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N Zisapel

Publications and source records attributed to N Zisapel.

At least 73 records · Page 4Linked to original sources
PubMed1988

Circadian variations in melatonin-binding sites in discrete areas of the male rat brain.

The binding of 125I-melatonin to synaptosomes prepared from whole brains of male rats of the CD strain and from the brain, hypothalamus and striatum of male rats of the Sabra-Wistar strain was assessed throughout a 24 h period. The animals were maintained under a daily schedule of 14 h light (05:00-19:00 h) and 10 h darkness. In whole brain preparations the density of binding sites at 18:00 h was higher by about 70% than at 02:00 h with no variations in apparent affinity of the binding sites throughout the daily period. Specific binding of 125I-melatonin was found in both hypothalamus and striatum of the male rat with a distinct diurnal variation in binding site density in the hypothalamus only. The density of 125I-melatonin-binding sites in the hypothalamus was maximal between 10:00 and 18:00 h and dropped sharply after the lights went off. The apparent 125I-melatonin-binding affinities in these regions were constant and very similar to those in whole brain preparations. The daily variations in densities of 125I-melatonin-binding sites in discrete brain areas may represent a diurnal rhythmicity in the responsiveness of the neuroendocrine axis to melatonin.

Animals↗
PubMed1988

Melatonin receptors revisited.

The pineal gland and its major product melatonin have a key role in conveying the environmental photoperiodic stimuli that impinge upon the mammalian reproductive axis. The brain, especially the medial preoptic and suprachiasmatic areas are thought to be the main sites of melatonin's neuroendocrine activity. The responsiveness of the mammalian reproductive system to the hormone is dependent on age, on the prevailing cyclical stage and on the circadian time. The existence of specific 125I-melatonin binding sites in synaptosomal fractions from rodent brain has recently been reported. The binding of 125I-melatonin is inhibited by melatonin and by the novel melatonin antagonist ML-23 but not by dopamine, serotonin or other structurally related compounds. The densities of 125I-melatonin binding sites at discrete brain regions vary significantly with age, circulating levels of steroid hormones or circadian time. These phenomena are compatible with the existence of melatonin receptors in the brain.

Animals↗
PubMed1988

Impact of circulating testosterone on iodomelatonin binding sites in the male rat brain.

The effects of castration and subsequent testosterone and estradiol treatment and of a single injection of ethylene-1,2-dimethanesulphonate (EDS) on the distribution of [2-125I]iodomelatonin ([ 125I]melatonin) binding sites in the male rat brain were investigated. Castration produced a marked testosterone-reversible decrease in [125I]melatonin binding in the male rat brain, particularly in the hypothalamus and hippocampus. In contrast, [125I]melatonin binding in the parietal cortex, medulla-pons and cerebellum was generally unaffected by castration. Estradiol did not reverse the effect of castration on [125I]melatonin binding. A single injection of EDS which causes the destruction of Leydig cells led to a marked decrease in [125I]melatonin binding in the brain of the rats between 3 and 7 days after treatment. This decrease correlated with the decline in serum concentrations of testosterone. Specific [125I]melatonin binding and serum concentrations of testosterone subsequently increased to control levels within 37 days after treatment in accord with the repopulation of Leydig cells. The results clearly show that testosterone regulates the density of melatonin receptors in the hypothalamus and hippocampus of the male rat.

Animals↗
PubMed1988

Melatonin receptors in discrete brain areas of the male rat. Impact of aging on density and on circadian rhythmicity.

The distribution of melatonin receptors in six discrete brain areas of mature (3-4 months old) and aged (greater than 24 months old) male rats was recorded every 4 h during a 24-hour light: dark cycle (L:D 14:10h). 125I-melatonin was used as a melatonin receptor probe. In the mature animals, specific binding of 125I-melatonin was found in all brain areas investigated, i.e. hypothalamus, medulla pons, hippocampus, cerebellum, parietal cortex and striatum. The density of 125I-melatonin-binding sites in the hypothalamus, medulla pons and hippocampus exhibited clear diurnal rhythms with different patterns and phases. No such rhythm was evident in the cerebellum, parietal cortex and striatum. The apparent affinity of the binding sites was similar in all the brain regions and did not change at any of the times recorded. In the old male rats, the density of 125I-melatonin binding sites in the hypothalamus was only 10% of that in the mature animals at 13 h after the onset of light and was vanishingly small throughout the 24-hour period. The 24-hour mean of the binding site density in the parietal cortex, hippocampus and medulla pons was significantly lower than in mature rats with no apparent diurnal variations. The age-related decrease in the density of melatonin-binding sites was less pronounced in the cerebellum and striatum. In all brain areas tested, apart from the hypothalamus, the decrease in receptor densities was not accompanied by changes in the apparent affinity towards the ligand.(ABSTRACT TRUNCATED AT 250 WORDS)

Aging↗
PubMed1988

N-(2,4-dinitrophenyl)-5-methoxytryptamine, a novel melatonin antagonist: effects on sexual maturation of the male and female rat and on oestrous cycles of the female rat [corrected].

N-(2,4-dinitrophenyl)-5-methoxytryptamine [corrected] has recently been synthesized and shown to antagonize the inhibitory effect of melatonin on the release of dopamine in vitro from the hypothalamus of female rats. In the present study the ability of N-(2,4-dinitrophenyl)-5-methoxytryptamine [corrected] to inhibit in vivo the following melatonin-mediated effects was investigated: (1) delayed sexual maturation of young male rats, (2) delayed sexual maturation of young female rats, (3) inhibition of ovulation in mature female rats and (4) re-establishment of oestrous cycles in adult female rats maintained in continuous light. The inhibitory effect of daily melatonin injections, given in the afternoon, on the growth of the prostate gland and seminal vesicles and on serum testosterone concentrations in young male rats was prevented by daily injections of N-(2,4-dinitrophenyl)-5-methoxytryptamine [corrected]. Daily injections of N-(2,4-dinitrophenyl)-5-methoxytryptamine [corrected] alone did not affect sexual maturation of young rats. In young male rats treated through the drinking water with melatonin, the growth of the accessory sex organs, but not that of the testes, was delayed and serum concentrations of testosterone were lower than in untreated rats. Administration of N-(2,4-dinitrophenyl)-5-methoxytryptamine [corrected] through the drinking water increased serum concentrations of testosterone but did not significantly affect the weights of the accessory sex organs. Simultaneous administration of N-(2,4-dinitrophenyl)-5-methoxytryptamine [corrected] and melatonin through the drinking water prevented completely, in a dose-dependent manner, the melatonin-mediated decrease in epididymal weights and in serum concentrations of testosterone and partially inhibited the delayed growth of the prostate glands and seminal vesicles. In young female rats treated with melatonin through the drinking water for 30 days, the growth of the ovaries was inhibited and serum concentrations of oestradiol were lower than in untreated rats. The growth of the uterus was not significantly affected. Administration of N-(2,4-dinitrophenyl)-5-methoxytryptamine [corrected] through the drinking water did not significantly affect uterine and ovarian weights or oestradiol concentrations. Simultaneous administration of melatonin and N-(2,4-dinitrophenyl)-5-methoxytryptamine [corrected] through the drinking water prevented completely the melatonin-mediated decrease in ovarian weights and in serum oestradiol concentrations. Ovulation during presumptive oestrus was prevented in adult female rats treated through the drinking water for 7 days with melatonin. Administration of N-(2,4-dinitrophenyl)-5-methoxytryptamine [corrected] alone did not significantly affect the average numbers of ova shed and corpora lutea present. Simultaneous administration of N-(2,4-dinitrophenyl)-5-methoxytryptamine [corrected] and melatonin prevented completely the melatonin-mediated inhibition of ovulation; the average number of ova shed was the same as in controls.(ABSTRACT TRUNCATED AT 400 WORDS)

5-Methoxytryptamine↗
PubMed1987

Impact of circulating estradiol on melatonin binding sites in discrete areas of the female rat brain.

The distribution of 125I-melatonin binding sites in 6 discrete brain regions of female rats at the estrous stage and the effects of ovariectomy and subsequent 17 beta-estradiol treatment on these binding sites were studied. Specific binding of 125I-melatonin was found in the hypothalamus, medulla-pons, hippocampus, cerebellum, striatum and parietal cortex of the female rats. Ovariectomy produced a large estradiol-reversible decrease in 125I-melatonin binding in the medulla-pons and hypothalamus. In contrast, 125I-melatonin binding sites in the other brain regions were generally unaffected by ovariectomy or estradiol. The estradiol-regulated changes in melatonin binding in the hypothalamus and medulla-pons may reflect the role of a specific estradiol-melatonin interaction in the coordination of the neuroendocrine reproductive axis.

Animals↗
PubMed1987

Regulation of melatonin's activity in the female rat brain by estradiol: effects on neurotransmitter release and on iodomelatonin binding sites.

The effects of ovariectomy and of 17 beta-estradiol (estradiol) treatment in vivo and in vitro on the ability of melatonin to inhibit dopamine release from the female rat hypothalamus and on [125I]-iodomelatonin binding sites in the brains and hypothalami of female rats were investigated. In long-term (2-4 weeks) ovariectomized (OVX) female rats, the inhibitory effect of melatonin in vitro on dopamine release from the hypothalami was abolished. After implantation of estradiol capsules, the ability of melatonin to inhibit dopamine release from the hypothalamus was reinstated and resembled that observed in intact female rats at estrus and in females exhibiting spontaneous persisting estrus. Similar reinstatement of the responsiveness to melatonin was observed in hypothalami of OVX rats shortly (2 h) after a single subcutaneous injection of estradiol (200 micrograms/animal). Incubation of hypothalami of OVX rats in vitro for 50-90 min with estradiol (0.1-1 nM) resulted in a partial (up to 50%), time and steroid concentration-dependent reinstatement of the ability of melatonin to inhibit the induced dopamine release. Conversely, incubations with estradiol in vitro reduced the ability of melatonin to inhibit dopamine release from the hypothalami of intact rats at proestrus. Such incubations had no effect on the release of dopamine from hypothalami of rats at estrus or of short-term OVX (3 days) rats. The changes in the responsiveness of the hypothalamus to melatonin were accompanied by profound changes in the binding of [125I]-iodomelatonin, to synaptosomes isolated from whole brains and from hypothalami of the OVX female rats. In OVX rats, the densities of the binding sites in the brains and particularly in the hypothalami decreased to 18 and 24% respectively, of the values observed in control females at estrus. The apparent dissociation constant of the remaining sites was significantly lower (ca. 90 nM) than that observed in the intact controls (ca. 300 nM). An almost complete reinstatement of the [125I]-iodomelatonin binding sites was observed in synaptosomes prepared from the hypothalami of OVX rats shortly (2 h) after a single subcutaneous injection of estradiol, or after incubation with estradiol in vitro. The estradiol-mediated reinstatement of [125I]-iodomelatonin binding sites was less pronounced in synaptosomes prepared from whole brains. The results clearly show that estradiol directly modulates the responses of the dopaminergic neurosecretory system in the hypothalamus to melatonin. This phenomenon may be primarily associated with the estradiol-induced changes in the density and function of melatonin receptors in the hypothalamus.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1986

Characterization of central melatonin receptors using 125I-melatonin.

The binding of 125I-melatonin, a potent analog of melatonin, to rat brain synaptosomal preparations was investigated. 125I-melatonin bound with high affinity (Kd = 38 nM) to a single class of sites (Bmax = 81 fmol/mg protein). Kinetic studies indicated that binding was time-dependent and reversible. Specific 125I-melatonin binding was inhibited by melatonin, and was unaffected by other structurally related compounds including serotonin. Binding of 125I-melatonin was greatly reduced if the synaptosomal preparations were pretreated by heat or trypsin but was unaffected by freeze-thawing. These results suggest that 125I-melatonin may serve as a valuable probe for studying melatonin receptors.

Animals↗
PubMed1986

Concerted enhancement of calcium influx, neurotransmitter release and protein phosphorylation by a phorbol ester in cultured brain neurons.

We have recently shown that the phorbol ester 12-O-tetradecanoyl-phorbol-13-acetate enhances the depolarization induced, calcium dependent release of [3H]dopamine from cultured brain neurons in the rat. In the present study the effects of 12-O-tetradecanoyl-phorbol-13-acetate on the kinetic parameters of depolarization induced calcium influx and on Ca2+ dependent neurotransmitter release and protein phosphorylation were investigated. Depolarization induced neurotransmitter release from the neurons occurs in two phases: an initial, fast release and a subsequent slow release. At low extracellular Ca2+, 12-O-tetradecanoyl-phorbol-13-acetate enhanced the quantity of fast release and in addition, increased the rate constant of the slow release. These effects mimicked the effects of increasing the extracellular Ca2+. Various phorbol derivatives known to activate the Ca2+ activated phospholipid dependent protein kinase (protein kinase C) were also able to enhance the stimulated release of [3H]dopamine from the neurons. 12-O-tetradecanoyl-phorbol-13-acetate induced the incorporation of 32Pi into a protein with an apparent molecular weight of 45,000 daltons regardless of depolarization or of the presence of Ca2+. In addition, 12-O-tetradecanoyl-phorbol-13-acetate induced in unstimulated neurons, Ca2+ dependent increase in the amount of 32Pi incorporated into a 43,000 dalton protein and decrease in the amount incorporated into a 55,000 dalton protein. These changes mimicked the Ca2+ dependent changes in protein phosphorylation which occur upon stimulation of the neurons. Kinetic studies of depolarization induced Ca2+ uptake by the neurons indicated that 12-O-tetradecanoyl-phorbol-13-acetate enhanced the maximal influx of Ca2+ through the voltage sensitive Ca2+ channels by 40%. The results indicate that 12-O-tetradecanoyl-phorbol-13-acetate acts primarily on the regulation of stimulated Ca2+ entry into the cells. Consequently neurotransmitter release at submaximal extracellular [Ca2+] is enhanced.

Animals↗
PubMed1985

Calcium permeability changes and neurotransmitter release in cultured rat brain neurons. I. Effects of stimulation on calcium fluxes.

The permeability of neuronal membranes to Ca2+ is of great importance for neurotransmitter release. The temporal characteristics of Ca2+ fluxes in intact brain neurons have not been completely defined. In the present study 45Ca2+ was used to examine the kinetics of Ca2+ influx and efflux from unstimulated and depolarized rat brain neurons in culture. Under steady-state conditions three cellular exchangeable Ca2+ pools were identified in unstimulated cells: 1) a rapidly exchanging pool (t1/2 = 7 s) which represented about 10% of the total cellular Ca2+ and was unaffected by the presence of Co2+, verapamil, or tetrodotoxin; 2) a slowly exchanging pool (t1/2 = 360 s) which represented 42% of the total cellular Ca2+ and was inhibited by Co2+, but not by verapamil or tetrodotoxin; 3) a very slowly exchanging pool (t1/2 = 96 min) which represented 48% of the total cell Ca2+ was observed only in the prolonged efflux experiments. The rate of exchange of 45Ca2+ in the unstimulated cells was dependent on the extracellular Ca2+ concentration (half-saturation at 70 microM). Depolarization of the neurons with elevated K+ causes a rapid and sustained 45Ca2+ uptake. The cellular Ca2+ content increased from 56 nmol/mg protein in unstimulated cells to 81 nmol/mg protein during 5 min of depolarization. The kinetics of the net 45Ca2+ uptake by the stimulated neurons was consistent with movement of the ion with a first order rate constant of 0.0096 s-1 (t1/2 = 72 s) into a single additional compartment. The other cellular Ca2+ pools were apparently unaffected by stimulation. The stimulated 45Ca2+ uptake was inhibited by Co2+ and by the Ca2+ channel blocker verapamil but not by the Na+ channel blocker tetrodotoxin. Ca2+ uptake into this compartment was dependent on the extracellular Ca2+ concentration (half-saturation at 0.80 mM Ca2+). Predepolarization of the cells with high K+ for 10-60 s prior to the addition of the radioactive calcium did not alter the rate of 45Ca2+ incorporation into the stimulated cells. It is concluded that the rapidly exchanging, the slowly exchanging, and the depolarization-induced Ca2+ pools observed in intact brain neurons are physically as well as kinetically distinct from each other. In addition, the depolarization-induced component observed in stimulated cells represents movement of the Ca2+ ions through a single class of voltage-sensitive Ca2+ channels. These Ca2+ channels are inhibited by Co2+ ions and by verapamil and are not inactivated during depolarization of the brain neurons.

Animals↗
PubMed1985

Calcium permeability changes and neurotransmitter release in cultured brain neurons. II. Temporal analysis of neurotransmitter release.

The coupling between depolarization-induced calcium entry and neurotransmitter release was studied in rat brain neurons in culture. The endogenous dopamine content of the cells was determined by high performance liquid chromatography utilizing electrochemical detection. The amount of dopamine in unstimulated cells was found to be about 16 ng/mg of protein. Depolarization of the neurons by elevated K+ caused a Ca2+-dependent release of dopamine from the cells. Following 1 min of depolarization, the cellular dopamine content and the amount of [3H]dopamine in cells preloaded with the radioactive transmitter were reduced by 35%. The release of [3H]dopamine by the neurons was measured at 1.5-6-s intervals by a novel rapid dipping technique. Depolarization in the presence of Ca2+ (1.8 mM) enhanced the rate of neurotransmitter release by 90-fold (0.072 +/- 0.003 s-1) over the basal release in the presence of Ca2+. The evoked release consisted of a major rapidly terminating phase (t1/2 = 9.6 s) which comprised about 40% of the neurotransmitter content of the cells and a subsequent slower efflux (t1/2 = 575 s) which was observed during following prolonged depolarization. Predepolarization of the cells in the absence of extracellular Ca2+ did not affect the kinetics of the evoked release. The fast evoked release could be re-elicited in the cells after 20 min "rest" in reference low K+ buffer. The effects of varying the extracellular Ca2+ concentrations on the kinetic parameters of the evoked release were measured. The amount of neurotransmitter released during the fast kinetic phase was very sensitive to the external Ca2+ (from 0% in the absence of Ca2+ to 40% of the neurotransmitter content at Ca2+ 0.3 mM). The rate constant of the fast release did not depend on the extracellular Ca2+, whereas the rate constant of the slow release increased from 0.0004 +/- 0.0001 s-1 at 0.4 mM Ca2+ to 0.0012 +/- 0.0002 s-1 at 0.8 mM Ca2+. The fast evoked release was inhibited by verapamil in a concentration-dependent manner. By contrast, verapamil enhanced the basal and the slow release independent of the presence of Ca2+. Both fast and slow phases of the evoked release were blocked by Co2+. Addition of Co2+ within the first 6 s after the onset of depolarization inhibited the fast release but failed to do so when added later on.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1985

Phorbol ester and calcium act synergistically to enhance neurotransmitter release by brain neurons in culture.

Preincubation of intact fetal brain neurons in culture with the phorbol ester TPA (12-O-tetradecanoyl phorbol-13-acetate) in the presence of calcium, resulted in the enhancement of the depolarization-induced, Ca2+-dependent neurotransmitter release by the cells. This effect was due to a marked decrease in the concentration of extracellular Ca2+ required to provoke the release. The concentration of Ca2+ needed to produce half-maximal release shifted from approx 0.1 mM in the absence of TPA to 0.018 mM in its presence. This activity of TPA was concentration-dependent (half-maximal effect at 4 nM TPA) and was also dependent on the presence of calcium during the preincubation period. The TPA-induced enhancement of the stimulated release was also observed when Ca2+ entry into the depolarized cells was partially inhibited by Co2+. The results suggest that TPA acts synergistically with Ca2+ to activate neuronal component(s) involved in Ca2+-dependent neurosecretion.

Animals↗
PubMed1985

Circadian variations in the inhibition of dopamine release from adult and newborn rat hypothalamus by melatonin.

The inhibitory effect of melatonin on evoked dopamine release from the hypothalamus was studied in adult male rats throughout a 24-hour period. The animals were maintained under a daily schedule of 14 h of light (0.00-14.00 h) and 10 h of darkness. The inhibition of dopamine release in vitro by 1 microM melatonin clearly exhibited a 24-hour rhythm with a peak at 5.00 h and almost no inhibition at 15.00 h. The concentrations of melatonin needed to produce this effect were similar throughout the 24-hour cycle, although the actual amount of inhibition at any given concentration of melatonin varied. Other indole derivatives, with the exception of 5-methoxy tryptophol, did not affect significantly the release of 3H-dopamine from the male rat hypothalami. The inhibition of dopamine release by melatonin was not observed in newborn rats but developed during the first week of life, reaching a plateau level between 6 and 7 days postnatally. However, the difference between the amount of inhibition by melatonin at 5.00 h and at 8.00 h existed from the time the inhibition was first observed. The change in amplitude of this difference was due not to differences in the apparent affinity towards melatonin but to increase in the maximal inhibition observed at 5.00 h. The data indicate that the hypothalamic sensitivity to melatonin exhibits a circadian rhythm, and that this develops postnatally prior to the development of circadian variations in melatonin levels, i.e. the 'melatonin rhythm'.(ABSTRACT TRUNCATED AT 250 WORDS)

Age Factors↗
PubMed1984

Calcium uptake and calcium-dependent phosphorylation during development of rat brain neurons in culture.

The emergence of the capacities for calcium uptake and calcium-regulated protein phosphorylation during the development of embryonic brain neurons in tissue culture was examined. In the maturing cells, the enhancement in 45Ca2+-uptake upon stimulation with high K+ increased by 3-4 fold during the second week in vitro, in parallel to an increase in the capacity for high K+-induced Ca2+-dependent release of prelabeled [3H]dopamine. The pattern of incorporation of [32Pi]phosphate into the major phosphoproteins in maturing cells under nonstimulating conditions also changed during cell development: the incorporation of 32Pi into two proteins of apparent molecular weights--55,000 and 43,000 dalton--increased, but decreased in a 45,000 dalton protein. Stimulation of mature cells (after 10-11 days in vitro) resulted in a Ca2+-dependent increase in the amount of 32Pi incorporated into the 43,000 dalton protein and a decrease in the amount incorporated into the 55,000 dalton protein. This calcium-regulated phosphorylation pattern was not observed until 6 days in vitro. Introduction of Ca2+ into the immature cells by means of the Ca2+ ionophore A23187 did not alter the phosphorylation pattern and did not cause neurotransmitter release. The amount of [35S]methionine incorporated into a 43,000 dalton protein which comigrated with the 43,000 dalton phosphoprotein also increased upon cell maturation. The results suggest that this phosphoprotein (which does not comigrate with nonphosphorylated actin on two-dimensional polyacrylamide gels) develops in the cells in parallel to the emerging processes of the stimulation-induced calcium entry and calcium-dependent neurosecretion.

Animals↗
PubMed1983

Inhibition by melatonin of dopamine release from rat hypothalamus: regulation of calcium entry.

The [Ca2+] dependency of dopamine release evoked by electrical field stimulation of hypothalamic tissue from female rats at the estrous stage was assessed in the presence of melatonin. At concentrations of 10(-9)-10(-6) M, melatonin inhibited the [Ca2+]-dependent dopamine release. Melatonin reduced tissue uptake of 45Ca2+ during stimulation either by an electrical field or by elevated K+ concentration. About 90% of this inhibition by melatonin of 45Ca2+ uptake, as well as of dopamine release, was not observed in the presence of the calcium ionophore A23187. Hence, the inhibitory effect on dopamine release by melatonin may stem from the reduction of calcium entry into the presynaptic nerve endings.

Animals↗
PubMed1983

Calcium-dependent protein phosphorylation and dephosphorylation in intact brain neurons in culture.

Preincubation of intact fetal rat brain neurons in culture with 32Pi results in the incorporation of 32Pi into about 20 specific proteins. Upon stimulation by electrical field stimulation or by K+-induced depolarization, highly significant calcium-dependent increase in phosphorylation of a protein of app. Mr 43 000 and decrease in phosphorylation of an app. Mr 55 000 protein occur. These changes can be attributed to the entry of Ca2+ into the cellular cytoplasm since they can occur upon selective permeabilization of the cell membrane to Ca2+ by the Ca2+-ionophore A23187 and are not observed upon stimulation of the cells in the presence of the Ca2+ channel blocker D-600. These data suggest that these phosphoproteins may be involved in the regulation of processes underlying neurotransmitter release.

Animals↗