Papers by Graziella Bernocchi
Experimental Neurology, Feb 1, 1986
Groups of 6- to 7-day-old and 10- to 11-day-old rats received cis-dichlorodiammineplatinum (cis-D... more Groups of 6- to 7-day-old and 10- to 11-day-old rats received cis-dichlorodiammineplatinum (cis-DDP) or inactive trans-DDP subcutaneously and were killed after 1, 6, 10, 15, or 21 days. In both age groups the acute effect (postinjection day 1) was most obvious in the germinal external granular layer (EGL), where many cells underwent shrinkage necrosis (pyknosis, apoptosis); the latter were more frequent within the fissures than on the surface of the cerebellar folia. Cell debris were often seen to be engulfed by macrophages, Bergmann cell fibers, and meningocytes. Bergmann cell bodies were swollen and the nuclei of Purkinje cells as well as of surviving cells of the EGL were enlarged. No comparable effects were observed in animals that had received an injection of trans-DDP in the same dose. Toward postinjection (p.i.) day 6, pyknotic cells disappeared from the EGL or, after the larger doses of cis-DDP, substantially decreased in number. Small nests of pyknotic cells appeared, however, at some places of the internal granular layer (IGL). The EGL was discontinuous or thinner in both age groups. In the IGL and molecular layer (ML) multiple focal fresh hemorrhages appeared together with some macrophages. The packing cell density in the IGL was less than in the controls, especially at the top of the cerebellar folia, i.e., contrary to the distribution of the primary damage in the EGL. Later, at p.i. day 10, ectopic nests of the IGL occurred occasionally in animals injected when 10 to 11 days old and severe atrophy of the ML and Purkinje cell population was observed at some places. At p.i. days 15 to 21, invasion of microglia-like cells appeared in the IGL and in some regions of the ML. Occasionally, subpial hemorrhages occurred at this interval. The acute damage caused by cis-DDP is thus similar to the effect of X rays or some, but not all, drugs with a cytostatic action. In addition, a more profound influence on dividing cells of the EGL within the fissures and distinct capillary lesions, indicated by hemorrhages, were found after cis-DDP. As in other experimental models, the acute cis-DDP damage of the immature cerebellum was partly repaired within a few p.i. days. Spatial difference of the repair process was inferred from the packing cell density in the IGL measured at the top and bottom of the cerebellar folia.
Acta Neuropathologica, 1980
ABSTRACT The cytoplasmic eosinophilia of the Purkinje cells appears in controls at 16 days wherea... more ABSTRACT The cytoplasmic eosinophilia of the Purkinje cells appears in controls at 16 days whereas in the offspring of malnourished mothers (fed 8% protein diet) it occurs at 22 days. The adult per cent values are reached by controls at 22 days and by malnourished animals at 30 days.This finding makes it difficult to refer to the cytoplasmic eosinophilia merely as fixation artifact.The possible interpretations for the occurrence of this parameter are discussed.
Journal of Chemical Neuroanatomy, Dec 1, 2012
Programmed cell death is regulated by prototypes of a large family of Bcl-2-like proteins such as... more Programmed cell death is regulated by prototypes of a large family of Bcl-2-like proteins such as Bax and Bcl-2. A neuroprotective role for cellular prion protein (PrPc) on programmed cell death has been reported, although the cytosolic accumulation of PrPc correlates with toxicity and death of some neurons by apoptosis. In order to understand the signalling function of PrPc in promoting survival or suppressing cell death, we analyzed the expression and co-localization of PrPc, Bax and Bcl-2 proteins in the developing cerebellum of rats treated at PD10 (postnatal day 10) with the chemotherapeutic drug cisplatin (cisPt) or the new platinum (Pt) compound [Pt(O,O'-acac)(γ-acac)(DMS)] (PtAcacDMS). Differences in the expression of PrPc, Bax and Bcl-2 were found in proliferating cells and immature Purkinje neurons. One day after administration (PD11), cisPt markedly increased the apoptosis of the proliferating cells of the EGL (external granular layer); at the same time, several apoptotic bodies with strong Bax immunoreactivity were noticed. After PtAcacDMS, changes in PrPc and apoptotic proteins, with respect to the controls, were found but Bax immunopositive apoptotic bodies were not detectable, which could mean that apoptotic cell death of proliferating cells is preserved. Co-localization was clearly detected in the Purkinje cell population and may explain better the mechanisms by which PrPc and the apoptotic proteins function, and particularly the role of PrPc. Considering the reactivity of Purkinje neurons to these proteins at PD11 and Pd17, at least PrPc expression increased after cisPt and PtAcacDMS treatments or, if PrPc decreased, balanced itself with Bcl-2. The noteworthiness of this finding is that it emphasizes that most of the post-mitotic Purkinje cells need to be rescued, otherwise they undergo degeneration and are not replaced. Based on the effects of both Pt compounds on Purkinje cell differentiation, it should be emphasized that PrPc, together with the synergistic action of the co-localized anti-apoptotic protein, acts as a neuroprotective protein countering cytotoxicity in the postnatal critical phases of cerebellum development.
Journal of neurosurgical sciences
Acute, severe injury of the rabbit spinal cord, induced by the weight-drop method, causes alterat... more Acute, severe injury of the rabbit spinal cord, induced by the weight-drop method, causes alterations of the enzyme activities related to cholinergic and energy metabolism. Morphological examinations at the trauma site show degenerative processes in neurons 0.5 hr posttrauma and a marked decrease in the number of living cells 24 hrs later. Both biochemical and cytochemical findings show that the tissue metabolic and morphologic derangement, caused by severe spinal cord injury, is mostly confined to the gray matter at an early stage (0.5 hr), whereas 24 hrs later the white matter is also involved. The decrease in choline acetyl-transferase and acetylcholinesterase activities in the gray matter parallels the impairment of complex IV (cytochrome c oxidase) of the respiratory chain and the presence of morphological alteration in neurons. The dramatic drop in the enzyme activities, observed 24 hrs after the induction of the severe trauma is clearly associated with the loss of cells.
Platinum compounds cause significant clinical neurotoxicity. Several studies highlight neurologic... more Platinum compounds cause significant clinical neurotoxicity. Several studies highlight neurological complications especially in paediatric oncology patients with Central Nervous System (CNS) and non-CNS malignancies. To understand the toxicity mechanisms of platinum drugs at cellular and molecular levels in the immature brain, which appears more vulnerable to injury than in the adult one, we compared the effects in vivo of the most used platinum compounds, i.e., cisdichlorodiammineplatinum (cisplatin, cisPt), and the new [Pt(O,O′-acac)(γ-acac)(DMS)] (PtAcacDMS). As models of developing brain areas, we have chosen the cerebellum and hippocampus dentate gyrus. Both areas show the neurogenesis events, from proliferation to differentiation and synaptogenesis, and therefore allow comparing the action of platinum compounds with DNA and non-DNA targets. Here, we focused on the changes in the intracellular calcium homeostasis within CNS architecture, using two immunohistochemical markers, the calcium buffer protein Calbindin and Plasma Membrane Calcium ATPase. From the comparison of the cisPt and PtAcacDMS effects, it emerges how essential the equilibrium and synergy between CB and PMCA1 is or how
Journal of Comparative Neurology, 2019
The extracellular matrix is essential for brain development, lamination, and synaptogenesis. In p... more The extracellular matrix is essential for brain development, lamination, and synaptogenesis. In particular, the basement membrane below the pial meninx (pBM) is required for correct cortical development. The last step in the catabolism of the most abundant protein in pBM, collagen Type IV, requires prolidase, an exopeptidase cleaving the imidodipeptides containing pro or hyp at the C-terminal end. Mutations impairing prolidase activity lead in humans to the rare disease prolidase deficiency characterized by severe skin ulcers and mental impairment. Thus, the dark-like (dal) mouse, in which the prolidase is knocked-out, was used to investigate whether the deficiency of prolidase affects the neuronal maturation during development of a brain cortex area. Focusing on the cerebellar cortex, thinner collagen fibers and disorganized pBM were found. Aberrant cortical granule cell proliferation and migration occurred, associated to defects in brain lamination, and in particular in maturation of Purkinje neurons and formation of synaptic contacts. This study deeply elucidates a link between prolidase activity and neuronal maturation shedding new light on the molecular basis of functional aspects in the prolidase deficiency.
Neurotoxicology and Teratology, 2011
In the field of experimental oncology, many efforts are being carried out to search new platinum-... more In the field of experimental oncology, many efforts are being carried out to search new platinum-based drugs overcoming the CNS toxicity and drug resistance. One of the adopted strategies is the synthesis of platinum compounds able to form Pt-DNA adducts different from the cisplatin ones or to react with other subcellular targets. In this context a novel Pt(II) complex, [Pt(O,O'-acac)(γ-acac)(DMS)](PtAcacDMS), was synthesized which reacts preferentially with protein thiols or thioethers. In this work we investigated the in vivo effects of cisplatin and PtAcacDMS on normal development. Moreover, to verify the dose-dependence of the effects, different groups of animals were treated with 5 μg/g or 10 μg/g body weight of cisPt and PtAcacDMS. We have focused our attention on the cerebellum because it provides a useful model system to evaluate the outcomes of perinatal treatment with chemotherapeutic agents on key CNS developmental processes such as neural cells proliferation, migration and differentiation. We have demonstrated the ability of both cisPt and PtAcacDMS to reach the brain tissue once injected. The brain platinum content after PtAcacDMS treatment was notably higher (approximately 4-fold as much) than after cisPt. The platinum accumulation in the brain was still considerable 7 days after PtAcacDMS administration. However, compared with cisplatin, PtAcacDMS induces less severe changes on fundamental events of neuroarchitecture development, such as no high apoptotic events, less altered granule cell migration and Purkinje cell dendrite growth, suggesting a low neurotoxicity of this new Pt complex for normal CNS. The mild damages could be attributable to the different subcellular target of this compound as well as to a greater efficiency of the cell repair system to recognize the drug-target adducts and to repair them. Together with the previously demonstrated antineoplastic effectiveness in vitro, the findings here reported suggest PtAcacDMS as a potential alternative to cisplatin indicating, at the same time, that the choice of platinum compounds with new subcellular targets could be a strategy to prevent neurotoxicity induced by cisplatin and overcome drug resistance induced by mutations in the intrinsic apoptotic pathway.
Pure and Applied Chemistry, 2012
The products obtained by forcing the reaction with nucleosides (guanosine, Guo, and adenosine, Ad... more The products obtained by forcing the reaction with nucleosides (guanosine, Guo, and adenosine, Ado) of potential anticancer drugs for nongenomic targets [PtCl(
Biochemical and Biophysical Research Communications, 2006
The cellular prion protein (PrP C) is a highly conserved protein throughout the evolution of mamm... more The cellular prion protein (PrP C) is a highly conserved protein throughout the evolution of mammals and therefore is thought to play important cellular functions. Despite decades of intensive researches, the physiological function of PrP C remains enigmatic. Differently, in particular pathological contexts, generally referred as transmissible spongiform encephalopathies, a conformational isoform of PrP C , i.e., PrP Sc , is considered the causative agent of these diseases. In this study, we investigated putative PrP C cellular functions through the identification of PrP C protein interactants. Using a bacterial two-hybrid approach, we identified a novel interaction between PrP C and a two-pore potassium channel protein, TREK-1. This interaction was further verified in transfected eukaryotic cells using co-immunoprecipitation and confocal microscopic analysis of the fluorescent transfected proteins. Importantly, in the cerebellar cortex, the endogenous PrP C and TREK-1 proteins exhibited co-localization signals in correspondence of the Purkinje cells. Furthermore, a deletion mapping study defined the carboxyl-terminal regions of the two proteins as the possible determinants of the PrP C-TREK-1 interaction. Our results indicated a novel PrP C interacting protein and suggested that this complex might be relevant in modulating a variety of electrophysiological-dependent cellular responses.
The Anatomical Record: Advances in Integrative Anatomy and Evolutionary Biology, 2010
A natural defect in rat cerebellum postnatal development has been found in the fissura prima, con... more A natural defect in rat cerebellum postnatal development has been found in the fissura prima, consisting in various complex configurations of the cerebellar layers. We investigated the genesis of fissure malformations through immunoreactions for PCNA, GFAP, GABAA a6, and calbindin to label proliferating cells of the external granular layer (egl), radial glial fibers, mature granule cells, and Purkinje cells, respectively. Results on critical stages of rat postnatal development provided interesting evidences on how the malformation develops in fissures prima and secunda. Early (postnatal day 10) at the site of malformation, the Bergmann radial glia was often retracted and showed distortions and irregular running. The interruption of GFAP-positive radial glial fibers could fit in with the presence of clusters of PCNA-unlabeled cells in the sites of fusion of the egl; the clusters of cells are granule cells since their soma is labeled by GABAA a6. Moreover, an altered migration of granule cell precursors to the internal granular layer was evident which, in turn, affected Purkinje cell differentiation and the growth of their dendrites. In summary, the changed relationship among glial fiber morphology, granule cell migration, and Purkinje cell differentiation suggests how the Bergmann glial fibers have a basic role in the foliation process, being the driving physical force in directing migration of the granule cells at the base of fissure.
Acta Neuropathologica, 1993
The effects on the substantia nigra of (~dihydroergocryptine (DEK), a drug with strong dopaminomi... more The effects on the substantia nigra of (~dihydroergocryptine (DEK), a drug with strong dopaminomimetic activity, were tested with a severe 1-methyl-4-phenyl-l,2,3,6-tetrahydropyridine (MPTP) treatment in monkeys. Compared with monkeys treated with MPTP alone, the animals which received DEK plus MPTP showed reduced neuronal death in the substantia nigra. The reactive astrocytes were increased in number. Moreover, several axons which were immunopositive to phosphorylated neurofilament proteins and with features similar to those of control animals were seen in the pars compacta. The findings suggest that DEK preserves neuronal morphology and brain architecture.
Acta Histochemica, 1983
The histochemical detection of SSADH activity corroborates some crucial steps of the maturation o... more The histochemical detection of SSADH activity corroborates some crucial steps of the maturation of Purkinje neurons in rat cerebellum. The SSADH-specific activity seems to appear early, within the first week of postnatal life, in a few Purkinje cells. Around the 12th day most cells were SSADH positive, even if some were still negative. This period is an important step of postnatal histogenesis when many histological and physiological data show important neuronal changes. At the end of cerebellar histogenesis the SSADH activity showed the same pattern as in adult rats. Our findings suggest that in the Purkinje cell population the maturation does not occur simultaneously thus confirming the concept of "heterogeneity" (different functional states).
Acta Histochemica, 1981
Cytochemical study of chromatin changes in Purkinje cell population as markers of rat cerebellar ... more Cytochemical study of chromatin changes in Purkinje cell population as markers of rat cerebellar histogenesis By GRAZIELLA BERNOCCRI and ELDA SCRERINI With 3 figures
Acta Histochemica, 1988
The intensity and distribution of histochemically demonstrable dihydrofolate reductase (FH2-R EC ... more The intensity and distribution of histochemically demonstrable dihydrofolate reductase (FH2-R EC 1.5.1.3.) in unfixed cryostat sections was studied in Purkinje neurons of adult vertebrates that have either simple neural circuits and cytoarchitectonics (Ictalurus nebulosus, Rana esculenta) or complex neural circuits and cytoarchitectonics (Coturnix coturnix japonica), compared with the rat as a control. The reaction was generally undetectable in Ictalurus nebulosus and in Rana esculenta; with positive reactions in only a few neurons. On the contrary, FH2-R in the Purkinje cell population of Coturnix coturnix japonica had several pattern (heterogeneity) as in the rat. These results suggest that the existence of FH2-R in Purkinje cell population may be correlated with the complexity of the neural circuits of the vertebrate's cerebellum and that the "heterogeneity" of the reaction may be related to the different functional states of the Purkinje cells within the cerebellum.
Acta Neuropathologica, 1987
The aim of this electron microscopy study was to further investigate the effects of cis-dichlorod... more The aim of this electron microscopy study was to further investigate the effects of cis-dichlorodiammineplatinum (cis-DDP) on the cerebellum of the immature rat. Ten-day-old animals were treated with cis-DDP subcutaneously and killed after 1, 7, 15 or 21 days. On postinjection day 1, cis-DDP effects were evident mainly in the external granular layer, with nuclear damage in many dividing cells, while their cytoplasm appeared to be less affected. Some binucleate cells were also present. On the contrary, in postmitotic or more differentiated cells, only cytoplasmic alterations were found. At later stages (postinjection day 7), the frequency of damaged cells in the external granular layer decreased, but there was a cellular deficit in the internal granular layer. Many postmitotic neurons underwent coagulative necrosis. Finally (postinjection days 15 and 21), the cellular deficit was partly compensated for by "reactive" structures, e.g., glial cell fibers, which underwent hypertrophy after initial edema. Moreover, packing densities of Bergmann astrocytes and oligodendrocytes were higher.
SUMMARY The pattern of specific Succinic-semialdehyde-dehydrogenase activity (SSADH), i.e. a degr... more SUMMARY The pattern of specific Succinic-semialdehyde-dehydrogenase activity (SSADH), i.e. a degradative step of gamma-amino-butyric acid, was analized in the Purkinje neuron populations of some species of vertebrates with different cerebellar cytoarchitectonics. The distribution of non-specific positivity (SDH activity and nothing dehydrogenase) was also considered and requires further investigation. Specific SSADH activity is not detectable in Purkinje neurons of catfish and frog, while it is found in a part of the Purkinje population of the quail. The results are discussed also from a phylogenetic standpoint on the basis of the knowledge of cerebellar anatomy and physiology in the three species studied.
International Journal of Molecular Sciences
Calcium-binding proteins (CBPs) can influence and react to Ca2+ transients and modulate the activ... more Calcium-binding proteins (CBPs) can influence and react to Ca2+ transients and modulate the activity of proteins involved in both maintaining homeostatic conditions and protecting cells in harsh environmental conditions. Hibernation is a strategy that evolved in vertebrate and invertebrate species to survive in cold environments; it relies on molecular, cellular, and behavioral adaptations guided by the neuroendocrine system that together ensure unmatched tolerance to hypothermia, hypometabolism, and hypoxia. Therefore, hibernation is a useful model to study molecular neuroprotective adaptations to extreme conditions, and can reveal useful applications to human pathological conditions. In this review, we describe the known changes in Ca2+-signaling and the detection and activity of CBPs in the nervous system of vertebrate and invertebrate models during hibernation, focusing on cytosolic Ca2+ buffers and calmodulin. Then, we discuss these findings in the context of the neuroprotectiv...
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Papers by Graziella Bernocchi