Histochemical studies on the rat submaxillary gland during post-natal development

Summary A post-natal development of the albino rat submaxillary gland was studied morphologically and histochemically. Throughout the developmental stages, conspicuous variations were observed morphologically; growth of acini, glandular tubules formation and advance of striated duct.For enzymatic histochemical observations, the localization and activity of hydrolytic and oxidative enzymes were apparently similar to those of the matured gland from about 5 weeks after birth in the intralobular striated ductal components. Granular tubules were clearly demonstrated from 7 weeks after birth showing almost the same activity as the adult gland.     

Histochemically demonstrable monoamines in the pituitary gland and median eminence of the female rat during the postnatal development

Summary The postnatal development of formaldehyde induced fluorescence (FIF) was studied in the pituitary glands of female rats. The effects of 3,4-dihydroxyphenylalanine (L-dopa), D,L-5-hydroxytryptophan (DL-5-HTP) and dopamine (DA) treatments on the FIF were followed during the postnatal period.The appearance of specifically fluorescing monoamines into the cells of the pars intermedia occurred postnatally and the level of the adult fluorescence was reached at 4–5 weeks' age. The intensity of the fluorescence was independent on the density of the fluorescing nerves. Among the fluorescing nerves droplet fibres were regularly observed from the age of 3 weeks, which confirms the theory that these fibres are caused by toxic factors when the blood-brain barrier is not functioning.There was no change postnatally in the number of fluorescing cells in the pars distalis.The fluorescing innervation of the median eminence, developed most rapidly at the age of 1–3 weeks and the level of the adult fluorescence was reached at the age of 4–5 weeks.The first specifically fluorescing cells after L-dopa treatment were observed at 6 days age. A remarkable increase in the number of fluorescing cells was seen between 12 and 18 days. After DL-5-HTP treatment fluorescent cells were seen but at later stages. These observations suggest that the cells in the pituitary gland, which store amine-precursors and monoamines developmentally differ from the APUD-cells. The rapid increase of the fluorescing cells between 12 and 18 days and the simultaneous development of the fluorescing innervation of the median eminence suggest the following correlations: the development of dopaminergic innervation of the median eminence — the secretion of releasing hormones — the activity of PAS-positive cells (FSH, LH and TSH secretion) — the uptake of L-dopa and DL-5-HTP into the PAS-positive cells.Dopamine was not uptaken into the cells of pars distalis. The walls of the blood vessels began to show fluorescence suggesting a barrier mechanism, which prevents the DA-uptake into the PAS-positive cells.This work was supported by the Grant for Young Research Workers, University of Helsinki.     

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 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     

Adenylate cyclase activity in rat submandibular gland during postnatal development

Adenylate cyclase activity was measured in homogenates of submandibular glands of 1 to 42 day old rats. During this period of time the gland reached its final stage of differentiation. Adenylate cyclase activity was higher in the glands of one day old rats than in those of 7 and 14 day old animals. Between 14 and 28 days of age the enzyme activity more than doubled and approached the level that characterized the glands of adult animals. Fluoride (10mM) stimulated the enzyme activity in all age groups but the stimulation was less in the case of one day old rats as compared to older animals. Isoproterenol (10^?4 M) stimulated adenylate cyclase by 50–60% in the gland of adult rats but had no effect on the enzyme activity in 7 to 28 day old animals. Administration of isoproterenol for 5 days to 9 day old rats increased the weight of the submandibular gland by 70 per cent. Total adenylate cyclase activity increased parallel with the weight of the gland but the specific activity of the enzyme remained unchanged. It is concluded that during the postnatal development of the submandibular gland the rapid increase in adenylate cyclase activity occurs after weaning and it coincides with an accelerated rate of functional differentiation of the acinar cells.     

Immunocytochemical demonstration of tissue-type plasminogen activator in endocrine cells of the rat pituitary gland

We immunocytochemically stained rat pituitary glands using antibodies against plasminogen activators of the tissue type (t-PA) and the urokinase type (u-PA). A large population of endocrine cells in the anterior lobe of the gland displayed intense cytoplasmic immunoreactivity with anti-t-PA. In some areas of the intermediate lobe we found a weak staining, and we observed weakly staining granular structures in the posterior lobe. Controls included absorption of the antibodies with highly purified t-PA. In addition, SDS PAGE followed by immunoblotting of pituitary gland extracts revealed only one band with an electrophoretic mobility similar to that of t-PA when stained with anti-t-PA IgG. No u-PA immunoreactivity was detected in the rat pituitary gland. Sequential staining experiments using antibodies against growth hormone and t-PA demonstrated that the t-PA-immunoreactive cells constitute a large subpopulation of the growth hormone-containing cells. These findings represent the first direct evidence for the presence of t-PA in cell types other than endothelial cells in the intact normal organism. In this article we discuss the implications of the results for a possible role of t-PA in the posttranslational processing of prohormones.     

Ultrastructural observations on the postnatal development of the rat prostate

Summary The postnatal development of the rat prostate has been studied with the electron microscope. Major developmental changes begin during the second week after birth and involve organelles associated with the formation of secretions. The amount of granular endoplasmic reticulum and the size of the Golgi complex increase greatly. Large vacuoles that probably contain secretory material are formed, and the lumen of the prostatic acini appears to contain secreted material. Large lysosomes with polymorphic interiors are present as early as 10 days after birth, and they become numerous by the end of the third week. Differences in fine structure between the different lobes of the prostate are detectable in 10–14 day old rats. The subsequent differentiation of the granular endoplasmic reticulum into the forms characteristic of the different prostatic lobes is described. The initial changes in the prostate occur in advance of sexual maturity of the animal, and the adult appearance of the gland is attained by 4–5 weeks after birth.This study was supported by Contract No. 69-2104, Program Project HD-02282, Health Sciences Advancement Award FR-02084, and a Research Career Development Award (1-K3-GM-28, 214-03) from the National Institutes of Health.The author wishes to acknowledge the technical assistance of Mrs. Stephanie Krah.     

Proliferating cells in the rat anterior pituitary during the postnatal period: immunoelectron microscopic observations using monoclonal anti-bromodeoxyuridine antibody

Proliferating cells in the male rat anterior pituitary at 1, 3, 5, and 8 weeks of age were labeled with bromodeoxyuridine (BrdU) and studied by light and electron microscopic immunocytochemistry using anti-BrdU. They decreased in number from 402±31/mm² at 1 week to 50±1.5/mm² at 8 weeks, while their cell area increased by about twofold during this period. They had a slightly higher nucleus/whole cell (N/C) ratio than non-proliferating cells. According to their ultrastructure we classified them into granular and agranular cells. The percentage of granular cells ranged from 73% to 82% of all the proliferating cells during the period studied. They had many granules of various sizes and shapes, and some contained growth hormone and prolactin. Agranular cells, constituting 18–27% of proliferating cells, were small and had a high N/C ratio, indicating their immaturity. Moreover, they showed several features of folliculo-stellate (FS) cells: they showed no secretory granules in the cytoplasm, extended thin cytoplasmic processes, and sometimes they constructed a follicle among them. These results suggest: (1) the majority of proliferating cells were mature cells producing anterior pituitary hormone(s) and (2) most of the agranular proliferating cells maybe FS cells. The possibility of the latter is discussed.     

L-serine and GABA uptake by synaptosomes during postnatal development of rat

Postnatal development changes in mechanisms of synaptosomal amino acid transport have been studied in rat cerebral cortex. Specific uptake of radiolabeled L-serine was examined and compared with that of radiolabeled GABA using synaptosomes-enriched fractions freshly prepared from cerebral cortex at different postnatal days from the birth to young adulthood. The preparations were incubated with 10 nM of [3H]L-serine and 10 nM of [3H]-GABA in either the presence or absence of NaCl, KCl or choline chloride, at 2 and 30 degrees C, for different periods up to 30 min. The uptake of [3H]l-serine was temperature dependent in synaptosomal fractions prepared from cerebral cortex of rats in postnatal days 5, 7, 13 and 21, but stronger dependence was observed in adult brain, irrespective of the presence of Na+, K+ or choline ions. At all postnatal ages studied, [3H]-GABA uptake showed a high activity in the presence of Na+ ions and at 30 degrees C. The values of Km were 90-489 microM in L-serine uptake. However, in the uptake of GABA the values of Km were 80-150 microM. The highest values of Vmax were obtained at 5 and 21 postnatal days for both transport systems. These results indicate that the uptake of l-serine and GABA are regulated differentially during postnatal development.     

Differential proteomic analysis of adrenal gland during postnatal development

The adrenal glands (AGs) are endocrine organs essential for life. They undergo a fetal to adult developmental maturation process, occurring in rats during the first postnatal month. The molecular modifications underlying these ontogenic changes are essentially unknown. Here we report the results of a comparative proteomic analysis performed on neonatal (Postnatal day 3) versus adult (Postnatal day 30) AGs, searching for proteins with a relative higher abundance at each age. We have identified a subset of proteins with relevant expression in each developmental period using 2‐DE and DIGE analysis. The identified proteins belong to several functional categories, including proliferation/differentiation, cell metabolism, and steroid biosynthesis. To study if the changes in the proteome are correlated with changes at the mRNA level, we have randomly selected several proteins with differential expression and measured their relative mRNA levels using quantitative RT‐PCR. Cell‐cycle regulating proteins (retinoblastoma binding protein 9 and prohibitin) with contrasting effects on proliferation are expressed differentially in neonatal and adult AG. Progesterone metabolizing enzymes, up‐regulated in the neonatal gland, might contribute to the hyporesponsiveness of the adrenal cortex characteristic of this developmental period. We have also observed in the adult gland a marked up‐regulation of enzymes involved in NAD(P)H production, thus providing the reducing power necessary for steroid hormone biosynthesis.     

Proliferative activity of cells of the intermediate lobe of the rat pituitary during the postnatal period.

Cellular proliferation was studied in the intermediate lobe (IL) of the pituitary gland of developing rats by labelling cells at the S-phase of the cell cycle with bromodeoxyuridine (BrdU). The number of BrdU-labelled cells in the IL decreased from birth until the 14th postnatal day and was low from that day until the end of the first month after birth. Throughout the postnatal period a large proportion of BrdU-labelled cells was found in the marginal layer (ML) of the IL, suggesting for the ML a role as a germinative layer of the IL during postnatal growth. Double immunostaining with anti-BrdU and anti-MSH showed that MSH cells actively proliferate as from the day of birth. Cells doubly immunostained with anti-BrdU and anti-S100 protein were first seen on the 14th postnatal day. From then onwards, most proliferating cells were labelled with either anti MSH or anti S-100 protein. This, together with the high proportion of proliferating cells found in the ML marks a clear difference with the pattern of cellular proliferation previously reported during a similar period in the anterior lobe of the rat pituitary.     

Structure of the non-lymphoid cells during the postnatal development of the rat lymph nodes

This study describes the postnatal development of the nonlymphoid cells with special reference to the fibroblastic reticulum cells (FRCs) and interdigitating cells (IDCs). The first lymphocytes of the neonatal lymph nodes are located in the developing deep cortex units (DCUs) identified by the Gomori's technique for reticulin fibres. Ultrastructural studies demonstrate that FRCs form the stroma of the DCUs. By light and electron microscopy, it is demonstrated that FRCs occupy the outer cortex in the following stages of development of the lymph nodes. Thus, FRCs form the stroma of the primary follicles and, later, are transformed in follicular dendritic cells (FDCs) of the germinal centres. Immature or pro-IDCs appear as migrating elements in the deep cortex of lymph nodes of the neonatal rats. The ultrastructure of the pro-IDCs resembles that of the mature IDCs but not that of the phagocytic cells. Pro-IDCs are transformed into mature IDCs whose cytoplasmic expansions contact lymphocytes via tight junctions. Some of these lymphocytes are likely apposed to FRCs of the DCUs. No cells containing Birbeck granules were found in the parenchyma of the lymph nodes during the postnatal development. The role of these nonlymphoid cells is discussed with respect to the immunologic function of mammalian lymph nodes.