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

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.

Adrenergic control of pinealocyte chondriome - an in vitro study.

Norepinephrine released from sympathetic innervation plays the main role in the regulation of melatonin secretion in mammalian pinealocytes. The present study was conducted for the following reasons: 1) to establish whether the pinealocyte chondriome is controlled by norepinephrine, 2) to determine the effect of adrenergic stimulation on mitochondria, and 3) to characterize adrenoceptors involved in the regulation of the chondriome. The static organ culture of the pineal gland was used. The explants were incubated for 5 consecutive days in control medium and between 20:00 and 08:00 in medium with the presence of 10 µM norepinephrine - adrenergic agonist; isoproterenol - beta-adrenoceptor agonist; cirazoline, methoxamine, M-6364 - alfa1 - adrenoceptors agonists or PMA - activator of PKC. The explants were then subjected to ultrastructural examination and morphometric analysis. The incubation of explants in the presence of norepinephrine or isoproterenol caused a decrease in the relative volume and the numerical density of mitochondria and induced an increase in the percentage of free mitochondria in pinealocytes. Significant changes in these parameters were not observed after treatment with methoxamine, cirazoline, M-6463 and PMA. The results obtained show that the chondriome of pig pinealocytes is controlled by norepinephrine acting via beta-adrenoceptors. Adrenergic stimulation, repeated for five consecutive days of organ culture, causes a decrease in the number of mitochondria and a shift in the distribution of mitochondria from the form of networks and filaments into the form of single particles. This indicates the intensive remodeling of the mitochondria network, which is closely linked to the metabolic status of the cell.

Regulation of melatonin secretion in the pineal organ of the domestic duck--an in vitro study.

The aim of study was to determine the mechanisms regulating melatonin secretion in the pineal organs of 1-day-old and 9-month-old domestic ducks. The pineals were cultured in a superfusion system under different light conditions. Additionally, some explants were treated with norepinephrine. The pineal glands of 1-day-old ducks released melatonin in a well-entrained, regular rhythm during incubation under a 12 hrs light:12 hrs dark cycle and adjusted their secretory activity to a reversed 12 hrs dark:12 hrs light cycle within 2 days. In contrast, the diurnal changes in melatonin secretion from the pineals of 9-month-old ducks were largely irregular and the adaptation to a reversed cycle lasted 3 days. The pineal organs of nestling and adult ducks incubated in a continuous light or darkness secreted melatonin in a circadian rhythm. The treatment with norepinephrine during photophases of a light-dark cycle resulted in: 1) a precise adjustment of melatonin secretion rhythm to the presence of this catecholamine in the culture medium, 2) a very high amplitude of the rhythm, 3) a rapid adaptation of the pineal secretory activity to a reversed light-dark cycle. The effects of norepinephrine were similar in the pineal organs of nestlings and adults. In conclusion, melatonin secretion in the duck pineal organ is controlled by three main mechanisms: the direct photoreception, the endogenous generator and the noradrenergic transmission. The efficiency of intra-pineal, photosensitivity-based regulatory mechanism is markedly lower in adult than in nestling individuals.

Calcium concrements in the pineal gland of the Arctic fox (Vulpes lagopus) and their relationship to pinealocytes, glial cells and type I and III collagen fibers.

The aim of the present study was to analyze the presence and morphology of the pineal concretions in the Arctic fox and their relationship to pinealocytes, glial cells and collagen fibers. Pineals collected from 7-8 month-old and 3-4 year-old foxes (6 in each age-group) were investigated. Sections of the glands were stained with HE, Mallory's method and alizarin red S as well as subjected to a combined procedure involving immunofluorescent staining with antibodies against antigen S, glial fibril acid protein (GFAP), type I and III collagen and histochemical staining with alizarin red S. The pineal concretions were found in 2 of 6 investigated Arctic foxes aged 3 years and they were not observed in animals aged 7-8 months. The acervuli were present in the parenchyma and the connective tissue septa. They were more numerous in the distal part than in the proximal part of the gland. The acervuli stained with alizarin red S revealed an intensive red fluorescence, what enabled the use of this compound in a combined histochemical-immunofluorescent procedure. A majority of cells in the fox pineal showed positive staining with antibodies against antigen S, a marker of pinealocytes. GFAP-positive cells were especially numerous in the proximal part of the gland. Both antigen S- and GFAP-positive cells were frequently observed close to the concrements. Collagen fibers of type I and III were found in the capsule, connective tissue septa and vessels. Immunoreactive fibers did not form any capsules or basket-like structures surrounding the concrements.

Pineal concretions in turkey (Meleagris gallopavo) as a result of collagen-mediated calcification.

The intra-pineal calcification is a well-known phenomenon in mammals, however it is almost completely unknown in birds. The aim of the present work was to analyze morphology and genesis of the pineal concretions in the turkey. The studies were performed on the pineals collected from one-year-old turkeys (Meleagris gallopavo). In addition to standard morphological methods, the alizarin red S and potassium pyroantimonate methods were employed for localization of calcium at the light and electron microscopy level. In light microscopy, calcified concretions with diameters from 300 microm to 2 mm and quantities from 3 to 6 per gland were observed in all the examined pineals. They were stained red with alizarin S and showed the presence of collagen in Mallory's staining. Two types of cells were noted inside the concretion: polygonal and elongated ones. Using electron microscopy, three parts were distinguished within the calcification area. The peripheral part contained densely packed collagen fibrils, some elongated cells and numerous pyroantimonate precipitates demonstrating the presence of calcium ions. In the intermediate part, the fibrils were covered by almost continuous sheets of pyroantimonate precipitates and fused side by side. The central part showed an appearance of calcified hard tissue and contained some polygonal (osteocyte-like) cells. The obtained data demonstrated that the formation of the pineal concretions in the turkey is associated with the mineralization of collagen. This process is completely different from the mechanisms responsible for the formation of the concretions in the mammalian pineal.

Histology and ultrastructure of the pineal organ in the domestic goose.

The pineal organs of 14-week-old domestic geese were investigated with light and electron microscopy. The pineals consisted of a wide distal part and a narrow middle-proximal one. The glands were attached to the intercommissural region via the choroid plexus. The pineal parenchyma was formed by round or elongated follicles. The follicular wall was composed predominantly by cells immunoreactive with antibodies against hydroxyindolo-O-methyltransferase (HIOMT) or glial fibrillary acid protein (GFAP). They formed two or more layers. HIOMT-positive elements were represented by elongated cells bordering the follicular lumen and oval cells located in the external layer of the follicular wall. These cells were identified in ultrastructural studies as rudimentary-receptor pinealocytes and secretory pinealocytes, respectively. Among rudimentary-receptor pinealocytes two types of cells, designed as A and B, were distinguished due to structural differences. Type A cells extended through the whole follicular wall and showed regular stratified distribution of organelles in well-recognizable zones with rough endoplasmic reticulum, the Golgi apparatus and mitochondria. Type B cells, like type A pinealocytes, contacted the pineal lumen and showed polarity of their internal structure. However, they were markedly shorter than the cells of type A and lacked stratified distribution of organelles. Secretory pinealocytes contained irregularly dispersed organelles. A prominent feature of all types of goose pinealocytes was the presence of numerous dense core vesicles. The population of GFAP-positive cells consisted of ependymal-like supporting cells and astrocyte-like cells.