Comparative characteristics of serotoninergic neurons in some nuclei of rat medulla

Using the immunohistochemical method, serotoninergic neurons were identified in eight medulla nuclei in the bulbar vasomotor centers of Wistar rats. Using morphometry, it was established that the proportion of serotoninergic neurons located in the projection of the studied nuclei varied at 17–26%; the value of this parameter only reached 34–40% in nuclei of the posterior raphe group. Single immunopositive cells able to perform integrative functions in the regulation of hemodynamics were identified between the nuclei, as well as between nuclei and conducting pathways.     

Preliminary observations on the distribution of serotoninergic and cerebellar-projecting neurons of the raphe nuclei of the brain stem

Cerebellar projection from raphe nuclei were investigated in rabbit by using retrograde transport of HRP and serotonergic mapping by direct fluorescence. A close topographical correlation between the HRP labeled cells and the serotonergic neurons has been observed. The current study has demonstrated the presence of paramedian and lateral cells whose cytoarchitecture is identical with midline cells of many raphe nuclei. All of the raphe nuclei except the linear nuclei, contained serotonergic perikarya. The midline and paramedian portions of the nuclei raphe obscurus, pallidus, magnus, and nucleus raphe dorsalis contained principally serotonergic neurons; the lateral portions of the medullary raphe nuclei and the nuclei raphe pontis and centralis superior contained a significant number of non-fluorescent cells. In these regions, fluorescent sections often revealed the size, shape, and orientation of the perikarya and dendrites; further verification of cytoarchitectural characteristics of these neurons depended heavily upon these clues.     

Ontogenesis of TRH mRNA in the rat pancreas

The rapid changes in TRH levels in the rat pancreas during the neonatal period make this organ an interesting model for the study of the regulation of TRH biosynthesis. Pancreatic RNAs were isolated by the guanidinium thiocyanate method and layered onto CsCl cushion. Northern blot preparations were hybridized with 32P labeled TRH cDNA probe. Pancreatic TRH mRNA was first detected in 19-day old fetuses and reached the highest level on day 0, then decreased, being barely detectable 14 days after birth. The neonatal injection of streptozotocin induced a dramatic drop of TRH mRNA levels 24 hours later. This result suggests that the peculiar evolution of TRH level in pancreas is partly due to the evolution of the expression of the TRH gene.     

Ontogenesis of hepatic growth hormone receptors in the rat

Detailed ontogenic studies of the binding of human (hGH) and bovine growth hormone (bGH) have been performed in liver preparations from male and female rats during the neonatal, weanling, pre- and post-pubertal periods. Specific binding of both hormones was readily detected at all ages, with no apparent interference due to occupancy by endogenous hormones. No sex difference in binding was observed prior to weaning (22 days) for hGH, which binds to both somatotrophic and lactogenic sites. However, after weaning a marked sex-related dissociation in the pattern of binding did occur, with female rats binding 3-4 times more hGH than in the pre-weaning period and male rats binding hGH to only half their pre-weaning levels. A very similar pattern was seen for binding of bGH (which binds only to somatotrophic sites) except that in male rats, the post-weaning levels did not fall. Binding patterns for either hGH or bGH prior to weaning did not mirror the known age-related pattern of circulating rat GH levels, suggesting the absence of a definitive auto-regulation system for the GH-GH receptor system under normal circumstances in vivo. The possible role of the weaning process per se in the post-weaning changes of GH binding seen in male and female rats still requires elucidation.     

Ontogenesis of the vitamin D receptor in rat heart

1,25-Dihydroxyvitamin D(3) through its receptor (vitamin D receptor; VDR) has important physiological effects such as calcium transport and cell growth and differentiation. Although the VDR is present in a variety of cell lines as well as in numerous tissues, including rat and human heart, no data are available about the presence of VDR in heart at different steps of rat life. In this study we evaluated the VDR expression using RT-PCR and immunohistochemical techniques in fetal (17, 18 and 20 gestational days), neonatal (4 and 8 days) and adult rat heart. Immunohistochemical techniques showed the VDR protein localisation in the nuclei of cardiac muscle fibres. Also, we demonstrated that VDR mRNA expression is changing over these different periods of development, showing significant differences in 20 days versus 18 days of fetal age. These changes in VDR expression may be related to other parameters associated with the development of the cardiac muscle and/or intracellular cardiac cell calcium homeostasis.     

Ontogenesis of enkephalin and humoral endorphin in the rat brain

Radioimmunoassay and column chromatography techniques were used to study the postnatal development of two different opioid ligands, humoral endorphin and enkephalin in the rat brain. Similar patterns were observed for both male and female animals during the period examined (from birth to the seventh week of life). Humoral endorphin content of the developing rat brain was found to increase in parallel to enkephalin, exhibiting a ‘lag’ period of 2 weeks. The most dramatic increase in opioid levels was detected during the third week of life; this stage was followed by a gradual change up to the adult levels.     

Functional recuperation of the serotoninergic innervation in the rat locus coeruleus

1. 5,6-dihydroxytryptamine (5,6-DHT) or a lesion of the raphe centralis superior (RCS) cause significant decreases in the serotonin (5-HT) content and significant increases in the tyrosine hydroxylase activity in the locus coeruleus (LC) of the rat. This suggests that noradrenaline (NA) synthesis is controlled by serotonin-containing neurons in the raphe system via their terminals in the LC. 2. Radioautography after intraventricular infusion of tritiated serotonin (3H-5-HT) and biochemical determinations of endogenous 5-HT content showed an almost complete disappearance of serotoninergic axonal varicosities and content in the LC region 10-15 days after intraventricular administration of 75 micrograms of 5,6-DHT. Two to 4 months after neurotoxin administration, 5-HT fibers had regrown in the LC but, contrary to the normal innervation pattern, the majority of them invaded the medial most portion of the nucleus and the adjacent subependymal region. The LC region regained almost all of its endogenous 5-HT content in the same time period. 3. Functional recuperation of these 5-HT fibers was demonstrated by the fact that the RCS had, after regeneration, the same functional control on NA synthesis as in the normal animal.     

Ontogenesis of the LH-RH-containing neuronal elements in the hypothalamus of the rat.

LH-RH-containing neuronal elements were detected earliest in the median eminence and in the vascular organ of the lamina terminalis of newborn rats. The peroxidase-labeled antibody method was used. Since the synthesis of the gonadotrophic hormones starts already on the 18th day of intrauterine life in the rat, this finding does not support the view that LH-RH could have inductive effect on the differentiation of the gonadotropic cells of the anterior pituitary gland.     

Ontogenesis of alpha-amylase in rat parotid gland during postnatal development

Changes in alpha-amylase (alpha-1,4-glucan-4-glucanohydrolase, EC 3.2.1.1) of parotid gland were investigated during postnatal development of the rat. Modifications in amylase activity after birth allow the distinction of three stages which can be correlated with the morphologic development of the parotid gland. Significant sexual differences in the evolution of alpha-amylase activity were found. During the first stage (from birth to the 20th day) there is a higher increase in females, while males have a more pronounced increment in the second stage (from the 20th to the 30th day). By means of gel electrophoresis of parotid extracts, four molecular forms of amylase can be separated. The slowest migrating band (Form 1) is not detected at the initial stage.     

Ontogenesis of pyruvate kinase in the brain and liver tissues of the rat

The ontogenetic study of pyruvate kinase in the brain and liver tissues was performed in different batches of rats, from the fœtus at the 13th day of gestation to the adult subject.

— According to the kinetic study, the shape of the curve is transformed from sigmoid to hyperbolic from the 13th day of fœtal life to adulthood in the brain. Hill cœfficient increases with the age of animal in the liver tissue.

— According to polyacrylamide gel isoelectrofocusing, a family of four, transitory or definite bands are detected in the fœtal brain: they are well defined by their pHi: M₄, M₃, M₂, M₁; at the adult stage, M₁ predominates, M₂ is minor. Three principal bands are distinguished in the liver: two are characteristic of fœtal life (Lf and M₂), one of adulthood (L).

— According to the immunochemical tests, there are antigenic determinants common to M₁, M₂, M₃ and M₄.

The confrontation of the first two methods prompts the conclusion that the kinetic of the enzyme (and perhaps its function) varies with the animals age and is linked to its molecular structure. With the third method, it allows to stress the precociousness of the appearance of the common antigenic determinants, simultaneously with immature enzymatic forms.The signification of the kinetic modifications as well as the succession of the isozymes of the M type in a determined order are discussed and the in vivo formation of hybrids is suggested.     

Ontogenesis of alpha-amylase in rat parotid gland during postnatal development

Summary Changes in -amylase (-1,4-glucan-4-glucanohydrolase, EC 3.2.1.1) of parotid gland were investigated during postnatal development of the rat. Modifications in amylase activity after birth allow the distinction of three stages which can be correlated with the morphologic development of the parotid gland. Significant sexual differences in the evolution of -amylase activity were found. During the first stage (from birth to the 20th day) there is a higher increase in females, while males have a more pronounced increment in the second stage (from the 20th to the 30th day). By means of gel electrophoresis of parotid extracts, four molecular forms of amylase can be separated. The slowest migrating band (Form 1) is not detected at the initial stage.Career scientist of the Consejo Nacional de Investigaciones Científicas y Téchnicas of Argentina     

Activation of PAF-synthesizing enzymes in rat brain stem slices after LTP induction in the medial vestibular nuclei

LysoPAF acetyltransferase (lysoPAF-AT) and PAF-synthesizing phosphocholinetransferase (PAF-PCT) are the two enzymes which catalyze the final reactions for the synthesis of PAF. Their activities, assayed in the homogenate of rat brain stem slices and under their optimal conditions, increased 5 min after high frequency stimulation of vestibular afferents, inducing LTP in the medial vestibular nuclei. The activity of phosphatidylcholine-synthesizing phosphocholinetransferase, was not affected. Sixty minutes from the induction of LTP, PAF-PCT activity, but not that of lysoPAF-AT, was still significantly higher with respect to 5 min test stimulated control. We used AP-5 to verify whether this increase was strictly dependent upon LTP induction, which requires NMDA receptor activation. In AP-5 treated slices, lysoPAF-acetyltransferase and PAF-synthesizing phosphocholinetransferase activities increased, but they were reduced after high frequency stimulation under AP-5. In conclusion, we have demonstrated that the activities of PAF-synthesizing enzymes are activated soon after the induction of LTP and that this effect is linked to the activation of NMDA-receptors. We suggest that the enzyme activation by AP-5, preventing LTP, might be due to glutamate enhancement but, in neurons showing LTP and under normal conditions, the activation of potentiation mechanisms is critical for the enhancement of enzyme activities.     

Radioautographic study of the hypothalamic serotoninergic structures in the perinatal period of the rat

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.     

Postnatal changes in the expression of serotonin 2A receptors in various brain stem nuclei of the rat.

Previously, we reported a critical period [around postnatal day (P) 12-13 in the rat] in respiratory network development when distinct neurochemical, metabolic, and physiological changes occur. Since serotonin 2A (5-HT(2A)) receptors play an important role in respiratory modulation, we hypothesized that they may undergo developmental adjustments during the critical period. Semi-quantitative immunohistochemical analyses were conducted in labeled neurons in a number of brain stem nuclei with or without known respiratory functions from P2 to P21 in rats. Our data indicate that the expressions of 5-HT(2A) receptors in neurons of the pre-B?tzinger complex, the nucleus ambiguus, and the hypoglossal nucleus were maintained within a relatively narrow range between P2 and P21, with a dip at P3-P4 and a significant reduction only at P12. This change was not observed in the nonrespiratory cuneate nucleus. These results suggest that reduced expressions of 5-HT(2A) receptors at P12 contributes to neurochemical imbalance within brain stem respiratory nuclei at that time and may be involved in decreased hypoxic ventilatory response at this critical period of development.