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A cyclic GMP-activated channel in dissociated cells of the chick pineal gland.

Phototransduction in the vertebrate retina is dependent in part on a cyclic GMP-activated ionic channel in the plasma membrane of rods and cones. But other vertebrate cells are also photosensitive. Cells of the chick pineal gland have a photosensitive circadian rhythm in melatonin secretion that persists in dissociated cell culture. Exposure to light causes inhibition of melatonin secretion, and entrainment of the intrinsic circadian oscillator. Chick pinealocytes express several 'retinal' proteins, including arrestin, transducin and a protein similar to the visual pigment rhodopsin. Pinealocytes of lower vertebrates display hyperpolarizing responses to brief pulses of light. Thus it is possible that some of the mechanisms of phototransduction are similar in retinal and pineal photoreceptors. We report here the first recordings of cyclic GMP-activated channels in an extraretinal photoreceptor. Application of GMP, but not cyclic AMP, to excised inside-out patches caused activation of a 15-25 pS cationic channel. These channels may be essential for phototransduction in the chick pineal gland.

Animals↗

The shape of synaptic ribbons in the rat pineal gland.

Under the transmission electron microscope, synaptic ribbons (SRs) of the mammalian pineal gland appear as rod-like organelles. Their three-dimensional structure is not precisely known. In the present study, pineal SRs were investigated using serial sections obtained from rats killed at noon and midnight. The shape of the SRs was reconstructed based on SR profile length and the number of sections in which the profiles were contained. The results obtained show that SRs are basically flat plate-like structures with polymorphic lateral edges. Reconstructions of SRs revealed that they had average dimensions of 300x150x35 nm and were 19.3% larger at night than at day; the difference in SR size points to perhaps major differences in synaptic function between day and night.

Animals↗

Molecular rhythms in the pineal gland.

Recent findings have clarified the mechanisms regulating the night- and pineal-specific transcription of serotonin N-acetyltransferase, the rate-limiting enzyme in melatonin formation. Norepinephrine, acting via beta-adrenoceptors and cAMP at night, stimulates the cAMP response element binding protein, which turns on the transcription of N-acetyltransferase and inducible cAMP early repressor, the major inhibitor of N-acetyltransferase transcription. The tissue-specific gene expression within the pineal gland and retina derives, in part, from a pineal/retina-specific transcription factor, cone-rod homeobox protein, which binds to a pineal regulatory element. This regulatory element is present in promoters of pineal-selective enzymes, such as N-acetyltransferase, hydroxyindole-O-methyl transferase, and pineal night-specific ATPase.

Adenosine Triphosphatases↗

Day-night variability in acid phosphatase activity of pineal gland macrophages.

Acid phosphatase (ACP) activity was investigated in the photoreceptive pineal gland of the goldfish by both light and electron microscopy. Intraluminal macrophages exhibited the greatest ACP reactivity that was generally confined to lysosomal bodies containing membranous debris. Quantitative light microscopic evaluation revealed significantly higher ACP reactivity in animals collected at mid-light compared to those collected at mid-dark suggesting that functional activities of macrophages in this photoneuroendocrine organ are cyclical over the 24-h photoperiod. Day-night differences in macrophage ACP activity may reflect rhythmic renewal and shedding of photoreceptor outer segment membranes.

Acid Phosphatase↗

Circadian expression of transcription factor Fra-2 in the rat pineal gland.

Physiological changes in Fos-like immunoreactivity in the rat pineal gland are shown here to be due primarily to changes in a 42/46-kDa Fos-related antigen (Fra). Studies are presented that indicate this 42/46-kDa Fra is Fra-2, a poorly understood member of the Fos family of transcription factors. Both Fra-2 mRNA and protein are absent during the day and increase robustly at night on a circadian basis; organ culture studies indicate that regulation is mediated by an adrenergic-->cyclic AMP mechanism. AP-1 binding activity changes in parallel to changes in the level of Fra-2 protein.

Amino Acid Sequence↗

Circannual analysis of pineal gland indoles and vasotocin-like immunoreactivity in male rats kept on constant photoschedules and temperature.

Levels of vasotocin-like immunoreactivity (iVT) and the indoles serotonin (5HT), 5-hydroxyindole acetic acid, N-acetyl-serotonin (NAS) and melatonin were measured over 24-hour periods at monthly intervals in the pineal gland of male rats kept under constant laboratory conditions of photoperiod and temperature. The 8- to 10-fold increase in pineal iVT, which occurred during mid-August, was not related to variations over the 24-hour light:dark (L:D) cycle. Tryptophan and 5HT exhibited seasonal variations over the 24 h L:D cycle with a single peak during the winter months. Twenty-four-hour levels of NAS tended to increase during the fall and spring. Seasonal effects on indole metabolism were further suggested by a marked decline during the fall in the correlation between NAS versus melatonin in individual pineal glands. Positive correlations between 5HT versus NAS and 5HT versus melatonin tended to be significant only during the summer months. These data suggest that environmental cues other than photo-period and temperature can regulate pineal neuroendocrine functions.

Animals↗

The ageing pineal gland and its physiological consequences.

Melatonin, the chief hormone of the pineal gland, is produced and secreted into the blood in a circadian manner with maximal production always occurring during the dark phase of the light:dark cycle. Whereas the 24h rhythm of melatonin production is very robust in young animals including humans, the cycle deteriorates during ageing. The rhythm of melatonin can be substantially preserved during ageing by restricting the food intake of experimental animals; this same treatment increases the life span of the animals. The exogenous administration of melatonin to non-food restricted animals also reportedly increases their survival. Moreover, melatonin has been shown to have immunoenhancing effects and oncostatic properties. The implication of these studies is that melatonin may have both direct and indirect beneficial effects in delaying ageing processes or it may retard the development of processes (e.g., immunodeficiency and tumor growth) which contribute to a reduced life span.

Aging↗

Effects of white light on the pineal gland of the chick embryo.

Chick embryos were directly exposed to a source of white light during incubation and sacrificed before hatching. The light caused a number of teratological effects such as high mortality, delay in development, celosomy, hepatomegaly, auricular dilation and micrognatia. The pineal gland of the illuminated embryos showed an increase in number and size of the intracytoplasmic lipid droplets of the follicular pinealocytes. These findings suggest that the pineal gland of the chick embryo is sensitive to light.

Abnormalities, Multiple↗

Ultrastructural localization of acetylcholinesterase in the guinea pig pineal gland.

The ultrastructural localization of acetylcholinesterase (AChE) activity in guinea pig pineal gland was studied using the copper-glycine procedure. A small number of pinealocytes and bundles of unmyelinated nerve fibers were labeled by the AChE reaction. The AChE-positive pinealocytes were located near blood vessels and distributed in small groups. The AChE reaction product was localized in the perinuclear cistern, in the cisternae of the endoplasmic reticulum (ER), and in the saccules of the Golgi apparatus. These findings suggest that the AChE-positive pinealocytes synthesize AChE. The AChE reaction product was also seen in the intercellular space between pinealocyte processes. Besides pinealocytes, AChE activity was localized on the axolemma of myelinated and unmyelinated nerve fibers and in the basement membrane surrounding unmyelinated nerve fibers. Pseudocholinesterase activity was confined to Schwann cells, which showed the reaction product in their perinuclear cistern, in the cisternae of the ER, and on the plasmalemma.

Acetylcholinesterase↗

The pinealocytes of the human pineal gland: A light and electron microscopic study.

The pinealocytes of the pineal gland of children and adults were studied at both light and electron microscopic levels. The pinealocytes were classified into light and dark pinealocytes on the basis of their shape, nuclear infolding, cytoplasmic contents and staining density. The light pinealocytes outnumber the dark pinealocytes and both of them have thin processes. The light pinealocytes have round or oval cell bodies and nuclei and have vesicles and ribbons. The dark pinealocytes showed more variations in their shape. The nuclear membrane of the dark cells showed numerous infoldings with deep invagination of parts of the cytoplasm within the nuclear folds, giving the appearance of nuclear pellets. The dark pinealocytes contain pigment in their cytoplasm. In addition to the light and dark pinealocytes a very small cell type with an extremely thin and elongated cell body and nucleus was found. The cells of this type were almost always associated with vacuoles filled with flocculent material and accumulations of presumptive secretion in the extracellular compartment. The findings of this study were discussed in the light of the published data about the pinealocytes of human and non-human species.

Adolescent↗

Passively propagated spikes from the soma of cells in pineal gland of guinea pigs.

The membrane properties of cells within the pineal gland of guinea pigs were studied using intracellular electrophysiological techniques. The electrotonic responses to intracellular current injection decayed with a single exponential in approximately 60% of cells but was preceded by a quicker component in the remainder. The membrane time constant was 2.8 ms. Depolarization beyond -29 mV activated an outward current that reversed at around the value of the resting potential. Hyperpolarization activated a slow inward current. Spikes occurred in response to activation of alpha-adrenoceptors. They were resistant to tetrodotoxin but were abolished by nifedipine and verapamil, suggesting that calcium carries the current during their upstroke. Spikes could not be evoked by depolarizing current pulses of 1-ms to 2-s duration. The responses to hyperpolarizing current steps or voltage-clamp steps applied during the peak of spikes evoked by nerve stimulation were indistinguishable from the responses to those applied between spikes. During nerve stimulation, fluctuations were observed in the current trace of cells under voltage clamp, indicating that the spikes could not be voltage clamped successfully. It is concluded that the spikes occurring in response to nerve stimulation are generated on the processes of the pinealocytes and are passive in the soma.

Animals↗

Role of eyes and pineal gland in melanophore response in the green frog. Rana esculenta.

The studies suggest that complete inhibition of MSH liberation from the pars intermedia is brought about in response to the receipt of photic cues through the eyes and, possibly, by the pineal gland. The punctuate condition of epidermal melanophores is brought about by the inhibition of MSH liberation alone, whereas the punctuate condition of dermal melanophores is brought about by the inhibition of MSH liberation and the liberation of melanin aggregating substance from the pineal gland in response to photostimulation.

Adaptation, Biological↗

Glial cells in the pineal gland of mice and rats. A combined immunofluorescence and electron-microscopic study.

Antigenic markers characteristic of astrocytes and their differentiative states (i.e., glial fibrillary acidic protein (GFAP), vimentin, and M1 and C1 antigens) were investigated in the pineal gland of mouse and rat using double immunolabeling techniques. In both species the so-called interstitial cells as characterized by TEM were shown to be astrocytes, since they expressed vimentin, but neither fibronectin (a marker for fibroblasts and endothelial cells) nor the neuron-specific L1 antigen or tetanus toxin receptors. Subpopulations of vimentin-positive pineal astrocytes were also GFAP- and C1- antigen-positive. M1- antigen-positive cells were not detected. It is concluded that a considerable proportion of interstitial cells in the pineal gland of rat and mouse are immature astrocytes which, in contrast to other parts of the central nervous system, persist into adulthood.

Animals↗

Effect of pentoxifylline on alpha- and beta-adrenoceptor sites in cerebral cortex medial basal hypothalamus and pineal gland of the rat.

The intraperitoneal injection of the methylxanthine derivative pentoxifylline (3,7-dimethyl-1-(5-oxo-hexyl)-xanthine] brought about, 3 hr later, a significant depression of alpha- and beta-adrenoceptor sites in the cerebral cortex, and of beta-adrenoceptor sites in medial basal hypothalamus and pineal gland, (assessed from the specific binding of radioactive dihydroergocryptine and dihydroalprenolol respectively). The changes in the density of binding sites were not accompanied by significant modifications of the Kd's. Sympathetic denervation of the pineal gland by superior cervical ganglionectomy (SCGx) abolished the changes of beta-adrenoceptor number in the pineal caused by pentoxifylline. The increase of alpha-adrenoceptor sites in the hypothalamus brought about by ganglionectomy was not affected by injection of pentoxifylline. Pentoxifylline did not compete in vitro for radioligand binding to brain membranes. These results suggest that methylxanthines depress brain adrenoceptor sites acutely, probably by down-regulation of receptors following the increase in catecholamine release caused by injection of the drug.

Animals↗

Immunohistochemical study of lymphocytes in rat pineal gland: selective accumulation of T lymphocytes.

Morphologic and immunohistochemical studies were performed to examine the existence of lymphocytes in the brain of rats. Special attention was paid to the time course of the appearance of lymphocytes in and around the pineal gland. Rabbit anti-rat T cell and anti-rat immunoglobulin sera were used for identification of T and B cells in tissue sections. Immunoperoxidase and immunofluorescence techniques were employed to identify cells reacting with anti-T and anti-immunoglobulin sera. No lymphocytes were found in the brain of rats until 20 days after birth. Small clusters of lymphocytes appeared in the pineal region by 30 days of age, after which they gradually increased in number, forming massive clusters in the pineal region by 120 days. Along with an increase in the number of lymphocytic cells, there was a gradual increase of cells reacting with anti-T cell serum. These T cells were only a minority of pineal lymphocytes in younger animals, but 90% or more cells were stained by anti-T cell serum at 120 days after birth. The remaining cells did not react with anti-immunoglobulin sera either. These findings suggest that the gradual increase of T lymphocytes in the rat pineal region is a simple reflection of the normal course of maturation of T cells, and the pineal gland in the rat may have some role in immune responses within the brain.

Aging↗

A possible role of the pineal gland in acute immobilization-related suppression of naloxone-induced LH release in ovariectomized estrogen-primed rats.

It has been recently reported that acute immobilization stress almost completely suppresses the luteinizing hormone (LH) release induced by naloxone, a mu-opioid antagonist, in ovariectomized estrogen-primed rats. The present study examined the possible involvement of the pineal gland in the acute immobilization-related suppression of the naloxone-induced LH release. An intraventricular (ICV) injection of 15 microg naloxone produced an abrupt increase in circulating LH concentrations in non-stressed rats. The naloxone-induced LH release was completely eliminated when tested 60 min after the end of a 30 min session of acute immobilization. The same stress conditions did not affect LH-releasing hormone (LHRH)-induced LH release, suggesting that the stress-related suppression of the naloxone-induced LH release was a suprapituitary event. In chronically-pinealectomized rats, but not in sham-pinealectomized rats, naloxone injected 60 min after the end of the stress session evoked a significant increase in serum LH concentrations. However, naloxone injected ICV during the acute immobilization did not elicit LH release in either pinealectomized or sham-operated rats. Under non-stressed conditions, the LH secretory response to naloxone was similar in pinealectomized and sham-operated animals. The same stress (30 min immobilization) significantly increased pineal melatonin content as well as plasma melatonin concentrations in rats bearing intact pineal glands, indicating that stress actually affected the pineal function. These results provide evidence for a role of the pineal in the suppression of the LH response to naloxone after stress, but not during stress.

Animals↗

Expression of the Otx2 homeobox gene in the developing mammalian brain: embryonic and adult expression in the pineal gland.

Otx2 is a vertebrate homeobox gene, which has been found to be essential for the development of rostral brain regions and appears to play a role in the development of retinal photoreceptor cells and pinealocytes. In this study, the temporal expression pattern of Otx2 was revealed in the rat brain, with special emphasis on the pineal gland throughout late embryonic and postnatal stages. Widespread high expression of Otx2 in the embryonic brain becomes progressively restricted in the adult to the pineal gland. Crx (cone-rod homeobox), a downstream target gene of Otx2, showed a pineal expression pattern similar to that of Otx2, although there was a distinct lag in time of onset. Otx2 protein was identified in pineal extracts and found to be localized in pinealocytes. Total pineal Otx2 mRNA did not show day-night variation, nor was it influenced by removal of the sympathetic input, indicating that the level of Otx2 mRNA appears to be independent of the photoneural input to the gland. Our results are consistent with the view that pineal expression of Otx2 is required for development and we hypothesize that it plays a role in the adult in controlling the expression of the cluster of genes associated with phototransduction and melatonin synthesis.

Age Factors↗

Otx2 homeobox gene controls retinal photoreceptor cell fate and pineal gland development.

Understanding the molecular mechanisms by which distinct cell fate is determined during organogenesis is a central issue in development and disease. Here, using conditional gene ablation in mice, we show that the transcription factor Otx2 is essential for retinal photoreceptor cell fate determination and development of the pineal gland. Otx2-deficiency converted differentiating photoreceptor cells to amacrine-like neurons and led to a total lack of pinealocytes in the pineal gland. We also found that Otx2 transactivates the cone-rod homeobox gene Crx, which is required for terminal differentiation and maintenance of photoreceptor cells. Furthermore, retroviral gene transfer of Otx2 steers retinal progenitor cells toward becoming photoreceptors. Thus, Otx2 is a key regulatory gene for the cell fate determination of retinal photoreceptor cells. Our results reveal the key molecular steps required for photoreceptor cell-fate determination and pinealocyte development.

Animals↗