Amitriptyline up-regulates connexin43-gap junction in rat cultured cortical astrocytes via activation of the p38 and c-Fos/AP-1 signalling pathway.

BACKGROUND AND PURPOSE: Intercellular communication via gap junctions, comprised of connexin (Cx) proteins, allow for communication between astrocytes, which in turn is crucial for maintaining CNS homeostasis. The expression of Cx43 is decreased in post-mortem brains from patients with major depression. A potentially novel mechanism of tricyclic antidepressants is to increase the expression and functioning of gap junctions in astrocytes. EXPERIMENTAL APPROACH: The effect of amitriptyline on the expression of Cx43 and gap junction intercellular communication (GJIC) in rat primary cultured cortical astrocytes was investigated. We also investigated the role of p38 MAPK intracellular signalling pathway in the amitriptyline-induced expression of Cx43 and GJIC. KEY RESULTS: Treatment with amitriptyline for 48 h significantly up-regulated Cx43 mRNA, protein and GJIC. The up-regulation of Cx43 was not monoamine-related since noradrenaline, 5-HT and dopamine did not induce Cx43 expression and pretreatment with α- and ß-adrenoceptor antagonists had no effect. Intracellular signalling involved p38 MAPK, as amitriptyline significantly increased p38 MAPK phosphorylation and Cx43 expression and GJIC were significantly blocked by the p38 inhibitor SB 202190. Furthermore, amitriptyline-induced Cx43 expression and GJIC were markedly reduced by transcription factor AP-1 inhibitors (curcumin and tanshinone IIA). The translocation of c-Fos from the cytosol and the nucleus of cortical astrocytes was increased by amitriptyline, and this response was dependent on p38 activity. CONCLUSION AND IMPLICATION: These findings indicate a novel mechanism of action of amitriptyline through cortical astrocytes, and further suggest that targeting this mechanism could lead to the development of a new class of antidepressants.

Primary cultures of rat cortical microglia treated with nicotine increases in the expression of excitatory amino acid transporter 1 (GLAST) via the activation of the α7 nicotinic acetylcholine receptor.

Although the clearance of glutamate from the synapse under physiological conditions is performed by astrocytic glutamate transporters, their expression might be diminished under pathological conditions. Microglia glutamate transporters, however, might serve as a back-up system when astrocytic glutamate uptake is impaired, and could have a prominent neuroprotective function under pathological conditions. In the current study, the effect of nicotine, well known as a neuroprotective molecule, on the function of glutamate transporters in cultured rat cortical microglia was examined. Reverse transcription polymerase chain reaction and pharmacological approaches demonstrated that, glutamate/aspartate transporter (GLAST), not glutamate transporter 1 (GLT-1), is the major functional glutamate transporter in cultured cortical microglia. Furthermore, the α7 subunit was demonstrated to be the key subunit comprising nicotinic acetylcholine (nACh) receptors in these cells. Treatment of cortical microglia with nicotine led to a significant increase of GLAST mRNA expression and (14)C-glutamate uptake in a concentration- and time-dependent manner, which were markedly inhibited by pretreatment with methyllycaconitine, a selective α7 nACh receptor antagonist. The nicotine-induced expression of GLAST mRNA and protein is mediated through an inositol trisphosphate (IP3) and Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) depend intracellular pathway, since pretreatment with either xestospongin C, an IP3 receptor antagonist, or KN-93, a CaMKII inhibitor, blocked GLAST expression. Together, these findings indicate that activation of nACh receptors, specifically those expressing the α7 subunit, on cortical microglia could be a key mechanism of the neuroprotective effect of nACh receptor ligands such as nicotine.

Noradrenergic regulation of period1 expression in spinal astrocytes is involved in protein kinase A, c-Jun N-terminal kinase and extracellular signal-regulated kinase activation mediated by α1- and ß2-adrenoceptors.

Our recent data suggest that noradrenaline (NA) regulates expression of Per1 mRNA in rat C6 cells, as a model of brain astrocytes, by two distinct NA-mediating pathways. Although C6 cells possess potential astrocyte-type characteristics, we hypothesize that astrocytes located in a distinct tissue or organ play specific roles consistent with their own unique functions in response to the surrounding environment. We have herein found in primary rat spinal astrocytes using real-time RT-PCR that NA induced robust transient increases in Per1, Cry1, Cry2 and Bmal1 mRNA expression. Cry1, Cry2 and Bmal1 expressions induced by NA were attenuated by transfection of Per1 small interference RNA (siRNA). The effect of NA on Per1 expression was partially blocked by either prazosin (a selective antagonist of α1-adrenoceptor) or ICI118551 (a selective antagonist of ß2-adrenoceptor), and completely blocked by the combination of both antagonists. Treatment with H89 (a protein kinase A [PKA] inhibitor), SP600125 (a c-Jun N-terminal kinase [JNK] inhibitor), or PD98059 (an extracellular signal-regulated kinase [ERK] inhibitor), partially inhibited NA-induced Per1 mRNA expression, and the combination of these three inhibitors inhibited expression to nearly a non-stimulated level. Furthermore, NA phosphorylated not only ERK but also JNK1, an effect that was detected by western blotting. These actions were inhibited only by prazosin, and not by ICI118551. In addition, we found that NA induced phosphorylation of transcription-related proteins such as cAMP response element binding protein (CREB) and c-Jun. These phosphorylation processes were regulated through distinct pathways: CREB phosphorylation was dependent on the PKA and JNK pathways but c-Jun phosphorylation was mediated by the ERK and JNK pathways. These results suggest that Per1 plays a key role in noradrenergic regulation on clock gene expression in spinal astrocytes and activation of α1 and ß2 adrenoceptors are of importance in regulation of Per1 mRNA expression via PKA/JNK-CREB and ERK/JNK-c-Jun cascades.

Down-regulation of norepinephrine transporter expression on membrane surface induced by chronic administration of desipramine and the antagonism by co-administration of local anesthetics in mice.

We have previously shown that chronic administration of the antidepressant desipramine, a norepinephrine transporter (NET) inhibitor to mice markedly enhanced convulsions induced by local anesthetics and that behavioral sensitization may be relevant to decreased [(3)H]norepinephrine uptake by the isolated hippocampus. The co-administration of local anesthetics with desipramine reversed the behavioral sensitization and down-regulation of NET function induced by desipramine. The present study aimed to elucidate whether chronic treatment with desipramine regulates the expression of NET protein examined in membrane fractions in various brain regions and whether co-administration of local anesthetics affects the desipramine-induced alteration of NET expression. Desipramine with or without local anesthetics was injected intraperitoneally once a day for 5 days. The animals were decapitated 48 h after the last administration of drugs and the whole cell fraction, membrane fraction and cell-surface protein fraction were prepared. [(3)H]nisoxetine binding was significantly reduced in the P2 fraction of the hippocampus by chronic administration of desipramine, and the reduction was overcome by co-administration of lidocaine with desipramine. Immunoreactive NET was detected by SDS-PAGE and immunoblotting in the murine hippocampus. NET protein expression in the whole cell fraction and membrane fraction was not affected by treatment with any drugs. However, administration of desipramine significantly reduced the amount of immunoreactive NET in the cell-surface protein fraction. This reduction was blocked by simultaneous injection of lidocaine, bupivacaine or tricaine. These results indicate that the NET down-regulation indicated by the reduction of [(3)H]nisoxetine binding was induced by administration of desipramine via decrease of NET localization on the cell surface. The antagonistic actions of local anesthetics against NET down-regulation by desipramine were related to alterations of the cell-surface localization of NET.

Transport of dopamine and levodopa and their interaction in COS-7 cells heterologously expressing monoamine neurotransmitter transporters and in monoaminergic cell lines PC12 and SK-N-SH.

The present study investigated the effects of levodopa, a precursor of dopamine (DA) therapeutically used for the treatment of Parkinson's disease, on DA transport in the two different systems, COS-7 cells heterologously expressing rat monoamine transporter cDNA and in monoaminergic cell lines PC12 and SK-N-SH. Levodopa enhanced uptake of [3H]DA and [3H]norepinephrine (NE) but not [3H]serotonin in the transfected COS-7 cells in a concentration-dependent manner. On the other hand, in PC12 and SK-N-SH cells where NET is functionally expressed, levodopa enhanced [3H]DA and [3H]NE uptake at low concentrations and inhibited the uptake at higher concentrations. The effects of levodopa on catecholamine transporters in the opposite direction suggest a different mechanism at the intra- and extracellular sites in a levodopa transport-dependent and independent manner.

Nitric oxide synergistically potentiates interleukin-1 beta-induced increase of cyclooxygenase-2 mRNA levels, resulting in the facilitation of substance P release from primary afferent neurons: involvement of cGMP-independent mechanisms.

We previously demonstrated that cultured rat dorsal root ganglion (DRG) cells respond to stimulation with interleukin-1 beta (IL-1 beta) by releasing substance P (SP), and this response is regulated via the cyclooxygenase (COX)-2 pathway. In this study, to ascertain the interaction between nitric oxide (NO) and prostaglandins in primary afferent neurons, we investigated the effect of NO on the IL-1 beta-induced release of SP in cultured DRG cells. An NO donor, S-nitroso-N-acetyl-DL-penicillamine (SNAP), did not in itself evoke SP release. However, it potentiated the IL-1 beta-induced release of SP. Similarly, while SNAP did not elicit the expression of COX-2 mRNA, it potentiated the expression induced by IL-1 beta in cultured DRG cells, and this potentiation was significantly suppressed by the NO scavenger, 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl 3-oxide (carboxy-PTIO). Moreover, SNAP also potentiated the expression of COX-2 protein induced by IL-1 beta in cultured DRG cells. The stimulatory effect of SNAP on the IL-1 beta-induced release of SP was completely inhibited on co-incubation with a selective COX-2 inhibitor, NS-398. 1H-[1,2,4]oxadiazolo[4,3-a]quinoxaline-1-one (ODQ), a potent inhibitor of soluble guanylate cyclase, did not suppress, and a membrane-permeable cGMP analogue, 8-Br-cGMP, did not mimic the stimulatory effects of SNAP in DRG cells. These results suggest that in cultured DRG cells, NO potentiates the IL-1 beta-induced increase in COX-2 expression via a soluble guanylate cyclase-cGMP-independent pathway, resulting in facilitation of SP release. The interaction between NO and COX in primary afferent neurons might contribute to the change in nociceptive perception in inflammatory hyperalgesia.

Buspirone and meperidine synergistically reduce the shivering threshold.

UNLABELLED: Mild hypothermia (i.e., 34 degrees C) may prove therapeutic for patients with stroke, but it usually provokes shivering. We tested the hypothesis that the combination of buspirone (a serotonin 1A partial agonist) and meperidine synergistically reduces the shivering threshold (triggering tympanic membrane temperature) to at least 34 degrees C while producing little sedation or respiratory depression. Eight volunteers each participated on four randomly-assigned days: 1) large-dose oral buspirone (60 mg); 2) large-dose IV meperidine (target plasma concentration of 0.8 microg/mL); 3) the combination of buspirone (30 mg) and meperidine (0.4 microg/mL); and 4) a control day without drugs. Core hypothermia was induced by infusion of lactated Ringer's solution at 4 degrees C. The control shivering threshold was 35.7 degrees C +/- 0.2 degrees C. The threshold was 35.0 degrees C +/- 0.8 degrees C during large-dose buspirone and 33.4 degrees C +/- 0.3 degrees C during large-dose meperidine. The threshold during the combination of the two drugs was 33.4 degrees C +/- 0.7 degrees C. There was minimal sedation on the buspirone and combination days and mild sedation on the large-dose meperidine day. End-tidal PCO2 increased approximately 10 mm Hg with meperidine alone. Buspirone alone slightly reduced the shivering threshold. The combination of small-dose buspirone and small-dose meperidine acted synergistically to reduce the shivering threshold while causing little sedation or respiratory toxicity. IMPLICATIONS: Mild hypothermia may be an effective treatment for acute stroke, but it usually triggers shivering, which could be harmful. Our results indicate that the combination of small-dose buspirone and small-dose meperidine acts synergistically to reduce the shivering threshold while causing little sedation or respiratory toxicity. This combination may facilitate the induction of therapeutic hypothermia in stroke victims.

Usefulness of selective myocardial contrast echocardiography in percutaneous transluminal septal myocardial ablation: a case report.

A 67-year-old woman with hypertrophic obstructive cardiomyopathy that was refractory to medical treatment underwent percutaneous transluminal septal myocardial ablation (PTSMA). The septal branch supplying the myocardium involved in the left ventricular outflow tract (LVOT) obstruction was identified by selective myocardial contrast echocardiography (MCE). MCE for the third and largest septal branch opacified the right side of the mid-septal myocardium and MCE for the second septal branch opacified the right side of the basal portion of the septal myocardium. Finally, contrast agent was injected into the first, small branch, which opacificied the myocardium protruding into the LVOT. Subsequently, septal myocardial ablation for this vessel with intracoronary alcohol was performed, followed by a reduction of the LVOT gradient and successful, dramatic improvement in the patient's clinical condition. Selective MCE was very useful to identify the appropriate septal branch for PTSMA and enabled maximal effect of this treatment with minimal myocardial damage.

[Neural-immune interactions in dorsal root ganglia].

Interleukin-1 (IL-1) beta is a proinflammatory cytokine that is produced by a large variety of cells, including macrophages, fibroblasts, mesangial cells and endothelial cells, and is believed to play important roles in the inflammatory responses, including hyperalgesia. Hyperalgesia is characterized by intensified pain with a reduced threshold to somatic stimulation, and it is involved in chronic inflammatory disease. Substance P (SP), an undecapeptide, has been shown to relay noxious signals as a neurotransmitter in primary afferent neurons. Thus it is expected that the change of neuropeptide activities in primary afferent neurons is attributed to inflammatory hyperalgesia by IL-1 beta. In our recent studies, IL-1 beta was found to stimulate SP release from cultured dorsal root ganglion cells via the cyclooxygenase system. These studies provide a new insight in the neural-immune intercommunication involved in the pain-regulation system observed in inflammation-induced hyperalgesia.

Changes in nocturnal melatonin secretion in perimenopausal women: correlation with endogenous estrogen concentrations.

Although age-related decrease in melatonin secretion in humans and animals is well documented, there is a paucity of data on the precise changes in melatonin secretion that occur during the perimenopausal period. The present study was designed to measure changes in nocturnal melatonin and to characterize the role played by estrogen in controlling nocturnal melatonin secretion in perimenopausal women. Nocturnal serum melatonin concentrations were determined every 2 hr in 46 premenopausal women, 44 postmenopausal women, and 11 premenopausal women with uterine leiomyoma scheduled for hysterectomy and bilateral salpingo-oophorectomy. Nocturnal serum melatonin secretion in premenopausal women declined moderately from 17 to 45 years of age, and increased during the period from 46 to 50 years of age. Among postmenopausal women, a steep, age-related decline in nocturnal melatonin secretion was found for up to 15 years postmenopause, followed by an extremely gradual decline thereafter. A significant negative correlation was observed between the peak serum melatonin concentration and the serum 17 beta-estradiol concentration in premenopausal women aged 40-50 years (r = -0.661, P<0.0005). Daily oral administration of conjugated estrogen (0.625 mg) to postmenopausal women suppressed nocturnal melatonin secretion (P<0.005). A low estrogen state, induced by oophorectomy of premenopausal women with uterine leiomyoma led to an increase in nocturnal melatonin secretion (P<0.0001). Our findings suggest that transient elevated nocturnal melatonin secretion during menopause may be related to the existence of a low estrogen environment. The age-related decrease in melatonin secretion observed in other conditions is most likely attributable to other age-related factors.

Effect of food deprivation and leptin repletion on the plasma levels of estrogen (E2) and NADPH-d reactivity in the ventromedial and arcuate nuclei of the hypothalamus in the female rats.

The exact role of leptin in fasting has not been completely elucidated. To determine whether leptin can act in fasting to influence plasma estrogen levels and nitric oxide synthase reactivity in food regulating centers of the brain, we fasted female rats for 4 days and treated them i.p. with vehicle or 100 microg of recombinant mouse leptin as 1 ml on the 3rd and 4th day twice daily (10.00 and 17.00 h). Proestrus blood was collected at 10.00, 14.00, 18.00 and at 22.00 h, plasma obtained and assayed for estrogen (E2) and leptin levels. Verification of ovulation occurrence was by examining the oviduct for extruded ovum. The rat brains were removed and processed for nitric oxide synthase reactivity in the ventromedial hypothalamus (VMH) and arcuate nucleus (ARC) using NADPH-diaphorase histochemistry, a marker for neurons expressing NOS enzyme. Leptin effect on dependable variables such as food intake, water intake and body weight gain was also investigated. Four days fasting significantly decreased body weight, estrogen and postfast leptin levels, nitric oxide reactivity in the VMH and ARC nucleus and stopped ovulation in many (4 out of 5) rats fasted and given vehicle. Leptin treatment significantly increased plasma estrogen and postfast leptin levels, restored ovulation in many (4 out of 5) rats and increased nitric oxide reactivity in the VMH and ARC. Leptin significantly inhibited food intake, water intake and gain in body weight during recommenced feeding. These observations suggest that leptin could act in the pituitary-ovarian axis during fasting to improve reproductive function by partly stimulating estrogen secretion.

Melatonin protects against age-related DNA damage in the brains of female senescence-accelerated mice.

We investigated whether melatonin reduces the age-related susceptibility of brain to oxidative DNA damage. Brain tissues and blood samples were obtained in the middle of dark period of the daily light:dark cycle from female senescence-accelerated mice (SAM-P/6) at ages 4, 8, and 12 months. Serum melatonin concentrations and the contents of deoxyguanosine (dG) and 8-hydroxydeoxyguanosine (8-OHdG) in DNA extracted from these brain homogenates were measured by high-performance liquid chromatography. Contents of 8-OHdG showed a significant age-related increase (P < 0.001) while that of dG did not. The 8-OHdG:dG ratio also exhibited a significant age-related increase (P < 0.001). Serum melatonin concentration decreased markedly between 8 (159.7 +/- 4.5 pg/mL) and 12 (46.8 + 4.5 pg/mL) months of age (P < 0.0001). Oral melatonin administration (2 microg/mL in water) starting at 8 months of age, which produced a significant increase in serum melatonin concentration at 12 months (187.6 +/- 18.3 pg/mL) compared with untreated animals (P < 0.0001) also resulted in significant decreases in brain 8-OHdG contents and 8-OHdG:dG ratios. These results indicate that administration of a physiologic dose of melatonin to SAM-P 6 mice may prevent the age-related oxidative DNA damage in the brain.

Interleukin-1beta induces substance P release from primary afferent neurons through the cyclooxygenase-2 system.

Substance P (SP) is synthesized in the dorsal root ganglion (DRG) and released from primary afferent neurons to convey information regarding noxious stimuli. The effects of the proinflammatory cytokine interleukin-1 (IL-1) beta on the release of SP were investigated using primary cultured rat DRG cells. Recombinant mouse IL-1beta added to the cells at 0.1 ng/ml increased the SP-like immunoreactivity (SPLI) in the culture medium after incubation for 6 h by approximately 50% as compared with that of nontreated DRG cells. The effect of IL-1beta was Ca(2+)-dependent and significantly inhibited by 100 ng/ml IL-1 receptor-specific antagonist (IL-1r antagonist), cyclooxygenase (COX) inhibitors such as 0.1 mM aspirin, 1 microg/ml indomethacin, and 1 microM NS-398 (specific for COX-2), and 1 microM dexamethasone. Furthermore, a 1-h incubation with IL-1beta markedly increased the inducible COX-2 mRNA level, which was inhibited by an IL-1r antagonist and dexamethasone, whereas IL-1beta showed no effect on the level of constitutive COX-1 mRNA. These observations indicated that IL-1beta induced the release of SP from the DRG cells via specific IL-1 receptors, the mechanism of which might involve prostanoid systems produced by COX-2. This could be responsible for the hyperalgesic action with reference to inflammatory pain in the primary afferent neuron to spinal cord pathway.

Changes in nocturnal pineal melatonin synthesis during the perimenopausal period: relation to estrogen levels in female rats.

To evaluate changes in melatonin synthesis during the perimenopausal period in the female rat and to determine the effects of estrogen on melatonin synthesis, pineal levels of tryptophan, melatonin and norepinephrine and activities of N-acetyltransferase (NAT) and hydroxyindole-O-methyltransferase (HIOMT) were determined. Homogenates for assay were prepared from the pineal glands of female virgin Sprague-Dawley rats between 4 and 24 months of age in the middle of the dark period of a daily light/dark cycle. Serum 17 beta-estradiol (E2) concentrations were also determined. Pineal melatonin levels significantly decreased from month 4 12 and significantly increased from month 12 16, decreasing thereafter. Serum E2 concentrations significantly decreased from month 12-16, and remained low thereafter. No significant changes in tryptophan or norepinephrine were seen. NAT activities paralleled the time course of changes in melatonin. HIOMT activities decreased gradually from month 4 24. Subcutaneous implantation of an E2 capsule between months 12 and 16 resulted in significant decreases in levels of melatonin and NAT activity at month 16. Ovariectomy at month 4 or 12 led to significant increases in the levels of melatonin and NAT activity. These findings represent a temporal increase in pineal melatonin synthesis during the perimenopausal period, and suggest that the increase in melatonin synthesis activity at that time might result from decreasing levels of endogenous estrogen. The effect of estrogen on melatonin synthesis appeared to involve modulation of NAT activity.

Melatonin inhibits the vasorelaxant action of peroxynitrite in human umbilical artery.

We evaluated the antioxidant property of melatonin as it relates to the vasorelaxant effect of peroxynitrite (ONOO ), a reaction product of superoxide anion radical (O2(-*)) and nitric oxide (NO), on the human umbilical artery. Helical sections of umbilical arteries were obtained from human placentas at elective cesarean delivery between weeks 37 and 39 of gestation. Changes in maximal tension induced by potassium chloride were measured in arterial sections with intact endothelium. Sections were treated with 3-morpholinosydomine (SIN-1), which releases O2(-*) and NO simultaneously, with or without pre-treatment either with hemoglobin (3 microM) or melatonin (0.1-10 microM). SIN-1 produced a significant dose-dependent relaxation of vascular tension. Pre-treatment with hemoglobin did not affect SIN-1-induced relaxation. Melatonin significantly reduced the vasorelaxant effect of SIN-1 in a concentration-dependent manner. These findings indicate that ONOO attenuates vascular tension in the human umbilical artery. Melatonin significantly suppressed the vasorelaxant effect of SIN-1, possibly due to its ability to scavenge ONOO-.

[PARAGRAPH as a neuromuscular blockade monitor].

To assess the clinical performance of a new neuromuscular blockade monitor "PARAGRAPH", we studied adults and pediatric patients during general anesthesia. PARAGRAPH enables anesthesiologists to perform various types of assessment of neuromuscular blockade, including TOF, DBS and single twitch even with children. The most remarkable point is its capability for analysis and printing by personal computer. We conclude this device is portable and easy to use, as well as a clinically reliable neuromuscular blockade monitor.

Coronary vasospasm at the site of myocardial bridge--report of two cases.

Two patients with angina pectoris and postmyocardial infarction angina due to coronary vasospasm at the site of myocardial bridge are described. Intracoronary injection of isosorbide dinitrate led to resolution of coronary vasospasm on acetylcholine provocation test, and vasospastic angina pectoris has been well controlled after treatment with calcium channel blockers.

Estrogen modulates the nocturnal synthesis of melatonin in peripubertal female rats.

Our objective was to evaluate the effects of estrogen deficit and of estrogen stimulation on the synthesis of pineal melatonin in female rats during the peripubertal period. The levels of melatonin and N-acetylserotonin (NAS) and the activities of N-acetyltransferase (NAT) and hydroxyindole-O-methyltransferase (HIOMT) were determined in homogenates of pineal glands obtained from peripubertal female Sprague-Dawley rats 4 to 12 weeks of age in the mid-dark during the daily light/dark cycle. Animals were ovariectomized at 4 weeks of age; daily administration of estradiol benzoate (E2B, 1.0 microg/d, s.c.) was initiated at 4 weeks of age. A peak in the pineal levels of melatonin and NAS and in NAT activity was observed in untreated (control) rats with intact ovaries at 6 weeks. HIOMT activity increased from Week 4 to 6 and remained unchanged thereafter. Ovariectomy at Week 4 led to significant increases in the levels of melatonin and of NAS and NAT in activity at Week 8. NAT activity Week 10 resembled that of control animals, but levels of melatonin and NAS were slightly elevated. Ovariectomy did not affect HIOMT activity. Subcutaneous injection of E2B significantly decreased the levels of melatonin and NAS and of NAT activity at Week 4, as compared with those in control rats. E2B suppressed the ovariectomy-induced elevation of levels of melatonin and NAS and of NAT activity, similar to the effect in control animals. E2B did not affect HIOMT activity. Our results suggest that estrogen modulates the nocturnal synthesis of melatonin in the pineal gland in peripubertal female rats. The effects of estrogen on melatonin synthesis appeared to be mediated by the modulation of NAT activity.