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139 Radioactive (“hot”) particles and early neuroembryogenesis
139
138 GENDER-RELATED DIFFERENCES IN THE BED NUCLEUS OF THE STRIA TERMlNALlS RATS
RADIOACTIVE (“HOT”) PARTICLES EARLY NEUROEMBRYOGENESIS
IN
V. Radeva and N. Stefanova Department of Defectology, University of Sofia “St. K Okhridsky”, 1574 Sotia, Bulgaria
N. Stefanova, A. Bozhilova-Pastirova, and W. Ovtscharoff Department of Anatomy and Histology, Medical University. 143I Sofia, Bulgaria The distribution of GABA-immunoreactive neurons in the subnuclei of the rat bed nucleus of the stria terminalis (BST) was studied by means of GABA immunohistochemistty. Twenty days old Sprague-Dawley rats were anaesthetizedwith sodium pentobarbital(40 mgikg, i.p.) and perfused transcardially with a fixative containing 4% parafomtaldehyde in 0.1 M phosphate buffer at pH 7.2. Brains were removed, posttixed in the same fixative (4’C, I h) then washed several times in PBS and stored overnight at 4°C. On the next day they were frozen and cut coronary at 40 urn thick sections on a microtome (Retchert-Yung, Cambridge Instruments). Sections through the BST were processed etther for Nissl staining, or for GABA immunohistochemistry. We used polyclonal anti-GABA antibodies (Incstar), (I :4000) and silver intensification with silver enhancer kit (Sigma). Parallelly, Nissl staining of the same brain area was done to determine the ratio of GABA-immunoreactive neurons to the total neuronal number in the BST, and to look for a certain sex difference in it. The same procedures were applied to hortnonally manipulated males in the “critical period” of sexual differentiation of the brain. Computer assisted quantitative analysis (Olympus-CUE 2 micro analysis system) of GABA-immunoreactive neurons showed statistically significant sex differences in the BST applymg Student’s t-test. Data were presented as means= S. E. M.
The present study considers some aspectsof the motphogenetic effect of “hot” particles (HP), (natural and artificial), containing radioactive isotopes, on the early neuroembtyogenesis of newts, Trim-us crislalus. at ultrastructural level. The embryos from the fecondation till larva stage were cultivated in aqua fontana around fixed HP. The influence of the HP on the neuroblasts and myoblasts during neurulation provokes destruction of the mitochondria and vacuolisation of their cristae, deformation and dilatation of the cisterns of the rough endoplasmic reticulum and Golgi apparatus. as well as increase in the number of lysosomes. The cell nuclei include atypical perichromatin granules and giant ones. The newly formed synapses are completely destroyed. The genesis of myolibrils is profoundly disturbed. The extent of the described effect dependson the radiation intensity, as well as on the distance between the HP and the investigated embryo
140
141
IDENTIFICATION DEVELOPING MARKER
AND
OF MOUSE
OF
RADIAL
ANTIGENS BRAIN
RECOGNIZED BY
THE
RC2
IN ANTIBODY,
MORPHOMETRIC ANALYSlS OF SYNAPTIC PLASTICITY OF SENSORIMOTOR CORTEX CINDER NEUROPEPTIDAL CORRECTION OF HYPO- AND HYPEROXIC DAMAGES
THE A
CLIA.
P. Leprince, G. Chanas-Sacre, R. Wattiez’, M. Lewm, B. Rogister, C. Mazy-Servais, J.P. Misson and G. Moonen. Laboratory of Human Physiology, University of Liege, and ‘Laboratory of Biological Chemistry, University of Mons-Hainaut, Belgium
A.Matstonis. IPavlov, G.Kuraev*, A.Mendzherttsky. P.Povilaitite Rostov Regional Bureau of Patholoa, Dcpattment of Experimental Pathomorphology. *Rostov Slate University, 314085, Rostov-on-Don, Russia.
The RC2 anttbody labels the radial glial cells in tissue sections of embryonic mouse brain and in cultures of dissociated cortical and cerebellar cells. An intense labeling of glial cells can be observed: in sections from the cell bodies in the periventricular zone to the pial surface, and in cultures mostly as a cytoskeletonassociated material along the cell intermediate filaments (IF). Less Intense staining is also observed in pial fibroblasts and in various cell nuclei. We have identified by Western blotting a serie of proteins of MW between 35 and 250 kDa that are labeled by RC2 antibody and are present in developing mouse brain only during the period ofneuronal migration. All ofthese proteins are only soluble tn SDS or Urea-containing solutions. They have been fractionated by ion-exchange chromatography and 2-D gel electrophoresis and several of them have been partially sequenced. Two of the most prominently expressed ones with a size of about 100 kDa have been found to be nuclear proteins (PSF and nucleolin). They are likely candidates to explain the pattern of nuclear labeling by RC2. Indeed, anti-PSF antibodies label only selected nuclei in dissociated cerebellar cells cultures. PSF mRNA expression in the mouse brain is restricted to the developmental period suggesting a specific role in neural ontogeny. Other proteins that are candtdates as the IF-associated antigens are now subjected to sequencing
Delta-sleep mducing peptide @SIP) shows antistressoty propertres owng to its modulating action on an excitatory and Inhibitory ncurotransmitter system. The impact of DSIP on the quantitative ultrastructural parameters of axosomatic synapses in rats was under the study The synapses located in the III-V layers of sensorimotor cortex which are commonly considered as GABAergic were examined. States of hypoxia and hyperosia of median heaviness were simulated in the specially designed chambers; DSIP was injected one hour before stress. Morphoruetric analysis eras performed with the use of computer software Sigma/Scan Image (Jandel Sc~entrlic). Obtained results clearly demonstrate the fact of modulating effect of DSIP on ultrastructure of inhibitory axosomatic synapses. It is manifested by reliable alterations of axonal endings’ areas in the same direction as under the stress conditions alone, while the values of such parameters as form factor of axonal endings, quantity and density of synaptic vesicles, length of active zone revert to the normal level. The role of axonal endtngs of different sir. and shape in realization of adapttve reactions at synaptic level is discussed
84
138 GENDER-RELATED DIFFERENCES IN THE BED NUCLEUS OF THE STRIA TERMlNALlS RATS
RADIOACTIVE (“HOT”) PARTICLES EARLY NEUROEMBRYOGENESIS
IN
V. Radeva and N. Stefanova Department of Defectology, University of Sofia “St. K Okhridsky”, 1574 Sotia, Bulgaria
N. Stefanova, A. Bozhilova-Pastirova, and W. Ovtscharoff Department of Anatomy and Histology, Medical University. 143I Sofia, Bulgaria The distribution of GABA-immunoreactive neurons in the subnuclei of the rat bed nucleus of the stria terminalis (BST) was studied by means of GABA immunohistochemistty. Twenty days old Sprague-Dawley rats were anaesthetizedwith sodium pentobarbital(40 mgikg, i.p.) and perfused transcardially with a fixative containing 4% parafomtaldehyde in 0.1 M phosphate buffer at pH 7.2. Brains were removed, posttixed in the same fixative (4’C, I h) then washed several times in PBS and stored overnight at 4°C. On the next day they were frozen and cut coronary at 40 urn thick sections on a microtome (Retchert-Yung, Cambridge Instruments). Sections through the BST were processed etther for Nissl staining, or for GABA immunohistochemistry. We used polyclonal anti-GABA antibodies (Incstar), (I :4000) and silver intensification with silver enhancer kit (Sigma). Parallelly, Nissl staining of the same brain area was done to determine the ratio of GABA-immunoreactive neurons to the total neuronal number in the BST, and to look for a certain sex difference in it. The same procedures were applied to hortnonally manipulated males in the “critical period” of sexual differentiation of the brain. Computer assisted quantitative analysis (Olympus-CUE 2 micro analysis system) of GABA-immunoreactive neurons showed statistically significant sex differences in the BST applymg Student’s t-test. Data were presented as means= S. E. M.
The present study considers some aspectsof the motphogenetic effect of “hot” particles (HP), (natural and artificial), containing radioactive isotopes, on the early neuroembtyogenesis of newts, Trim-us crislalus. at ultrastructural level. The embryos from the fecondation till larva stage were cultivated in aqua fontana around fixed HP. The influence of the HP on the neuroblasts and myoblasts during neurulation provokes destruction of the mitochondria and vacuolisation of their cristae, deformation and dilatation of the cisterns of the rough endoplasmic reticulum and Golgi apparatus. as well as increase in the number of lysosomes. The cell nuclei include atypical perichromatin granules and giant ones. The newly formed synapses are completely destroyed. The genesis of myolibrils is profoundly disturbed. The extent of the described effect dependson the radiation intensity, as well as on the distance between the HP and the investigated embryo
140
141
IDENTIFICATION DEVELOPING MARKER
AND
OF MOUSE
OF
RADIAL
ANTIGENS BRAIN
RECOGNIZED BY
THE
RC2
IN ANTIBODY,
MORPHOMETRIC ANALYSlS OF SYNAPTIC PLASTICITY OF SENSORIMOTOR CORTEX CINDER NEUROPEPTIDAL CORRECTION OF HYPO- AND HYPEROXIC DAMAGES
THE A
CLIA.
P. Leprince, G. Chanas-Sacre, R. Wattiez’, M. Lewm, B. Rogister, C. Mazy-Servais, J.P. Misson and G. Moonen. Laboratory of Human Physiology, University of Liege, and ‘Laboratory of Biological Chemistry, University of Mons-Hainaut, Belgium
A.Matstonis. IPavlov, G.Kuraev*, A.Mendzherttsky. P.Povilaitite Rostov Regional Bureau of Patholoa, Dcpattment of Experimental Pathomorphology. *Rostov Slate University, 314085, Rostov-on-Don, Russia.
The RC2 anttbody labels the radial glial cells in tissue sections of embryonic mouse brain and in cultures of dissociated cortical and cerebellar cells. An intense labeling of glial cells can be observed: in sections from the cell bodies in the periventricular zone to the pial surface, and in cultures mostly as a cytoskeletonassociated material along the cell intermediate filaments (IF). Less Intense staining is also observed in pial fibroblasts and in various cell nuclei. We have identified by Western blotting a serie of proteins of MW between 35 and 250 kDa that are labeled by RC2 antibody and are present in developing mouse brain only during the period ofneuronal migration. All ofthese proteins are only soluble tn SDS or Urea-containing solutions. They have been fractionated by ion-exchange chromatography and 2-D gel electrophoresis and several of them have been partially sequenced. Two of the most prominently expressed ones with a size of about 100 kDa have been found to be nuclear proteins (PSF and nucleolin). They are likely candidates to explain the pattern of nuclear labeling by RC2. Indeed, anti-PSF antibodies label only selected nuclei in dissociated cerebellar cells cultures. PSF mRNA expression in the mouse brain is restricted to the developmental period suggesting a specific role in neural ontogeny. Other proteins that are candtdates as the IF-associated antigens are now subjected to sequencing
Delta-sleep mducing peptide @SIP) shows antistressoty propertres owng to its modulating action on an excitatory and Inhibitory ncurotransmitter system. The impact of DSIP on the quantitative ultrastructural parameters of axosomatic synapses in rats was under the study The synapses located in the III-V layers of sensorimotor cortex which are commonly considered as GABAergic were examined. States of hypoxia and hyperosia of median heaviness were simulated in the specially designed chambers; DSIP was injected one hour before stress. Morphoruetric analysis eras performed with the use of computer software Sigma/Scan Image (Jandel Sc~entrlic). Obtained results clearly demonstrate the fact of modulating effect of DSIP on ultrastructure of inhibitory axosomatic synapses. It is manifested by reliable alterations of axonal endings’ areas in the same direction as under the stress conditions alone, while the values of such parameters as form factor of axonal endings, quantity and density of synaptic vesicles, length of active zone revert to the normal level. The role of axonal endtngs of different sir. and shape in realization of adapttve reactions at synaptic level is discussed
84