Behavior of male and female mice submitted to action of <Emphasis Type="Italic">p</Emphasis>-chlorophenylalanine in prenatal ontogenesis

Effect of serotonin (5-HT) deficit produced by administration of p-chlorophenylalanine at a dose of 400 mg/kg to pregnant female mice at the 8th day of gestation on the subsequent behavior of their offspring (hybrids F1(C57BL/CBA)) was studied. The 5-HT deficit in prenatal ontogenesis leads to the following behavioral changes: (1) females and males of the experimental group have a higher level of the explorative activity in the “open field” than control animals; (2) in females of the experimental group at the age of 90 days, unlike control females and males of experimental and control groups, the explorative activity is extinguished at the threefold testing in the “open field”; (3) females of the experimental group have a decreased level of anxiety in tests “elevated plus-maze” and the “dark-light” chamber. Males of the experimental group, on the contrary, have an elevated level of anxiety. The obtained data show that the 5-HT deficit at the prenatal period affects various aspects of behavior. The degree of the changes produced by the prenatal 5-HT deficit can have different manifestations depending on sex of the animals.     

Effect of serotonin on the formation of neurons producing gonadotropin-releasing hormone in Wistar rats

The effect of serotonin on the formation of neurons producing gonadotropin-releasing hormone (GnRH) during embryogenesis of Wistar rats was studied. The neurons producing GnRH were detected immunocytochemically on days 18 and 21 of embryonic development and on day 15 of postnatal development of rats with normal serotonin metabolism and rats in which the synthesis of serotonin was inhibited by p-chlorophenylalanine. The total number of GnRH neurons in serotonin deficiency was larger than in the case of its normal metabolism at all developmental stages studied. This is an indirect evidence for the inhibitory effect of serotonin on the formation of GnRH neurons. To confirm the morphogenetic effect of serotonin, we studied the rate of formation of GnRH neurons by injecting bromodeoxyuridine in the formation period of these neurons. It was found that serotonin deficiency had no effect on the time of formation of GnRH neurons: over 97% of neurons formed on days 11 to 15 of embryonic development both in the experimental and control groups. Note that, in serotonin deficiency, the maximum number of GnRH neurons formed one day later than in the normal state. Thus, serotonin inhibits the proliferation of GnRH neuron progenitor cells and thereby has a morphogenetic effect on the development of these neurons.     

Biosynthesis of serotonin in Raphé nuclei of rat brain: effect of p-chlorophenylalanine

The activity of tryptophan hydroxylase (EC 1.99.1.4) in the region of the raphé nuclei of rat brain was higher than that of any other brain area. The content of serotonin and the rate of serotonin synthesis were also highest in the raphé nuclei. Following the administration of p-chlorophenylalanine the injection of tryptophan and pargyline increased the content of serotonin in the region of the raphé nuclei of rat brain. The results suggest that the raphé nuclei retained the capacity to hydroxyl-late tryptophan to some extent after the injection of p-chlorophenylalanine.     

Effect of Growth Conditions on Serotonin Content of Tetrahymena pyriformis*

SYNOPSIS The serotonin content of Tetrahymena was measured under several growth conditions and after exposure to drugs. Serotonin was maximal during stationary phase, then declined to a level typical of logarithmically growing cells. Addition of 5-hydroxytryptophan to the culture increased cellular serotonin content, but neither L-tryptophan, reserpine, p-chlorophenylalanine, nor desmethylimipramine altered the serotonin content.     

LHRH-like system in the brain of Xenopus laevis daud

Summary Rabbit antiserum to synthetic LHRH was used with the immunofluorescence technique to identify the LHRH-secreting neurons and their axonal pathways in the brain of Xenopus laevis. Three groups of immunoreactive neurons were identified: the first, in the telencephalon, is a paired group of cells scattered near the two telencephalic ventricles; the second group lies near the preoptic recess; the third group occurs in the ventral wall of the infundibulum. Two principal neuronal pathways were observed: Fibres originating from the dorsally located telencephalic neurons converge on the cephalic median plane where they form a single bundle behind the telencephalic furrow. This bundle descends towards the anterior border of the preoptic recess where it divides into two nerve bundles which pass on either side of the preoptic recess, run above the optic chiasma then cross the infundibular floor and finally terminate in the median eminence. The second pathway is more direct. The more ventrally located telencephalic LHRH cells give rise to this second pathway. Their axons converge with the other LHRH fibres near the lateral border of the preoptic recess. Most of the LHRH nerve fibres terminate in the median eminence although some terminate near the paired pars tuberalis. No reaction was observed after the use of antiserum absorbed with synthetic antigen.Equipe de Recherche associée C.N.R.S. n 492. This work was financed by the D.G.R.S.T., Contract n 7470046     

Migration and differentiation of neurons producing gonadotropin-releasing hormone under conditions of serotonin excess in the brain of mouse embryos

The work has been carried out on mice of the Tg8 line with knockout of gene of monoamineoxidase A with an increase of serotonin and noradrenaline content in the brain, and on mice of the C3H line with unchanged genome and normal concentration of monoamines. An immunocytochemical study has been performed of development of neurons producing gonadotropin-releasing hormone (GnRH) under conditions of excess of serotonin and noradrenaline in the mice in embryogenesis. The GnRH-neurons were revealed at the 18th day of embryonic development in telencephalon along trajectory of their migration from olfactory bulbs to the retrochiasmatic area. In telencephalon of mouse embryos of the Tg8 line, a redistribution of the GnRH-neurons along their migration trajectory was observed as compared with embryos of the C3H line mice. The percent of the GnRH-neurons in the Tg8 mouse embryos in caudal parts of the migration trajectory was lower than in rostral parts, the opposite distribution of the neurons being observed in the C3H line mouse embryos; at the excess of serotonin and noradrenaline in the Tg8 line mouse embryos, the total amount of GnRH-neurons in the brain was lower than in the C3H mice. In males of the Tg8 line mice under conditions of excess of serotonin and noradrenaline the optical density of neurons, which correlated with the GnRH concentration in the cell, was higher than in control mice. Thus, in the Tg8 mice under conditions of the serotonin and noradrenaline excess, migration of the GnRH-neurons to their final anlage in hypothalamus is accelerated as well as the total number of the GnRH-neurons decreases, which indicates a decrease of proliferation of cells-precursors and the earlier differentiation of neurons.     

Migration and differentiation of gonadotropin-releasing hormone-producing neurons in the brain of mouse fetus exposed to excess of serotonin

The work has been carried out on mice of the Tg8 line with knockout of gene of monoamineoxidase A with an increase of serotonin and noradrenaline content in the brain, and on mice of the C3H line with unchanged genome and normal concentration of monoamines. An immunocytochemical study has been performed of development of neurons producing gonadotropin-releasing hormone (GnRH) under conditions of excess of serotonin and noradrenaline in the mice in embryogenesis. The GnRH-neurons were revealed at the 18th day of embryonic development in telencephalon along trajectory of their migration from olfactory bulbs to the retrochiasmatic area. In telencephalon of mouse embryos of the Tg8 line, a redistribution of the GnRH-neurons along their migration trajectory was observed as compared with embryos of the C3H line mice. The percent of the GnRH-neurons in the Tg8 mouse embryos in caudal parts of the migration trajectory was lower than in rostral parts, the opposite distribution of the neurons being observed in the C3H line mouse embryos; at the excess of serotonin and noradrenaline in the Tg8 line mouse embryos, the total amount of GnRH-neurons in the brain was lower than in the C3H mice. In males of the Tg8 line mice under conditions of excess of serotonin and noradrenaline the optical density of neurons, which correlated with the GnRH concentration in the cell, was higher than in control mice. Thus, in the Tg8 mice under conditions of the serotonin and noradrenaline excess, migration of the GnRH-neurons to their final anlage in hypothalamus is accelerated as well as the total number of the GnRH-neurons decreases, which indicates a decrease of proliferation of cells-precursors and the earlier differentiation of neurons.     

Gonadal steroids affect LHRH preoptic cell number in a sex/role changing fish

In diandric sex-reversing fishes, sexually active males and females (primary phase) regularly transform into an alternative reproductive morph, terminal-phase males, that are morphologically and behaviorally distinct. The transformation from primary to terminal phase is associated with a twofold increase in the number of luteinizing hormone-releasing hormone (LHRH) immunopositive cells in the forebrain preoptic area, a region involved in both the initial development and daily control of reproductive physiology and behavior. We now show that implants of 11-ketotestosterone induce increases in LHRH cell number in both primary phase sexes to the level observed in field-collected terminal phase males. Conversely, gonadal steroids had no effect on the number of LHRH preoptic cells in terminal phase males, suggesting that this is indeed a terminal stage in the development of this species. These results demonstrate that transition to the terminal phase by both sexes involves a parallel and convergent change in LHRH cell number, which utilizes an evolutionarily conserved mechanism of sexual differentiation: the inductive effects of gonadal steroid hormones.     

Improved Production of Human Immunoglobulin γ1 Chain Fc Polypeptides in Escherichia coli and Their Properties

Exposure to some xenobiotics (pentobarbital, 3-terf-butyl-4-methoxyphenol (BHA), chloretone (acetone chloroform), 1, l-bis-(p-chlorophenyl)-2,2,2-trichloroethane (DDT) and polychlorinated biphenyls (PCB)) for a 5 hr period increased the concentrations of brain serotonin and 5-hydroxyindole acetic acid (5HIAA). The decrease in the brain serotonin level elicited by /7-chlorophenylalanine (PCPA), an inhibitor of serotonin synthesis, was prevented by the concomitant administration of chloretone. The administration of both chloretone and pargyline (an inhibitor of monoamine oxidase) caused significant elevation of the brain 5HIAA level as compared with that in a pargyline control, however, the concentration of brain serotonin was not different between pargyline alone and chloretone plus pargyline. These results show that the increase in the brain serotonin level caused by chloretone is not due to acceleration of brain serotonin synthesis, but to retardation of the degradation of brain serotonin, and the increase in brain 5HIAA caused by chloretone may be due to the reduced removal of 5HIAA from the brain. Chloretone plus pargyline caused significant elevation of hypothalamus catecholamines, as compared to in the pargyline control, so the catecholamine turnover rates may be accelerated by the administration of chloretone.     

Early development of an identified serotonergic neuron in Helisoma trivolvis embryos: Serotonin expression,de-expression,and uptake

In early-stage embryos of Helisoma trivolvis, a bilateral pair of identified neurons (ENC1) express serotonin and project primary descending neurites that ramify in the pedal region of the embryo prior to the formation of central ganglia. Pharmacological studies suggest that serotonin released from ENC1 acts in an autoregulatory pathway to regulate its own neurite branching and in a paracrine or synaptic pathway to regulate the activity of pedal ciliary cells. In the present study, several key features of early ENC1 development were characterized as a necessary foundation for further experimental studies on the mechanisms underlying ENC1 development and its physiological role during embryogenesis. ENC1 morphology was determined by confocal microscopy of serotonin-immunostained embryos and by differential-interference contrast (DIC) microscopy of live embryos. The soma was located at an anteriolateral superficial position and contained several distinguishing features, including a large spherical nucleus with prominent central nucleolus, large granules in the apical cytoplasm, a broad apical dendrite ending in a sensory-like structure at the embryonic surface, and a ventral neurite. ENC1 first expressed serotonin immunoreactivity around stage E13, followed immediately by the appearance of an immunoreactive neurite (stage E14). Both the intensity of immunoreactivity and primary neurite length were consistently greater in the right ENC1 at early stages. Serotonin uptake, as indicated by 5,7-dihydroxytryptamine-induced fluorescence, first occurred between stages E18 and E25. At later stages of embryogenesis (after stage E65), serotonin immunoreactivity disappeared, whereas serotonin uptake and normal cell morphology were retained. © 1998 John Wiley & Sons, Inc. J Neurobiol 34: 361–376, 1998     

Effect of estrogen administration on endogenous and luteinizing hormone-releasing-hormone-induced luteinizing hormone secretion and follicular development in the lactating sow

The objectives of this study were to investigate whether estradiol treatment during lactation modifies 1) the patterns of endogenous LH, FSH, and prolactin (PRL) release; 2) the sensitivity of the pituitary to exogenous injections of LHRH; and 3) the responsiveness of the ovarian follicles to gonadotropin. Plasma LH, FSH, and PRL were determined in samples taken repeatedly from 18 sows on Days 24-27 of lactation. Ovaries were then recovered, and follicular development was assessed by measuring the follicular diameter (FFD) and follicular fluid estradiol-17 beta concentration (FFE) of the ten largest follicles dissected from each ovary. Sows were randomly allocated to one of four treatments: 1) Group C (4 sows) received no treatment; 2) Group LHRH (5 sows) received 800 ng of LHRH every 2 h throughout the sampling period; 3) Group E2 (4 sows) received subcutaneous implants containing estradiol-17 beta 24 h after start of sampling; 4) Group LHRH + E2 (5 sows) were administered a combination of LHRH and estradiol-17 beta implants. Between-animal variability for plasma LH, FSH, and PRL was considerable. LH concentration and LH pulse frequency increased (p less than 0.05) after LHRH treatment in the LHRH and LHRH + E2 groups; however, an acute inhibition of LH secretion was observed in the latter group immediately after estradiol implant application. In the absence of LHRH treatment, estradiol caused chronic inhibition of LH secretion. Follicular development was greater in the LHRH and LHRH + E2 groups compared to the C and E2 groups (p less than 0.05 for both FFD and FFE).(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects ofp-chlorophenylalanine on cortical monoamines and on the activity of noradrenergic neurons

The catecholamines noradrenaline, dopamine, adrenaline, the indoleamine 5-hydroxy-tryptamine (5-HT; serotonin), and some of their major metabolites were assayed, using high performance liquid chromatography (HPLC), in the neocortex of normal rats as well as in animals in which 5-HT synthesis had been inhibited withp-chlorophenylalanine. Besides important depletions in serotonin and in 5-hydroxyindole-3-acetic acid, noradrenaline levels were significantly reduced, but the content in 3-methoxy-4-hydroxyphenylglycol was increased, indicating an augmented utilization of this amine. The levels of dopamine and 3-methoxytyramine were also reduced, although homovanillic acid and 3,4-dihydroxyphenylacetic acid levels remained constant. The spontaneous unitary activity of identified noradrenergic neurons in the Locus coeruleus was increased, indicating an hyperactivity of this system. These results can be interpreted in relation to functional interactions between the catecholamines and serotonin; i.e.: a decrease in endogenous serotonin results in the loss of a negative feedback control of noradrenaline release.Special Issue dedicated to Prof. Eduardo De Robertis.     

Involvement of the Serotonergic System in Analgesia Induced by the Influence of Low-Intensity Microwaves on an Antipain Acupuncture Point

In experiments on mice, we studied changes in the level of analgesia induced by irradiation of the antipain acupuncture point (AP) E36 by low-intensity microwaves under conditions of modification of the serotonin level in the brain; this level was modified by injection of 300 mg/kg DL-p-chlorophenylalanine (pCPA). The duration of the nociceptive behavioral reaction (licking the limb) caused by injection of the formalin solution into the foot dorsal surface increased 24, 48, and 72 h after pCPA injection by 99.9, 84.4, and 114.4%, as compared with those in animals subjected to microwave irradiation of the AP E36 with no preliminary pCPA injection. It is concluded that the brain serotonergic system is actively involved in the analgesia effects induced by irradiation of the AP by low-intensity microwaves.     

The effect of the entomopathogenic fungus Conidiobolus coronatus on the composition of cuticular and internal lipids of Blatta orientalis females

The composition of cuticular and internal lipids in females of the cockroach Blatta orientalis L. exposed to the entomopathogenic fungus Conidiobolus coronatus is investigated. The compositions of the fatty acids, n‐alkanes, alcohol, sterols and methyl esters in the lipids are chemically characterized. Although contact with virulent colonies of the fungus does not induce insect mortality, significant changes in the lipid profiles, both cuticular and internal, are found. The cuticular extracts of a control group of B. orientalis females contain 24 compounds varying in carbon chain length from C6 to C22. The main cuticular fatty acids identified are: C16:1, C16:0, C18:1 and C18:0_. The cuticular lipids of B. orientalis females after exposure to C. coronatus contain only 14 free fatty acids from C8 to C20. The highest concentrations identified are C16:0, C18:2 and C18:1. Analysis by gas chromatography‐mass spectrometry identifies the presence of a homologous series of n‐alkanes containing from 25 to 31 carbon atoms. In the case of the insects after fungal exposure, the content of the n‐alkanes in the cuticular lipid is two‐fold higher compared with the controls. Of the cuticular lipids, 11 alcohols are found, ranging from C12:0 to C20:0. There is no presence of alcohols in the internal lipids of the control B. orientalis females and in all of the extracts from the B. orientalis females after fungal exposure. In the samples analyzed, the most common sterol is cholesterol. This is present in the cuticular lipids and the internal lipids of all of the insects sampled. The cuticular and internal lipids of females contain five fatty acid methyl esters, ranging in size from C15 to C19.     

Serotonin and norepinephrine

Rats which had been primed with the serotonin depletor,p-chlorophenylalanine, and sacrificed months later were found to have the same resting levels of brain serotonin and norepinephrine as unprimed controls. However, when treated with the monoamine oxidase inhibitor, tranylcypromine, the former showed a significantly lower accumulation of these biogenic amines than their tranylcyprominetreated unprimed counterparts. These findings indicate that brain serotonin, which had been lowered at the outset byp-chlorophenylalanine, had returned to normal levels but that the priming procedure might have resulted in a long-term decrease in the turnover rates of serotonin as well as norepinephrine. Primed animals may prove suitable as models of disturbed biogenic amine metabolism with possible relevance to schizophrenia and other brain dysfunctions.     

Neonatal PACAP administration in rats delays puberty through the influence of the LHRH neuronal system

The onset of puberty is a concerted action of many factors which leads to cyclic LHRH release in rats. It has been demonstrated that; in common with vasoactive intestinal polypeptide (VIP), pituitary adenylate cyclase activating polypeptide (PACAP) is also involved in the differentiation of the central nervous system. In our previous work, it was shown that a single PACAP injection into neonatal female rats delayed puberty. In the present work, neonatal administration of PACAP delayed the vaginal opening and decreased the weight of anterior pituitaries, the number of expelled ova at the first ovulation and the intensity of LHRH immunostaining in the septo-preoptico-infundibular system. PACAP antiserum had a reverse effect on LHRH immunoreactivity. The other studied parameters in the latter group remained unchanged compared to control rats. It was concluded that neonatal PACAP administration delayed the onset of puberty through the influence of the LHRH neuronal system.     

Influence of Catecholamines on Migration and Differentiation of GnRH Neurons in Rats

The GnRH producing neurons are the key link of neuroendocrine regulation of the adult reproductive system. Synthesis and secretion of GnRH are, in turn, under the afferent catecholaminergic control. Taking into account that catecholamines exert morphogenetic effects on target cells during ontogenesis, this study was aimed at investigation of the effects of catecholamines on development of GnRH neurons in rats during ontogenesis. We carried out comparative quantitative and semiquantitative analyses of differentiation and migration of GnRH neurons in fetuses of both sexes under the conditions of normal metabolism of catecholamines (administration of saline) or their pharmacologically induced deficiency (administration of -methyl-para-tyrosine). The inhibition of catecholamine synthesis from day 11 of embryogenesis led to an increasing number of GnRH neurons in rostral regions of the trajectory of their migration over the brain: in the area of olfactory tubercles on day 17 and in the area of olfactory bulb on days 18 and 21. In addition, the optical density of GnRH neurons located in the rostral regions of migration was higher in the fetuses after administration of -methyl-para-tyrosine during embryogenesis, as compared to the control. It has been concluded that catecholamines stimulate the migration of GnRH neurons and affect their differentiation.     

Benefit of Dietary Supplementation with Bacillus subtilis BYS2 on Growth Performance,Immune Response,and Disease Resistance of Broilers

A strain of Bacillus subtilis (B. subtilis) BYS2 was previously isolated from Mount Tai, which is located in Tai’an City in the Shandong Province of China. The strain was then stored in the Environmental Microbiology Laboratory at Shandong Agricultural University. To evaluate the effect of the bacterium preparation in broiler production, we fed the bacterium (10⁶ CFU/g) to 1-day-old broilers and continued this feeding for 6 weeks to analyze its effect on growth and immune performance. We found that the average weight of the bacterium-fed group increased by 17.19% at weeks 5 compared to the control group (P < 0.05). The height of the villi in the duodenum and jejunum and the ratio of villi to crypt were significantly increased in the bacterium-fed group at weeks 5 (P < 0.05). Also, the IgG in the serum of broilers in the experimental group increased by 31.60% (P < 0.05) and IgM 30.52% (P < 0.05) compared with those in the control group. The expressions of the major pattern recognition receptors (PRRs), antiviral proteins, pro-inflammatory cytokines, and β-defensins were significantly higher than those in the control group (P < 0.05). Meanwhile, the bursa immune organ indices of broilers in the experimental group were significantly higher than those in the control group (P < 0.05). Also, after 5 weeks of continuous feeding, when infected with Escherichia coli (E. coli) O1K1 and Newcastle disease virus (NDV) F48E8, the content of bacteria and virus in tissues and organs of the experimental group decreased significantly, and the survival rate of infected chickens increased by 31.1% and 17.7%, respectively (P < 0.05). These results show that the anti-infective B. subtilis BYS2 could, to some extent, replace antibiotics to promote growth, improve innate immunity, and enhance disease resistance in broilers.

     

Changes in populations of LHRH-immunopositive cell bodies following gonadectomy

One day after castration of male rats, plasma LH rose and the number of LHRH immunopositive neuronal perikarya decreased. As plasma LH continued to rise six days and three weeks post-castration, the number of LHRH immunopositive neurons also increased. The largest population of LHRH immunopositive neurons was detected three weeks post-castration and the cell group that showed the greatest increase was in the rostral preoptic area. In females, the largest population of LHRH immunopositive neurons was observed one day post-ovariectomy; at this time plasma LH levels were not significantly elevated above diestrous levels. Six days post-ovariectomy, LH levels were elevated and the number of LHRH immunopositive cells decreased. As LH levels continued to rise three weeks post-ovariectomy, the population increased in size. In males, primarily LHRH cells of the rostral preoptic area increased in in number; in females, the cell groups that increased were scattered over the diagonal band of Broca, preoptic and anterior hypothalamic areas. Although LHRH neurons demonstrated these variations following gonadectomy, there was no evidence of alteration(s) in molecular processing of precursor hormone.