Neurotrophic Factor Receptors trkB and trkC in Experimental Model of Lesion in Rat Brain Structures in Schizophrenia.

INTRODUCTION: The maldevelopmental model of schizophrenia postulates pathological alterations in embryonal neurogenesis as the etiopathogenetic basis of schizophrenic psychoses. The neurotrophic factor hypothesis explains these neuropathological abnormalities as the result of alterations of the neurotrophin system caused by different mechanisms such as a genetic, infectious and traumatic factors. The tyrosine-kinase containing receptors trkB and trkC mediate growth-promoting effects of neurotrophins and respond to changes in neurotrophic factors availability. AIM: The aim of the present study was to establish the expression pattern of trkB and trkC in rat brain structures by a developmental model of schizophrenia. MATERIALS AND METHODS: On cryostat coronal brain sections of control and lesioned rats (after infusion of ibotenic acid solution bilaterally into the hippocampal formation), immunoreactions for trkB and trkC were performed. RESULTS: We found diminished expression of trkB and trkC in the hippocampal formation of lesioned animals compared to the controls. Quantitative measurements of immunohistochemical reactions intensity and statistical analysis confi rmed the reduced immunoreactivity for antigens under study (trkB and trkC) in the positive hippocampal neurons of 56-day-old lesioned rats compared to the control animals. CONCLUSION: The observed downregulation of neurotrophic factor receptors expression may compromise the function and plasticity of hippocampal formation in schizophrenic brains.

Brain-derived neurotrophic factor and its serum levels in schizophrenic patients.

INTRODUCTION: Neurotrophins have an important role in regulating the development and maintenance of the peripheral and central nervous systems' function. Thus, the neurotrophin hypothesis of schizophrenia has postulated that the changes in the brain of schizophrenic patients are the result of disturbances of developing processes involving these molecules. AIM: We analyse in the present study the changes in the serum levels of brain-derived neurotrophic factor (BDNF) in schizophrenic patients as possible epiphenomena of underlying alterations of the neurotrophic factor in central nervous system, reflecting its role in the pathophysiology of schizophrenia. PATIENTS AND METHODS: Twenty-one schizophrenic patients satisfying the DSM-IV criteria for diagnosis of schizophrenia were enrolled in the study. The control group consisted of 28 age-matched mentally healthy subjects. Serum BDNF levels were determined in patients and normal controls using ELISA (Chemicon International, USA & Canada). The data were analyzed statistically with Student's t- test in SPSS 9.0. RESULTS: The serum BDNF levels were lower in the schizophrenic patients than in the control subjects, reaching statistically significant difference (t = 2.72, p = 0.009). Female patients had lower serum BDNF levels than the male patients but the difference fell short of statistical significance (t = 0.1, p = 0.9). CONCLUSIONS: The BDNF reduction in serum indicates a potential deficit in neurotrophic factor release in patients with schizophrenia and support the concept that BDNF might be associated with schizophrenia.

Serotonin-producing cells in human gastric mucosa--immunohistochemical and electron microscopic study.

UNLABELLED: The great many hormones released by the endocrine cells of the glands and lining epithelium of gastric mucosa determine its significance for the processes in the gastrointestinal tract. One of these hormones, serotonin, plays an important role in the regulation of the motility, secretion and sensation in the gastrointestinal tract. The aim of the present study was to conduct immunohistochemical and electron microscopic studies of serotonin-producing EC cell of gastric mucosa. MATERIAL AND METHODS: Gastric mucosa biopsies were obtained and studied immunihistochemically for serotonin expression in the mucosa endocrine cells. Electron microscopic study was performed to specify the processes of synthesis, accumulation and release of secretory product by those cells. RESULTS: The immunohistochemical study revealed a considerable number of serotonin-containing EC cells scattered in the lining epithelium and between the glands in the corpus and pyloric region of the stomach. The electron microscopic study followed the stages of formation of the secretory granules from the initial accumulation of granular substance, its membrane packing and formation of mature granules to their disintegration in the secretory process. CONCLUSIONS: Serotonin as a neurotransmitter and gastrointestinal hormone appears to be a key to understanding a number of symptoms of gastrointestinal disorders like nausea, vomiting, pain, diarrhea and constipation. A detailed study of serotonin functions in the gastrointestinal tract realised through different types of receptors, and of the development of specific antagonists and agonists to these receptors would open up new opportunities for a more efficient treatment of gastrointestinal disorders.

11beta hydroxysteroid dehydrogenase type 2 in the adrenal gland by testosterone withdrawal of adult rats.

AIM: 11beta hydroxysteroid dehydrogenase (11beta HSD) catalyzes the interconversion of glucocorticoids to inert metabolites in man and rodents and plays a crucial role in regulating corticosteroid hormone action. The physiological role and regulation of 11beta HSD type 2 in the adrenal gland remains obscure. Therefore, the aim of the present study was to establish the pattern of 11beta HSD type 2 expression in rat adrenal gland under conditions of testosterone withdrawal. MATERIAL AND METHODS: We performed immunohistochemical analyses of adrenal gland sections of ethane dimethanesulphonate (EDS)-treated adult rats. RESULTS: In controls, strong positive 11beta HSD type 2 signals were detected in the adrenal cortex cells, but not in the medulla. We observed the lowest 11beta HSD type 2 expression intensity 7 days after initial treatment with ethane dimethanesulphonate (EDS) followed by progressive increase in the immunoreactivity toward days 14 and 21. Maximal staining intensity of 11beta HSD type 2 in the adrenocorticocytes was found by day 35 after EDS treatment. CONCLUSIONS: By using the EDS model the present study provides new data about 11beta HSD type 2 expression in the adrenal gland under conditions of testosterone withdrawal of adult rats. Our results elucidate further the functional significance of 11beta HSD system in rat adrenal gland and the regulatory role of testosterone in its activity.

Prenatal and postnatal differentiation of the small intestine in rat.

UNLABELLED: The gastrointestinal tract in the early prenatal development is an endoblastic mesenchyme-lined tube. The endoblast differentiates and gives origin to all epithelial structures (covering epithelium, glands). The mesenchyme develops into connective tissue, blood vessels, the smooth muscle cells of lamina muscularis mucosae and muscular tunic. Neuroectoblast cells participate in these processes--individual cells with future endocrine function, nerve cells and fibers that form nerve plexuses and vegetative ganglia. AIM OF THE PRESENT STUDY: To trace the changes in the small intestine development during the prenatal period in rat embryos and fetuses, and during the postnatal period in newborn rats. We specifically studied the beta-actin expression in the cytoskeletal structures of the covering epithelium and in the contractile elements of the differentiating smooth muscle cells. The presence and localization of the enteroendocrine EC cell was studied using the immunohistochemical expression of serotonin in them. MATERIAL AND METHODS: Material from rat embryos and fetuses aged 8-11, 12-15, 16-20 days of gestation and small intestine fragments from newborn rats was studied using routine hematoxylin-eosin staining, enzymohistochemically for succinate dehydrogenase and immunohistochemically for beta-actin and serotonin. RESULTS: In the early embryogenesis (8-11 day of gestation), the primitive gut of rat embryos is an endoblastic tube of 2-3 layers of cuboidal cells covered with a thin layer of mesenchyme. In the subsequent stages of embryonic and fetal development the processes of differentiation run at different rates in the different tissues. The maturation process in the small intestine wall of one-day-old newborn rats is incomplete. The mucosa presents with shallow crypts and loosely set villi. Differentiated resorptive and enteroendocrine EC cells are found in the lining epithelium. CONCLUSION: The changes we found in the beta-actin expression in the contractile elements of the differentiating smooth muscle cells and the cytoskeletal structures of the lining epithelium probably reflect the induction interference between the derivatives of the mesenchyme and endoblast.

c-erbAalpha and c-erbaAbeta in the Leydig cell repopulation by ethane-1,2-dimethanesulphonate model of mature rats.

INTRODUCTION: The regulatory effect of thyroid hormones on the proliferation and maturation of the Leydig cells (LC) in testis is still poorly understood. To date, it remains obscure whether the thyroid hormones have direct effect on the LC, as far as in rat testis the thyroid hormones receptors are localized predominantly in the Sertoli cells. A single intraperitoneal dose of cytotoxin ethane-1,2-dimethanesulphonate (EDS) injected into mature rats caused a rapid, selective elimination of the adult LC associated with temporary impairment of fertility. Regeneration of the LC population by EDS model is a result of the differentiation of LC progenitors as well as of the proliferation of the newly formed LC whereas the process is similar with the development of adult LC in the prepubertal testis. AIM: The present study aimed to establish the immunohistochemical expression of high affinity triiodothyronine nuclear receptors c-erbAalpha and c-erbAbeta in the regenerating LC after treatment with EDS of mature rats. MATERIAL AND METHODS: Mature male Wistar rats were divided into two groups: (1) a group of rats receiving a single intraperitoneal (i.p.) injection of EDS (75 mg/kg body weight) and (2) a group of control animals. The animals were killed 24 hours, 7, 14, 21 and 35 days after treatment. Testicular fragments were prepared for routine histological and immunohistochemical examinations. RESULTS: The immunohistochemical analysis revealed similar changes in the immunoreactivity for both c-erbAalpha and c-erbAbeta after EDS administration. On day 1 after EDS treatment, the intensity of the immune reactions for c-erbAalpha and c-erbAbeta in the LCs decreased simultaneously with their number. Seven days after EDS administration there was neither LCs nor c-erbAalpha nor c-erbAbeta-immunoreactivity. The first positive stained LCs were found 14 days after EDS when LCs progenitors were detected. The most prominent c-erbAalpha- and c-erbAbeta-immunostaining in the regenerating LCs was evident 21 days after EDS; this coincided with the increased number of LCs progenitors and their transformation into adult LCs population. Thirty-five days after EDS c-erbAalpha and c-erbAbeta-positive LCs were abundant their number and localization in the testicular interstitium being very similar to that in the control rats. CONCLUSION: The observed change in the intensity of the immune reactions for c-erbAalpha and c-erbAbeta in LC repopulation after EDS treatment corresponds to the process of differentiation of progenitors into mature LC. The results obtained support the idea about the regulatory role of thyroid hormones in the differentiation of LC in prepubertal rat testis.

INSLF3-LGR8 ligand-receptor system in testes of mature rats after exposure to ethane dimethanesulphonate (short communication).

The cytotoxic agent ethane-1,2-dimethanesulphonate (EDS) specifically destroys the Leydig cells (LC) in the adult testis, followed by a complete regeneration. The process of LC renewal after exposure to EDS shows homology to the development of the adult-type LC population in prepubertal testis. INSL3, also known as Leydig insulin-like peptide or relaxin-like factor, is a peptide hormone, a novel member of the insulin/relaxin family, and seems to be localized predominantly in the gonadal tissues. INSL3 mRNA is expressed in the LC in a constitutive fashion and INSL3 thus seems to be a useful marker of LC differentiation status. The present study was aimed at establishing the chronology and dynamic of expression of INSL3 and its specific receptor LGR8 in the LC repopulation after exposure to mature rats to EDS. As material, testes of mature Wistar rats that received single intraperitoneal injection of EDS (75 mg/kg body weight) were used. The animals were killed 1, 7, 14, 21 and 35 days after the initial treatment. The pattern of INSL3-LGR8 expression in newly formed LC after EDS administration was established using a high sensitive immunohistochemical polymer detection kit. After treatment with EDS, the immunoreactivity for INSL3 and LGR8 disappeared from the testis and reappeared again at the time of regeneration of the first LC, 14 days after EDS. The INSL3-LGR8 positive cells grew in number concomitantly with the increase of the LC repopulation. Thirty-five days after EDS destruction a larger number of immunopositive LC were seen in form of clusters corresponding with the regeneration of adult type LC population. The present findings support the hypothesis that EDS-treated rats can serve as a model for studying the LC development in the prepubertal testis and indicate a specific role of hormonal factors like INSL3 in this process.

Computertomographic findings of structural brain anomalies in schizophrenic patients in support of the neurodevelopmental hypothesis of the disorder.

AIM: The aim of the study was to make a comparative CT examination of schizophrenic patients and find lifetime criteria for recognition of brain changes in schizophrenia. MATERIAL AND METHODS: Twenty-two schizophrenic inpatients (mean age 32.86 +/- 2.65 yrs) satisfying the DSM-IV criteria for schizophrenia were examined. The control group comprised 27 clinically healthy subjects (16 men, 11 women, mean age 46.44 +/- 2.32 yrs) all of Bulgarian ancestry. All subjects underwent CT examination without venous enhancement at an examination angle of + 15 degrees-20 degrees in relation to the orbitomeatal line. Cortical atrophy was assessed according to criteria determining the external and internal liquor spaces (after Meese and Groome). RESULTS: There is a consistent low-grade enlargement of the brain ventricles. The variables have increased values (decreased for CMI) in the schizophrenic patients compared with the controls. The patients show moderately increased width of the lateral sulcus and brain convexity sulci. CONCLUSION: The brain tissue loss and enlarged extracerebral space suggest that the observed evidence of cortical loss in schizophrenic patients reflects a pathological process operating before completion of the brain growth.

Bcl-2/Bax ratio, mitochondrial membranes and aerobic enzyme activity in cardiomyocytes of rats after submaximal training.

INTRODUCTION: Mitochondria are an active and continuous source of reactive oxygen species (ROS) during respiration. The ROS increased production during endurance training is a result of an augmented electron transport through the respiratory chains, making in this way the mitochondria a potential target for oxidative damage. The Bcl-2 protein family plays a central role in the transition of apoptotic signals towards the mitochondria in stress-induced apoptosis. AIM: The present work studied the effect of endurance training on the expression of the apoptotic proteins Bcl-2 and Bax in rat cardiomyocytes, as well as the concomitant changes in the ultrastructure of the mitochondria and activity of some enzymes residing there. MATERIAL AND METHODS: Two groups of male Wistar rats were used. One was the control and the other was trained on treadmill with submaximal loading for eight weeks. At the end of the trial, samples of the myocardium of all the experimental animals were obtained. Immunohistochemical reactions for Bcl-2 and Bax and enzymehistochemical reactions for succinate dehydrogenase and NADH2-cytochrome C-reductase were done. The results were analyzed using specialized software. Transmission electron microscopical study was carried out too. RESULTS: In the myocardium of the trained animals the expression of Bcl-2 and Bcl-2/Bax ratio were significantly higher compared to the controls. The mitochondria had intact outer and inner membranes, with no signs of swelling. Mitochondria with denser packed cristae were found predominantly. No significant differences were found in the activity of the investigated enzymes in the cardiomyocytes of the animals from both groups. CONCLUSIONS: In the myocardium of the experimental animals endurance training for eight weeks does not lead to activation of apoptotic processes via the mitochondrial pathway. This type of exercise training could be used for cardioprotection in order to elevate apoptotic threshold of cardiomyocytes.

Effects of submaximal training and anabolic androgenic steroids administration on steroidogenic enzyme activity in rat Leydig cells.

The purpose of the present study was to investigate the single and combined effects of submaximal training and anabolic androgenic steroids (AAS) treatment on the activity of 3beta hydroxysteroid dehydrogenase (3betaHSD) in rat Leydig cells (LC). Forty male Wistar rats were distributed into 4 groups. Half of them exercised on treadmill. After 2 weeks half of the trained and sedentary rats received weekly either 10 mg x kg(-1) Nandrolone Decanoate (ND) or Placebo (Pl) i.m. for 6 weeks. The day after the last exercises all the groups: 1) sedentary + Pl (SP); 2) sedentary + ND (SND); 3) trained + Pl (TP) and 4) trained + ND (TND) were decapitated. On fresh cryostat sections of the testes of each animal enzymehistochemical reaction for the activity of 3betaHSD was carried out. Our results demonstrate that in sedentary rats ND treatment decreased the activity of 3betaHSD in the LC in comparison to SP. Endurance training also decreased the activity of 3betaHSD in TP group compared to SP. On sections of the testes of group TND a pronounced reduction in the enzyme activities of 3betaHSD in the LC was found in comparison with the other groups. In conclusion we suggest that submaximal endurance training and/or administration of AAS downregulate the steroidogenic enzyme activity of rat Leydig cells.

Immunoreactivity for ferritin in Leydig cells of human testis.

The aim of the present study was to demonstrate the immunocytochemical localization of ferritin in human Leydig cells. Testes from patients orchidectomized for carcinoma of the prostate were used. The immunoreactivity for ferritin was visualized in the Leydig cells by amplification immunocytochemical technique. The Sertoli cells and some of the germ cells show moderate immunoreactivity for the examined antigen. Our result represent an immunocytochemical verification for presence of ferritin in the human Leydig cells (also in other cellular components) and suggest the role of this factor in the local auto- and/or paracrine control of the testicular functions.

Immunolocalization of neurotrophic factors and their receptors in the Leydig cells of rat during postnatal development.

The aim of the present study was to localize the neurotrophic factors glial cell line-derived neurotrophic factor (GDNF), nerve growth factor receptor (NGFr) and low sensitive receptor for NGF (p75LNGFR) in rat testes during the postnatal development. Rat testes obtained at different stages of the postnatal development--day 5, 10, 15, 20, 24 and 27 after birth were used. Amplification immunocytochemical technique, which includes a combination of the peroxidase-antiperoxidase method (PAP) and the avidin-biotin peroxidase complex (ABC) methods, was applied. Immunoreactivity for the tested antigens was established in the Leydig cell with characteristic fluctuation in the intensity of the immune reaction at the different stages of the postnatal development. Positive immunostaining was seen in the Sertoli and some of the germ cells (spermatocytes and spermatids). The results obtained show that neurotrophic factors play a role in the processes of postnatal differentiation of the Leydig cells and in the regulation of their functional activity.

Dexamethasone and oxytocin effects on rat Leydig cells--morphological and enzyme-histochemical characteristics.

In the present study the fine structure of rat Leydig cells was examined by electron microscopy. Oxytocin and dexamethasone induced changes in the activities of 3beta hydroxysteroid dehydrogenase, NADH2 cytochrome-C-reductase and glucose-6-phosphate dehydrogenase in these cells were studied in an in vivo experiment. Two groups of male Wistar rats were used to test the effects of oxytocin--rats from group I received a single injection of oxytocin; rats from group II were given a 10-day course of oxytocin. Pregnant female rats were injected with dexamethasone on day 17 and 18 post conception. The testes of 19 and 20-day old embryos were removed. It was established that both short and long term courses of oxytocin increased the activities of the above-mentioned enzymes. On the contrary, prenatal administration of dexamethasone decreased enzymatic activity in Leydig cells. Electron microscopy revealed clusters of fetal Leydig cells. Our results indicated the role of oxytocin in the local regulation of steroidogenesis and the importance of glucocorticoids in the differentiation and activity of Leydig cells.