Chick gonadogenesis following early surgical bursectomy. I. Histological, morphometric and histochemical data on the embryonic left ovary.

Left ovaries of bursectomized chick embryos were examined on the 17th day of incubation in comparison to normal and sham-operated controls, by histological and histochemical observations. The results show that in bursectomized embryos the cortex appears irregularly developed, with a significant decrease in the mean thickness and in the percentage of the secondary sex cords in the total cortical area. Furthermore, the germinal epithelium appears thicker and the subcortical medulla and the tunica albuginea more compact. The greater activity of the enzyme delta 5-3 beta-hydroxysteroid dehydrogenase (delta (5)3 beta HSD) found in ovaries of bursectomized embryos (histochemical method) could be related to an endocrine dismetabolism affecting the cortical development. On the basis of these results and those of other authors, some hypotheses are advanced. In particular, an action of the bursal factor on GTH receptors could be the factor responsible of the enhanced steroidogenic activity altering the hormonal environment.

Chick gonadogenesis following early surgical bursectomy. II. Ultrastructural data on the embryonic left and right female gonads.

We examined the ultrastructural characteristics of the medullary steroidogenic cells in left and right female gonads of surgically bursectomized chick embryos killed on the 17th day of incubation. The steroidogenic cells of the bursectomized embryos have a more developed system of cisternae in the smooth endoplasmic reticulum than controls, and their mitochondria show some alterations in the density of the matrix and in the shape of the cristae. On the basis of these results, an enhancement of the steroidogenic activity in both gonads is suggested.

Immunohistochemical localization of oestrogen receptor alpha in the various cell categories of chick embryo ovary.

The immunohistochemical (IHC) localization of oestrogen receptor alpha (ERα) was studied in the developing left ovary of 14.5-day-old chick embryos. The study was focused in particular on distinguishing in cortex and medulla the different cell categories that proved positive to the reaction, in order to gain further understanding of gonadal cell interactions during ovarian development. Immunostained cells were observed in both the cortex and medulla, but the reactivity for ERα was discontinuous, probably due to variable cell requirements. In the cortex, positivity was observed in cells of the ovarian surface epithelium, in germ cells and in prefollicular cells. In the medulla, positivity was found in the following cell categories: interstitial cells, poorly differentiated somatic cord cells, including those delimiting lacunae, germ cells and their accompanying cells of epithelial origin. Furthermore, the IHC results showed that the intracellular localization of the antigen was cytoplasmic, nuclear, or both. The significance of ERα presence and intracellular localization was discussed in relation and as supplementary to previous research by various Authors. In particular, as regards the unusual cytoplasmic immunoreactivity, a gradual shift of ERα localization from cytoplasmic to nuclear during the embryonic period is suggested.

Immunohistochemical localization of nNOS in the head kidney of larval and juvenile rainbow trout, Oncorhynchus mykiss.

The aim of this investigation was to assess whether in teleosts, as in mammals, nitric oxide (NO) is involved in the regulation of cellular activity in the adrenal homolog. Larval and juvenile stages of the rainbow trout, Oncorhynchus mykiss, were used, in which the adrenal homolog consists of chromaffin adrenergic and interrenal steroidogenic cells localized mainly in the head kidney where there are also ganglion cells and nerve fibres that innervate the gland. In 12-month-old juveniles, the immunohistochemical reaction for neuronal nitric oxide synthase (nNOS), which catalyzes the synthesis of NO, revealed the presence of this enzyme in some nerve fibres and ganglion cells and only rarely in chromaffin cells. The latter are identified by the immunohistochemical reaction for tyrosine hydroxylase (TH) and phenylethanolamine-N-methyltransferase (PNMT). In larvae at 27 days postfertilization, numerous cells dispersed in the head kidney are nNOS positive, whereas the TH and PNMT positive cells are very rare. At hatching (31 days postfertilization), the positivity for nNOS in the cells of the head kidney disappears and reappears at 60 days posthatching in some nerve cells and fibres. These results suggest an involvement of NO in the regulation of adrenal function as in mammals and the nature of nNOS positive cells present in the head kidney of larvae of 27 days is discussed.

Adrenaline-, noradrenaline- and small granule- containing cells in the adrenal gland of Discoglossus pictus (Amphibia, Anura).

In the adrenal gland of Discoglossus pictus, various types of chromaffin cells are described: noradrenaline cells, adrenaline cells and small granule-containing cells (on the basis of electron density and shape of the granules). The chromaffin cells occur in small groups, and have cytoplasmic processes which may surround them in the form of parallel layers. Their nerve supply is sparse. The possible function of SGC-cells, in relation to those described in other vertebrates, is discussed.

Comparative data on acetylcholinesterase cytochemistry in various chromaffin cell types of Discoglossus pictus (Amphibia, Anura).

The cytochemical localization of acetylcholinesterase (AChE) activity was studied in the adrenal chromaffin cells of Discoglossus pictus. Reaction end-products were associated with all types of chromaffin cells, i.e. adrenaline (A), noradrenaline (N) and small granule chromaffin (SGC-A, SGC-N) cells, and nervous elements present in the gland. The SGC-A and SGC-N showed the same intensity of AChE reaction in A and N cells, respectively. On the whole, the A and SGC-A cells were more reactive than the N and SGC-N cells. The functional role of the SGC cells is discussed on the basis of the cytochemical results.

Ultrastructural and histochemical study on the interrenal cells of the male stickleback (Gasterosteus aculeatus, Teleostea), in relation to the reproductive annual cycle.

The steroidogenic interrenal cells in the adrenal homologue of the male stickleback (Gasterosteus aculeatus) were studied in relation to the reproductive cycle by means of histological and ultrastructural observations, and using histochemical methods for the localisation of 3beta-hydroxysteroid dehydrogenase (3betaHSD) and 17beta-hydroxysteroid dehydrogenase (17betaHSD). To determine the various stages of the reproductive cycle, the testes were also examined by histological and histochemical methods (3betaHSD). The results indicate that in this teleost the interrenal cells can undergo a cycle in which phases characterised by different cytological aspects are observed. During this cycle there is a renewal of organelles, in particular mitochondria and SER. Periodic degenerative processes are also found. Organelle cytology showed that the cell cycle has at least 3 different aspects during the year. An analogy with some cytological aspects of the adrenal zonation in mammals is possible. It is postulated that the interrenal gland activity could substitute or supplement androgen production by the testes.

Fine localization of the acetylcholinesterase activity in somatic and germ cells during the morphogenesis of chick ovarian cortex.

The ultracytochemical localization of the enzyme acetylcholinesterase (AChE) was studied in the germinal epithelium and cortex during chick ovarian morphogenesis at the 7, 12.5, 14 and 19 day of incubation. The results evidenced at day 7 the presence of the enzyme in some somatic cells, both "dark" and "light" and in some gonocytes. The reaction appears also in some tracts between these types of cells. At day 12.5 the reaction is present in some somatic cells only towards the deepest zone of the cortex, in many oogonia and oocytes and in some tracts between germ and somatic cells. At day 14 and day 19 the various cell categories of the cortex are negative for the reaction. The significance of the presence of the enzyme is discussed in relation to an embryonic cholinergic system active during morphogenesis. Regarding the presence of the enzyme in germ cells, the positivity in endoplasmic reticulum cisternae associated to mitochondria may suggest an implication of the enzymatic activity in the proliferation and transformation of these organelles.

Acetylcholinesterase activity in the adrenal chromaffin vesicles of Urodela.

The presence of acetylcholinesterase (AChE) activity in the adrenal chromaffin cells of Necturus maculosus and Ambystoma maculatum (Amphibia, Urodela) has been demonstrated by cytochemical method at the electron microscope level. The enzymatic activity is localized in RER and perinuclear cisternae, on the plasma membrane and within the chromaffin vesicles, both in adrenaline (A) and noradrenaline (N) cells. Moreover N cells appear to be more reactive than A cells and Necturus more reactive than Ambystoma. The possible function of the AChE activity inside the vesicles is discussed as a mechanism of protons donor or as peptidasic activity acting on various peptides present in the vesicle.

Immunohistochemical and ultrastructural evidence of adrenal chromaffin cell subtypes in sea bass Dicentrarchus labrax (L.).

Histology demonstrated chromaffin cells in the head kidney and opisthonephros of the sea bass. Immunohistochemistry showed the catecholamine-synthesizing enzymes tyrosine hydroxylase and dopamine beta-hydroxylase in these cells. Phenlethanolamine N-methyltransferase was detected in a fraction of the chromaffin cells in the head kidney (59%) and in the opisthonephros (54%). Distinct noradrenaline- and adrenaline-synthesizing cells are therefore suggested. Ultrastructural analysis confirmed the existence of two main chromaffin cell types, distinguished by different types of the secretory granules. Cells that contain vesicles with a round, electron-dense core were interpreted as noradrenaline cells, while cells with electron-lucent vesicles were identified as adrenaline cells. The ultrastructure of these chromaffin cell subtypes does not differ in the head kidney and opisthonephros. A minor population of chromaffin cells was identified, which typically show smaller vesicles with an electron-dense core. This population may account for a limited number of not fully differentiated chromaffin cells.

The adrenal chromaffin cells of Salmo gairdneri Richardson (Teleostei, Salmonidae).

The chromaffin cells of the adrenal homologue of Salmo gairdneri R. have been studied by light and electron microscopy. The chromaffin tissue was localised in the head kidney adjacent to the wall of the proximal part of the cardinal veins and their main branches. Specific histochemical techniques failed to demonstrate different types of chromaffin cells. With the electron microscope two different types of chromaffin cells were observed. The first type, characterised by numerous dense cytoplasmic granules of average diameter 90 nm, was interpreted as a noradrenalin cell; the second type was characterised by the presence of moderately electron-dense granules of average diameter of 85 nm, and was interpreted as an adrenalin cell. The reaction for acetylcholinesterase activity was present on nerve terminals and sometimes in the vacuolar membrane systems of both chromaffin cells.

The adrenal gland of Euproctus (Urodela, Salamandridae): comparison of three species and phylogenetic inferences.

Adrenal glands of three species of Euproctus (E. asper, E. montanus and E. platycephalus) were compared. Differences were observed as regards: (1) the distribution of the adrenal tissue, which is more sparse in E. asper than in the other two species; (2) the amount and size of the adrenal islets: in E. asper, they are numerous and small, in E. platycephalus, they are decreased in number and larger, in E. montanus they are few and very large; (3) the distance of the islets from the medial border of the kidney is variable in E. asper; in E. montanus they are mostly in contact with the medial edge, whereas in E. platycephalus they are distant from it. The adrenals of E. montanus and E. platycephalus are more similar to each other than to that of E. asper; the gland in this species may be considered a more primitive type. Such structural relationships are in agreement with phylogenetic inferences concerning the three species.

Cytological and biochemical studies on chromaffin cells in the head kidney of Gasterosteus aculeatus (Teleostei, Gasterosteidae).

Adrenal chromaffin cells in the head kidney of the stickleback Gasterosteus aculeatus were examined by ultrastructural analysis and determination, by HPLC, of the catecholamine content (adrenaline, noradrenaline, and dopamine). Chromaffin cells are characterized by the presence of membrane-bound vesicles and two cell types were identified: (i) those with vesicles containing a strongly electron-dense core; and (ii) those with vesicles that are completely electron lucent or containing smaller and less-dense granules. HPLC analysis revealed the presence of adrenaline and noradrenaline in both right and left head kidneys. Dopamine was not detected. Cytological and biochemical data suggest that the cells with electron-dense granules contain noradrenaline and those with electron-lucent vesicles contain adrenaline.