Effects of Deoxynivalenol and Zearalenone on the Histology and Ultrastructure of Pig Liver.

The purpose of this study was to determine the effects of single and combined administrations of deoxynivalenol (DON) and zearalenone (ZEN) on the histology and ultrastructure of pig liver. The study was performed on immature gilts, which were divided into four equal groups. Animals in the experimental groups received DON at a dose of 12 µg/kg body weight (BW) per day, ZEN at 40 µg/kg BW per day, or a mixture of DON (12 µg/kg BW per day) and ZEN (40 µg/kg BW). The control group received vehicle. The animals were killed after 1, 3, and 6 weeks of experiment. Treatment with mycotoxins resulted in several changes in liver histology and ultrastructure, including: (1) an increase in the thickness of the perilobular connective tissue and its penetration to the lobules in gilts receiving DON and DON + ZEN; (2) an increase in the total microscopic liver score (histology activity index (HAI)) in pigs receiving DON and DON + ZEN; (3) dilatation of hepatic sinusoids in pigs receiving ZEN, DON and DON + ZEN; (4) temporary changes in glycogen content in all experimental groups; (5) an increase in iron accumulation in the hepatocytes of gilts treated with ZEN and DON + ZEN; (6) changes in endoplasmic reticulum organization in the hepatocytes of pigs receiving toxins; (7) changes in morphology of Browicz-Kupffer cells after treatment with ZEN, DON, and DON + ZEN. The results show that low doses of mycotoxins used in the present study, even when applied for a short period, affected liver morphology.

Pinopsin and photoreception in the pineal organ of the domestic turkey during post-hatching development.

The avian pineal organ is photosensitive because of the presence of photopigments, of which pinopsin seems to be one of the most important. This organ is subject to far-reaching changes during post-hatching development, but evidence regarding pinopsin presence and direct photoreception during this time is lacking. This study was carried out to demonstrate the following: 1) the structures showing immunoreactivity to pinopsin in the turkey pineal organ, 2) the changes of these structures during development, 3) the pinopsin localization in pinealocytes in monolayer cultures, and 4) the role of direct photoreception in the regulation of melatonin secretion in pineal organs in adult turkeys. Pinopsin immunoreactivity was localized in the apical extensions of columnar cells limiting the follicular lumen, in fiber-like structures located between columnar cells in the inner part of follicle wall, in string-shapes or small spherical structures distributed in the outer part of follicle wall and in amorphous material inside the follicle lumen. In young birds, immunoreactivity was also sporadically noted in cell bodies of rudimentary receptor pinealocytes. The distribution of pinopsin showed prominent age-dependent changes, including a subsequent increase in pinopsin-positive structures in the outer part of the follicle wall and a prominent reduction in the number and size of positive apical extensions in 40- and 56-week-old turkeys. These data demonstrate that the role of secretory pinealocytes in pineal photoreception increases with age. In monolayer cultures, all pinealocytes showed strong reactions in club- or bulbous-shaped prolongations. The pineal organs of adult birds were less sensitive to light exposition at night than those of young turkeys, which points to differences in light sensitivity between rudimentary receptor and secretory pinealocytes. However, direct photoreception could play an important role in the regulation of melatonin secretion in adult turkeys.

Developmental morphology of the turkey pineal organ. Immunocytochemical and ultrastructural studies.

Our previous study showed that the turkey pineal organ, in contrast to that of the chicken, is characterized by a follicular structure throughout the entire period of post-hatching life. Despite the preservation of the follicular organization, the histological structure of the pineal follicles in turkeys changes prominently with age. The present research was performed to investigate the cellular composition and organization of the follicle wall as well as the ultrastructure of parenchymal cells in the turkey pineal organ during the period of post-hatching development. Pineal organs were collected from female turkeys at 2 days, 2 weeks, 4 weeks, 10 weeks, 20 weeks, 30 weeks, 40 weeks, and 56 weeks post-hatching. The organs were prepared for immunocytochemical studies using antibodies against N-acetylserotonin O-methyltransferase (ASMT), glial fibrillary acidic protein (GFAP) and proliferating cell nuclear antigen (PCNA) and for ultrastructural examination. The results showed that regardless of age, the pineal follicle was formed by ASMT-immunopositive cells, among which rudimentary photoreceptor and secretory pinealocytes were identified. The second component of the follicle wall consisted of GFAP-immunopositive cells, as represented by ependymal-like and astrocyte-like cells. Rudimentary photoreceptor pinealocytes and ependymal-like cells formed the inner part of the follicle wall, while secretory pinealocytes and astrocyte-like cells created the outer part. Three forms of the pineal follicle structure characteristic of young (two days to ten weeks), young adult (20-30 weeks) and adult (40-56 weeks) turkeys were distinguished. These forms primarily differed in the relative dimensions of the inner and outer parts of the follicle wall. Ultrastructural studies showed prominent changes in the organization of rudimentary receptor pinealocytes during the investigated period of life. These cells developed until the age of 20 weeks, at which time they appeared as strongly elongated cells with a stratified, highly regular distribution of organelles. In adult turkeys, rudimentary receptor pinealocytes showed pronounced regressive changes; however, we never observed their transformation into cells of the secretory type. Secretory pinealocytes increased in number and size during the post-hatching period, which was especially pronounced after 20 weeks of age. The most prominent changes in the supporting cells included the intensification of GFAP-immunoreactivity due to the accumulation of filaments in the cytoplasm and the development of astrocyte-like cells. The increase in the number of secretory pinealocytes and astrocyte-like supporting cells resulted in the formation of two distinct parts of the follicle wall in the pineal organs of young adult and adult turkeys.

The Effects of Deoxynivalenol and Zearalenone on the Pig Large Intestine. A Light and Electron Microscopy Study.

The contamination of feed with mycotoxins results in reduced growth, feed refusal, immunosuppression, and health problems. Deoxynivalenol (DON) and zearalenone (ZEN) are among the most important mycotoxins. The aim of the study was to examine the effects of low doses of these mycotoxins on the histological structure and ultrastructure of the large intestine in the pig. The study was performed on 36 immature gilts of mixed breed (White Polish Big × Polish White Earhanging), which were divided into four groups administrated per os with ZEN at 40 µg/kg BW, DON at 12 µg/kg BW, a mixture of ZEN (40 µg/kg BW) and DON (12 µg/kg BW) or a placebo. The pigs were killed by intravenous overdose of pentobarbital after one, three, and six weeks of treatment. The cecum, ascending and descending colon samples were prepared for light and electron microscopy. Administration of toxins did not influence the architecture of the mucosa and submucosa in the large intestine. ZEN and ZEN + DON significantly decreased the number of goblet cells in the cecum and descending colon. The mycotoxins changed the number of lymphocytes and plasma cells in the large intestine, which usually increased in number. However, this effect differed between the intestine segments and toxins. Mycotoxins induced some changes in the ultrastructure of the mucosal epithelium. They did not affect the expression of proliferative cell nuclear antigen and the intestinal barrier permeability. The obtained results indicate that mycotoxins especially ZEN may influence the defense mechanisms of the large intestine.

Feline Infectious Peritonitis: Immunohistochemical Features of Ocular Inflammation and the Distribution of Viral Antigens in Structures of the Eye.

Feline infectious peritonitis (FIP) is a serious, widely distributed systemic disease caused by feline coronavirus (FCoV), in which ocular disease is common. However, questions remain about the patterns of ocular inflammation and the distribution of viral antigen in the eyes of cats with FIP. This study characterized the ocular lesions of FIP including the expression of glial fibrillary acidic protein and proliferating cell nuclear antigen by Müller cells in the retina in cases of FIP and to what extent macrophages are involved in ocular inflammation in FIP. Immunohistochemistry for FCoV, CD3, CD79a, glial fibrillary acidic protein, calprotectin, and proliferating cell nuclear antigen was performed on paraffin sections from 15 naturally occurring cases of FIP and from controls. Glial fibrillary acidic protein expression was increased in the retina in cases of FIP. Müller cell proliferation was present within lesions of retinal detachment. Macrophages were present in FIP-associated ocular lesions, but they were the most numerous inflammatory cells only within granulomas (2/15 cats, 13%). In cases of severe inflammation of the ciliary body with damage to blood vessel walls and ciliary epithelium (3/15, 20%), some macrophages expressed FCoV antigens, and immunolabeling for calprotectin on consecutive sections suggested that these FCoV-positive macrophages were likely to be recently derived from blood. In cases of severe and massive inflammation of most ocular structures (4/15, 26%), B cells and plasma cells predominated over T cells and macrophages. These results indicate that gliosis can be present in FIP-affected retinas and suggest that breakdown of the blood-ocular barrier can allow FCoV-bearing macrophages to access the eye.

Adrenergic regulation of cytoplasmic structures related to secretory processes in pig pinealocytes-an ultrastructural, quantitative study.

Two structures, considered as secretory in nature, are present in the pinealocytes in of the domestic pig show the presence of two structures, which are considered as secretory in nature - the dense core vesicles (DCV) and the membrane bounded (dense) bodies (MBB). The latter are extremely numerous in pig pinealocytes (they occupy 6-20% of the cytoplasm), and the number of MBB changes under different physiological and experimental conditions. Norepinephrine is the main neurotransmitter that regulates the secretion of pineal melatonin. The present study was carried out to 1) clarify whether the DCV and their source - the Golgi apparatus (GA) - as well as the MBB are controlled by norepinephrine, 2) determine the effect of adrenergic stimulation on these structures, and 3) identify the receptors involved in the regulation of these structures. The studies were performed using a static organ culture of pig pineal explants. The explants were incubated in a control medium between 08:00 and 20:00 and in a medium with 10µM norepinephrine or alpha- or beta-adrenoceptor agonists between 20:00 and 08:00 on five consecutive days. The tissues were subsequently prepared for ultrastructural analysis. The results distinctly showed that the DCV, GA and MBB in pig pinealocytes are under adrenergic control. The stimulation of the beta-adrenoceptors resulted in an increase in the numerical density of the DCV and a decrease in the relative volume of the GA in the perikarya, while the incubation with agonists of the alpha1-adrenoceptors was ineffective. The relative volume of the MBB in the perikarya significantly decreased after treatment with both beta-agonists and alpha1-agonists, which suggested the involvement of two types of adrenoceptors in the regulation of these structures.

Histological structure of duodenum in gilts receiving low doses of zearalenone and deoxynivalenol in feed.

Deoxynivalenol (DON) and zearalenone (ZEN), produced by microfungi of the Fusarium family, are among the most commonly occurring mycotoxins. They are considered important factors affecting human and animal health as well as livestock productivity. The aim of this study was to determine the effect of low doses of these mycotoxins on the histological structure of the pig duodenum. The study was performed on 72 gilts, with initial weights of approximately 25kg, divided into 4 equal groups. Group I received per os ZEN (40µg/kg BW), group II-DON (12µg/kg BW), group III-ZEN (40µg/kg BW) and DON (12µg/kg BW), and group IV-vehicle. The pigs were killed after 1, 2, 3, 4, 5 and 6 weeks of the treatment, and the duodenum samples were prepared for histological investigations. The slides were digitalized and subjected to morphometrical analysis. The treatment with DON and ZEN did not change the architecture of the mucosa or the ratio between goblet and adsorptive cells in the epithelium. The administration of DON induced an increase in the number of lymphocytes in the mucosal epithelium. Both mycotoxins, administered alone or together, increased the quantity of lymphocytes, plasma cells and macrophages with black-brown granules in the lamina propria. The time-courses of changes in the number of defense system cells evoked by DON and ZEN were different. In conclusion, dietary exposure to low doses of Fusarium mycotoxins should be considered an important risk factor for subclinical inflammation in the small intestine.

The effects of low doses of two Fusarium toxins, zearalenone and deoxynivalenol, on the pig jejunum. A light and electron microscopic study.

Immature gilts were administered per os with zearalenone (ZEN) at 40 µg/kg BW (group Z, n = 9), deoxynivalenol (DON) at 12 µg/kg BW (group D, n = 9), a mixture of ZEN and DON (group M, n = 9) or a placebo (group C, n = 9) over a period of six weeks. The pigs were sacrificed after one, three, or six weeks of the treatment (12 pigs per each time-point). Histological investigations revealed an increase in the mucosal thickness and the crypt depth as well as a decrease in the ratio of the villus height to the crypt depth in groups D and M after six weeks of exposure to the mycotoxins. The number of goblet cells in the villus epithelium was elevated in groups Z and M after one week and in group D after three weeks. The administration of ZEN increased the lymphocyte number in the villus epithelium after 1 week and the plasma cell quantity in the lamina propria after one, three, and six weeks of the experiment. DON treatment resulted in an increase in the lymphocyte number in the villus epithelium and the lamina propria after six weeks, and in the plasma cell quantity in the lamina propria after one, three, and six weeks of exposure. In group M, lymphocyte counts in the epithelium and the lamina propria increased significantly after six weeks. Neither mycotoxin induced significant adverse changes in the ultrastructure of the mucosal epithelium and the lamina propria or in the intestinal barrier permeability. Our results indicate that immune cells are the principal target of low doses of ZEN and DON.

Adrenergic activation of melatonin secretion in ovine pineal explants in short-term superfusion culture occurs via protein synthesis independent and dependent phenomena.

The ovine pineal is generally considered as an interesting model for the study on adrenergic regulation of melatonin secretion due to some functional similarities with this gland in the human. The present investigations, performed in the superfusion culture of pineal explants, demonstrated that the norepinephrine-induced elevation of melatonin secretion in ovine pinealocytes comprised of two subsequent periods: a rapid increase phase and a slow increase phase. The first one included the quick rise in release of N-acetylserotonin and melatonin, occurring parallel to elevation of NE concentration in the medium surrounding explants. This rapid increase phase was not affected by inhibition of translation. The second, slow increase phase began after NE level had reached the maximum concentration in the culture medium and lasted about two hours. It was completely abolished by the treatment with translation inhibitors. The obtained results showed for the first time that the regulation of N-acetylserotonin synthesis in pinealocytes of some species like the sheep involves the on/off mechanism, which is completely independent of protein synthesis and works very fast. They provided strong evidence pointing to the need of revision of the current opinion that arylalkylamines N-acetyltransferase activity in pinealocytes is controlled exclusively by changes in enzyme abundance.

Influence of electric, magnetic, and electromagnetic fields on the circadian system: current stage of knowledge.

One of the side effects of each electrical device work is the electromagnetic field generated near its workplace. All organisms, including humans, are exposed daily to the influence of different types of this field, characterized by various physical parameters. Therefore, it is important to accurately determine the effects of an electromagnetic field on the physiological and pathological processes occurring in cells, tissues, and organs. Numerous epidemiological and experimental data suggest that the extremely low frequency magnetic field generated by electrical transmission lines and electrically powered devices and the high frequencies electromagnetic radiation emitted by electronic devices have a potentially negative impact on the circadian system. On the other hand, several studies have found no influence of these fields on chronobiological parameters. According to the current state of knowledge, some previously proposed hypotheses, including one concerning the key role of melatonin secretion disruption in pathogenesis of electromagnetic field induced diseases, need to be revised. This paper reviews the data on the effect of electric, magnetic, and electromagnetic fields on melatonin and cortisol rhythms-two major markers of the circadian system as well as on sleep. It also provides the basic information about the nature, classification, parameters, and sources of these fields.

Diurnal profiles of melatonin synthesis-related indoles, catecholamines and their metabolites in the duck pineal organ.

This study characterizes the diurnal profiles of ten melatonin synthesis-related indoles, the quantitative relations between these compounds, and daily variations in the contents of catecholamines and their metabolites in the domestic duck pineal organ. Fourteen-week-old birds, which were reared under a 12L:12D cycle, were killed at two-hour intervals. The indole contents were measured using HPLC with fluorescence detection, whereas the levels of catecholamines and their metabolites were measured using HPLC with electrochemical detection. All indole contents, except for tryptophan, showed significant diurnal variations. The 5-hydroxytryptophan level was approximately two-fold higher during the scotophase than during the photophase. The serotonin content increased during the first half of the photophase, remained elevated for approximately 10 h and then rapidly decreased in the middle of the scotophase. N-acetylserotonin showed the most prominent changes, with a more than 15-fold increase at night. The melatonin cycle demonstrated only an approximately 5-fold difference between the peak and nadir. The 5-methoxytryptamine content was markedly elevated during the scotophase. The 5-hydroxyindole acetic acid, 5-hydroxytryptophol, 5-methoxyindole acetic acid and 5-methoxytryptophol profiles were analogous to the serotonin rhythm. The norepinephrine and dopamine contents showed no significant changes. The DOPA, DOPAC and homovanillic acid levels were higher during the scotophase than during the photophase. Vanillylmandelic acid showed the opposite rhythm, with an elevated level during the daytime.

Light and electron microscopy of the European beaver (Castor fiber) stomach reveal unique morphological features with possible general biological significance.

Anatomical, histological, and ultrastructural studies of the European beaver stomach revealed several unique morphological features. The prominent attribute of its gross morphology was the cardiogastric gland (CGG), located near the oesophageal entrance. Light microscopy showed that the CGG was formed by invaginations of the mucosa into the submucosa, which contained densely packed proper gastric glands comprised primarily of parietal and chief cells. Mucous neck cells represented <0.1% of cells in the CGG gastric glands and 22-32% of cells in the proper gastric glands of the mucosa lining the stomach lumen. These data suggest that chief cells in the CGG develop from undifferentiated cells that migrate through the gastric gland neck rather than from mucous neck cells. Classical chief cell formation (i.e., arising from mucous neck cells) occurred in the mucosa lining the stomach lumen, however. The muscularis around the CGG consisted primarily of skeletal muscle tissue. The cardiac region was rudimentary while the fundus/corpus and pyloric regions were equally developed. Another unusual feature of the beaver stomach was the presence of specific mucus with a thickness up to 950 µm (in frozen, unfixed sections) that coated the mucosa. Our observations suggest that the formation of this mucus is complex and includes the secretory granule accumulation in the cytoplasm of pit cells, the granule aggregation inside cells, and the incorporation of degenerating cells into the mucus.

Morphological studies of the pineal gland in the common gull (Larus canus) reveal uncommon features of pinealocytes.

The avian pineal is a directly photosensory organ taking part in the organization of the circadian and seasonal rhythms. It plays an important role in regulation of many behavior and physiological phenomena including migration. The aim of the study was to investigate morphology of the pineal organ in the common gull (Larus canus). The light and electron microscopic studies were performed on the pineals of juvenile birds living in natural conditions of the Baltic Sea coast, which have been untreatably injured during strong storms in autumn and qualified for euthanasia. The investigated pineals consisted of a wide, triangular, superficially localized distal part and a narrow, elongated proximal part, attached via the choroid plexus to the intercommissural region of the diencephalon. The accessory pineal tissue was localized caudally to the choroid plexus. Based on the histological criteria, the organ was classified as the solid-follicular type. Two types of cells of fotoreceptory line were distinguished: rudimentary-receptor pinealocytes and secretory pinealocytes. Both types of cells were characterized by unusual features, which have been not previously described in avian pinealocytes: the presence of paracrystalline structures in the basal processes and their endings, the storage of glycogen in the form of large accumulations and the arrangement of mitochondria in clusters. Further studies on other species of wild water birds dwelling in condition of cold seas are necessary to explain if the described features of pinealocytes are specific for genus Larus, family Laridae or a larger group of water birds living in similar environmental conditions.

Peptidergic and nitrergic innervation of the pineal gland in the domestic pig: an immunohistochemical study.

The presence and co-localization of vasoactive intestinal polypeptide (VIP), peptide N-terminal histidine C-terminal isoleucine (PHI), pituitary adenylate cyclase-activating peptide (PACAP), somatostatin (SOM), calcitonin gene-related peptide (CGRP), substance P (SP) and the neuronal isoform of nitric oxide synthase (NOS) were studied in neuronal structures of the pig pineal gland. Paraformaldehyde-fixed pineals of 3-month-old gilts were sliced into serial cryostat sections, which were subjected to a set of double immunofluorescence stainings. Based on the co-existence patterns of neuropeptides, five populations of nerve fibres supplying the pig pineal were distinguished: (1) PHI-positive, (2) PACAP-positive, (3) SOM-positive, (4) SP/CGRP-positive and (5) SP-positive/CGRP-negative. Only a subpopulation of PHI-positive fibres contained VIP at the level detectable by immunofluorescence. NOS was found in some intrapineal PHI- and VIP-positive fibres. PHI-, VIP- and NOS-positive nerve fibres were more numerous in the peripheral than in the central part of the pineal. PACAP-positive fibres were equally distributed within the gland. The density of SOM-positive fibres was higher in the ventro-proximal than in the dorso-distal part of the pineal. SOM was also detected in some neuronal-like cells or specialized pinealocytes situated in the central region of the gland. Two populations of fibres containing SP were found: CGRP-positive, present in the distal and central parts of the pineal as well as CGRP-negative, localized in the proximal compartment of the gland.

N-bromoacetyltryptamine strongly and reversibly inhibits in vitro melatonin secretion from mammalian pinealocytes.

OBJECTIVES: Cell-permeable and specific inhibitors of melatonin secretion are sill lacking among tools of the pineal research. Recently, a large effort has been made in the development of arylalkylamine N-acetyltransferase inhibitors, but in most cases the new drugs were tested exclusively using cell-free assays or non-pineal cells. The aim of the present study was to characterize the effect of N-bromoacetyltryptamine (BAT), the first synthesized cell-permeable inhibitor of arylalkylamine N - acetyltransferase, on melatonin secretion from rat and pig pineal glands. METHODS: The studies were performed in the superfusion cultures of rat and pig pineal explants. Melatonin secretion was determined by radioimmunoassay (RIA). RESULTS: BAT strongly inhibited the non-stimulated and norepinephrine - stimulated melatonin secretion from the pig and rat pineal explants, with ED50 0.3 - 0.7 microM. The adrenergic stimulation did not modify significantly the inhibitory potency of BAT on the melatonin release. The decline in melatonin secretion induced by the BAT - treatment was biphasic in both rat and pig pinealocytes, with an initial rapid phase followed by a slow one. The half-time of BAT-induced decline in the non - stimulated and norepinephrine-stimulated melatonin secretion was ca. 25 - 35 minutes. The inhibitory effect of BAT was reversible in pinealocytes of both investigated mammals. CONCLUSIONS: The results show that BAT is a potent and reversible inhibitor of the melatonin secretion in the mammalian pineal gland and open the way for the use of this inhibitor in investigations on the pinealocyte physiology performed in vitro.

Post-hatching development of the turkey pineal organ: histological and immunohistochemical studies.

OBJECTIVE: The study was performed to analyze structural changes of the turkey pineal during the post-hatching development. MATERIAL AND METHODS: The pineals were collected from male turkeys at the age of 1 day, 2, 8, 22, 56 weeks and subjected to histological investigations including morphometrical analyses. The pinealocytes were identified immunohistochemically using antiserum against hydroxyinolo-O-methyltransferase (HIOMT). RESULTS AND CONCLUSIONS: Independently of age, the pineal consisted of a narrow proximal part and a club-shaped top. The narrow part extended into the stalk attached to the diencephalon. The pineal parenchyma was formed by the follicles, surrounded by the connective tissue. The caudal part of the organ contained the pineal lumen, which prolonged into the stalk lumen. Up to the age of two weeks the stalk lumen was open to the third ventricle, later--closed. The proximal part of the stalk showed age-dependent reduction. During the investigated period of life, the pineal increased in size due to creation of new follicles, enlargement of the follicles and development of the stroma. In immature turkeys, the follicular wall was formed by elongated cells bordering the lumen and sparse, peripherally localized, round cells. This pseudostratified organization was transformed during ontogenesis into thick, multilayer structure (characteristic for 22- and 56-week-old turkeys) composed by the layer of elongated cells and several layers of round cells, located peripherally. The rudimentary-receptor and secretory pinealocytes were demonstrated based on HIOMT-immunoreactivity. The secretory pinealocytes were sparse in young birds and predominated in mature turkeys. Intra-pineal calcified concrements occurred in 56-week-old turkeys.

Diurnal rhythms of pinealocyte ultrastructure, pineal serotonin content and plasma melatonin level in the domestic pig.

The study was conducted to investigate diurnal changes in pinealocyte ultrastructure, pineal serotonin content and plasma melatonin concentration in the domestic pig. The immature pigs (n=24) were kept under a cycle of 12 h light : 12 h dark, with a photophase between 0800 and 2000. During the photophase the animals were exposed to direct sunlight. After four weeks the gilts were slaughtered at 0900, 1400, 2100 and 0200. The pineals were removed and divided into two parts - one for quantitative ultrastructural study (by a point count method) and one for serotonin assay. Simultaneously, blood samples were taken for melatonin assay. The relative volume of mitochondria in pinealocyte perikarya was significantly higher at 1400 than at 0200 and 0900 as well as at 2100 than at 0200. The relative volume of Golgi apparatus was higher at 0900 and 1400 than at 0200. The relative volume of dense bodies of the MBB-1 type in pinealocyte perikarya was significantly lower at 1400 and 2100 than at 0900. In contrast, the relative volume of MBB-2 was higher at 1400 than at 0900 and 0200. The numerical density of DCV in perikarya was significantly higher at 0200 than at 1400. No significant differences were found in rough endoplasmic reticulum, lysosomes and multivesicular bodies. The pineal serotonin content showed a prominent rhythm with the maximum at 1400. The plasma melatonin concentration was significantly higher at 0200 than at 0900, 1400 and 2100. The obtained results demonstrate that both pinealocyte ultrastructure and pineal biochemistry in the pig undergo significant changes in the course of the diurnal rhythm.

Qualitative and quantitative studies on the ultrastructure of ovine pinealocytes during postnatal development.

OBJECTIVE: The study was performed to analyze the ultrastructure of ovine pinealocytes during the period of postnatal development. MATERIAL AND METHODS: The pineals of newborn, 10-week and one-year old females of the domestic sheep were prepared for ultrastructural investigations. The point count analysis was used in quantitative studies of the pinealocyte substructures. RESULTS: The prominent feature of pinealocytes in the newborns was the presence of well developed rough endoplasmic reticulum and numerous polysomes. The pinealocyte cytoplasm contained also smooth endoplasmic reticulum, mitochondria, Golgi apparatus, lysosomes, dense core vesicles, multivesicular bodies and lipid droplets. Pinealocytes of the 10-week and 1-year old sheep were characterized by the occurrence of numerous vesicles and short cisterns of smooth endoplasmic reticulum, abundant microtubules and lipid droplets. Pinealocytes of the adult sheep were distinguished by well developed Golgi apparatus, numerous dense core vesicles and multivesicular bodies. The relative volume of rough endoplasmic reticulum in pinealocytes was significantly higher in the newborn sheep than in two other groups. The relative volume of mitochondria was significantly higher in pinealocytes of the 10-week old sheep than the newborns and one-year old animals. The relative volume of Golgi apparatus was significantly higher in the one-year old animals than in two other groups. No differences concern lysosomes. The relative volume of lipid droplets as well as the numerical density of dense core vesicles and multivesicular bodies increased significantly with age. CONCLUSION: The ultrastructure of ovine pinealocytes undergoes the marked changes during postnatal development. The changes concern mainly substructures involved in secretory activity.

Specific distribution pattern of nerve fibers containing catecholamine-synthesizing enzymes, neuropeptide Y (NPY) and C-terminal flanking peptide of NPY (CPON) in the pineal gland of the chinchilla (Chinchilla laniger)--an immunohistochemical study.

The sympathetic nerve fibers originating from the superior cervical ganglia and supplying the pineal gland play the most important role in the control of the pineal activity in mammals. NPY and CPON are also present in the majority of the pinealopetal sympathetic neurons. In this study, immunohistochemical techniques were used to demonstrate the existence and coexistence of tyrosine hydroxylase (TH), dopamine beta-hydroxylase (DbetaH) as well as NPY and CPON in the nerve fibers supplying the chinchilla pineal gland. Ten two-year-old female chinchillas housed in natural light conditions were used in the study. The pineals were fixed by perfusion. ABC immunohistochemical technique and immunofluorescence labelling method were employed. TH-immunoreactive (TH-IR) varicose nerve fibers were observed in the pineal gland as well as in the posterior commissural area. Within the chinchilla pineal gland, TH-IR nerve fibers were located in the capsule and connective tissue septa. Numerous varicose TH-IR branches penetrated into the parenchyma and formed a network showing the highest density in the proximal region of the gland. In the central and distal parts of the pineal parenchyma, a subtle network, composed of thin varicose nerve branches, was observed. Double immunostaining revealed that the majority of TH-IR nerve fibers was positive for DbetaH or NPY. TH- and DbetaH-positive neuron-like cells were observed in the proximal region of the gland. The pattern of pineal innervation immunoreactive to CPON was similar to the innervation containing NPY, TH and DbetaH. The chinchilla intrapineal innervation containing TH, DbetaH, NPY and CPON is characterized by the higher density in the proximal part of the gland than in the middle and distal ones. The specific feature of the chinchilla pineal is also the presence of single TH/DbetaH-immunoreactive neuron-like cells in the proximal part of the gland.

Demonstration of nerve fibers immunoreactive to peptide N-terminal histidine C-terminal isoleucine (PHI) and vasoactive intestinal peptide (VIP) in the pig pineal gland.

The pineal functions are modulated by some neuropeptides including PHI and VIP. The presence of PHI-immunoreactive and VIP-immunoreactive nerve fibers in the pineal gland has been shown in several mammalian species. Both peptides influence the pineal serotonin N-acetyltransferase activity and melatonin synthesis. The aim of the present study was to examine the localization of PHI- and VIP-immunoreactive nerve fibers in the pig pineal gland. Four three-month old female pigs housed in natural light conditions, with free access to food and water, were used in the study. The pineals were fixed by perfusion with 4% paraformaldehyde in 0.1 M phosphate buffer. An immunohistochemical ABC streptavidin-biotin-complex method was used for the demonstration of PHI and VIP. PHI- and VIP-immunopositive nerve fibers were found in the pineal gland as well as in the habenular and posterior commissural areas. In the pineal gland, the density of PHI-immunoreactive nerve fibers was considerably higher than that of the fibers containing VIP. PHI- and VIP-immunopositive nerve fibers were more abundant in the cortical than in the medullary part of the gland. The nerve fibers formed bundles in the pineal capsule, from where they penetrated to the connective tissue septa and formed a dense meshwork surrounding blood vessels. In the parenchyma, PHI- and VIP-immunoreactive nerve terminals created baskets around clusters of pinealocytes. No PHI- or VIP-immunopositive cells were found in the pig pineal gland.