scispace - formally typeset
Search or ask a question

Showing papers on "Pinealocyte published in 2004"


Journal ArticleDOI
TL;DR: Almost exclusively, TPH2 mRNA is expressed in raphe, in a pattern overlapping the histologically defined raphe nuclei, and is likely to be the gene giving rise to the majority of TPH activity in these cells.

211 citations


Journal ArticleDOI
TL;DR: In vivo the nightly secretion of melatonin desensitizes endogenous MT2 melatonin receptors in the mammalian SCN thereby providing a temporally integrated profile of sensitivity of the mammalian biological clock to a melatonin signal.
Abstract: The hormone melatonin phase shifts circadian rhythms generated by the mammalian biological clock, the suprachiasmatic nucleus (SCN) of the hypothalamus, through activation of G protein-coupled MT2 melatonin receptors. This study demonstrated that pretreatment with physiological concentrations of melatonin (30-300 pM or 7-70 pg/mL) decreased the number of hMT2 melatonin receptors heterologously expressed in mammalian cells in a time and concentration-dependent manner. Furthermore, hMT2-GFP melatonin receptors heterologously expressed in immortalized SCN2.2 cells or in non-neuronal mammalian cells were internalized upon pretreatment with both physiological (300 pM or 70 pg/mL) and supraphysiological (10 nM or 2.3 ng/mL) concentrations of melatonin. The decrease in MT2 melatonin receptor number induced by melatonin (300 pM for 1 h) was reversible and reached almost full recovery after 8 h; however, after treatment with 10 nM melatonin full recovery was not attained even after 24 h. This recovery process was partially protein synthesis dependent. Furthermore, exposure to physiological concentrations of melatonin (300 pM) for a time mimicking the nocturnal surge (8 h) desensitized functional responses mediated through melatonin activation of endogenous MT2 receptors, i.e., stimulation of protein kinase C (PKC) in immortalized SCN2.2 cells and phase shifts of circadian rhythms of neuronal firing in the rat SCN brain slice. We conclude that in vivo the nightly secretion of melatonin desensitizes endogenous MT2 melatonin receptors in the mammalian SCN thereby providing a temporally integrated profile of sensitivity of the mammalian biological clock to a melatonin signal.

132 citations


Journal ArticleDOI
TL;DR: The circadian control of the cAMP signaling cascade indicates that the clock has a more general and profound impact on retinal functions than previously thought and may play a central role in the integration of circadian signals that control physiology and behavior.
Abstract: Melatonin is synthesized in retinal photoreceptor cells and acts as a neuromodulator imparting photoperiodic information to the retina The synthesis of melatonin is controlled by an ocular circadian clock and by light in a finely tuned mechanism that ensures that melatonin is synthesized and acts only at night in darkness Here we report that the circadian clock gates melatonin synthesis in part by regulating the expression of the type 1 adenylyl cyclase (AC1) and the synthesis of cAMP in photoreceptor cells This gating is effected through E-box-mediated transcriptional activation of the AC1 gene, which undergoes robust daily fluctuations that persist in constant illumination The circadian control of the cAMP signaling cascade indicates that the clock has a more general and profound impact on retinal functions than previously thought In addition, rhythmic control of AC1 expression was observed in other parts of the central circadian axis, the suprachiasmatic nucleus and pineal gland, but not in other brain areas examined Thus, clock control of the cAMP signaling cascade may play a central role in the integration of circadian signals that control physiology and behavior

112 citations


Journal ArticleDOI
TL;DR: The hypothesis argues that early in chordate evolution, at a point before the genes required for melatonin synthesis were acquired, retinaldehyde—which is essential for photon capture—was depleted by reacting with naturally occurring arylalkylamines and xenobiotic aries, resulting in the formation of adducts analogous to formation in the retina of the toxic bis-retinyl ethanolamine (A2E).
Abstract: A theory is presented that explains the evolution of the pinealocyte from the common ancestral photoreceptor of both the pinealocyte and retinal photoreceptor. Central to the hypothesis is the previously unrecognized conflict between the two chemistries that define these cells-melatonin synthesis and retinoid recycling. At the core of the conflict is the formation of adducts composed of two molecules of retinaldehyde and one molecule of serotonin, analogous to formation in the retina of the toxic bis-retinyl ethanolamine (A2E). The hypothesis argues that early in chordate evolution, at a point before the genes required for melatonin synthesis were acquired, retinaldehyde--which is essential for photon capture--was depleted by reacting with naturally occurring arylalkylamines (tyramine, serotonin, tryptamine, phenylethylamine) and xenobiotic arylalkylamines. This generated toxic bis-retinyl arylalkylamines (A2AAs). The acquisition of arylalkylamine N-acetyltransferase (AANAT) prevented this by N-acetylating the arylalkylamines. Hydroxyindole-O-methyltransferase enhanced detoxification in the primitive photoreceptor by increasing the lipid solubility of serotonin and bis-retinyl serotonin. After the serotonin --> melatonin pathway was established, the next step leading toward the pinealocyte was the evolution of a daily rhythm in melatonin and the capacity to recognize it as a signal of darkness. The shift in melatonin from metabolic garbage to information developed a pressure to improve the reliability of the melatonin signal, which in turn led to higher levels of serotonin in the photodetector. This generated the conflict between serotonin and retinaldehyde, which was resolved by the cellular segregation of the two chemistries. The result, in primates, is a pineal gland that does not detect light and a retinal photodetector that does not make melatonin. High levels of AANAT in the latter tissue might serve the same function AANAT had when first acquired- prevention of A2AA formation.

106 citations


Journal ArticleDOI
TL;DR: It is demonstrated that neuronal activity of both PVN and SCN is necessary to stimulate melatonin synthesis during the dark period and glutamatergic signalling within the PVN plays an important role inmelatonin synthesis.
Abstract: The rhythm of melatonin synthesis in the rat pineal gland is under the control of the biological clock, which is located in the suprachiasmatic nucleus of the hypothalamus (SCN). Previous studies demonstrated a daytime inhibitory influence of the SCN on melatonin synthesis, by using gamma-aminobutyric acid input to the paraventricular nucleus of the hypothalamus (PVN). Nevertheless, a recent lesion study suggested the presence of a stimulatory clock output in the control of the melatonin rhythm as well. In order to further investigate this output in acute in vivo conditions, we first measured the release of melatonin in the pineal gland before, during and after a temporary shutdown of either SCN or PVN neuronal activity, using multiple microdialysis. For both targets, SCN and PVN, the application of tetrodotoxin by reverse dialysis in the middle of the night decreased melatonin levels. Due to recent evidence of the existence of glutamatergic clock output, we then studied the effect on melatonin release of glutamate antagonist application within the PVN in the middle of the night. Blockade of the glutamatergic input to the PVN significantly decreased melatonin release. These results demonstrate that (i) neuronal activity of both PVN and SCN is necessary to stimulate melatonin synthesis during the dark period and (ii) glutamatergic signalling within the PVN plays an important role in melatonin synthesis.

88 citations


Journal ArticleDOI
TL;DR: The presence of ET-1 mRNA, ECE-1 and ET(1) in 22 brain regions confirms ET expression and processing in human brain and suggests receptor-mediated action akin to a neurotransmitter role forET-1.

60 citations


Journal ArticleDOI
TL;DR: The presence of two putative photoisomerases within the pineal gland and in retinal layers associated with biological clock function provides two candidate opsinlike genes that may serve in the visual cycle regulation of the circadian clock.
Abstract: PURPOSE. The chick retina and pineal gland exhibit circadian oscillations in biochemical and physiological processes in vivo and in vitro, which entrain to 24-hour light- dark cycles. However, the phototransduction mechanisms responsible for entrainment are largely unknown. The present study characterizes two candidate opsinlike genes that may play a role in entrainment of the clocks in these tissues. METHODS. Bioinformatics, cladistic techniques, and in situ hybridization and Northern blot analyses were conducted to characterize, localize, and determine the circadian expression of the candidate opsinlike genes in the retina and pineal gland. RESULTS. Two candidate photosensors and/or photoisomerases were predominantly distributed within the pineal gland and retina: the retinal pigmented epithelium-derived rhodopsin homologue (peropsin, gRrh) and retinal G-protein- coupled receptor opsin (RGR opsin, gRgr). Northern blot and in situ analyses revealed mRNA expression for both opsins in the pineal gland, retina, and brain tissue. The mRNA for both genes within the pineal gland and retina is regulated on a circadian basis such that they are highest in late subjective day. Digoxigenin in situ analyses showed retinal gRgr message within the inner nuclear layer (INL) and retinal ganglion cell layer (RGL), whereas gRrh message was distributed predominantly in the RGL. In the pineal gland, gRgr message was sparsely distributed among pinealocytes in follicles, but not within the follicles themselves, whereas gRrh was localized in interstitial areas indicative of astrocytic and/or endothelial origin. CONCLUSIONS. The presence of two putative photoisomerases within the pineal gland and in retinal layers associated with biological clock function provides two candidate opsinlike genes that may serve in the visual cycle regulation of the circadian clock.

52 citations


Journal ArticleDOI
TL;DR: A general reduction of circadian control of transcript abundance in the ovine pineal gland is suggested, consistent with a marked evolutionary divergence in the mechanism regulating melatonin production between terrestrial ruminants and fossorial rodents.

45 citations


Journal ArticleDOI
TL;DR: Results suggest that melatonin may influence retinal function by binding to receptors on RPE and photoreceptor cells, and by acting on neurons of the inner retina that do not use dopamine or GABA as a neurotransmitter.

45 citations


Journal ArticleDOI
TL;DR: In this article, the authors investigated the day/night expression profiles of all major clock genes and their products in the pineal gland of one melatonin-proficient and one melanatonin-deficient mouse strain and found that PER1 may have a trigger function for initiating the cycles of the clockwork's transcriptional/translational feedback loops.
Abstract: In vertebrates, the rhythmic transcription of clock genes, regulated by their own gene products, provides the basis for self-sustaining circadian clockworks. These endogenous clocks are lost in most mammalian tissues, but not in the central pacemaker of the hypothalamic suprachiasmatic nucleus (SCN). An interesting model system to understand this phylogenetic shift in function of clock gene products is the rodent pineal gland, as its intrinsic clockwork was replaced during evolution by an input-dependent oscillator. By means of immunohistochemistry, immunoblotting and real time PCR, we investigated the day/night expression profiles of all major clock genes and their products in the pineal gland of one melatonin-proficient and one melatonin-deficient mouse strain. All clockwork elements known to be indispensable for a sustained rhythm generation in the SCN were also found in the pineal organ of both mouse strains. Only mPer1 mRNA and PER1 protein accumulation coincides with timecourses of many other pineal genes and their products, which are cyclicAMP inducible. Here, presented data together with the known mechanisms for regulation of the mPer1 gene in the rodent pineal gland forward the idea that in this tissue PER1 may have a trigger function for initiating the cycles of the clockwork's transcriptional/translational feedback loops.

45 citations


Journal ArticleDOI
TL;DR: Observations in the present study extend the number of pineal genes that are known to be regulated by Fra‐2, and provide the first indication that a member of the NGFI‐B group of nuclear receptors is involved in controlling gene expression in the pineal gland.
Abstract: NGFI-B (Nur77/Nr4a1) is a member of a nuclear steroid receptor subgroup that includes the related factors Nurr1 (Nr4a2) and NOR-1 (Nr4a3). These proteins do not have recognized ligands and in fact function independently as orphan receptors with transcriptional regulatory activity. In the present study, expression of the NGFI-B gene in the rat pineal gland was found to exhibit a robust circadian rhythm, with elevated levels of NGFI-B mRNA occurring at night. The rhythm of NGFI-B mRNA is translated into a circadian rhythm of NGFI-B protein, which accumulates in the nucleus of pinealocytes. In addition, there is a parallel marked nocturnal increase in pineal DNA binding activity to a NGFI-B response element (NBRE, AAAGGTCA). Pharmacological studies indicate that NGFI-B mRNA and protein levels are elevated via activation of adrenergic receptors. NGFI-B protein levels are also elevated by dibutyryl cyclic AMP, as in other systems. In the pineal gland, regulation of NGFI-B expression also involves the AP-1 protein Fra-2, based on studies with a transgenic Fra-2 knockdown rat, in which pineal NGFI-B expression increases. This set of observations extends the number of pineal genes that are known to be regulated by Fra-2, and also provides the first indication that a member of the NGFI-B group of nuclear receptors is involved in controlling gene expression in the pineal gland.

Journal ArticleDOI
TL;DR: The results indicate that virtually all of the nAChRs in the rat pineal gland are the α3β4 nA ChR subtype and that the Pineal gland can therefore serve as an excellent and convenient model in which to study the pharmacology and function of these receptors in a native tissue.
Abstract: The rat pineal gland contains a high density of neuronal nicotinic acetylcholine receptors (nAChRs). We characterized the pharmacology of the binding sites and function of these receptors, measured the nAChR subunit mRNA, and used subunit-specific antibodies to establish the receptor subtype as defined by subunit composition. In ligand binding studies, [3H]epibatidine ([3H]EB) binds with an affinity of approximately 100 pM to nAChRs in the pineal gland, and the density of these sites is approximately 5 times that in rat cerebral cortex. The affinities of nicotinic drugs for binding sites in the pineal gland are similar to those at alpha3beta4 nAChRs heterologously expressed in human embryonic kidney 293 cells. In functional studies, the potencies and efficacies of nicotinic drugs to activate or block whole-cell currents in dissociated pinealocytes match closely their potencies and efficacies to activate or block 86Rb+ efflux in the cells expressing heterologous alpha3beta4 nAChRs. Measurements of mRNA indicated the presence of transcripts for alpha3, beta2, and beta4 nAChR subunits but not those for alpha2, alpha4, alpha5, alpha6, alpha7, or beta3 subunits. Immunoprecipitation with subunit-specific antibodies showed that virtually all [3H]EB-labeled nAChRs contained alpha3 and beta4 subunits associated in one complex. The beta2 subunit was not associated with this complex. Taken together, these results indicate that virtually all of the nAChRs in the rat pineal gland are the alpha3beta4 nAChR subtype and that the pineal gland can therefore serve as an excellent and convenient model in which to study the pharmacology and function of these receptors in a native tissue.

Journal ArticleDOI
TL;DR: In NE-stimulated pinealocytes, blockade of β-adrenergic receptors caused a rapid reduction in MKP-1 mRNA, but it had a minimal effect on MKp-1 protein and appeared to function primarily in prolonging the β- adrenergic- Stimulation responses.
Abstract: In this study, we investigated the mechanisms through which norepinephrine (NE) regulates MAPK phosphatase-1 (MKP-1) expression in rat pinealocytes. Stimulation with NE (a mixed α- and β-adrenergic agonist) caused a rapid increase in MKP-1 mRNA and protein that peaked around 1 h post stimulation, and the response was sustained for at least 4 h. Selective activation of β-adrenergic receptors with isoproterenol for 1 h caused a similar increase in MKP-1 mRNA and protein as observed with NE, but at 3 h, the isoproterenol response was much lower relative to NE. In contrast, selective activation of α-adrenergic receptors caused only small increases in MKP-1 mRNA and protein and appeared to function primarily in prolonging the β-adrenergic-stimulated responses. In NE-stimulated pinealocytes, blockade of β-adrenergic receptors caused a rapid reduction in MKP-1 mRNA, but it had a minimal effect on MKP-1 protein. In contrast, blockade of α-adrenergic receptors specifically reduced NE-induced MKP-1 protein but not ...

Journal ArticleDOI
TL;DR: A hypothalamic regulation of the pineal gland via central orexinergic nervous inputs is suggested, supported by previous studies on rodents.
Abstract: Orexins/hypocretins, two isoforms of the same prepropeptide, are widely distributed throughout the brain and are involved in several physiological and neuroendocrine regulatory patterns, mostly related to feeding, sleep, arousal, and cyclic sleep-wake behaviors. Orexin-A and orexin-B bind with different affinities to two G-protein-coupled transmembrane receptors, orexin-1 and orexin-2 receptors (OR-R1 and OR-R2, respectively). Because of the similarities between the human and the swine brain, we have studied the pig to investigate the orexinergic system in the diencephalon, with special emphasis on the neuroanatomical projections to the epithalamic region. By using antibodies against orexin-A and orexin-B, immunoreactive large multipolar perikarya were detected in the hypothalamic periventricular and perifornical areas at the light and electron microscopic levels. In the region of the paraventricular nucleus, the orexinergic neurons extended all the way to the lateral hypothalamic area. Immunoreactive nerve fibers, often endowed with large varicosities, were found throughout the hypothalamus and the epithalamus. Some periventricular immunoreactive nerve fibers entered the epithalamic region and continued into the pineal stalk and parenchyma to disperse among the pinealocytes. Immunoelectron microscopy confirmed the presence of orexinergic nerve fibers in the pig pineal gland. After extraction of total mRNA from the hypothalamus and pineal gland, we performed RT-PCR and nested PCR using primers specific for porcine orexin receptors. PCR products were sequenced, verifying the presence of both OR-R1 and OR-R2 in the tissues investigated. These findings, supported by previous studies on rodents, suggest a hypothalamic regulation of the pineal gland via central orexinergic nervous inputs.

Journal ArticleDOI
TL;DR: This study reveals that the stimulatory effect of opioid receptor on melatonin synthesis is mediated via the activation of adenylate cyclase system.
Abstract: Mammalian pinealocytes synthesize and secrete melatonin. The synthesis of melatonin is regulated by several biogenic amine, amino acid and peptide transmitters. In our previous study, the delta- and mu-opioid receptors have been identified and characterized in bovine pinealocytes. In order to elaborate the function of different types of opioid receptors in regulating melatonin synthesis, we used a selective mu-opioid receptor agonist, Tyr-[D-Ala(2), N-methyl-phe(4), glycol(5)] (DAMGO), a selective delta-opioid receptor agonist, Enkephalin [D-Pen(2), D-Pen(5)], (DPDPE) and a selective kappa-opioid receptor agonist, ((+)-(5alpha, 7alpha, 8beta)-N-methyl-N-[7- (1-pyrrolidinyl)-1-oxaspiro [4,5] dec-8-yl]-benzene acetamide) (U69593) to investigate the activity of N-acetyltransferase (NAT) activity and melatonin secretion. The results of the present study show that both DAMGO and DPDPE stimulated NAT activity and increased the level of melatonin in cultured bovine pinealocytes. These stimulatory effects were blocked by naloxone, an opioid receptor antagonist. However, the kappa-opioid receptor agonist U69593 was unable to alter either the activity of NAT or the level of melatonin. In order to clarify the mechanism of how the activation of mu- and delta-opioid receptors in bovine pinealocytes leads to an increase in NAT activity, cyclic AMP levels were measured after bovine pinealocytes were treated with morphine, DAMGO and DPDPE. The results indicated that these stimulatory effects acted via induction of cAMP production. This study reveals that the stimulatory effect of opioid receptor on melatonin synthesis is mediated via the activation of adenylate cyclase system.

Journal ArticleDOI
TL;DR: Results indicate that activation of p38MAPK is under photoneural control in the rat pineal gland and that protein kinase A and intracellular Ca(2+) signaling pathways are involved in NE regulation of p 38MAPK.
Abstract: In this study, we investigated adrenergic and photoneural regulation of p38MAPK phosphorylation in the rat pineal gland. Norepinephrine (NE), the endogenous neurotransmitter, dose-dependently increased the levels of phosphorylated MAPK kinase 3/6 (MKK3/6) and p38MAPK in rat pinealocytes. Time-course studies showed a gradual increase in MKK3/6 and p38MAPK phosphorylation that peaked between 1 an d2ha nd persisted for 4 h post NE stimulation. In cells treated with NE for 2 and 4 h, the inclusion of prazosin or propranolol reduced NE-induced MKK3/6 and p38MAPK phosphorylation, indicating involvement of both - and -adrenergic receptors for the sustained response. Whereas treatment with dibutyryl cAMP or ionomycin mimicked the NE-induced MKK3/6 and p38MAPK phosphorylation, neither dibutyryl cGMP nor 4phorbol 12-myristate 13-acetate had an effect. The NEinduced increase in MKK3/6 and p38MAPK phosphorylation was blocked by KT5720 (a protein kinase A inhibitor) and KN93 (a Ca 2 /calmodulin-dependent kinase inhibitor), but not by KT5823 (a protein kinase G inhibitor) or calphostin C (a protein kinase C inhibitor). In animals housed under a lighting regimen with 12 h of light, MKK3/6 and p38MAPK phosphorylation increased in the rat pineal gland at zeitgeber time 18. The nocturnal increase in p38MAPK phosphorylation was blocked by exposing the animal to constant light and reduced by treatment with propranolol, a -adrenergic blocker. Together, our results indicate that activation of p38MAPK is under photoneural control in the rat pineal gland and that protein kinase A and intracellular Ca 2 signaling pathways are involved in NE regulation of p38MAPK. (Endocrinology 145: 5194 –5201, 2004)

Journal ArticleDOI
TL;DR: It is revealed, that PACAP is present in nerve fibers in the chicken pineals and pineal cells contain functioning PACAP-sensitive receptors and cAMP-mediated intracellular mechanisms in ChPG are not components of the pineal circadian clock, but intermediaries between the clock-mechanism and MT release and may also be components of clock-independent MT release modifiers.

Journal ArticleDOI
J. R. Man1, S. Rustaeus1, D. M. Price1, Constance L. Chik1, Anthony K. Ho1 
TL;DR: Investigation of whether p38(MAPK) played a role in the adrenergic regulation of arylalkylamine-N-acetyltransferase (AA-NAT) induction and melatonin (MT) synthesis shows that inhibition of p38 (MAPK), although having no effect on cAMP-mediated AA- NAT transcription, appears to increase AA-NAT activity either by increasing translation or by reducing degradation of the AA- NATO protein.
Abstract: We have previously shown that inhibition of p38MAPK increases adrenergic-stimulated p42/44MAPK activation in rat pinealocytes. In this study we investigated whether p38MAPK played a role in the adrenergic regulation of arylalkylamine-N-acetyltransferase (AA-NAT) induction and melatonin (MT) synthesis. Treatment of pinealocytes with norepinephrine (NE) caused a time-dependent increase in the levels of AA-NAT mRNA, AA-NAT protein, and enzymatic activity as well as MT production. Cotreatment with SB202190, a selective p38MAPK inhibitor, although having no effect on AA-NAT activity or protein level 3 h after NE treatment, caused a sustained increase in AA-NAT activity and protein level after 6 h of NE treatment. The increases in NE-stimulated AA-NAT activity and protein level by SB202190 occurred in the absence of an increase in AA-NAT mRNA. Similar results were obtained when AA-NAT was induced by (Bu)2cAMP or when SB203580 was used to inhibit p38MAPK. In comparison, SB202474, the inactive analog, had no effe...

Journal Article
TL;DR: It was observed that the pinealocyte cell activity of rats exposure to constant darkness was increased but decreased in rats exposed to constant light, while the number of pinealocytes observed was extensive and some of the observed Pinealocytes were determined to contain double nucleoli.
Abstract: OBJECTIVE: This study was aimed to examine the pineal gland of rats exposed to constant light and darkness at light and electron microscopic level. DESIGN: For this purpose 18 male Wistar rats were used. Animals were divided into three groups. Rats in group I (Control) were kept under 12 hrs light: 12 hrs dark conditions. Rats in group II were exposed to constant darkness, while rats in group III were exposed to constant light for 6 weeks. At the end of the experiment, all animals were killed by decapitation. The pineal glands of rats were removed, then processed for light and electron microscopy. RESULTS: In our study, extensive number of pinealocytes was observed in the structure of pineal gland of rats exposed to constant darkness and some of the observed pinealocytes were determined to contain double nucleoli. Furthermore, mitochondria and lipid droplets in the cytoplasm of pinealocytes were increased and rough endoplasmic reticulum sacs were enlarged in this group. Whereas, in rats those exposed to the constant light, a decrease in pinealocyte intensity was associated with increase in the connective tissue between parenchymal cells. Additionally, mitochondria and lipid droplets in the cytoplasm of cells were decreased. CONCLUSIONS: It was observed that the pinealocyte cell activity of rats exposed to constant darkness was increased but decreased in rats exposed to constant light.

Journal ArticleDOI
TL;DR: The results indicate that, in the female rat, estrogen directly modulates pineal sensitivity to adrenergic stimulation in a complex, dose‐dependent manner that may be related to differential expression and activity of two estrogen receptor subtypes within pineal cells.
Abstract: The effect of different doses of 17beta-estradiol (E2) on the pineal response to beta-adrenoceptor stimulation in female rats was examined. Pinealocytes from 21-day-old ovariectomized rats were exposed to different estrogen doses and treated with beta-adrenergic agonists. Estrogen treatment produced a dose-dependent, biphasic response to beta-adrenoceptor-induced accumulation of cAMP. This effect was inhibitory at estrogen doses up to 0.1 nM and fitted to a negative exponential curve, while at doses from 0.1 to 100 nM the effect was stimulatory and fitted to a standard positive hyperbola. For in vivo studies, ovariectomized rats were treated with equivalent estrogen concentrations plus a single dose of progesterone (250 microg per rat), and their pineals exposed in vitro to beta-adrenergic agonists. Low doses of E2 (0.1-100 ng per rat) reduced both pineal cAMP accumulation and N-acetyltransferase activity after beta-adrenoceptor stimulation, while a high dose (10 microg per rat) induced the opposite response. Apparently, the final estrogen target was the pineal beta-adrenergic receptor, as a low dose of E2 (which had diminished cAMP accumulation after beta-adrenoceptor stimulation) also reduced its maximal binding capacity (Bmax) and its dissociation constant (Kd). We also found that the female rat pineal gland contains two different ER subtypes, alpha and beta, which respond to estrogen exposure with nucleocytoplasmic shuttling. These results indicate that, in the female rat, estrogen directly modulates pineal sensitivity to adrenergic stimulation in a complex, dose-dependent manner that may be related to differential expression and activity of two estrogen receptor subtypes within pineal cells.

Book
01 Jan 2004
TL;DR: It is reported that the phase of the circadian melatonin rhythm of the explanted chicken pineal gland can be entrained easily to photoperiods whose length approximates 24 h, even if the light period is extremely short, i.e., 3L:21D.
Abstract: The pineal gland plays a key role in the control of the daily and seasonal rhythms in most vertebrate species. In mammals, rhythmic melatonin (MT) release from the pineal gland is controlled by the suprachiasmatic nucleus via the sympathetic nervous system. In most non-mammalian species, including birds, the pineal gland contains a self-sustained circadian oscillator and several input channels to synchronize the clock, including direct light sensitivity. Avian pineal glands maintain rhythmic activity for days under in vitro conditions. Several physical (light, temperature, and magnetic field) and biochemical (Vasoactive intestinal polypeptide (VIP), norepinephrine, PACAP, etc.) input channels, influencing release of melatonin are also functional in vitro, rendering the explanted avian pineal an excellent model to study the circadian biological clock. Using a perifusion system, we here report that the phase of the circadian melatonin rhythm of the explanted chicken pineal gland can be entrained easily to photoperiods whose length approximates 24 h, even if the light period is extremely short, i.e., 3L:21D. When the length of the photoperiod significantly differs from 24 h, the endogenous MT rhythm becomes distorted and does not follow the light-dark cycle. When explanted chicken pineal fragments were exposed to various drugs targeting specific components of intracellular signal transduction cascades, only those affecting the cAMP-protein kinase-A system modified the MT release temporarily without phase-shifting the rhythm in MT release. The potential role of cGMP remains to be investigated.

Journal ArticleDOI
TL;DR: Results suggest that GABA is stored and secreted from astrocytes and a subpopulation of microglia in pineal glands, and Vesicular inhibitory amino acid transporter (VIAAT) is a potential marker for the GABAergic phenotype.

Journal ArticleDOI
TL;DR: The hypothesis of the presence of photosensitive cells in the encephalon of lower vertebrates is supported and the labelling with antisera against different components of the phototransductory cascade strengthens the idea that such cells employ a biochemical mechanism similar to that in the retinal visual photoreceptor cells, rods and cones.
Abstract: Summary. The existence of cells capable of detecting changes of the photoperiod within the deep brain, the socalled deep brain photoreceptors, was proposed in the early years of the twentieth century. By using immunocytochemistry with antisera against phototransductory proteins on paraffin and vibratome sections, we have localized several positive areas in the brain of the teleost Phoxinus phoxinus. These areas were restricted to two encephalic regions: the epithalamus and the hypothalamus. Immunopositive (rod-opsin- and α-transducin-like) pinealocytes and parapinealocytes, as well as some sparse neurons in the habenula, were seen in the epithalamus. The immunoreaction of the hypothalamus was represented by α-transducin-like positive (magnocellular and parvicellular) neurons of the Nucleus Preopticus, as well as by α-transducin- and arrestin-like positive fibers corresponding to the hypothalamic-hypophyseal tract and a few fibers running towards the basal telencephalon. These findings corroborate the data published on other teleost fish and fully support the hypothesis of the presence of photosensitive cells in the encephalon of lower vertebrates. The labelling with antisera against different components of the phototransductory cascade also strengthens the idea that such cells employ a biochemical mechanism similar to that in the retinal visual photoreceptor cells, rods and cones. Although the function is still unclear, the detection of the photoperiod seems to be the most likely role for these extraretinal photoreceptors.

Journal ArticleDOI
TL;DR: The hypothesis that the mechanism through which light information is conveyed to the pacemaker in the SCN is separate from and independent of the mechanism of light information from retinal efferents to the pineal gland is supported.
Abstract: The suprachiasmatic nucleus (SCN) of the anterior hypothalamus contains a light-entrainable circadian pacemaker. Neurons in the SCN are part of a circuit that conveys light information from retinal efferents to the pineal gland. Light presented during the night acutely increases mRNA levels of the circadian clock genes Per1 and Per2 in the SCN, and acutely suppresses melatonin levels in the pineal gland. The present study investigated whether the ability of light to increase Per1 and Per2 mRNA levels and suppress pineal melatonin levels requires sodium-dependent action potentials in the SCN. Per1 and Per2 mRNA levels in the SCN and pineal melatonin levels were measured in Syrian hamsters injected with tetrodotoxin (TTX) prior to light exposure or injection of N-methyl-d-aspartate (NMDA). TTX inhibited the ability of light to increase Per1 and Per2 mRNA levels and suppress pineal melatonin levels. TTX did not, however, influence the ability of NMDA to increase Per1 and Per2 mRNA levels, though it did inhibit the ability of NMDA to suppress pineal melatonin levels. These results demonstrate that action potentials in the SCN are not necessary for NMDA receptor activation to increase Per1 and Per2 mRNA levels, but are necessary for NMDA receptor activation to decrease pineal melatonin levels. Taken together, these data support the hypothesis that the mechanism through which light information is conveyed to the pacemaker in the SCN is separate from and independent of the mechanism through which light information is conveyed to the SCN cells whose efferents suppress pineal melatonin levels.

Journal ArticleDOI
TL;DR: Examination of the effect of dark exposure on two "circadian clock" genes Period1 and Period2 mRNA levels in the rat SCN, and on Period1, Period2, and arylalkylamine N-acetyltransferase (Aa-Nat, the rate-limiting enzyme in melatonin synthesis) gene expression in the pineal gland demonstrates that dark information may reach the Pineal during the daytime.

Journal Article
TL;DR: The obtained results demonstrate that both pinealocyte ultrastructure and pineal biochemistry in the pig undergo significant changes in the course of the diurnal rhythm.
Abstract: The study was conducted to investigate diurnal changes in pinealocyte ultrastructure, pineal serotonin content and plasma melatonin concentration in the domestic pig. The immature pigs (n=24) were kept under a cycle of 12 h light : 12 h dark, with a photophase between 0800 and 2000. During the photophase the animals were exposed to direct sunlight. After four weeks the gilts were slaughtered at 0900, 1400, 2100 and 0200. The pineals were removed and divided into two parts - one for quantitative ultrastructural study (by a point count method) and one for serotonin assay. Simultaneously, blood samples were taken for melatonin assay. The relative volume of mitochondria in pinealocyte perikarya was significantly higher at 1400 than at 0200 and 0900 as well as at 2100 than at 0200. The relative volume of Golgi apparatus was higher at 0900 and 1400 than at 0200. The relative volume of dense bodies of the MBB-1 type in pinealocyte perikarya was significantly lower at 1400 and 2100 than at 0900. In contrast, the relative volume of MBB-2 was higher at 1400 than at 0900 and 0200. The numerical density of DCV in perikarya was significantly higher at 0200 than at 1400. No significant differences were found in rough endoplasmic reticulum, lysosomes and multivesicular bodies. The pineal serotonin content showed a prominent rhythm with the maximum at 1400. The plasma melatonin concentration was significantly higher at 0200 than at 0900, 1400 and 2100. The obtained results demonstrate that both pinealocyte ultrastructure and pineal biochemistry in the pig undergo significant changes in the course of the diurnal rhythm.

Book ChapterDOI
TL;DR: The melatonin agonist LY-156735, 2 was recently reported to alleviate the symptoms of shift lag and to help resynchronize circadian rhythms at a dose of 5 mg/day in human clinical trials.
Abstract: Publisher Summary Melatonin is widely used as an over-the-counter therapy for the treatment of sleep disorders. This chapter discusses the use of melatonin and synthetic melatonin agonists for the treatment of sleep, major depression and anxiety. The hormone melatonin 1 (N-acetyl-5-methoxytryptamine) is produced and secreted from the pinealocytes of the pineal gland. Melatonin is synthesized in a circadian mode as it is produced at high levels at night and low levels during the day. Melatonin has high affinity for two G-protein coupled receptors (GPCRs), designated MT1 and MT2, and activates the inhibition of adenylyl cyclase via these receptors. Melatonin receptors MT1 and MT2 have amino acid sequences which are 60% identical. In situ analysis has revealed that MT1 mRNA is present in human suprachiasmatic nucleus (SCN). The distinct function of MT1 and MT2 has yet to be clearly defined; however, their role in the regulation of sleep and in circadian rhythm is well established. Melatonin has been studied extensively as close indole containing analogs have also been reported in the literature. The melatonin agonist LY-156735, 2 was recently reported to alleviate the symptoms of shift lag and to help resynchronize circadian rhythms at a dose of 5 mg/day in human clinical trials.

Journal Article
TL;DR: The ultrastructure of ovine pinealocytes undergoes the marked changes during postnatal development, mainly substructures involved in secretory activity.
Abstract: OBJECTIVE: The study was performed to analyze the ultrastructure of ovine pinealocytes during the period of postnatal development. MATERIAL AND METHODS: The pineals of newborn, 10-week and one-year old females of the domestic sheep were prepared for ultrastructural investigations. The point count analysis was used in quantitative studies of the pinealocyte substructures. RESULTS: The prominent feature of pinealocytes in the newborns was the presence of well developed rough endoplasmic reticulum and numerous polysomes. The pinealocyte cytoplasm contained also smooth endoplasmic reticulum, mitochondria, Golgi apparatus, lysosomes, dense core vesicles, multivesicular bodies and lipid droplets. Pinealocytes of the 10-week and 1-year old sheep were characterized by the occurrence of numerous vesicles and short cisterns of smooth endoplasmic reticulum, abundant microtubules and lipid droplets. Pinealocytes of the adult sheep were distinguished by well developed Golgi apparatus, numerous dense core vesicles and multivesicular bodies. The relative volume of rough endoplasmic reticulum in pinealocytes was significantly higher in the newborn sheep than in two other groups. The relative volume of mitochondria was significantly higher in pinealocytes of the 10-week old sheep than the newborns and one-year old animals. The relative volume of Golgi apparatus was significantly higher in the one-year old animals than in two other groups. No differences concern lysosomes. The relative volume of lipid droplets as well as the numerical density of dense core vesicles and multivesicular bodies increased significantly with age. CONCLUSION: The ultrastructure of ovine pinealocytes undergoes the marked changes during postnatal development. The changes concern mainly substructures involved in secretory activity.

Journal ArticleDOI
TL;DR: The results suggest that in the chicken retinally perceived light provides a powerful and important signal for synchronization of circadian rhythmicity in the pineal gland.

Journal ArticleDOI
TL;DR: It is suggested that the photoperiod determines the effectiveness of nocturnal AA-NAT induction by long-term modulation of the intrapineal pathway that transmits the cAMP signal to the AA- NAT gene.