M. V. Ugryumov

Development of central and peripheral serotonin-producing systems in rats in ontogenesis

The work deals with study of development of central and peripheral serotonin-producing systems in rat ontogenesis before and after formation of the blood-brain barrier. By the method of highly efficient liquid chromatography it has been shown that the serotonin level in peripheral blood before formation of the blood-brain barrier (in fetuses and neonatal rats) is sufficiently high for realization of physiological effect on target cells and organs. At the period of formation of the blood-brain barrier the serotonin level in brain sharply rises, whereas the serotonin concentration and amount in blood plasma and duodenum increase insignificantly. Completion of formation of the blood-brain barrier is accompanied by a significant increase of the serotonin content in duodenum, probably for maintenance of the high serotonin level in blood. To evaluate secretory activity, the mean rate of daily serotonin increment in the studied tissues was calculated. In brain, this parameter was maximal at the period of formation of the blood-brain barrier-from the 4th to the 16th postnatal days. This allows thinking hat brain before formation of the blood-brain barrier is the most important source of serotonin in peripheral blood.

Activity of Monoamine Oxidase in the Nigrostriatal System at Presymptomatic and Early Symptomatic Stages of Parkinsonism in Mice.

Activities of monoamine oxidases A and B were examined on the models of presymptomatic and early symptomatic stages of Parkinson’s disease developed in mice treated with MPTP, a specific neurotoxin affecting dopaminergic neurons. Activity of monoamine oxidases A, the key enzyme of dopamine degradation, is increased in neuronal somas during the symptomatic stage, and it is augmented in the axons during both stages. Neuronal activity of monoamine oxidases A is higher during the symptomatic stage than that during the presymptomatic stage, which can explain depletion of intercellular dopamine and appearance of motor disturbances. Activity of monoamine oxidase B in the striatum is reduced during the presymptomatic stage, but returns to the control level during the symptomatic stage. Variation in monoamine oxidase activity seems to reflect the compensatory mechanisms triggered in degrading nigrostriatal dopaminergic system.

Dopamine uptake in the substantia nigra and striatum in the presymptomatic and early symptomatic stages in Parkinsonian mice

The degeneration of dopaminergic (DAergic) neurons of the nigrostriatal system results in the development of Parkinsons disease (PD). It is assumed that the increase of DA release from survived DA� ergic neurons might be one of the compensatory processes serving to maintain the DA concentration in the extracellular space at a normal level and thereby providing a longlasting period of asymptomatic develop� ment of the disease. The goal of this study was to estimate DA release in the substantia nigra (SN) containing somas of DAergic neurons and in the striatum containi ng their axonal terminals under the different extents of the degradation of the nigrostriatal system corresponding to either presymptomatic or symptomatic stage of parkinsonism in mice. In the ex vivo studies of the brain slices under perfusion, it has been shown that the values of both spontaneous and K + �stimulated release of DA in the SN at both stages of parkinsonism were the same as those in the controls suggesting the increase of DA release from the survived somas of DAergic neurons. In the striatum at presymptomatic and symptomatic stages, no compensatory modification of spon� taneous DA release was observed whereas K + �stimulated release of DA from the survived axonal terminals increased significantly. However, the total amount of DA released at symptomatic stage was below the control level that was in contrast to that at presymptomatic stage. This observation is probably among the reasons of the appearance of the initial symptoms of parkinsonism.

Chronic models of the preclinical and early clinical stages of Parkinson’s disease in mice

Degradation of the dopaminergic nigrostriatal system is a central process of the pathogenesis of Parkinson’s disease, which is a chronic neurodegenerative disorder. A specific feature of the disease is longterm asymptomatic progress over decades, which occurs due to the functioning of compensatory processes in the brain. Specific motor symptoms appear after substantial lesion of the nigrostriatal system and depletion of compensatory reserves. At this stage, traditional treatment of patients has low efficiency. In the present study, we developed new neurotoxic models of the maximally prolonged preclinical stage and early clinical stage of Parkinson’s disease in mice and performed a thorough evaluation of motor behavior and the morphofunctional state of the nigrostriatal system in these animals. Further comparative study of these models will help to identify: (a) specific peripheral biomarkers of each stage as a basis for the development of the early preclinical diagnostics of Parkinson’s disease; (b) the mechanisms of neuroplasticity that are responsible for asymptomatic progress of the disease; (c) molecular triggers of impaired motor behavior during the transition from the preclinical stage to the clinical stage.

Time course of degeneration of dopaminergic neurons and respective compensatory processes in the nigrostriatal system in mice

160 Neurons are known to die continuously during the whole life of a human or animals [1]. However, until advanced age or even death, this neuronal loss does not influence negatively the functioning of the brain, including its involvement in neural or neuroendocrine regulation of the most important body functions. Even an increase in the rate of demise of specific neurons induced by unknown reasons in neurodegenerative diseases, including Alzheimer’s disease, Parkinson’s disease (PD), and others, does not impair brain func� tions, such as cognitive functions in Alzheimer’s dis� ease or motor functions in PD, for several decades [1]. This may be explained by compensation of functional insufficiency caused by neuronal loss related to the mechanisms of brain plasticity. However, at a certain threshold level of neuronal degeneration, compensa� tory capabilities of the brain become insufficient or exhausted, which results in impairments of specific body functions [2]. From the viewpoint of basic neu� rophysiology, it is important to understand the tempo� ral and functional relationships between two linked processes, neurodegeneration and compensation of functions of dead neurons, which continuously occur in the brain. This basic knowledge is very important for medicine too, because it may allow us to understand the mechanisms of regulation of both processes and to cope with them using prospective drugs. The aim of the present study was to examine the development of the linked processes of degeneration

Endocrine function of dopaminergic neurons of the whole rat brain in ontogeny: control of prolactin secretion.

81 The endocrine function of dopaminergic (DA ergic) neurons in the brain of adult animals is limited to secretion of DA by neurons of the arcuate nucleus of the hypothalamus to the median eminence area where the blood–brain barrier (BBB) is absent and to the pituitary portal circulation system where DA is involved in inhibitory control of prolactin secretion by lactotrophs of the anterior pituitary [1]. Due to a small volume of the portal vessels, even secretion of a small DA quantity into them results in a relatively high, “physiologically active” concentration, which is suffi cient for physiological influence on the target cells [2]. However, mixing of the portal blood with a larger vol ume of the peripheral blood leads to a multifold dilu tion of DA to a concentration that has no effect on the target cells or organs [2].

Dopamine release in the substantia nigra and striatum at presymptomatic and early symptomatic stages in parkinsonian mice

The degeneration of dopaminergic (DA-ergic) neurons of the nigrostriatal system results in the development of Parkinson’s disease (PD). It is assumed that the increase of DA release from survived DA-ergic neurons might be one of the compensatory processes serving to maintain the DA concentration in the extracellular space at a normal level and thereby providing a long-lasting period of asymptomatic development of the disease. The goal of this study was to estimate DA release in the substantia nigra (SN) containing somas of DA-ergic neurons and in the striatum containing their axonal terminals under the different extents of the degradation of the nigrostriatal system corresponding to either presymptomatic or symptomatic stage of parkinsonism in mice. In the ex vivo studies of the brain slices under perfusion, it has been shown that the values of both spontaneous and K+-stimulated release of DA in the SN at both stages of parkinsonism were the same as those in the controls suggesting the increase of DA release from the survived somas of DA-ergic neurons. In the striatum at presymptomatic and symptomatic stages, no compensatory modification of spontaneous DA release was observed whereas K+-stimulated release of DA from the survived axonal terminals increased significantly. However, the total amount of DA released at symptomatic stage was below the control level that was in contrast to that at presymptomatic stage. This observation is probably among the reasons of the appearance of the initial symptoms of parkinsonism.

The brain is one of the most important sources of dopamine in the systemic circulation in the perinatal period of ontogenesis in rats

This study was designed to test the authors’ hypothesis that dopamine passes from dopamine-synthesizing cells in the brain to the systemic circulation prior to the formation of the blood-brain barrier during ontogenesis. High-performance liquid chromatography studies demonstrated that peripheral blood dopamine levels before formation of the blood-brain barrier-in rat fetuses and neonates-are significantly higher than after formation of the barrier in adult rats, providing indirect evidence in support of the hypothesis. Furthermore, formation of the blood-brain barrier is accompanied by a significant increase in dopamine levels in the rat brain. Direct evidence for the hypothesis was obtained in the form of a sharp decrease in blood dopamine levels in fetuses after lesioning of dopamine-synthesizing neurons in the brain by encephalectomy.

Non-Dopaminergic Neurons Expressing Dopamine Synthesis Enzymes: Differentiation and Functional Significance

The development and functional significance of neurons in the arcuate nucleus expressing tyrosine hydroxylase and/or aromatic L-amino acid decarboxylase were studied in rat fetuses, neonates, and adults using immunocytochemical (single and double immunolabeling of tyrosine hydroxylase and aromatic L-amino acid decarboxylase) methods with a confocal microscope and computerized image analysis, HPLC with electrochemical detection, and radioimmunological analysis. Single-enzyme neurons containing tyrosine hydroxylase were first seen on day 18 of embryonic development in the ventrolateral part of the arcuate nucleus. Neurons expressing only aromatic L-amino acid decarboxylase or both enzymes of the dopamine synthesis pathway were first seen on day 20 of embryonic development, in the dorsomedial part of the nucleus. On days 20–21 of embryonic development, dopaminergic (containing both enzymes) neurons amounted to less than 1% of all neurons expressing tyrosine hydroxylase and/or aromatic L-amino acid decarboxylase. Nonetheless, in the ex vivo arcuate nucleus and in primary neuron cultures from this structure, there were relatively high levels of dopamine and L-dihydroxyphenylalanine (L-DOPA), and these substances were secreted spontaneously and in response to stimulation. In addition, dopamine levels in the arcuate nucleus in fetuses were sufficient to support the inhibitory regulation of prolactin secretion by the hypophysis, which is typical of adult animals. During development, the proportion of dopaminergic neurons increased, reaching 38% in adult rats. Specialized contacts between single-enzyme tyrosine hydroxylase-containing and aromatic L-amino acid decarboxylase-containing neurons were present by day 21 of embryonic development; these were probably involved in transporting L-DOPA from the former neurons to the latter. It was also demonstrated that the axons of single-enzyme decarboxylase-containing neurons projected into the median eminence, supporting the secretion of dopamine into the hypophyseal portal circulation. Thus, dopamine is probably synthesized in the arcuate nucleus not only by dopaminergic neurons, but also by neurons expressing only tyrosine hydroxylase or aromatic L-amino acid decarboxylase.

The state of the tuberoinfundibular dopaminergic system during modeling of parkinsonism in mice

Parkinson’s disease (PD) is one of the most widespread neurodegenerative diseases; it develops as a result of the death of dopaminergic (DA-ergic) neurons of the nigrostriatal system of the brain. It has been shown in autopsy material that the neurodegenerative process spreads to other populations of neurons of the peripheral and central nervous system. In PD patients the tuberoinfundibular DA-ergic system (TIDAS) undergoes degradation; however, no experimental models exist that can reproduce the combined degradation of the nitrostriatal system and the TIDAS. In this work, we analyzed the state of the TIDAS during presymptomatic and symptomatic stages of parkinsonism induced by the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). These stages of parkinsonism develop as a result of subthreshold and threshold degradation of the nigrostriatal DA-ergic system. To evaluate the morphofunctional state of the TIDAS, we determined: (a) the number of neurons with tyrosine hydroxylase (TH), a key enzyme of DA synthesis; (b) the sizes of the terminals of TH-positive axons; (c) the levels of TH in the neuronal somas and the terminals of axons; and (d) the contents of DA, noradrenaline, and 3,4-dihydroxyphenyl acetic acid. According to the data that was obtained during the presymptomatic stage, for the TIDAS no changes were found in all the measured parameters. During the symptomatic stage, we observed (a) a decrease in DA content by 50%; (b) a decrease in the level of TH protein in axonal terminals and an increase in neuronal somas; and (c) an increase in the turnover of DA by 149% as compared to the control. The data we obtained suggest that the functional activity of the TIDAS was insufficient; this was seen in the form of alterations of DA metabolism, a decrease in the level of synthesis, probably in axonal terminals, and compensatory acceleration of the functional cycle from synthesis to enzymatic degradation. Although no changes in the number of neurons and axons that express TH was found in the TIDAS, the question of organic degradation of the TIDAS in parkinsonism is still open because TH is a marker of not only DA-ergic neurons that degenerate in the presence of neurotoxins but also of non-DA-ergic neurons that are insensitive to neurotoxins. Thus, during experimental modeling of Parkinsonism, we observed at least the metabolic characteristics of TIDAS degradation only at the symptomatic stage of the disease.