[Endocrine functions of the brain in adult and developing mammals].

The main prerequisite for organism's viability is the maintenance of the internal environment despite changes in the external environment, which is provided by the neuroendocrine control system. The key unit in this system is hypothalamus exerting endocrine effects on certain peripheral organs and anterior pituitary. Physiologically active substances of neuronal origin enter blood vessels in the neurohemal parts of hypothalamus where no blood-brain barrier exists. In other parts of the adult brain, the arrival of physiologically active substances is blocked by the blood-brain barrier. According to the generally accepted concept, the neuroendocrine system formation in ontogeny starts with the maturation of peripheral endocrine glands, which initially function autonomously and then are controlled by the anterior pituitary. The brain is engaged in neuroendocrine control after its maturation completes, which results in a closed control system typical of adult mammals. Since neurons start to secrete physiologically active substances soon after their formation and long before interneuronal connections are formed, these cells are thought to have an effect on brain development as inducers. Considering that there is no blood-brain barrier during this period, we proposed the hypothesis that the developing brain functions as a multipotent endocrine organ. This means that tens of physiologically active substances arrive from the brain to the systemic circulation and have an endocrine effect on the whole body development. Dopamine, serotonin, and gonadotropin-releasing hormone were selected as marker physiologically active substances of cerebral origin to test this hypothesis. In adult animals, they act as neurotransmitters or neuromodulators transmitting information from neuron to neuron as well as neurohormones arriving from the hypothalamus with portal blood to the anterior pituitary. Perinatal rats--before the blood-brain barrier is formed--proved to have equally high concentration of dopamine, serotonin, and gonadotropin-releasing hormone in the systemic circulation as in the adult portal system. After the brain-blood barrier is formed, the blood concentration of dopamine and gonadotropin-releasing hormone drops to zero, which indirectly confirms their cerebral origin. Moreover, the decrease in the blood concentration of dopamine, serotonin, and gonadotropin-releasing hormone before the brain-blood barrier formation after the microsurgical disruption of neurons that synthesize them or inhibition of dopamine and serotonin synthesis in the brain directly confirm their cerebral origin. Before the blood-brain barrier formation, dopamine, serotonin, gonadotropin-releasing hormone, and likely many other physiologically active substances of cerebral origin can have endocrine effects on peripheral target organs--anterior pituitary, gonads, kidney, heart, blood vessels, and the proper brain. Although the period of brain functioning as an endocrine organ is not long, it is crucial for the body development since physiologically active substances exert irreversible effects on the targets as morphogenetic factors during this period. Thus, the developing brain from the neuron formation to the establishment of the blood-brain barrier functions as a multipotent endocrine organ participating in endocrine control of the whole body development.

[Vasopressinergic neurons in rats during ontogenesis: reaction to salt-loading and its modulation by noradrenergic afferents].

Salt-loading in adult mammals stimulates vasopressin secretion by vasopressinergic neurons of the supraoptic nucleus that is under control by a number of hormones and neurotransmitters including noradrenalin. This study was aimed to determine at what period of ontogenesis the vasopressinergic neurons begin to respond to salt-loading and when the noradrenergic control of this process is switched on. Rats on the 21st embryonic day (E), the 3rd postnatal day (P) and P13 were salt-loaded, sometimes under simultaneous treatment with prasozin, an inhibitor of al -adrenoreceptors. Thereafter, the hypothalamic nuclei of the animals were processed for immunocytochemistry and in situ hybridization. Salt-loading provoked increased synthesis of vasopressin mRNA and, most probably, vasopressin itself in rats in all studied age groups. Under salt-loading, the intraneuronal content of vasopressin increased significantly at E21 and P3, whereas it did not change at P13. No change in the intracellular contents of vasopressin mRNA and vasopressin was observed in foetuses following salt-loading and treatment with prasozin though the same treatment provoked an increase of both parameters at P3. These data show that noradrenalin provides an inhibitory control of vasopressin expression at least since P3. Thus, vasopressinergic neurons begin to respond to salt-loading at the since P3. Thus, life by the increased expression of vasopressin that is postnatally under the inhibitory control by noradrenalin.

[The brain is one of the most important sources of dopamine in general circulation during perinatal ontogenesis in rats].

This study was aimed to test our hypothesis that dopamine synthesized in the neurons of the brain is delivered to the general circulation in rats during prenatal and early postnatal periods, i.e. before the establishment of the blood-brain barrier. Using the high performance liquid chromatography, it was demonstrated that the dopamine concentration and content in the peripheral blood in fetuses and neonatal rats (i.e. before the establishment of the blood-brain barrier) greatly exceeded those in adult rats. Moreover, the establishment of the blood-brain barrier was accompanied by the significant increase of the dopamine concentration in the brain. A drop of the dopamine concentration in fetal plasma after the microsurgical lesion of the forebrain and mesencephalon (encephalectomy) are considered as direct evidence in favour of our hypothesis.

[Dopamine synthesis by non-dopaminergic neurons of the rat fetus arquate nucleus]. / Sintez dofamina nedofaminergicheskimi neironami arkuatnogo iadra plodov krys.

Our hypothesis was tested in respect to dopamine synthesis by non-dopaminergic neurons expressing individual complementary enzymes of the DA synthetic pathway. According to the hypothesis, L-dihydroxyphenylalanine (L-DOPA) synthesised in tyrosine hydroxylase(TH)-expressing neurons for conversion to dopamine. The mediobasal hypothalamus of rats on the 21st embryonic day was used as an experimental model. The fetal substantia nigra containing dopaminergic neurons served as control. Dopamine and L-DOPA were measured by high performance liquid chromatography in cell extracts and incubation medium in presence or absence of L-tyrosine. L-tyrosine administration increased L-DOPA synthesis in the mediobasal hypothalamus and substantia nigra. Moreover, L-tyrosine provoked an increase of dopamine synthesis in substantia nigra and a decrease in the mediobasal hypothalamus. This is, probably, due to an L-tyrosine-induced competitive inhibition of the L-DOPA transport to monoenzymatic AADC neurons after its release from the monoenzymatic TH neurons. This study provides a convincing evidence of dopamine synthesis by non-dopaminergic neurons expressing TH or AADC, in cooperation.

[Involvement of accessory neurosecretory nuclei of hypothalamus in the formation of hypothalamohypophyseal system during prenatal and postnatal development in rats]. / Uchastie dopolnitel'nykh neirosekretnykh iader gipotalamo-gipofizarnoi sistemy v pre- i postnatal'nom ontogeneze u krys.

We studied the development of direct axonal connections of the accessory neurosecretory hypothalamic nuclei with the posterior pituitary lobe on the fixed rat brain from day 15 of embryogenesis until day 10 of postnatal development using the retrograde diffusion method of the lipophilic fluorescent carbocyanine dye 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate along the neuronal membranes. The marker was applied onto the posterior pituitary lobe and, after incubation in a fixative, fluorescing bodies of nerve cells were visualized in the hypothalamus. Neuronal axons of the retrochiasmatic nucleus were the first of the accessory nuclei to ingrow in the posterior pituitary lobe (on days 16-17 of embryogenesis). Neurons of the circular and dorsolateral nuclei and the nuclei of the median bundle of the forebrain sent their axons to the posterior pituitary lobe starting from the first days of postnatal development. No direct connections of the anterior commissure and perifornical accessory nuclei with the posterior pituitary lobe were found in perinatal development. These facts are discussed in the light of concepts about the different functional role of accessory peptidergic hypothalamic nuclei in rats.

[Differentiation of dopaminergic neurones in situ, in vitro, and in the transplant]. / Differentsirovka dofaminergicheskikh neironov in situ, in vitro i v transplantate.

Four stages were distinguished in differentiation of the hypothalamic dopaminergic (DA) neurones: a) origin of the neurone, b) expression of enzymes and dopamine specific synthesis and of the dopamine transmembrane transportation mechanisms, c) development of permanent and transient efferent connections, d) forming of afferent innervation and synaptogenesis. The differentiating DA neurones revealed sexual dimorphism in the neurone origin dynamics and in expression of the enzyme synthesis. The maternal and placenta factors did not affect the differentiation of the DA neurones. The period from the 6th to 10th foetal week was found to be optimal for transplantation of the neurones to the striatum of parkinsonian patients. Grafted DA neurones seem to get involved in regulation of the target neurones in the striatum.

[The formation and regulation of the lactotrophic function of the hypophysis in the prenatal period of rat development. II. Dopamine inhibitory control of prolactin secretion]. / Stanovlenie i reguliatsiia laktotrofnoi funktsii gipofiza v prenatal'nom periode razvitiia krys. II. Dofaminovyi ingibitornyi kontrol' sekretsii prolaktina.

Using the technique of radioimmunoassay, we studied the secretion of prolactin and its control by dopaminergic system in 22-day-old rat fetuses under normal conditions and after pharmacological inhibition of dopamine receptors. In order to elucidate the origin of prolactin and dopamine participating in this process, we used decapitation and encephalectomy of fetuses in utero. Decapitation of fetuses did not result in any changes of baseline prolactin secretion into blood in males and insignificantly decreased it in females as compared with nonoperated controls. We conclude that prolactin detected in blood plasma of nonoperated fetuses does not originate in the pituitary, and any prolactin synthesized in the pituitary is not secreted into blood. Inhibition of dopamine receptors in decapitated fetuses did not result in any changes of prolactin level in blood. This provided evidence that in nonoperated fetuses, it is pituitary prolactin which is secreted in response to haloperidol, while the secretion of nonpituitary prolactin is not controlled by dopamine. Encephalectomy increased prolactin level in plasma and resulted in a drastic decrease of its level in the pituitary. The block of dopamine receptors did not affect the level of prolactin in blood plasma or pituitary of encephalectomized fetuses. We conclude that the inhibitory dopaminergic control of prolactin secretion by the pituitary during the prenatal period is accomplished just as in adult animals by dopaminergic neurons of hypothalamus.

[Effects of male sex hormones on specific uptake and release of 3H-serotonin in the rat hypothalamus in vitro]. / Vliianie muzhskikh polovykh gormonov na spetsificheskii zakhvat i vydelenie 3H-serotonina gipotalamusom krysy in vitro.

With the use of "isotopic method" a study was made of the main parameters of functional activity of serotoninergic elements of hypothalamus--the specific uptake and release of 5-OT. The animals used were sexually mature rats castrated on the first postnatal day. In sexually mature intact males the specific uptake of 3H-5-OT by serotoninergic structures of the anterior hypothalamus was significantly lower than in females. Castration of animals on the first day of life resulted in the increase of specific 5-OT uptake in sexually mature males up to that observed in females. There were no differences between the sexes in the rate of spontaneous release of 5-OT. However, response to K(+)-depolarization in the anterior hypothalamus of intact males was significantly lower than that in females. In the hypothalamus of males castrated neonatally the amplitude of the response to the effect of the depolarizing agent was increase up to the level observed in females. By the results obtained it is indicated that elimination of the effect of male hormones on the first postnatal day results in the increase of 5-OT uptake and release in the hypothalamus of sexually mature rat males.

[The formation of the catecholaminergic system of the hypothalamus in rats. Dopamine uptake and release]. / Formirovanie katekholaminergicheskoi sistemy gipotalamusa u krys. Zakhvat i vydelenie dofamina.

The development of the hypothalamic catecholaminergic (CA) system during ontogenesis in rats has been studied with the isotopic biochemical technique in vitro. It has been demonstrated, that at the 15th fetal day, the CA system was functionally inactive at least in its ability for the uptake and K(+)-stimulated release of catecholamines. Since the 16th fetal day, hypothalamic neuronal elements gained an ability for specific uptake and K(+)-stimulated release of the exogenous radioactively labeled dopamine increased significantly. The specific uptake doubled from the 20th fetal till the 9th postnatal day, whereas K(+)-stimulated release doubled between the 9th 45th postnatal days.

[The uptake of 3H-serotonin by neural elements of the rat hypothalamus in ontogeny]. / Zakhvat 3H-serotonina nervnymi élementami gipotalamusa krys v ontogeneze.

Development of serotoninergic system of rat hypothalamus has been studied. The level of its maturity was measured by specific absorption of 3H-serotonin by hypothalamus of fetuses (days 16-20 of intra-uterine development), newborn (9 days after birth) and adult rats. Inhibitors of specific serotonin capture by serotoninergic neural elements (cocaine, fluoxetin and cytalopram) were used in control experiments. Specific uptake of 3H-serotonin by hypothalamus was detected on day 16 of intra-uterine development. It increased twice by day 18 and remained unchanged until birth. No statistically significant increase of 3H-serotonin uptake was detected in newborn and adult animals.

[Ca2+-dependent secretion of serotonin by the rat hypothalamus during ontogenesis]. / Ca2+-zavisimaia sekretsiia serotonina iz gipotalamusa krys v ontogeneze.

A study was made of functional maturity of the terminal part of serotoninergic system of rat hypothalamus in perinatal period: the maturity was estimated by the ability to release serotonin. The release of specifically taken up serotonin (3H-5-OT) by the tissue of hypothalamus of 16-20-day-old rat fetuses, 8-9-day males and adult males was studied in the perfusion system. Spontaneous release of the labelled amine was recorded in the earliest studied period--on the 16th day of the prenatal period, but the response to K+ depolarization was absent at this time. For the first time the increase of the rate of 3H-5-OT release in response to depolarization was noted on the 17th day of development. In the absence of Ca2+ depolarizing stimulus produced no increase in the release of the labelled product. Similar results were obtained with perfusion of fetal hypothalamus on the 18t hand 20th days of development. In neonatal animals the release of 5-OT in response to depolarization was equal to that in adults. The data obtained point to a possible functioning of serotoninergic elements of hypothalamus in the perinatal period in rats.

[Morphofunctional characteristics of serotonin-like neurons in the hypothalamus of rats in ontogeny]. / Morfofunktsional'naia kharakteristika serotoninopodobnykh neironov v gipotalamuse krys v ontogeneze.

An attempt has been made to reveal 5-HT immunopositive (IP) neurones in the hypothalamus of intact foetuses (18th day of gestation) and neonatal (9-day) rats under normal conditions and after their treatment with drugs involved into 5-HT metabolism or into regulation of its uptake by serotoninergic neurones. 5-HTIP cells were not observed in intact animals as well as after L-tryptophan treatment, whereas two large colonies of these neurones were found in the anterio-lateral hypothalamus and dorsomedial nucleus after subsequent injections of monoamine oxidase inhibitor, pargyline, and amino acid precursor of 5-HT synthesis, L-tryptophan. Significantly less intensive reaction was observed after injections of another precursor of 5-HT synthesis, 5-hydroxytryptophan, or pargyline only. Immunostaining evoked by pargyline or L-tryptophan can be prevented by preliminary injections of fluoxetine, a specific inhibitor of 5-HT uptake by serotoninergic neurones. These data suggest that the immunostaining of hypothalamic neurones is due to their capacity to take up specifically 5-HT from the environment rather than to its intraneuronal synthesis from L-tryptophan. However, 5-HT synthesis from 5-hydroxytryptophan in the same cells may also take place. The uptake of extracellular 5-HT by catecholaminergic neurones is absent, since nomifensine, a specific inhibitor of this uptake, does not affect immunostaining.

[Radioautographic study of the hypothalamic serotoninergic structures in the perinatal period of the rat]. / Radioavtograficheskoe issledovanie serotoninergicheskikh struktur gipotalamusa v perinatal'nom periode krysy.

The distribution of 3H-serotonin-binding structures in hypothalamus of 16 and 18 day old fetuses and of 9 day old rats was studied after intraventricular injection of 3H-serotonin. Rare 3H-serotonin-binding little differentiated cells were found predominantly in the intermediate zone of the 3rd ventricle in the retrochiasmatic area wall on the 16th and 18th days of prenatal development. In addition, an aggregate of heavily labeled neurones was observed in the suprachiasmatic area. At the same time 3H-serotonin-binding fibers first appeared, predominantly in the optic chiasma and perichiasmatic area. Radioactively labelled cells, which can be characterized by their morphology as differentiated neurones, were located in the dorsomedial nucleus on the 9th day of postnatal development. The number of serotonin-binding fibers markedly increased but the pattern of their distribution was, on the whole, similar to that in fetuses. The data obtained suggest that the main stages of structural organization of serotoninergic system of hypothalamus in rats are realized during perinatal period.

[Immunocytochemical and autoradiographic research on the growth of serotoninergic fibers to the cerebral ventricles in the perinatal period in rats]. / Immunotsitokhimicheskoe i avtoradiograficheskoe issledovaniia prorastaniia serotoninergicheskikh volokon k zheludochkam mozga v perinatal'nom periode u krys.

Anatomical relationships between serotoninergic (5-HT) fibers and cerebral ventricles were studied in rats from the 16th fetal day until the 9th postnatal day with immunocytochemistry and radioautography. In the latter case, 5-HT neuronal elements were detected according to their specific uptake of intraventricularly injected 3H-5-HT. On the 16th fetal day, occasional 5-HT fibers first spread from the main place of their origin in the raphe nuclei to the dorsocaudal portion of the 3rd ventricle and aqueduct. Two days later, a more extensive network of 5-HT fibers appeared around the dorsal portion of the 3rd ventricle, whereas fibers only rarely penetrated fibers became noticeable in the lateral and 3rd ventricles. The functional significance of hypothalamic and ventricular 5-HT is discussed from the standpoint of its being either a modulator of growth and differentiation of the developing brain, or a factor involved in some specific neuroendocrine functions.