Derivatives of [3H]serotonin in organ cultures of hamster pineal gland (original) (raw)
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Temporal Effects of Norepinephrine on Pineal Serotonin In Vitro
Journal of Pineal Research, 1986
The purpose of this study was to examine the temporal effects of norepinephrine (NE) stimulation o n pineal serotonin (5HT) in vitro. Rat pineal glands were individually incubated in nutrient media for 6 h. After a 2-h preincubation, the glands were exposed to tryptophan (TRYP) or TRYP + norepinephrine M each) for 4 additional h. Media and pineals were then analyzed for 5HT and 5HIAA content. Initial exposure to TRYP increased 5HT levels in the media. Norepinephrine increased media 5HT concentrations further and also caused 5HIAA levels to rise significantly. These findings suggest that 5HT secretion and/ or oxidation is an early response to stimulation of the pineal gland by NE. Following prolonged NE exposure, media 5HT declined to control values while 5HIAA remained high. Similarly, intrapineal levels of 5HT fell after prolonged stimulation with NE. Thus, pineal responses to initial and continued stimulation with NE are differentiable as early and late effects upon 5HT. Early effects may include secretion and/or oxidation of 5HT followed later by utilization of 5HT in N-acetylation pathways such as that leading to melatonin synthesis. The finding that 5HT is secreted in response to NE suggests that the indoleamine may be a hormone of the pineal gland. Based upon its known influence on physiologic rhythms, daily secretion of 5HT from the pineal may be an important part of the gland's time-keeping function.
Journal of Pineal Research, 1986
Norepinephrine (NE, 10−6 M) stimulated melatonin accumulation in the incubation medium of rat (but not Syrian hamster) pineals taken at the end of the light phase. However, NE elevated melatonin accumulation in the medium of pineals taken after 20 min of light exposure of animals of either species at 6 h into the 10-h dark phase. A dose response to 10−7−10−5 M NE was observed in both the medium and pineals upon incubation of pineals taken from rats at 4 h into the light phase and from hamsters after 20 min light exposure at 6 h into the dark phase. Approximately 95% of the melatonin present was in the medium. The incubation time was 4 h in all cases. Subcutaneous injection of 1 μg/g NE (either at the end of the light phase or after 30 min of light at 6 h into the dark phase) did not stimulate in vivo Syrian hamster pineal melatonin content determined 1 or 2 h after injection, whether the hamsters were placed in light or darkness after the injection. However, after 30 min of light beginning at 6 h into dark, injection of 5 μg/g desipramine (DMI, a blocker of catecholamine uptake into nerve endings) allowed a dramatic hamster pineal melatonin response to additional injection of 1 μg/g NE, observed at 1 and 2 h in light after injection. A small effect of DMI alone was seen. DMI also potentiated the effect of NE (each 10−6 M) on melatonin accumulation in the medium of incubated hamster pineals taken after a short light exposure at night. No significant stimulatory effect of NE and/ or DMI was seen in vivo or in vitro near the middle of the light phase. Measurement of melatonin in the incubation medium is a useful method for studying pineal function. The Syrian hamster pineal has rhythm of sensitivity to NE (sensitivity evident at night) and even at night is protected by neuronal uptake from circulating NE-induced stimulation of melatonin production. NE appears to be the neurotransmitter for stimulation of pineal melatonin production in the Syrian hamster. The sensitivity rhythm and uptake protection might provide specificity of control of the nightly melatonin signal by reducing the chance of a melatonin response during the day or a response to circulating catecholamines from general sympathetic stimuli.
Journal of Pineal Research, 1984
The nighttime rise in pineal melatonin levels can he blocked by administration of the 0-adrenergic receptor antagonist, propranolol, in both Syrian hamsters and rats. Although the administration of P-adrenergic receptor agonists such as norepinephrine or isoproterenol stimulates pineal melatonin production in the rat, these drugs are without apparent effect on indole production in the Syrian hamster. To determine whether this lack of stimulatory effect in the Syrian hamster is characteristic of this species, a comparison of the effects of norepinephrine and isoproterenol o n pineal serotonin N-acetykransferase activity and melatonin content was conducted. In contrast to their lack of effect in the Syrian hamster, norepinephrine and isoproterenol stimulated pineal serotonin N-acetyltransferase activity and melatonin content in the Djungarian hamster. Hourly injection of norepinephrine during a continuation of light into the normal dark period stimulated increases in the activity of serotonin N-acetyltransferase and melatonin content in the Djungarian hamster but was without effect o n these pineal parameters in the Syrian hamster.
Norepinephrine-induced elevation of acetyl coenzyme A in rat pineal glands in culture
Journal of Pharmacological Methods, 1982
A micromethod for the determination of acetyl coenzyme A was developed and the relationship between the concentration of acetyl coenzyme A and the activity of rat pineal serotonin N-acetyltransferase was studied. Acetyl coenzyme A was determined by converting it into N-acetylserotonin using rat liver serotonin Nacetyltransferase. Subsequently, the hydroxy group of N-acetylserotonin was Omethylated by hydroxyindole-0-methyltransferase and S-[methyl-3Hladenosyl-/methionine to form [3H]melatonin, which was then conveniently separated from S-[methyl-3H]adenosyl-/-methionine by thin-layer chromatography. The amount of radioactivity in melatonin is a measure of acetyl coenzyme A concentration. This method is sensitive and specific, since it can detect as low as 5 pmol of acetyl coenzyme A but not structurally related substances such as coenzyme A, adenosine diphosphate, cysteamine, o-panthothenic acid, or sodium acetate. After treating cultured rat pineal glands with I-norepinephrine (IO PM) for 6 hr, the concentration of acetyl coenzyme A was increased significantly from 3.26 +-0.37 to 10.24 5 0.93 pmol/gland, while the activity of serotonin N-acetyltransferase increased 68-fold. This result suggests that acetyl coenzyme A may play an important role in the norepinephrine-induced induction of serotonin N-acetyltransferase. Sensitivity and adaptability of this method can be utilized to measure acetyl coenzyme A in discrete regions of rat brain and in experimental conditions in which micromeasuremenf of acetyl coenzyme A may be required.
Journal of Chromatography B: Biomedical Sciences and Applications, 1986
Simultaneous determination of melatonin and serotonin in rat pineal gland is described using reversed-phase high-performance liquid chromatography with fluorimetric detection. These indoles were analysed isocratically within 15 min. In this work, veratric acid (3,4dimethoxybenzoic acid), which has fluorescence characteristics (h,, = 290 nm, h,, = 350 nm) around the wavelength of native fluorescence of melatonin (& = 285 nm, h,, = 345 nm), was used as an internal standard. This method was applied to the determination of melatonin and serotonin in male and female rat pineal gland. No significant differences between the two groups were observed in the pineal melatonin and serotonin contents. The pineal melatonin and serotonin contents were compared with the oestrous and the dioestrous phases of female rats. They were not widely different from each other.
Neuroscience Letters, 1994
Day-and nighttime content of catecholamines, serotonin and their metabolites were measured in the pineal gland of Sprague-Dawley rats and Djungarian hamsters. In addition, monoamine turnover rates were determined in the hamster pineal gland following administration of ~-methyl-p-tyrosine. Animals were decapitated in the middle of the light or dark period, respectively, and pineal tissue was analysed by high performance liquid chromatography with electrochemical detection. Pineals of both species exhibited day/night-differences in serotonin, 5-hydroxyindole-3-acetic acid and dopamine content. The hamster pineal gland further showed day/night differences in its content of epinephrine and 3-methoxy-4-hydroxyphenylethyleneglycol. The dopamine turnover rate was augmented at night, while norepinephrine turnover was constant. Immunohistochemical incubations of pineal paraffine sections showed fibers and terminals stained by antisera to tyrosine-hydroxylase (TH) and dopamine-fl-hydroxylase, and a few perikarya-like structures exhibiting TH immunoreactivity. The results support the view that dopamine, rather than only functioning as a norepinephrine precursor, is actively involved in the control of melatonin synthesis.
2002
OBJECTIVES: This study was performed to search for changes in rat pineal function attributed to age and immunization with Freund's adjuvant. METHODS: Young (2 months) and old (18-20 months) Wistar rats were injected s.c. with Freund's adjuvant or its vehicle. Eighteen days later, at the acute phase of arthritis, pineal concentration of serotonin (5-HT), 5-hydroxyindoleacetic acid (5-HIAA), norepinephrine (NE) and melatonin was measured by high pressure liquid chromatography at 4 different time intervals throughout the nocturnal activity span. RESULTS: Old rats had the lowest pineal 5-HT and 5-HIAA content, the decrease in 5-HIAA exceeding that of 5-HT; consequently, old rats had the lowest 5-HIAA/5-HT ratio, an index of pineal 5-HT turnover. Although immunization did not affect globally pineal 5-HT or 5-HIAA levels, significant interactions "immunization x age" and "immunization x time" were found, i.e., immunization augmented pineal 5-HT content at the beginning of the activity span in young rats and at second half of the activity span in young and old rats, and increased pineal 5-HIAA concentration in young rats at the second part of the activity span only. Freund's adjuvant treatment increased pineal 5-HT turnover exclusively in old rats, an effect mainly seen during the second part of the activity span. Old rats exhibited the lowest pineal NE and melatonin levels, immunization further depressing them. CONCLUSION: The effect of immunization with Freund's adjuvant on a number of pineal pre-and postsynaptic parameters are age-dependent.
Peptidergic Modulation of Serotonin Release from Cultured Rat Pinealocytes
Journal of Neuroendocrinology, 2003
This paper describes the effects of b-adrenergic and peptidergic inputs on serotonin (5-HT) synthesis, outflow and metabolism into melatonin in cultured dissociated rat pinealocytes. The spontaneous outflow of 5-HT from pinealocytes was high as demonstrated by the elevated levels of extracellular 5-HT accumulated in the medium (about 5 ng/h/70,000 pineal cells). The b-adrenergic agonist isoproterenol (ISO) used at concentrations up to 10−6 M induced a moderate (+20-40%) increase in intra-and extracellular 5-HT levels together with a large release of melatonin. At a higher ISO stimulation (10−5 M), the intra-and extracellular levels of 5-HT were significantly (−25-30%) reduced whereas melatonin secretion was dramatically increased. This is interpreted as a large 5-HT mobilization for melatonin synthesis and release, consequently reducing both the intracellular pool and outflow of 5-HT. The peptides vasoactive intestinal peptide (VIP) and pituitary adenylate cyclase activating peptide (PACAP) up to 10−7 M induced always a moderate (+20-30%) increase in intra-and extracellular levels of 5-HT. However, the use of nM concentrations of VIP or PACAP together with 10−6 M ISO induced a decrease in 5-HT outflow (−25-30%) and a dramatic increase in melatonin secretion as did 10−5 M ISO alone. Neuropeptide Y (NPY) is another pineal peptide which induced a stimulation of 5-HT outflow (+30-40%) although its effect on melatonin release was marginal. The above results are discussed in term of the multineuronal regulation of the synthetic and secretory activities of the rat pineal gland.
Subcutaneous injections of 25 #g of 5-methoxytryptamine (5-MT) in oil into intact and pinealectomized male hamsters given between 4.30 p.m. and 5 p.m. (light on from 5 a.m. to 7 p.m.; 14 L/10 D) for 54 consecutive days caused involution of the testes. 5-MT, however, is more effective when the pineal is present. These results indicate that melatonin is not the only pineal factor inducing gonadal atrophy in the hamster. 5-MT seems even more effective than melatonin in so far as it is, contrary to melatonin under the same experimental conditions, also effective in the absence of the pineal. Like melatonin, 5-MT appears to be implicated in the control of the reproductive function.