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PubMed1994

Circadian variations of neuropeptide Y-like immunoreactivity in the rat pineal gland.

Neuropeptide Y, which is present in the sympathetic nerves innervating pineal glands, has been shown to affect N-acetyltransferase activity and melatonin production. Using enzyme immunoassay, we measured daily patterns of neuropeptide Y (NPY) in rat pineal glands in light-dark cycles (LD) and in constant darkness (DD). NPY within rat pineal glands displayed daily rhythms in LD with a peak at zeitgeber time 16. Similar rhythms in the NPY content were also observed in DD. These rhythms have a similar phase-relationship to that of the rhythm of NAT activity, suggesting that NPY in rat pineal glands plays a role in circadian rhythms of NAT activity.

Animals↗
PubMed1988

Effects of continuous light exposure on the rat retina and pineal gland.

We examined the effects of intense continuous light (400 ftc for 10 days) on the morphology and immunoreactivity of the rat retina and pineal gland. The light treatment caused severe degeneration in the retina, with loss of most photoreceptor cells, and produced a marked decrease in S-antigen immunoreactivity in this organ. Unlike the retina, the light treatment had minimal effects on the histological structure of the pineal gland. However, pineal glands of light-treated rats exhibited a substantial reduction in their S-antigen immunoactivity, as demonstrated immunohistochemically and quantitatively verified by the rocket immunoelectrophoresis technique: pineal glands of light exposed rats had approximately two-thirds of the S-antigen immunoactivity of the untreated controls. Light treatment was also found to reduce both uveoretinitis and pinealitis in rats immunized with S-antigen. Yet pinealitis was not affected by light exposure in rats, as they were enucleated before treatment. This study thus provides new information to support the notion that the mammalian pineal gland is directly associated with light detection by the retina.

Animals↗
PubMed1992

Vasoactive intestinal peptide stimulates N-acetyltransferase and hydroxyindole-O-methyltransferase activities and melatonin production in cultured rat but not in Syrian hamster pineal glands.

The purpose of this study was to compare the responses of the Syrian hamster and rat pineal glands in organ culture to vasoactive intestinal peptide (VIP). The endpoints in these studies were the activities of pineal N-acetyltransferase (NAT) and hydroxyindole-O-methyltransferase (HIOMT), as well as pineal and medium melatonin levels. When rat pineal glands were incubated with either VIP (1 microM) or isoproterenol (1 microM), a beta-adrenergic agonist, a significant increase in NAT and HIOMT activities and melatonin levels were observed within 3 hr. Conversely, during the day, VIP (1 microM) was ineffective in stimulating these parameters in hamster pineal gland after incubation times of either 2, 4, 6, or 8 hr. In another experiment, hamster pineal glands were collected from animals killed in the late dark period (after 30 min light exposure). In these glands, isoproterenol promoted NAT activity and melatonin production; however, VIP was ineffective in stimulating either NAT or HIOMT activities; likewise, VIP had no stimulatory effect on pineal melatonin levels at night. Finally, when hamster pineal glands at night were incubated with either 0, 10 nM, 100 nM, 10 microM, or 100 microM VIP, no changes in any parameter of melatonin synthesis were measured. The results indicate that the hamster pineal gland, unlike that of the rat, may not respond to VIP with an increased melatonin production.

Acetylserotonin O-Methyltransferase↗
PubMed1998

Rhythmic variation in beta1-adrenergic receptor mRNA levels in the rat pineal gland: circadian and developmental regulation.

In the rat pineal gland noradrenaline is released in large quantities from sympathetic nerve endings at the onset of darkness, thereby driving rhythmic melatonin synthesis with elevated levels at night-time. Upon release, noradrenaline interacts with postsynaptic beta1-adrenergic receptors to activate the cyclic AMP signalling pathway. Well characterized third messengers of this signalling cascade affect cyclic AMP-inducible genes that are crucially involved in initiation, maintenance and termination of hormone production. Among these third messengers are CREB (cyclic AMP responsive element binding protein) as an activating and ICER (inducible cyclic AMP early repressor) as an inhibitory transcription factor. Because a cyclic AMP-inducible promoter element is present on the beta1-adrenergic receptor gene, the expression of the receptor itself may be under control of the cyclic AMP-signalling pathway. By in situ hybridization, Northern blot analysis and RT-PCR we demonstrate a day/night rhythm in beta1-adrenergic receptor mRNA in the rat pineal gland with elevated levels during the dark period. As this rhythm persists, under constant darkness but is abolished upon removal of the sympathetic innervation, it is truly circadian. A marked day/night difference in the levels of beta1-adrenergic receptor mRNA becomes evident only after postnatal day 10, coinciding with the appearance of a functional cyclic AMP signalling pathway in the rat pineal gland. Furthermore, targeting ICER expression by transfection of pinealocytes with an antisense ICER construct, clearly indicates that the levels of the beta1-adrenergic receptor mRNA are regulated by the cyclic AMP-signalling pathway in a feedback mechanism.

Animals↗
PubMed1992

Immunohistochemical localization of nerve growth factor in the rat pineal gland.

Sympathetic nerve fibers arising from the superior cervical ganglia are the main innervation of the rat pineal gland. Since most organs innervated by these ganglia contain nerve growth factor (NGF), the hypothetical existence of NGF in the pineal gland was investigated. The peroxidase anti-peroxidase technique was applied for the immunohistochemical demonstration of NGF using a polyclonal antiserum on Bouin-fixed, paraffin-embedded pineal glands from adult, young and 6-hydroxydopamine (6-OHDA)-treated rats. Few immunopositive cells were observed in the adult pineal gland. A more conspicuous population of immunoreactive cells was noted in young animals (20-45 days old), especially in those chemically denervated with 6-OHDA. NGF immunoreactive cells displayed a stellate shape resembling the interstitial or glial cells previously described in the rat pineal gland. Since NGF plays a trophic effect on sympathetic neurons during development and adulthood, we postulate that its presence in the pineal gland may exert a trophic role on its sympathetic innervation.

Animals↗
PubMed1980

Studies on the presence of vasopressin, oxytocin and vasotocin in the pineal gland, subcommissural organ and fetal pituitary gland: failure to demonstrate vasotocin in mammals.

The demonstration of vasotocin in the mammalian pineal gland, subcommissural organ and fetal pituitary gland by bioassay has led to hypotheses regarding the function of this hormone in various reproductive processes. Preliminary examinations of the pineal gland and subcommissural organ with a specific radioimmunoassay failed to show vasotocin immunoreactivity. The presence of vasotocin, vasopressin and oxytocin in the pineal gland, subcommissural organ and fetal neurohypophysis was therefore investigated, using three specific radioimmunoassays. Frog and chicken pituitary glands were used to validate the vasotocin radioimmunoassay. Direct measurements in diluted homogenates of pituitary glands from frogs, chickens, mid-term fetal sheep and near-term fetal seals revealed the presence of vasotocin only in the frog and chicken pituitary glands, while vasopressin and oxytocin were found in the two fetal pituitary homogenates. Vasopressin and ocytocin were measured in homogenates of rat and bovine pineal glands and in preparations of the subcommissural organ of rats and rabbits after extraction with Vycor glass powder, but no specific vasotocin immunoreactivity was observed. These results indicate a discrepancy between the reported biological activity of vasotocin in the pineal gland, subcommissural organ and fetal pituitary gland and the immunoreactivity of this material, which can at present only be explained by the presence of a peptide which is structurally closely related to, but not identical with, vasotocin.

Animals↗
PubMed1992

Third ventricular pineal gland grafts in the gerbil: an electron microscopical and immunohistochemical investigation.

The superficial pineal gland was grafted into the third ventricle of adult Mongolian gerbils. Donor pineal glands from both neonatal and 3-4 week old gerbils were able to survive for at least 6 months. The pinealocytes of the grafted superficial pineal glands maintained the morphology and the S-antigen immunoreactivity of the in situ pineal complex. Synaptic ribbons and spherules were present but rare. Unlike the in situ pinealocytes, glycogen accumulations were common in the graft pinealocytes. Site specific modulation of structure was indicated as the ventricular surface of the grafts became covered with cerebrospinal fluid (CSF)-contacting pinealocytes typical of those seen in the deep pineal. The CSF-contacting pinealocytes of the graft had numerous processes that extended along the ventricular surface of the graft. The blood vessels of the grafts had non-fenestrated endothelium and wide perivascular areas typical of those seen in the in situ pineal complex. Tyrosine hydroxylase-immunopositive nerve fibers were present in the grafted tissue indicating reinnervation of the graft. The source of the fibers was not determined. The nerve fibers were present both within the perivascular area and within the parenchyma where they were associated with pinealocytes. The results demonstrate that the cerebral ventricles are an ideal location for the survival of superficial pineal gland grafts. It is suggested that pineal grafts may be a means to further study pineal development and innervation.

Animals↗
PubMed1994

Arachidonate 12-lipoxygenase of rat pineal glands: catalytic properties and primary structure deduced from its cDNA.

When a crude extract of rat pineal glands (the 1000 x g supernatant of a homogenate) was incubated with arachidonic acid, 12-hydroxy-5,8,10,14-eicosatetraenoic acid was found as a major product. The 12-lipoxygenase of rat pineal gland also reacted with linoleic and alpha-linolenic acids at 35% and 101% the rate of arachidonate 12-oxygenation, respectively. Upon Western blot analysis using polyclonal antibody against porcine leukocyte 12-lipoxygenase, the cytosol fraction of rat pineal gland showed a positive band with a molecular weight of approx. 74 kDa. A full-length cDNA for this enzyme was cloned from a cDNA library of rat pineal gland and the identity of the 12-lipoxygenase cDNA was confirmed by its expression in E. coli. The amino acid sequence of the enzyme was deduced from the nucleotide sequence of the cDNA, encoding 663 amino acids with a calculated molecular weight of 75,305. The enzyme showed 72% identity of amino acid sequence with porcine leukocyte 12-lipoxygenase and 73% with bovine tracheal 12-lipoxygenase, but only 59% with human platelet 12-lipoxygenase. Taken together, the high reactivity with C-18 fatty acids, the immunoreactivity and the amino acid homology data indicate that the rat pineal 12-lipoxygenase is more closely related to leukocyte 12-lipoxygenase than to platelet 12-lipoxygenase. Upon RNA blot analysis, by far the highest content of 12-lipoxygenase mRNA was observed in the pineal gland and negligible amounts of mRNA were detected in other parts of the brain. The predominant presence of 12-lipoxygenase mRNA in pineal gland was confirmed by in situ hybridization of rat brain. Significant amounts of 12-lipoxygenase mRNA were also detected in rat spleen, aorta, lung and leukocytes.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗
PubMed2006

Dopamine transporter immunoreactive terminals in the bovine pineal gland.

The dopaminergic system has been proposed to be one of the innervations in controlling the mammalian pineal gland function. The dopamine receptors have been characterized in the pineal and the biphasic effects of dopamine on melatonin production have been demonstrated. Recently, the site of dopamine transporter (DAT), a plasma membrane transport protein of dopaminergic neuron, also has been characterized in the bovine pineal gland. The aim of the present study was to identify the dopaminergic innervation in the bovine pineal gland. The localization and expression of DAT have been performed by using an immunohistochemical method and a reverse transcriptase polymerase chain reaction (RT-PCR) technique. DAT-immunoreactivity was found in the nerve terminals throughout the gland, but not in pinealocytes or neuronal-like cells. Some DAT-immunoreactive nerve fibers were observed along the pineal stalk. DAT mRNA product from RT-PCR was found in the bovine substantia nigra, but not in the pineal gland. The colocalization of DAT with tyrosine hydroxylase (TH) or dopamine beta hydroxylase (DBH) immunoreactivities was observed in nerve terminals. However, no colocalization of DAT with DBH was found in some terminals/fibers. The present results showed the central dopaminergic innervation in the bovine pineal gland distinctively from noradrenergic nerve fibers, and their perikarya origin was located possibly outside of the gland.

Animals↗
PubMed1991

Autoradiographic localization of dopaminergic and noradrenergic receptors in the bovine pineal gland.

Dopamine and norepinephrine are involved in regulation of melatonin synthesis in the pineal gland. In bovine pineal gland, D1- and D2-dopaminergic and alpha 1-adrenergic receptors have been characterized pharmacologically in several laboratories, while beta 1-adrenergic receptors have been studied using physiological technique. The current study presents a quantitative autoradiographic analysis of these four dopaminergic and noradrenergic receptors in bovine pineal gland. The density order of the receptors is D1 greater than alpha 1 greater than D2 greater than or equal to beta 1. The Bmax of dopamine D1 receptors is about 5 to 6 times higher than the Bmax for alpha 1-adrenergic receptors and about 20 times higher than the Bmax values for beta 1-adrenergic and D2-dopaminergic receptors. Dopamine D1 receptors are significantly denser in the pineal cortex than in the medulla. Both dopamine receptors are more concentrated in the distal area than in the proximal area (close to the habenula), whereas both noradrenergic receptors are homogeneously distributed along the longitudinal axis. Only D1-dopaminergic receptors display a heterogeneous distribution between the superior and the inferior areas, being denser in the inferior area. The observation of a much higher concentration of D1-dopaminergic receptors relative to the other receptors suggests an important role for dopamine in the regulation of bovine pineal physiology.

Animals↗
PubMed1994

Vascularization of the pineal gland in the crow.

The blood vascularization in the pineal gland of the crow was investigated in detail using a vascular corrosion cast technique and by scanning electron microscopy. The pineal gland received two afferent arteries on either side, each artery arising from the A. cerebralis caudalis (CC) which supplied its branches to the hemisphere. The pineal gland of the crow was so highly vascularized as to be suggestive of its high metabolic and endocrine activities. The efferent veins drained into the Sinus occipitalis and the Sinus sagittalis dorsalis. There was a direct vascular connection between capillaries of the stalk of the gland and those of the Plexus choroideus.

Animals↗
PubMed1990

Ultrastructural study of the human pineal gland in aged patients including a centenarian.

An ultrastructural study of human pineal glands obtained at autopsy from 7 patients older than 70 years was conducted in order to clarify the functional anatomy of the pineal in the aged. By light microscopy, the pineal glands from aged patients were parenchymatous and almost indistinguishable from those of younger controls. Electron microscopy of the pineal parenchymal cells revealed deep nuclear indentations, synaptic ribbons and ribbon fields, Golgi apparatus, lipofuscin granules and microtubular sheaves in all subjects, cilia with a 9 + 0 pattern in a few, and lamellated structures suggestive of the outer segment of photoreceptor cells very rarely. Microtubules were numerous in the cytoplasmic processes and bulbous endings. Fibrous astrocytes located between the pinealocytes showed long and thin cytoplasmic processes containing numerous glial filaments. Two types of nerve bouton were present in the pineal parenchyma, one of which contained clear vesicles forming synapse-like contacts with pinealocytes. There were no significant age-related changes in these features in a qualitative comparison with pineal glands from 5 adult patients younger than 70 years. These findings indicate that even in advanced age, the human pineal gland maintains some functions, such as intercellular communication and photoreception, in common with the pineal in lower vertebrates.

Adult↗
PubMed1989

Retinoids in the bovine pineal gland.

Using high performance liquid chromatography, the level of retinoids was determined in the bovine pineal gland, retina, retinal pigment epithelium, cortical and subcortical brain tissues, skeletal muscle and the liver. Except the retina, the bovine pineal gland possesses levels of retinol and retinyl esters significantly higher than other brain tissues and muscle. However, unlike the retina, the bovine pineal gland does not possess a significant level of retinal, the chromophore for visual pigments in the retina. This finding suggests that the bovine pineal gland does not possess rhodopsin nor the property of phototransduction which has been fully established in the retina and pineal glands of lower vertebrates.

Animals↗
PubMed1998

Development and control of the circadian pacemaker for melatonin release in the chicken pineal gland.

UNLABELLED: Melatonin (MT) release from explanted pineal glands of 3- to 20-week-old chicken was investigated in a 5-day perifusion system. Both the chicken and the explanted glands were exposed to various environmental lighting regimens. OBSERVATIONS: (1) The explanted chicken pineal is sensitive to direct light. Continuous illumination during the in vitro period abolishes the circadian rhythm of the MT secretion in 3 days. Continuous darkness has limited effect. (2) Reverse illumination completely reverses the MT cycle in 2 days. (3) Rhythmic illumination with short (6-h) periods only slightly modulates the MT release pattern: the basic, 24-h periodicity is preserved. (4) The circadian MT pacemaker develops normally and becomes synchronized to the day even if the chicken has never experienced alteration in the environmental illumination (those hatched and bred under continuous illumination). The explanted pineal from these chickens exhibits normal MT cycle and light sensitivity. Conclusion, Chicken pineal contains a complete, genetically coded circadian pacemaker with a fixed frequency. The pacemaker is synchronized to the day by the altered environmental illumination and by at least one other, unknown environmental factor. With altered illumination, in vitro, the 24-h periodicity of the pacemaker cannot be changed significantly, but its phase can be shifted. In contrast to conclusions obtained from in vivo observation in mammals, light seems to stimulate MT secretion from the avian pineal in vitro. For development and daily synchronization of the circadian MT pacemaker in the chicken pineal gland, periodic changes in the environmental illumination are not necessary.

Animals↗
PubMed1993

Induction of the c-fos proto-oncogene in the rat pineal gland during stress.

To investigate the effects of stressful stimuli on pineal gland activity, male Wistar albino rats (200-250 g, 2-4 per group) were submitted to 30 min of forced immobilization or to unilateral vibrissotomy 30 min before sacrifice. In situ hybridization was performed with a 35S-labelled 50-base oligonucleotide probe complementary to nucleotides 270-319 of rat c-fos on sections containing the pineal gland. Autoradiograms were quantified using a JAVA microdensitometer. Stressful stimuli induced a significant increase in the expression of c-fos mRNA in the pineal gland (restraint = 144.3 +/- 14.4 cpm/mm2; hemivibrissotomy = 206.7 +/- 29.5 cpm/mm2) as compared to no restraint animals (30.6 +/- 5.1 cpm/mm2), animals displaying tonic-clonic seizures after an ip (64 mg/kg) injection of pentylenetetrazole (34.0 +/- 4.7 cpm/mm2), or competition (70.6 +/- 11.4 cpm/mm2) and RNAase-treated (52.7 +/- 9.1 cpm/mm2) controls. These results raise the possibility that stressful stimuli may interfere with pineal gland function.

Animals↗
PubMed1989

In vitro synthesis of dopamine and noradrenaline in the isolated rat pineal gland: day-night variations and effects of electrical stimulation.

Isolated rat pineal glands were incubated in vitro in a medium containing [14C]dopamine or [14C]tyrosine, and the tissue contents of 14C-labelled and total dopamine and noradrenaline were determined by HPLC followed by electrochemical detection and scintillation spectrometry. During incubation with [14C]dopamine, the labelled amine accumulated in pineal glands and was partially converted into [14C]noradrenaline. Nomifensine, a neuronal amine uptake blocker, largely inhibited the accumulation of [14C]dopamine and the formation of [14C]noradrenaline. These experiments demonstrated dopamine beta-hydroxylase activity in the sympathetic nerves of the pineal gland. During incubation with [14C]tyrosine, formation of [14C]dopamine and [14C]noradrenaline was observed in the pineal tissue, indicating that noradrenaline can also be synthesized from dopamine, endogenously formed in the gland. Electrical stimulation of the stalk region of the pineal gland during incubation with [14C]dopamine enhanced the accumulation of [14C]dopamine and synthesis of [14C]noradrenaline. Electrical stimulation also enhanced the formation of [14C]dopamine during incubation with [14C]tyrosine. Compared to that at midday, the tissue content of endogenous noradrenaline at midnight was enhanced by 50% and that of dopamine by 450%. The in vitro accumulation of [14C]dopamine, as well as the synthesis of [14C]dopamine and [14C]noradrenaline, was also increased at midnight. In conclusion, sympathetic nerves in the rat pineal gland contain tyrosine hydroxylase and dopamine beta-hydroxylase, the two enzymes required for the synthesis of noradrenaline.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗