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L J Grota

Publications and source records attributed to L J Grota.

At least 37 records · Page 2Linked to original sources
PubMed1985

Immunohistochemical assessment of melatonin binding in the pineal gland.

Melatonin binding in the pineal gland of albino rats is estimated using an immunohistochemical procedure. Binding is saturable, has relatively high affinity (Apparent KD = 2.7 nM), and competition studies indicate binding of indoleamines possessing an N-acetyl group on the terminus of the side chain (N-acetylserotonin and melatonin). These data are consistent with the interpretation that immunohistochemically determined melatonin in unfixed pineal tissue is assessing binding of N-acetylated indolealkylamines to pineal cell components. In albino rats maintained on 12-hour light: 12-hour dark cycles, melatonin binding exhibits a diurnal rhythm with low levels of saturation (30%) early in the light and saturation by endogenous melatonin near the onset of darkness. An annual rhythm of melatonin binding was observed in albino rats with low levels during the summer and high levels during the winter. Other rats were maintained on 12-hour light:dark cycles and fed for 2 hours either early in the light period or early in the dark period. For both morning- and evening-fed animals, melatonin binding was high prior to feeding and dropped immediately after feeding. Changes in melatonin binding that occur in response to alterations of feeding and time of year suggest the possibility that this binding reflects a functional site for melatonin.

Animals↗
PubMed1985

Serum melatonin response to melatonin administration in the Syrian hamster.

Chronic daily administration of melatonin (MT) can have potent effects on reproduction in the hamster. Various theories have been elaborated to explain these effects but little information has been available on circulating levels of MT following MT administration. We have examined the serum MT response in the male hamster to a single dose of 25 micrograms MT administered in the morning or in the afternoon--the same timing and dose used by others to produce reproductive effects. With both morning and afternoon administration, serum MT increased above 1,000 pg/ml and remained above the highest basal levels during most of the 24-hour cycle. These levels are clearly supraphysiologic ones. The decline in serum MT showed two distinct components following morning administration. Half-life of the initial component which probably represents rapid distribution into tissues was 17.3 min. A half-life of 25.1 h was calculated for the second component. We conclude that use of a 25-micrograms dose of melatonin to study pineal effects may be misleading.

Animals↗
PubMed1985

Scheduled feeding and 24-hour rhythms of N-acetylserotonin and melatonin in rats.

Male rats, kept under a lighting condition of 14-h light, 10-h dark, were subjected to scheduled feeding regimens. Food was available either in the early light phase or the early dark phase. The 24-h rhythms of serum corticosterone and of N-acetylserotonin (NAS) and melatonin (MT) in the pineal and serum were determined. It was found that whereas serum corticosterone and NAS rhythms responded to the feeding schedules, the rhythms of pineal NAS and of serum and pineal MT remained synchronized with the light-dark cycle. These findings indicated that the pineal was not the major source of circulating NAS. Whereas environmental lighting was the dominant "Zeitgeber" for the NAS rhythms in the pineal and the MT rhythms in the pineal and serum, for serum NAS rhythm, food presentation was the stronger Zeitgeber.

Animals↗
PubMed1984

N-Acetylserotonin in the central nervous system.

N-acetylserotonin has been identified by immunohistochemistry in specific brain areas separate from melatonin and serotonin. N-acetylserotonin is widely distributed within the brain stem, cerebellum and hippocampus and in the brain stem it is contained within the reticular formation nuclei and motor nuclei. Like serotonin, N-acetylserotonin appears to be derived from tryptophan as tryptophan hydroxylase inhibition leads to a lowering in immunoreactive N-acetylserotonin in brain and blood. Beta adrenergic drugs influence N-acetylserotonin neurons with beta adrenergic agonists causing a rise in immunoreactive N-acetylserotonin. The presence of N-acetylserotonin in brain has been confirmed by gas chromatography, mass spectrometry and radioimmunoassay. At this point little is known of the possible role of N-acetylserotonin in the brain. In the hippocampus N-acetylserotonin is present in granule cells and its appearance parallels the appearance of those cells. High affinity binding of tritiated N-acetylserotonin is found in brain and in various brain areas and this radioligand appears to label serotonergic receptors. Preliminary iontophoretic studies performed on hippocampal slices indicate an inhibitory action of N-acetylserotonin on glutamate induced firing of pyramidal cells. Taken together these findings suggest that N-acetylserotonin may have a role in the central nervous system distinct from that of being a precursor for melatonin. If this hypothesis is correct it would suggest that indoleamines have certain similarities to catecholamines. Thus for the catecholamines, dopamine, norepinephrine and epinephrine form a synthetic sequence and yet have independent roles as neurotransmitters and/or hormones. The three indoleamines serotonin, N-acetylserotonin and melatonin also form a synthetic sequence and these three substances may also have independent roles as neurotransmitters and/or hormones.

Acetyltransferases↗
PubMed1984

Relationship between pineal N-acetyltransferase activity, pineal melatonin and serum melatonin in rats under different lighting conditions.

Pineal N-acetyltransferase (NATase) activity has often been used as an indicator of pineal melatonin (MT) production, and the rhythms of pineal and serum MT levels are assumed to be parallel. However, these issues require further examination in view of some recent findings. In the present study, the 24-hour rhythms of pineal NATase, pineal MT content and serum MT concentrations were examined in groups of rats exposed to four artificial lighting regimens: (a) 16 h of light, 8 h of darkness (LD 16:8) for 2 weeks; (b) LD 12:12 for 2 weeks; (c) LD 4:20 for 2 weeks, and (d) LD 4:20 for 4 weeks. Under the above lighting conditions, the rhythms of pineal NATase, serum and pineal MT contents when tested demonstrated a closely parallel relationship. These results supported the idea that pineal NATase accurately reflected MT synthesis and that the rhythms of pineal and serum MT content were in phase under long and short photoperiods, in spite of extrapineal sites of MT synthesis. Further, the results also demonstrated that the timing of the MT peaks in the serum and the pineal may depend on the length of the adaptation period to the environmental lighting. When the rats were housed under a short photoperiod (LD 4:20), the peak MT levels appeared in the middle of the dark period after 2 weeks but shifted towards the end of the dark period after 4 weeks.

Acetyltransferases↗
PubMed1983

B-adrenergic regulation of N-acetylserotonin (NAS) synthesis in the rat cerebellum.

The synthesis of N-acetylserotonin (NAS) in the pineal gland is dependent upon the activity of the enzymes tryptophan-hydroxylase, 1-aromatic amino acid decarboxylase and N-acetyltransferase. Pineal N-acetyltransferase activity is regulated by the level of B-adrenergic activation. N-acetylserotonin (NAS) has also been identified in extra-pineal brain tissue. In order to investigate whether extra-pineal brain NAS levels are regulated by tryptophan hydroxylase and B-adrenergic activity, the effects of tryptophan hydroxylase inhibitors (parachlorophenylalanine and 6-fluoro-tryptophan) and adrenergic drugs (l-isoproterenol and propranolol) were examined. NAS was evaluated in the cerebellum of the rat using quantitative NAS-immunohistochemistry. A significant decrease in NAS-immunofluorescence was observed after tryptophan hydroxylase inhibition. Treatment with l-isoproterenol, a B-adrenergic agonist, resulted in a significant increase in NAS-immunofluorescence intensity. This effect was blocked by propranolol, a B-adrenergic blocking agent. These data indicate that the synthesis of NAS, in the cerebellum utilizes the established serotonin pathway and that NAS synthesis in the cerebellum is regulated by a B-adrenergic mechanism similar to that in the pineal gland.

Animals↗
PubMed1983

Antibodies to indolealkylamines II: site of conjugation of melatonin to protein using formaldehyde.

The site of coupling on the hapten was investigated for melatonin, formaldehyde, and protein conjugates. Model reactions of glycine ethyl ester and piperidine with formaldehyde and melatonin suggested that coupling first occurred through the indole nitrogen and then a stable bond was formed at C-2. Cross-reaction studies of antisera stimulated by melatonin, formaldehyde, and protein conjugates support the hypothesis that C-2 is the site of conjugation of melatonin to protein.

Animals↗
PubMed1983

An immunohistochemical method for the localization of N-acetylserotonin (NAS) in the central nervous system. Description, validation, and application of the technique.

Antisera to N-acetylserotonin (NAS) were raised in rabbits by coupling NAS to bovine serum albumin (BSA) through a p-carboxybenzyl (PCB) bridge at the indole N. The specificity and applicability of these antisera in immunohistochemistry is reported. The anti-NAS antiserum and a fluorescein-labeled immunoglobulin were employed to investigate the topographic distribution of immunoreactive NAS (INAS) in the hindbrain (mesencephalon, cerebellum, pons, and medulla oblongata). Positive identification of INAS was confirmed in the granular layer of the cerebellum, the tractus spinalis nervi trigemini and the reticular formation. INAS was also identified in Purkinje cells, cerebellar nuclei, nucleus principalis nervi trigemini, nucleus tractus mesencephali, cochlear and vestibular nuclei, the locus coeruleus, and other brain stem regions. The pattern of INAS distribution is independent of serotonin (5-HT) and norepinephrine (NE), although certain loci could contain both INAS and serotonin or INAS and norepinephrine.

Animals↗
PubMed1982

Identification and quantification of n-acetylserotonin (NAS) in the developing hippocampus of the rat.

1. A specific anti-NAS antibody and fluorescein-labelled second antibody were employed to investigate the presence of NAS in the dentate gyrus of the hippocampus as a function of age. 2. Immunoreactive NAS (INAS) was present in the granular cell layer of the dentate gyrus as early as 20 days postconception. 3. INAS appears to be present primarily in cell bodies. 4. Cross-reactivity and inhibition experiments confirm the positive identification of INAS. 5. INAS is age-dependent and increases with age reaching adult levels by day 30 post-conception/17 days after birth. 6. The appearance and subsequent increase in INAS correlates with the development of dentate granular cells and their subsequent synapse development suggesting a role for NAS in the normal functioning of these cells.

Animals↗
PubMed1982

24-hour rhythm of hypothalamic melatonin immunofluorescence correlates with serum and retinal melatonin rhythms.

The 24-hour rhythm of retinal and hypothalamic melatonin immunofluorescence was determined in male albino rats and compared to the 24-hour rhythm of serum melatonin determined by radioimmunoassay. Under a 12-hour (0:00-12:00): 12-hour dark cycle, the 24-hour rhythm of melatonin immunocytochemical fluorescence in the retina was biomodal (crests at 1:00 and 10:00-13:00). In serum, melatonin has a single crest late in the dark period (18:00). In the hypothalamus melatonin immunofluorescence showed increments corresponding to the crests in retina (13:00) and serum (18:00). In a 2-hour light: 22-hour dark cycle, the retinal rhythm was suppressed, the serum rhythm unchanged, and the hypothalamic rhythm had a single crest corresponding to the crest of the serum melatonin rhythm. These data indicate that the 24-hour hypothalamic rhythm of melatonin immunofluorescence may be secondary to the serum and retinal 24-hour rhythms of melatonin.

Animals↗
PubMed1981

Serotonin regulation of corticoid secretion in infant rats.

The plasma corticosterone response to various doses of serotonin and 5-hydroxytryptophan was studied in the 3-day-old rat. The maximum response to both drugs occurred 60 min after injection. Increased corticosterone concentration was observed at lower doses of serotonin than of 5-hydroxytryptophan. The effects of serotonin injection on Days 1-7 after birth on the plasma corticosterone response to 3 min of novelty stimulation at weaning was also studied. Serotonin treatment resulted in reduced adrenocortical reactivity which was reduced further when fluoxetine was given prior to the serotonin. Shocking rat pups on Days 1-7 also reduced adrenocortical reactivity and fluoxetine prior to shock further reduced adrenocortical reactivity when administered twice per day.

5-Hydroxytryptophan↗
PubMed1981

A radioimmunoassay for N-acetylserotonin in biological tissues.

We have developed a specific radioimmunoassay for N-acetylserotonin. Cross-reactivity of 23 related or selected compounds is less than 1 percent; sensitivity is 10 to 25 picograms per tube; within assay coefficient of variation is 7.5 to 8.4%; and between assay coefficient of variation is from 7.1 to 11.1%. Satisfactory parallelism has been demonstrated for rat, hamster and rabbit serum, rat and hamster serum extract and rat brain retina and pineal extract. An extracted sample of rat serum gave one peak on HPLC which corresponded to authentic N-acetylserotonin. Daytime levels of N-acetylserotonin have been established for each of the tissues studied. This radioimmunoassay provides a sensitive and specific method for determination of N-acetylserotonin levels in biological tissues.

Animals↗
PubMed1980

Determination of immunoreactive melatonin in the colon of the rat by immunocytochemistry.

Melatonin immunofluorescence in the rat colon was measured with a cadmium sulfide photometer in conjunction with an immunohistological double antibody procedure. The method proved to be very reliable with 5-10% intra- and interassay reliabilities. In addition, manipulation of the primary antibody concentration, the incubation time, and the nature of the first antibody produced predicted changes in fluorescence intensity. Treatment with p-chlorophenylalanine led to a marked reduction of melatonin-specific fluorescence.

Animals↗