Diurnal variation of dopamine content in the rat pineal gland

Levels of norepinephrine and dopamine in the rat pineal gland were determined by a radioenzymatic assay with modifications to separate the reaction products. Catecholamines were converted to 3-O-methylated derivatives in the presence of catechol-O-methyltransferase (EC 2.1.1.1) and S-adenosyl-L-[methyl-³H]-methionine. Following solvent extraction of the labelled normetanephrine and 3-methoxytyramine, the amines were separated by high-performance liquid chromatography. Contents of both catecholamines in the pineal gland varied with a 24-hr rhythm. The content of norepinephrine was maximal at about 6 A.M. (lights on from 8 A.M. to 8 P.M.) and declined gradually thereafter. In contrast to the level of norepinephrine, the dopamine level was highest at about 0 A.M. and fell rapidly to reach a trough after the lights were turned on. These observations suggest that the diurnal variation of norepinephrine is generated by changes in the contents of dopamine in sympathetic nerve terminals innervating the pineal.     

Diurnal and circadian variations in indole contents in the goose pineal gland

ABSTRACT

The diurnal and circadian profiles of pineal indoles, except melatonin, are poorly characterized in birds. Moreover, there are no data on the effect of sudden changes in the light–dark cycle on these profiles. Therefore, we investigated the diurnal (Experiment I) and circadian variation (Experiment II) of nine pineal indoles (tryptophan, 5-hydroxytryptophan, serotonin, N-acetylserotonin, melatonin, 5-hydroxyindole acetic acid, 5-methoxytryptophol, 5-methoxyindole acetic acid, 5-methoxytryptamine) in geese, as well as the changes in the profiles of these substances in geese subjected to a reversed light–dark cycle (Experiment III). For the first 12 weeks of life, all geese were kept under a diurnal cycle of 12 h of light and 12 h of darkness (12L:12D). In Experiment I (n = 48), they were kept under these conditions for another 14 days before being sacrificed at 2-h intervals for sampling of the pineal glands. In Experiment II, the geese (n = 48) were divided into three groups (12L:12D, 24L:0D, 0L:24D) for 10 days before sampling at 6-h intervals. In Experiment III, 24 geese were exposed to a reversed light–dark cycle before sampling at 14:00 and 02:00 on the first, second and third days after light–dark cycle reversal. To determine the content of the indoles in the goose pineals, HPLC with fluorescence detection was used. We found that, with the exception of tryptophan, all the investigated indoles showed statistically significant diurnal variation. When geese were kept in constant darkness, most of the indoles continued to show this variation, but when geese were kept in constant light, the indoles did not show significant variation. When the light–dark cycle was reversed (12L:12D to 12D:12L), the profiles of NAS, melatonin, 5-MTAM and 5-MTOL reflected the new cycle within 2 days. The content of serotonin in geese in 12L:12D was higher than that observed in other birds under these conditions, which suggests that this compound may play a special role in the pineal physiology of this species. In conclusion, our results show that the daily variations in the metabolism of melatonin-synthesis–related indoles in the goose pineal gland are generated endogenously and controlled by environmental light conditions, as in other birds. However, comparison of the results obtained with the goose to those obtained with other species (chicken, duck) unambiguously shows that the profiles of pineal indoles differ markedly between species, in both the quantitative proportions of the compounds and the characteristics of the diurnal changes. These findings provide strong arguments for the need for comparative studies.     

Diurnal and seasonal variations of glycogen content in the rat brain.

The effect of seasonal and diurnal variations on glycogen content in the brain was investigated in 3-month old Wistar rats. In the first experimental series, the glycogen content in the brain was determined every 2-months, throughout a whole year. In each experiment, the material was taken at the same time of a day. The results indicated that the brain glycogen content did not change significantly, during the periods of a year examined. In the second series of experiments, the glycogen content in the rat brain was assayed every 3 hours, during a 24-hour-period. A low content of glycogen in the rat brain was found during the night, with a pronounced decrease of this value between 9.00 p.m. and midnight. A possible relationship between the diurnal changes in the brain glycogen content and catecholamine metabolism in the central nervous system is discussed.     

Proteomic analysis of day-night variations in protein levels in the rat pineal gland

The pineal gland secretes the hormone melatonin. This secretion exhibits a circadian rhythm with a zenith during night and a nadir during day. We have performed proteome analysis of the superficial pineal gland in rats during daytime and nighttime. The proteins were extracted and subjected to 2-DE. Of 1747 protein spots revealed by electrophoresis, densitometric analysis showed the up-regulation of 25 proteins during nighttime and of 35 proteins during daytime. Thirty-seven of the proteins were identified by MALDI-TOF MS. The proteins up-regulated during the night are involved in the Krebs cycle, energy transduction, calcium binding, and intracellular transport. During the daytime, enzymes involved in glycolysis, electron transport, and also the Krebs cycle were up-regulated as well as proteins taking part in RNA binding and RNA processing. Our data show a prominent day-night variation of the protein levels in the rat pineal gland. Some proteins are up-regulated during the night concomitant with the melatonin secretion of the gland. Other proteins are up-regulated during the day indicating a pineal metabolism not related to the melatonin synthesis.     

Circadian variations of vasopressin level and vasopressin-converting aminopeptidase activity in the rat pineal gland

Vasopressin levels and vasopressin-converting aminopeptidase activity were measured in the rat pineal gland during the 24 hr light-dark cycle. A rhythmic variation in peptide levels and peptidase activity occurred. At the onset of light at 6.00 hr, the peptidase displayed a significant, short-lasting (approximately 3 hr) increase of about 35% in activity, while a decrease of 28% in pineal vasopressin levels was observed. The changes in peptidase activity and peptide level were not triggered by light per se, since they persisted to occur at the same time point in animals which were not exposed to light, indicating the circadian nature of the rhythmicity. These changes were specific to the pineal gland, since other tissues, like hippocampus and pituitary gland, did not show these daily variations. The data suggest a relationship between vasopressin levels and vasopressin-converting aminopeptidase activity.     

Circadian and seasonal variations in pineal gland intercellular canaliculi in the white rat.

Seventy Wistar rats are used to study the changes in pineal intercellular canaliculi over a 21-hour period and for two different photoperiods (pre-autumn, first week of September, and winter, first week of February). The study considers these changes at pineal body, cortical and medullar level separately, and compares the values obtained. The results show variations in canalicular surface at different point times (10:00, 14:00, 18:00) and for both photoperiods. The variations are found to favour the cortical layer, and are also observed between nocturnal and diurnal hours. Canalicular surface to greater during the diurnal hours of both photoperiods. Interesting histological findings are described that suggest an important function of the intercellular canaliculi in pineal gland metabolic exchange.     

Alteration in the D-amino acid content of the rat pineal gland under anesthesia

Summary In a previous report (Hamase, K. et al., Biochim Biophys Acta 1134: 214–222 (1997)), we showed that the rat pineal gland contains D-leucine (D-Leu) as well as D-aspartic acid (D-Asp). In this communication we report alterations in the content of these D-amino acids during anesthesia. The D-Asp content was significantly increased from 2.8 to 5.0, 4.8 and 5.8 nmol/pineal gland by administration of ether, urethane and pentobarbital, respectively. In contrast, the D-Leu content was decreased by administration of urethane or pentobarbital. The D-Leu content decreased from 4.2 to 2.2 pmol/pineal gland 4 hours after administration of urethane, although the content remained unchanged until 1.5 hours after administration. The content of the L-enantiomers of these amino acids were not affected by anesthesia. The urethane-induced decrease in D-leucine content was almost completely suppressed by a-agonist, (-)-isoproterenol, whereas the agonist itself had no effect.     

Determination of rat pineal gland melatonin content by a radioimmunoassay.

Melatonin content in individual rat pineal glands was measured by radioimmunoassay (RIA). The RIA used can very reliably detect as little as 50 pg of melatonin. The various precursors, analogues, and the metabolite of melatonin (6-hydroxymelatonin) which were tested for cross-reactivity were not recognized by the antibody. The effects on melatonin levels in rat pineal glands following the administration of L-tryptophan, 5-hydroxy-L-tryptophan, serotonin, N-acetylserotonin, melatonin and pargyline are also presented.     

Quantitative variations in nucleolar components within pineal gland cells during the rat estrous cycle

Quantitative changes in the size of pinealocyte nucleoli have been reported in various studies on this cell type. However, the significance of quantitative changes in the nucleolar components is unknown. The present study is an attempt to analyze ultrastructural and morphometric modifications occurring in the pinealocyte nucleolar components during the estrous cycle in female rats. The fibrillar centers showed an increase during estrus consistent with a decrease in pinealocyte nucleolar activity and melatonin pineal levels. The fibrillar components and granular components tended to display a reciprocal relationship. An increase in the dense fibrillar component took place at metaestrus and diestrus when melatonin synthesis increased in pinealocytes. Maximum values of granular and interstitial components were found at the proestrus phase before the day of ovulation.     

Seasonal variations of gonadotropins and prolactin in the laboratory rat. Role of maternal pineal gland

The laboratory rat, a non-photoperiodic rodent, exhibits seasonal fluctuations of melatonin. Melatonin has been found to be readily transferred from the maternal to the fetal circulation. No data exist on the possible influence of maternal pineal gland upon seasonal variations of the offspring. The aim of the present study was to asses the influence of the maternal melatonin rhythm on the offspring postnatal development of the reproductive hormones LH, FSH and prolactin. Male offspring from control, pinealectomized (PIN-X) and PIN-X + melatonin (PIN-X+MEL) mother Wistar rats were studied at 21, 31, and 60 days of age. Seasonal age-dependent variations were found for all hormones studied in control offspring but PIN-X offspring showed a tendency to have reduced duration or altered seasonal variations. Maternal melatonin treatment to PIN-X mothers partially restored the effect of pinealectomy. The chronological study of LH, FSH, and prolactin in PIN-X offspring also showed an altered pattern as compared to control-offspring. Melatonin treatment to the mothers partially restored the developmental pattern of reproductive hormones. Results of this study indicate that maternal pineal gland of the laboratory rat is involved in the seasonal postnatal development variations of reproductive hormones of the offspring.     

Diurnal variations in the catecholamine content in rat tissues. Effects of exogenous noradrenaline

Endogenous noradrenaline (NA) content of the interscapular brown adipose tissue (IBAT), and of the hypothalamus, as well as adrenaline (A) and NA content of the adrenals were measured in the morning and in the evening in rats treated with saline (controls) or with NA (1.6 mg kg-1 pp). NA content in IBAT and in hypothalamus of control animals varies diurnally. NA content in IBAT was significantly higher in the evening than in the morning. In the hypothalamus, these variations were different of those found in the IBAT. Exogenous NA applied both in the morning and in the evening increases significantly NA content in IBAT both in the morning and in the evening, but more in the evening. Unlike IBAT, NA content of the hypothalamus after NA administration does not change essentially. In the adrenal gland of control rats, A content only is changed during the day, being markedly lower in the evening. After NA injection both A and NA are not changed significantly.     

Cholinergic signaling in the rat pineal gland

Summary 1. Innervation of the mammalian pineal gland is mainly sympathetic. Pineal synthesis of melatonin and its levels in the circulation are thought to be under strict adrenergic control of serotoninN-acetyltransferase (NAT). In addition, several putative pineal neurotransmitters modulate melatonin synthesis and secretion.2. In this review, we summarize what is currently known on the pineal cholinergic system. Cholinergic signaling in the rat pineal gland is suggested based on the localization of choline acetyltransferase (ChAT) and acetylcholinesterase (AChE), as well as muscarinic and nicotinic ACh binding sites in the gland.3. A functional role of ACh may be regulation of pineal synaptic ribbon numbers and modulation of melatonin secretion, events possibly mediated by phosphoinositide (PI) hydrolysis and activation of protein kinase C via muscarinic ACh receptors (mAChRs).4. We also present previously unpublished data obtained using primary cultures of rat pinealocytes in an attempt to get more direct information on the effects of cholinergic stimulus on pinealocyte melatonin secretion. These studies revealed that the cholinergic effects on melatonin release are restricted mainly to intact pineal glands since they were not readily detected in primary pinealocyte cultures.     

Alterations in γ-aminobutyric acid content in the rat superior cervical ganglion and pineal gland

The endogenous γ-aminobutyric acid (GABA) content of the rat pineal gland and superior cervical ganglion (SCG) was measured by high pressure liquid chromatography. It was found that GABA levels in both tissues increased after decapitation of the animals. The GABA content of tissues frozen within 20 seconds after decapitation was the same as that of tissues removed from animals killed by microwave irradiation. Amino-oxyacetic acid, a GABA-transaminase inhibitor, increased the endogenous GABA content of both of these tissues. Dimethylphenylpiperizinium or isoniazid administration did not alter GABA levels in these tissues. Isoproterenol increased the GABA content of the SCG but did not change the pineal gland GABA levels. The ability of the pineal gland to take up and accumulate ³H-GABA was significantly reduced in rats that had been ganglionectomized. A fluctuation in endogenous GABA levels in the pineal gland was seen to occur when measures were taken at different times of the day. These results tend to suggest that GABA may have some functional role in the pineal gland and the superior cervical ganglion.     

Diurnal variations of potassium content in lucerne plants

Diurnal changes in K content in leaf blades, petioles, stems and roots of eleven lucerne genotypes were followed. Significant positive correlations between changes in K content in petioles and upper half of stems and significant negative correlations between changes of K content in leaf blades and lower half of stems reflected rapid K movement. The velocity — up to 60 μmol g−1 (f.m.)h−1 — of changes in K content from leaf blades to lower part of stems and the other way round showed that long distance phloem transport occurred. Only moderate increase of K content contemporarily took place in roots. When total K amount in the whole plant was calculated then K uptake alternatively with K release were noticed during the day. Average K release reached 1.48 μmol g−1 (f.m.) h−1. The rate of K movement correlated with irradiance and physiological activity of plants. The time course of K movement was uniform in plants of the same strain and it differed partially in different strains.     

Rhythmic control of endocannabinoids in the rat pineal gland

Endocannabinoids modulate neuroendocrine networks by directly targeting cannabinoid receptors. The time-hormone melatonin synchronizes these networks with external light condition and guarantees time-sensitive and ecologically well-adapted behaviors. Here, the endocannabinoid arachidonoyl ethanolamide (AEA) showed rhythmic changes in rat pineal glands with higher levels during the light-period and reduced amounts at the onset of darkness. Norepinephrine, the essential stimulus for nocturnal melatonin biosynthesis, acutely down-regulated AEA and other endocannabinoids in cultured pineal glands. These temporal dynamics suggest that AEA exerts time-dependent autocrine and/or paracrine functions within the pineal. Moreover, endocananbinoids may be released from the pineal into the CSF or blood stream.     

12- and 15-lipoxygenases in rat pineal gland

A whole organ or a homogenate of rat pineal gland was incubated with arachidonic Acid. Two predominant metabolites were identified by mass spectrometry to be 12-hydroxy-5,8,10,14-eicosatetraenoic acid and 10-hydroxy-11,12-epoxy-5,8,14-eicosatrienoic acid. 15-Hydroxy-5,8,11,13-eicosatetraenoic acid was also formed in a smaller amount. In addition, peroxy acids appeared rapidly only at the initial stage of reaction. In various parts of rat brain the 12-lipoxygenase activity was by far the highest in pineal gland, and less than 5% of the activity was found in pituitary gland and hypothalamus.