and/or with bursts. Biochemical results show that manganese significantly decreased tissue levels of norepinephrine and serotonin with increased metabolism of dopamine in the striatum. Our data provide evidence that manganese intoxication is associated with impaired neurotransmission of monoaminergic systems, which is at the origin of changes in basal ganglia neuronal activity and the manifestation of motor and non-motor deficits similar to those observed in atypical Parkinsonism

Manganese progressively reduced exploratory activity.

<p>Exploratory activity histograms represent the number of horizontal (A), stereotypic (B), and vertical movements (C) recorded during the first session of 10 min in the “open field” actimeter before and during all period of treatment. Values are the mean ± SEM. Data from manganese-treated rats (<i>n</i> = 12) and controls (<i>n</i> = 11) were compared using the two way repeated measures ANOVA and Holm-Sidak <i>post hoc</i>. *<i>p</i><0.05, **<i>p</i><0.01.</p

Manganese progressively reduced motor coordination.

<p>Motor coordination histogram represents the time that rat stay on the bar of rotarod before and during all period of treatment. Values are the mean ± SEM. Data from manganese-treated rats (<i>n</i> = 5) and controls (<i>n</i> = 5) were compared using the two way repeated measures ANOVA and Holm-Sidak <i>post hoc</i>. **<i>p</i><0.01, ***<i>p</i><0.001.</p

Manganese decreased the firing rate of GP neurons and increased the proportion of bursty and irregular neurons.

<p>(ABC) Representative examples of spike trains recorded in the GP (a) with interspike interval histogram (b) and density histogram (c), showing a regular pattern in control rats (A) and irregular and bursty patterns in manganese (Mn)-treated animals (B and C respectively). (D) Firing rate histograms with values as the mean ± SEM. Firing rate data from manganese-treated rats and controls were compared using Student <i>t</i>-test. **<i>p</i><0.01. (E) Firing pattern histograms showing the proportion of GP cells discharging regularly, irregularly or with bursts. Changes in the proportion of different firing patterns were analyzed using a chi square test. ***<i>p<</i>0.001 in comparison with controls.</p

Manganese decreased the firing rate of STN neurons and increased the proportion of irregular neurons.

<p>(AB) Representative examples of spike trains recorded in the STN (a) with interspike interval histogram (b) and density histogram (c), showing a regular pattern in control rats (A) and irregular pattern in manganese (Mn)-treated animals (B). (<b>C</b>) Firing rate histograms with values as the mean ± SEM. Firing rate data from manganese-treated rats and controls were compared using Student <i>t</i>-test. (D) Firing pattern histograms showing the proportion of STN cells discharging regularly, irregularly or with bursts. Changes in the proportion of different firing patterns were analyzed using a chi square test. *<i>p</i><0.05, ***<i>p<</i>0.001 in comparison with controls.</p

Biochemical analysis of manganese-induced changes in the tissue level of monoamines.

<p>Manganese-induced a decrease in the tissue level of NE and 5-HT in the frontal cortex and increase of DA metabolism in the striatum. Tissue contents of DA, DOPAC, HVA, 5-HT, and 5-HIAA in the striatum and NE, 5-HT, and 5-HIAA in the frontal cortex measured by HPLC in manganese-treated rats and their controls. Values are concentrations in ng/g of wet tissue presented as the mean ± SEM. Statistical analysis using Mann–Whitney test was performed;</p><p>*<i>p</i><0.05 in comparison with control.</p