Cellular localization of 3H-oestradiol in the hypothalamus

Summary The hypothalamus of male and female rats, given 0.3 g/100 g body weight of 6.7-³H-oestradiol-17 and killed 1 hour after the injection, was examined by autoradiography in order to 1) localize the areas and the cells involved in the uptake of the hormone, and 2) study the intracellular localization of the labelled material.Only nerve cells contained radioactive material while glial and ependymal cells were not significantly labelled. In the anterior hypothalamus, labelled nerve cells were concentrated in areas corresponding to nucleus preopticus medialis and nucleus preopticus, pars suprachiasmatica. The nucleus supraopticus was unlabelled. In the medial basal hypothalamus, neurons corresponding to the nucleus arcuatus and the lateral part of the nucleus ventromedialis showed marked labelling. No significant labelling was observed in the nucleus paraventricularis, pars magnocellularis.Although the individual nerve cells varied in their extent of labelling, the major proportion of the silver grains were consistently concentrated over the nuclei. Castration was not found to influence the results. The findings were essentially the same in male and female rats and appear to suggest that oestradiol exerts a direct effect on nerve cells in certain hypothalamic areas.This work was supported by grants from the Norwegian Cancer Society, Nordisk Insulinfond and Anders Jahres Fond. The skilful assistance of Miss Helga Friedl and Mrs. Jane Larsen is gratefully acknowledged.     

Light microscope radioautographic study of glycoprotein secretion in the hypothalamic-neurohypophysial system of the rat,after L-fucose-3H injection

Summary Fucose-³H was injected into the cerebral ventricle of rats that were killed at several time intervals after injection. Semi-thin sections of the supraoptic nucleus and neurohypophysis were processed for radioautography and analysed quantitatively. Silver grains indicating the site of fucose-labeled glycoproteins were first located at the perinuclear region of the secretory neurons. The highest silver-grain density in these cells was observed at 2 h after injection, declining afterwards. Silver grains over the neurohypophysis were observed from 2 h on, reached a peak at 1 day after injection and decreased in the subsequent time intervals. The distributions of the silver grains over the neurohypophysis fitted Poissonian distributions and these were shown to be heterogeneous at the several time intervals. Pituicytes were not labeled. The percentage of silver grains over the Herring bodies increased with time. In rats deprived of water after fucose-³H injection there was a great increase in the release of labeled glycoproteins from the neurohypophysis. These results indicate that the glycoproteins synthesized by the secretory neurons of the hypothalamic nuclei are secreted in the neurohypophysis.     

Fine structure of pars neuro-intermedia of the loach,Misgurnus fossilis L.

Summary Two types of glandular cells are present in the pars intermedia of the loach, Misgurnus fossilis. Basophils are characterized by the presence in their cytoplasm of numerous secretory granules containing electron-dense and homogeneous material and by scarce endoplasmic reticulum. Weak acidophils contain in their cytoplasm abundant endoplasmic reticulum and numerous granules of different electron densities.The distal part of the neurohypophysis is composed of several types of neurosecretory axons, strongly branched pituicytes, numerous capillaries, and connective tissue elements. The axon terminals form the neuroglandular, neurovascular and neurointerstitial contacts. Some axon terminals are closely apposed to the basement membrane separating neurohypophysis from meta-adenohypophysis. At points of absence of continuity of this membrane, some neurosecretory axons become directly contiguous with cytoplasmic membranes of the intermedia cells.The investigation was partly supported by a research grant from the Zoological Committee of the Polish Academy of Sciences.     

Radioautographic study of the rat brain and pituitary after injection of 3H dexamethasone

The central nervous system and the pituitary of adrenalectomized male rats injected with 3H-dexamethasone were examined by radioautography. At 1 hr after the injection, radioactivity concentration was high in the medial basal hypothalamus, the pituitary and the pineal gland. In the hypothalamus, radioactive material was found to be selectively concentrated in neurons in the ventral part of nucleus arcuatus and in the infundibular region. In the anterior pituitary, a large proportion of cells showed silver grains both in the cytoplasm and over the cell nuclei. However, in a small number of cells, the radioactive material was associated with the cell nuclei. Less radioactivity was present in the intermediate and posterior lobes. The pineal gland contained more silver grains than did other regions of the brain. The results obtained in the present study suggest essentially an action of dexamethasone in the medial basal hypothalamus and at the level of the pituitary.     

THE CYTONUCLEOPROTEINS OF AMEBAE : I. Some Chemical Properties and Intracellular Distribution

Autoradiographs of whole Amoeba proteus host cells fixed after the implantation of single nuclei from A. proteus donors labeled with any one of 8 different radioactive amino acids showed that the label had become highly concentrated in the host cell nucleus as well as in the donor nucleus and that the cytoplasmic activity was relatively low. When these amebae were sectioned, the radioactivity was found to be homogeneously distributed throughout the nuclei. The effect of unlabeled amino acid "chaser," the solubility of the labeled material, and the long-term behavior of the labeled material gave evidence that the radioactivity was in protein. At equilibrium, the host cell nucleus contained approximately 30 per cent of the radioactivity distributed between the two nuclei. This unequal nuclear distribution is attributed to the presence of two classes of nuclear proteins: a non-migratory one that does not leave the nucleus during interphase, and a migratory one, called cytonucleoprotein, that shuttles between nucleus and cytoplasm in a non-random manner. It is estimated that between 12 per cent and 44 per cent of the cytonucleoproteins are present in the cytoplasm of a binucleate cell at any one moment. Nuclei of Chaos chaos host cells also concentrated label acquired from implanted radioactive A. proteus nuclei.     

Autoradiographic localization of [3H]hydroxytamoxifen to uterine oestrogen- and antioestrogen-binding sites

Summary Immature rats were injected subcutaneously with 0.36 g of [³H]hydroxytamoxifen ([³H]TAM(OH)) or 0.24 g of [³H]oestradiol in oil, and 4 h later uteri were processed for thaw-mount autoradiography. The specificity of [³H]TAM(OH) localization was determined by injecting a 200-fold excess of unlabelled TAM(OH) or a 20-, 200- or 2000-fold excess of oestradiol 1 h before injection of [³H]TAM(OH). After injection of [³H]TAM(OH) or [³H]oestradiol, autoradiograms showed concentration of radioactivity in nuclei of stromal, epithelial and myometrial cells, but this labelling varied among the cell types depending upon which compound was injected. After [³H]TAM(OH) injection, the decreasing order of labelling intensity was stroma, myometrium, epithelium; after [³H]oestradiol injection the decreasing order was stroma, epithelium, myometrium. Injection of TAM(OH) before [³H]TAM(OH) eliminated nuclear labelling in all the uterine cell types. Injection of oestradiol before [³H]TAM(OH) decreased nuclear labelling and resulted in the concentration of label in the cytoplasm of luminal epithelium which was not present when [³H]TAM(OH) was injected alone. Cytoplasmic labelling increased initially as the oestradiol competition dose increased, but the increase in labelling did not continue with increasing concentrations of oestradiol. The results indicate that antioestrogen and oestrogen localize to nuclei of the same uterine cell types, but that cellular uptake differs among the tissue compartments. The results also suggest that a high concentration of antioestrogen-binding sites exist in the cytoplasm of the uterine luminal epithelium.     

Tritiated thymidine autoradiographic study on the origin and renewal of argentaffin cells in the pyloric gland of hamsters

Summary The origin and renewal of the argentaffin cells in the pyloric glands of hamsters were studied by flash, cumulative and pulse labelling autoradiography with ³H-thymidine. The argentaffin cells were identified by the Diazo Method using Fast Red B Salt.By flash labelling autoradiography, it was shown that the argentaffin cells located from the middle to the lower level of the pyloric mucosa were not labelled with ³H-thymidine, indicating that this cell type has no proliferative activity. On the 10th and the 20th day of cumulative labelling, 31% and 63% of the argentaffin cells in the gland were found to be labelled, respectively. The labelled argentaffin cells were concentrated in the upper part of the gland (around the region of the isthmus), and no label was found over nuclei of the cells at the lowermost level of the gland. These labelled cells were shown to undergo a downward migration in the days following pulse labelling. They were replaced by unlabelled (and weakly or very weakly labelled) cells which arose at the region of the isthmus. The argentaffin cells in the pyloric gland are thought to arise from epithelial precursor cells at the region of the isthmus.The labelled argentaffin cells in the gland were found to decrease in number almost exponentially after pulse labelling. This indicates that the life span of argentaffin cells is not fixed, but their renewal conforms to the random loss system. The half time of turnover of this cell population was 15 days on average.Supported by a Grant-in-Aid for Cancer Research from the Ministry of Education, Science and Culture, Japan     

Selective retention of oestradiol by cell nuclei in specific brain regions of the ovariectomized rat

—Cell nuclei were isolated from four regions of the brains of ovariectomized female rats 2 hr after the injection of [³H]oestradiol. By light microscopy, the nuclear pellets contained highly purified nuclei of neuronal and glial cells with little cytoplasmic contamination. Tritium was concentrated in cell nuclei from the preoptic-hypothalamic area, to a lesser extent in nuclei from the amygdaloid region and hippocampus, and least of all in cerebral cortical nuclei. In comparison with whole homogenates (= 1-0), the nuclear concentrations of radioactivity were 12·9, 4·7, 1·9 and 0·8, respectively. Approximately 40 per cent of the radioactivity in homogenates of the preoptic-hypothalamic area was present in cell nuclei, and upon TLC more than 85 per cent of the radioactive material in the nuclei exhibited the R_F of oestradiol-17β. Pretreatment of ovariectomized females with 1 mg of unlabelled oestradiol 30 min before the injection of labelled hormone abolished the nuclear uptake of [³H]oestradiol in all four regions of the brain. A concurrent injection of 10 μg of unlabelled oestradiol-17β significantly reduced nuclear uptake, while a similar injection of testosterone or oestradiol-17α had no significant effect. One mg of oestradiol-17α, but not testosterone, did reduce nuclear uptake. The retention of [³H]oestradiol by the preoptic-hypothalamic area decreased exponentially in the tissue from 30 min to 4 h after an intraperitoneal injection; however, nuclear binding reached a peak at 1-2 h and still showed high retention at 4 h. These results, together with observations in other laboratories of morphological changes induced by oestrogens, establish that certain regions of the brain are bona fide targets for the action of oestradiol.     

Quantitative electron microscopic autoradiography on the mouse thyroid gland after administration of 3H-L-DOPA

Summary The cellular and subcellular distribution of radioactivity in the mouse thyroid gland different times (20 min — 8 hours) after intravenous administration of ³H-L-DOPA was studied by means of quantitative electron microscopic autoradiography.High concentrations of autoradiographic silver grains occur over parafollicular cells and adrenergic nerves while the labelling of follicular cells and lumina is low or absent and similar to the labelling of connective tissue cells at all observation times.Over the parafollicular cells high levels of radioactivity can be recorded already 20 min after administration of the labelled amino acid. The grain counts are highest at 1 hour and decrease then at 2.5 and 8 hours.The intracellular distribution of label is similar at all observation times; thus, the concentration of silver grains over the typical cytoplasmic granules of the parafollicular cells is 4–5 times higher compared to the concentration over the remainder of the cytoplasm and the nucleus.Treatment with a decarboxylase inhibitor prior to the injection of ³H-L-DOPA results in a low and uniform labelling of all thyroid cells. This finding, taken together with the observation that also pretreatment with reserpine abolishes the autoradiographic reaction over the cytoplasmic granules, gives strong support to the idea that the great majority of silver grains observed over parafollicular cells represents dopamine formed by decarboxylation of the labelled precursor.This study was supported by grant K71-12X-3352-01 from the Swedish Medical Research Council. The author wishes to express his gratitude to Mrs. Gunnel Bokhede and Miss Dala Sjögren for expert technical assistance.     

Purkinje cell maturation in the light of RNA synthesis

Wistar rats 1- to 90-day-old received an injection of 3H-uridine and were killed 20 min to 44 h later. Autoradiographic examination revealed the highest grain count densities in Purkinje cell nuclei around postnatal day (PD) 6 while the incidence of labelled nuclei stayed at the peak values till PD 15. Silver staining of Purkinje cell nuclei showed that the expression of nucleolar r-RNA coding genes is maximal at PD 15; in some cells it even slightly exceeds adult values. After PD 15, the percentage of labelled Purkinje cell nuclei declined; this was more pronounced in the nucleolar region than outside the nucleolus. The percentage of cells with cytoplasmic labelling culminated on PD 15. The highest grain counts were found in Purkinje cell cytoplasm on PD 6 at 44 h p.i. interval. Reversal in nuclear grain counts at 2 and 6 h p.i. intervals observed between PD 15 and PD 25 suggests faster degradation, or processing and export, of a newly synthesized nuclear RNA in these age groups. Frequency distribution analysis of grain count densities revealed a small group of Purkinje cells with higher incorporation of 3H-uridine both in the nucleolar region and the whole nucleus at PD 15. In situ hybridization of 3H-r-RNA revealed a slight binding excess to DNA of some Purkinje cell nuclei but not in granule cells of 1-month-old rats. These data, together with those published recently by Brodsky et al. (1985), indicate an uneven structural organization and partial overexpression of the genom coding r-RNA synthesis in the population of Purkinje cells.     

KINETICS OF THE INCORPORATION OF SOME TRITIATED NUCLEIC ACID PRECURSORS AND [3H]LYSINE INTO MOUSE BRAIN CELLS FOLLOWING INTRAPERITONEAL INJECTION

Abstract— Intraperitoneal injection into white mice of the same amount of radioactivity (0.5 mCi) of [³H]uridine and [³H]lysine demonstrated by autoradiography that there was a much greater labelling of nerve cells from lysine than from uridine. For uridine, the choroid plexus cell nuclei gave maximal labelling within 1 h, with a decrease after 6 h. The plexus nuclei of lysine-injected animals gave almost the same amount of labelling during the experimental period of 48 h. In nerve cells, labelling from uridine increased in the nuclei up to 18 h after injection and there was an almost parallel increase in the labelling in the cytoplasm and neuropil. These results are compared with earlier reports on the results from intravenous injection of uridine. In lysine-injected animals the nerve cell nuclei and cytoplasm showed a fairly constant amount of label over 48 h, but the neuropil counts increased steeply. The activity of the blood was determined by scintillation counting during the 48-h period, and, as with uridine injection, was found to be almost constant over this period. A small series of animals was injected with 0.5 mCi of [³H]uracil, [³H]guanine, [³H]guanosine or [³H]cytidine for comparison. The autoradiograms from animals injected with these bases showed very slight labelling; that from guanosine was heavy in plexus nuclei, slight in nerve cells, and from cytidine it was heavy in plexus cells and moderate in nerve cells.     

Factors promoting vitellogenic competence and yolk deposition in the cockroach ovary: The post-ecdysis female

Incorporation of labelled histidine into a follicle cell product was shown to be juvenile hormone-dependent in Periplaneta americana. After 30 min the labelling was localized in the follicle cell cytoplasm, followed by labelling of intercellular spaces between the follicle cells after 60 min, and finally labelling of cortical yolk spheres after 120 min. Synthesis of the follicle cell product was observed exclusively in ovarian follicles participating in yolk formation. DNA synthesis was observed in follicle cell nuclei of ovarian follicles from the early stages of follicle differentiation to chorion formation. The evidence indicates that, although ovarian follicles exhibit protein synthetic responses to juvenile hormone in adult females, incorporation of thymidine into follicular nuclei suggests that changes in the cell cycle after adult ecdysis do not necessarily affect sensitivity to juvenile hormone.     

Sensitivity of intranuclear glucocorticoid complex from normal rat liver and Zajdela ascites hepatoma to ionic strength and nuclease treatment

After 30 min of intraperitoneal injection of dexamethasone to adrenectomized rats about 15% of the label are incorporated into liver homogenate and only 1% of the cytosol-bound hormone is detected in the cell nuclei. The binding of the "in vitro" injected hormone by the nuclei does not obey the second-order reactions (the Scatchard plots). This is probably due to the existence of various ancillary mechanisms, which control the translocation of the hormone complex into cell nuclei at the level of cytoplasm and nuclear membranes. DNAase I, micrococcal nuclease and endogenous nucleolysis markedly increase the part of the nuclear hormone complex resistant to 0.4 M NaCl. In hepatocyte nuclei obtained by the collagenase method, the content of the 0.4 M NaCl-resistant receptor complex is also increased. The resistance of 0.4 M NaCl was also found in 80% of the glucocorticoid-insensitive nuclear complex from Zajdela ascite hepatoma cells. The changes in interaction of the hormone-receptor complex with nuclear acceptor sites eventually resulting in impaired sensitivity of host tissues to hormonal control can be due to the damage of chromatin structure induced by different influences and tumour growth.     

A negative image of the quiescent centre in regenerating root apices of Zea mays

Usually the presence of the quiescent centre in roots is demonstrated by the absence of labelled nuclei following treatment of the root with appropriate radioactive markers. By modification of the pulselabelling technique, a negative image of the quiescent center, showing more intense labelling from [³H]thymidine than the surrounding area, was obtained in regenerating root apices of Zea mays L.     

The effect of actinomycin D on RNA synthesis and nucleolar ultrastructure in the liver of rats treated with fluorouracil

Summary Rats were given cytidine-³H and 10 min later 50 mg fluorouracil. They were killed after 25 hours. Actinomycin D was given at various times before sacrifice. The collapse of the nucleolus and the segregation of its components, seen in rats sacrificed one hour after administration of actinomycin D only, was prevented by prior treatment with fluorouracil. In rats treated with fluorouracil and given actinomycin 12 or 20 hours prior to death, there was a more or less pronounced collapse of the nucleolus but no typical segregation of its components. Radioautographs of livers from untreated rats or rats given actinomycin only at the times mentioned, and killed 25 hours after administration of cytidine-³H, were labelled mainly over the cytoplasm. Radioautographs from rats, treated with fluorouracil only, or fluorouracil plus actinomycin, showed labelling over the nucleoli, but depressed labelling over the cytoplasm. Biochemical analysis of RNA labelling showed high ribosomal peaks in untreated rats and rats treated with actinomycin only. Rats treated with fluorouracil, or fluorouracil plus actinomycin showed no labelling of the 29S and 18S ribosomal peaks. The results indicate that fluorouracil blocks or delays the formation of ribosomal RNA and that the inhibition, at least in part, takes place in the nucleolus.This work was supported by grants from the Swedish Medical Research Council (Project K68-12X-623-04), the Swedish Cancer Society (Project 6831), the Medical Faculty of Uppsala and the Swedish Society for Medical Research.     

P32 UPTAKE BY NUCLEI

1. A method for isolating nuclei in quantity from mammalian tissues is described. 2. The rate of uptake of radioactive phosphorus by nuclei is found to be quite rapid. The phosphorus was shown not to be taken up by exchange. 3. Nuclei of tumors accumulate more radioactive phosphorus than normal liver nuclei. This was shown to be due to mitotic activity and not a form of metabolism peculiar to tumor cells. 4. The specific activities of nuclei and cytoplasm are compared. 5. 60 to 70 per cent of the nuclear radioactive phosphorus is present as nucleoprotein from 1 hour to 5 days after it is administered. In the lymphoma nuclei 90–95 per cent of the phosphorus is in the nucleoprotein fraction from 1–5 days after it is administered. 6. The specific activities of the nucleoprotein, lipid, and acid-soluble fractions of liver and tumor nuclei are compared. 7. From the rate of P₃₂ uptake by nuclei it is calculated that a new lymphoma nucleus is synthesized on the average once every 27 hours. This is in agreement with the observed rate of growth of the tumor. 8. In the lymphoma nucleus it is calculated that 7 x 10⁴ molecules of tetranucleotide are synthesized per second. 9. Irradiation with 200 r. x-rays alters the distribution of P₃₂ in the lymphoma cell, markedly increasing the concentration in the nucleus shortly after irradiation. The P₃₂ concentration in the cytoplasm decreases with time after irradiation. It is suggested that the altered distribution is correlated with the inhibition of mitosis produced by the x-rays. 10. Continual synthesis of nucleoprotein takes place even in nuclei of cells which do not undergo mitosis.     

Tritiated thymidine autoradiographic study of cell migration and renewal in the pyloric mucosa of golden hamsters

Summary The kinetics of cell proliferation, migration and renewal in the pyloric mucosa of golden hamsters were studied by flash, cumulative and pulse labelling autoradiography following ³H-thymidine injections.By flash labelling autoradiography, it was shown that the labelled epithelial cells are exclusively confined to a zone several cells wide in the region of the isthmus between the gastric pits and the pyloric glands. In the cumulative and pulse labelling experiments, this cell proliferation in the isthmus region was shown to be for replacement of both the surface epithelial and the glandular cells. The surface epithelial cells of the pyloric mucosa arising in the upper portion of the isthmus come to line the pits and the surface, and are sloughed off into the gastric lumen within a week. The mucin-containing glandular cells, which arise more deeply in the isthmus region, migrate downwards and are apparently lost at the deepest level of the glands. The life span of the mucin-containing glandular cells was estimated at about 14 days. This cell type appears to undergo renewal of the first produced, first lost pipe line variety. However, a small number of glandular cells was found to survive for more than 20 days (up to 30 days), suggesting the existence of a sub-population of cells with different kinetics in the pyloric glands.Supported by a Grant-in-Aid for Cancer Research from Ministry of Education, Science and Culture, Japan     

Distribution of the glio-interstitial system in molluscs

Summary Ten hamsters received repeated injections of ³H-thymidine for 4 days and were allowed to survive for 7, 28, 42 and 100 days. Changes in spatial distribution of the labelled cells and in labelling indices of each cell line in the gastric glands were studied at various days after ³H-thymidine injections, and the fate of the mucous neck cell, the replacement of the chief cell and the mode of cell migration were discussed.After 4 days of repeated injections of ³H-thymidine, the labelled parietal cells and the mucous neck cells were concentrated at the neck area. Starting from the neck area, they migrated an average of 3 micra downwards per day. By 42 days, they reached the middle level of the glands, where the labelled mucous neck cells decreased but the labelled chief cells increased in number. The differentiation of the chief cell then appears to take place at the middle level of the glands through transformation of the migratory mucous neck cells. After 4 days of the labelling, about 1.8% of the chief cells located in the lower part of the glands was found to undergo in situ replication. This indicates that the renewal of this cell type is partly assured by its own mitotic activity.The foveolar cell — the future surface epithelium — seems to migrate upwards along the long axis of the glandular tubule in the pipe line system, which means first produced, first migrates. After migrating out from the neck area, the parietal cell and the mucous neck cell (the future chief cell) take an average of 200 days to reach the lower end of the glands. In the process of migration, however, the cells produced contemporaneously at the neck area became scatteringly spread from the neck towards the bottom of the gland. The time required for the newly-formed cells to reach the lower end of the gland varied between 100 and 300 days. In the gastric glands the cells first produced at the neck area do not first reach the lower end of the glands. This mode of random migration is referred to as the stochastic flow system. As one of the probable factors which disturb the pipe line flow of downward cell migration, cellular movements perpendicular to the long axis of the glandular tubule were suggested to occur at random at an any level of the gastric glands.Supported by a Grant-in-Aid for Cancer Research from the Ministry of Education, Science and Culture, Japan     

Immunocytochemical localization of bovine serum albumin (BSA) in the liver and testis of rats injected with testosterone-BSA, hydrocortisone-BSA or corticosterone-BSA

Through observations of colloidal gold with silver enhancement, we have demonstrated that 2-nm colloidal gold labeled-testosterone-bovine serum albumin (BSA) conjugate or hydrocortisone-BSA conjugate injected intravenously enters the hormone-target cell nuclei of rats (Nishimura and Ichihara, 1997; Nishimura and Nakano, 1997, 1999). To confirm immunocytochemically whether the nature of BSA in the steroid hormone-BSA conjugates (steroid-BSAs) remains intact in the hormone-target cell nuclei, testosterone-BSA, hydrocortisone-BSA or corticosterone-BSA was injected into the vascular system of rats, then the liver and testes of rats killed 2 h postinjection were reacted with FITC-conjugated anti-BSA antibody, and examined under fluorescence microscopy and confocal laser scanning microscopy. In the liver of rat injected with testosterone-BSA, the fluorescence was observed in the nuclei of endothelial cells, but not in the nuclei of hepatocytes, hepatic stellate cells and Kupffer cells. In the liver of rat injected with hydrocortisone-BSA, intense fluorescence was seen in the nuclei of hepatic stellate cells, but did not seem to be present in the nuclei of the other three kinds of cells. In the liver of rat injected with corticosterone-BSA, the fluorescence seemed to be in a few nuclei of hepatic stellate cells, and appeared as speckles in a few nuclei of the hepatocytes and Kupffer cells. In some seminiferous tubules of rat injected with testosterone-BSA, fluorescence was observed in the nuclei of spermatocytes and spermatids. These results suggest that BSA conjugated with steroid hormone can enter the hormone-target cell nuclei with its antigenicity kept intact, and that the fate of steroid-BSAs is decided at the cell membrane level.     

Cytochemical observations of coelomocytes from the earthworm, Lumbricus terrestris.

Synopsis Coelomocytes of the earthworm,Lumbricus terrestris, were stained by cytochemical techniques to determine the biochemical composition of the seven different cell types and subtypes. The enzymes acid phosphatase and -glucuronidase are present in all types of coelomocytes, but are especially abundant in basophils and neutrophils; the differences in enzyme amounts correlate well with the differences in phagocytic activity of the various cell types. No peroxidase is present. The cytoplasmic basophilia of basophils is due primarily to ribonucleic acid. Basophils also contain large deposits of glycogen, with neutrophils and chloragogen cells containing somewhat lesser amounts. The predominant granules of the two types of acidophils and of granulocytes are composed of a basic protein and a neutral mucopolysaccharide or glycoprotein. A second granule population, present in low numbers in acidophils and granulocytes, but in larger numbers in basophils and neutrophils, is small in size and lipid-positive and may, in part, represent lysosomes.Lipid is especially abundant in the vesicles and granules of the two types of chloragogen cells. Some granules of chloragogen cells also contain ferrous and ferric iron and a substance with pseudoperoxidase activity. The cytoplasm contains protein, glycogen, and a neutral mucopolysaccharide. In addition, acid mucopolysaccharides are variably present in the cytoplasm of chloragogen cells, the only coelomocytes to contain this class of substances.