Hippocampal sclerosis is the most frequent pathology encountered in mesial temporal structures re... more Hippocampal sclerosis is the most frequent pathology encountered in mesial temporal structures resected from patients with intractable temporal lobe epilepsy and it mainly involves hippocampal neuronal loss and gliosis. These alterations are accompanied by changes in the expression of a variety of molecules in the surviving neurons, as well as axonal reorganization in both excitatory and inhibitory circuits. The alteration of a subpopulation of GABAergic interneurons that expresses the calcium binding protein parvalbumin (PV) is thought to be a key factor in the epileptogenic process. We investigated the distribution and density of parvalbumin-immunoreactive (PV-ir) neurons in surgically resected hippocampal tissue from epileptic patients with and without sclerosis. Using quantitative stereological methods, we show for the first time that there is no correlation between total neuronal loss and PV-ir neuronal loss in any of the hippocampal fields. We also observed higher values of the total neuronal density in the sclerotic subiculum, which is accompanied by a lower density of PV-ir when compared with non-sclerotic epileptic and autopsy hippocampi. These findings suggest that, the apparently normal subiculum from sclerotic patients also shows unexpected changes in the density and proportion of PV-ir neurons.
Highlights • MIMO2 had myelin-potentiating effects at 10 mg/kg.• MIMO2 showed no side-effects.• M... more Highlights • MIMO2 had myelin-potentiating effects at 10 mg/kg.• MIMO2 showed no side-effects.• MIMO2 reduced neuropathologies.
Dendritic cells (DCs) are professional antigen-presenting cells, derived from a bone marrow proge... more Dendritic cells (DCs) are professional antigen-presenting cells, derived from a bone marrow progenitor, which modulate the balance between suppression and induction of the immune response. DCs have been widely characterized in peripheral organs, especially in lymph nodes where antigen presentation mostly occurs. DCs have also been identified in the meninges and choroid plexus, as well as within the brain parenchyma. Several key aspects of DCs function in the brain are, however, still unexplored. Transgenic mice expressing fluorescent proteins in cell subsets provide valuable tools for in vivo investigations by two-photon fluorescence (TPF) microscopy. We recently demonstrated that in thy1GFP-M mice, engineered for green fluorescent protein (GFP) expression in neurons, also DCs express GFP. This murine line is, therefore, suited for the visualization of brain DCs. We here analyzed with TPF microscopy GFP-tagged brain DCs in healthy thy1GFP-M mice and during infection with Trypanosoma brucei (Tb). This parasite is the etiological agent of human African trypanosomiasis or sleeping sickness, whose encephalitic stage is fatal if untreated, and in which pathogenetic mechanisms of the neuroimmune response remain to be clarified. Our in vivo observations showed, in normal conditions, GFP-DCs in the subarachnoid space and meninges, where these cells were mainly static and occasionally in a probing-like motion. A motile behavior of GFP-DCs was also observed in the upper cortical layers, supporting a role of immunosurveillance of DCs in the healthy brain. Striking changes of motility and quantity of GFP-DCs were observed in the brain of Tb-infected thyGFP-M mice. During the early meningoencephalitic stage, GFP-DCs invaded the parenchyma with rapid and wide displacements, and also occurred in static adhesion to or crawling on the inflamed endothelium. With disease progression, a drastic decrease in the number of GFP-DCs was observed at the brain surface, and GFP-DCs appeared arranged in static clusters of cells exhibiting numerous processes, likely to increase the cell membrane surface on which antigens are exposed. Preliminary in vivo observations of thy1GFP-M mice infected with transgenic fluorescent Tb have shown direct interactions between GFP-DCs and the parasites; further analyses are ongoing. Taken together, the present in vivo investigations not only reveal a motile behavior of DCs at the brain surface and in the upper cortical layers, but also suggest a relevant role of brain DCs in African trypanosome infection. In particular, these cells could play a role in the transition from immune resistance to immune tolerance during this severe brain infection
Human African trypanosomiasis, also known as sleeping sickness, is a severe disease caused by the... more Human African trypanosomiasis, also known as sleeping sickness, is a severe disease caused by the parasite Trypanosoma brucei (T.b.), in which systemic infection evolves into meningoencephalitis1. One of the most effective strategies adopted by several parasites to attack the host immune system is to interfere with dendritic cells (DCs)2, which play a key role of immune surveillance. In African trypanosomiasis, DCs have been analyzed in peripheral organs3,4 but have not been hitherto examined in the brain. We recently demonstrated that thy1GFP-M transgenic mice represent a novel tool for the study of brain DCs because they express green fluorescent protein (GFP) not only in neurons but also in DCs5. Here, we investigated in vivo, by two-photon microscopy, DCs and their interaction with T.b. in infected thy1GFP-M mice at different time points during the meningoencephalitic stage of the disease
Dendritic cells (DCs) are a subset of leukocytes which promote the immune response by antigenpres... more Dendritic cells (DCs) are a subset of leukocytes which promote the immune response by antigenpresentation to T cells. In the central nervous system (CNS), DCs are poorly explored during inflammation and infection. Both DCs and brain\u2013derived antigens are drained by cerebrospinal fluid in the afferent lymphatic vessels of cervical lymph nodes, where antigen presentation mainly occurs. We recently demonstrated that in the thy1GFP-M transgenic mice DCs express green fluorescent protein ( GFP ) . These mice represent, therefore, a novel animal model for the study of DCs in vivo by two-photon microscopy ( TPM) . Here, we investigated DCs in an experimental model of CNS infection represented by African trypanosomiasis, also known as sleeping sickness. The causative agent of this disease, which is deadly if untreated, is the parasite Trypanosoma brucei ( Tb ) . After infection, the disease evolves from a first, hemolymphatic stage, to a second, menigoencephalitic stage, in which both T cells and parasites cross the blood-brain barrier and enter the brain parenchyma by mechanisms that still require full clarification. Our aim was understand whether DCs lead upstream events for T cells/Tb neuroinvasion. In vivo transcranial acquisition by TPM in thy1GFP-M mice infected with fluorescent trypanosomes ( DsRed Tb ) showed, during the hemolymphatic stage, direct interactions in the blood vessels between DCs and Ds-Red Tb, possibly representing antigen capture events. Moreover, during the early meningoencephalitic stage, we found that DCs are recruited from the blood and invade the brain parenchyma, where they exhibit random motions. At an advanced phase of the disease, DCs are mainly arranged in static clusters which incorporate the parasite. Confocal analysis showed that the sub-capsular zone of cervical lymph nodes, where DCs are rare or absent in basal conditions, is invaded by migratory DCs during the late meningoencephalitic stage; at the same time point, both migratory and resident DCs preferentially contact CD8+ T cells, probably to present Tb antigens and to prime the immune-response. The present findings provide for the first time evidence of a dynamic role of DCs during CNS infection and point to a role of DCs in mechanisms of pathogen neuroinvasion
Dendritic cells (DCs) present self or no-self antigens to T cells, to modulate the immune respons... more Dendritic cells (DCs) present self or no-self antigens to T cells, to modulate the immune response. DCs are widely studied in peripheral organs, but little is known about their function in the brain. We recently demonstrated (Laperchia et al., 2013) that DCs can be monitored in vivo by two-photon microscopy in thy1GFP-M transgenic mice, in which a subset of DCs is tagged by green fluorescent protein (GFP-DCs). Here we investigated by two-photon microscopy the migratory pattern of brain GDP-DCs both in basal condition and during the meningo-encephalitic stage of an experimental model of African trypanosomiasis, also known as sleeping sickness, induced by the infection with parasites Trypanosome brucei brucei. Trough a chronically implanted brain window in thy1GFP-M mice we found, in basal conditions, GFP-DCs floating in the cerebrospinal fluid or static at the pia mater/parenchyma interface. At an early stage of the meningoencephalitis, circulating GFP-DCs were in contact with the parasites, maybe representing an antigen capture process. Subsequently, DCs roll and crawl on the inflamed endothelium and were massively recruited from the blood stream to the brain parenchyma, where exhibited rapid and wide displacements. At a late stage, the number of motile GFP-DCs was significantly reduced and, interestingly, GFP-DCs were mainly arranged in static clusters that incorporate the parasite. Our results show for the first time the migratory pattern of DCs during invasion of the inflamed brain and suggest a role of brain DCs in the passage from brain immune-resistance to immune-tolerance during a parasitic infection
The perirhinal cortex (PRh) is strategically located between the neocortex and memory-related str... more The perirhinal cortex (PRh) is strategically located between the neocortex and memory-related structures such as the entorhinal cortex and the hippocampal formation. The pattern of strong reciprocal connections between these areas, together with experimental evidence that PRh damage induces specific memory deficits, has placed this cortical region at the center of a growing interest for its role in learning and memory mechanisms. The aim of the present study is to clarify the involvement of PRh in learning and retention in a novel experimental model of spatial working memory, the water T-maze. The data show that pre-acquisition neurotoxic PRh lesions caused task-learning deficits. This impairment was observed during the acquisition phase as well as the retrieval phase. On the other hand, a post-acquisition PRh neurotoxic lesion failed to impair the acquisition and the retrieval of the water T-maze task performed 32 day after lesion. These results suggest a possible key role of PRh in the acquisition but not in the retention of a working memory task. Furthermore, these results show that the water T-maze may be a suitable learning paradigm to study different components of learning and memory.
Astrocytic networks and gap junctional communication mediated by connexins (Cxs) have been repeat... more Astrocytic networks and gap junctional communication mediated by connexins (Cxs) have been repeatedly implicated in seizures, epileptogenesis, and epilepsy. However, the effect of seizures on Cx expression is controversial. The present study focused on the response of Cxs to status epilepticus (SE), which is in turn an epileptogenic insult. The expression of neuronal Cx36 and astrocytic Cx30 and Cx43 mRNAs was investigated in the brain of rats in the first day after pilocarpine-induced SE. In situ hybridization revealed a progressive decrease in Cx43 and Cx30 mRNA levels, significantly marked 24 h after SE onset in neocortical areas and the hippocampus, and in most thalamic domains, whereas Cx36 mRNA did not exhibit obvious changes. Regional evaluation with quantitative real-time-RT-PCR confirmed Cx43 and Cx30 mRNA downregulation 24 h after SE, when ongoing neuronal cell death was found in the same brain regions. Immunolabeling showed at the same time point marked a decrease in Cx43...
Dendritic cells (DCs) are a subset of leukocytes which promote the immune response by antigenpres... more Dendritic cells (DCs) are a subset of leukocytes which promote the immune response by antigenpresentation to T cells. In the central nervous system (CNS), DCs are poorly explored during inflammation and infection. Both DCs and brain\u2013derived antigens are drained by cerebrospinal fluid in the afferent lymphatic vessels of cervical lymph nodes, where antigen presentation mainly occurs. We recently demonstrated that in the thy1GFP-M transgenic mice DCs express green fluorescent protein ( GFP ) . These mice represent, therefore, a novel animal model for the study of DCs in vivo by two-photon microscopy ( TPM) . Here, we investigated DCs in an experimental model of CNS infection represented by African trypanosomiasis, also known as sleeping sickness. The causative agent of this disease, which is deadly if untreated, is the parasite Trypanosoma brucei ( Tb ) . After infection, the disease evolves from a first, hemolymphatic stage, to a second, menigoencephalitic stage, in which both T...
The perirhinal cortex (PRh) is strategically located between the neocortex and memory-related str... more The perirhinal cortex (PRh) is strategically located between the neocortex and memory-related structures such as the entorhinal cortex and the hippocampal formation. The pattern of strong reciprocal connections between these areas, together with experimental evidence that PRh damage induces specific memory deficits, has placed this cortical region at the center of a growing interest for its role in learning and memory mechanisms. The aim of the present study is to clarify the involvement of PRh in learning and retention in a novel experimental model of spatial working memory, the water T-maze. The data show that pre-acquisition neurotoxic PRh lesions caused task-learning deficits. This impairment was observed during the acquisition phase as well as the retrieval phase. On the other hand, a post-acquisition PRh neurotoxic lesion failed to impair the acquisition and the retrieval of the water T-maze task performed 32 day after lesion. These results suggest a possible key role of PRh in the acquisition but not in the retention of a working memory task. Furthermore, these results show that the water T-maze may be a suitable learning paradigm to study different components of learning and memory.
Hippocampal sclerosis is the most frequent pathology encountered in mesial temporal structures re... more Hippocampal sclerosis is the most frequent pathology encountered in mesial temporal structures resected from patients with intractable temporal lobe epilepsy and it mainly involves hippocampal neuronal loss and gliosis. These alterations are accompanied by changes in the expression of a variety of molecules in the surviving neurons, as well as axonal reorganization in both excitatory and inhibitory circuits. The alteration of a subpopulation of GABAergic interneurons that expresses the calcium binding protein parvalbumin (PV) is thought to be a key factor in the epileptogenic process. We investigated the distribution and density of parvalbumin-immunoreactive (PV-ir) neurons in surgically resected hippocampal tissue from epileptic patients with and without sclerosis. Using quantitative stereological methods, we show for the first time that there is no correlation between total neuronal loss and PV-ir neuronal loss in any of the hippocampal fields. We also observed higher values of the total neuronal density in the sclerotic subiculum, which is accompanied by a lower density of PV-ir when compared with non-sclerotic epileptic and autopsy hippocampi. These findings suggest that, the apparently normal subiculum from sclerotic patients also shows unexpected changes in the density and proportion of PV-ir neurons.
Highlights • MIMO2 had myelin-potentiating effects at 10 mg/kg.• MIMO2 showed no side-effects.• M... more Highlights • MIMO2 had myelin-potentiating effects at 10 mg/kg.• MIMO2 showed no side-effects.• MIMO2 reduced neuropathologies.
Dendritic cells (DCs) are professional antigen-presenting cells, derived from a bone marrow proge... more Dendritic cells (DCs) are professional antigen-presenting cells, derived from a bone marrow progenitor, which modulate the balance between suppression and induction of the immune response. DCs have been widely characterized in peripheral organs, especially in lymph nodes where antigen presentation mostly occurs. DCs have also been identified in the meninges and choroid plexus, as well as within the brain parenchyma. Several key aspects of DCs function in the brain are, however, still unexplored. Transgenic mice expressing fluorescent proteins in cell subsets provide valuable tools for in vivo investigations by two-photon fluorescence (TPF) microscopy. We recently demonstrated that in thy1GFP-M mice, engineered for green fluorescent protein (GFP) expression in neurons, also DCs express GFP. This murine line is, therefore, suited for the visualization of brain DCs. We here analyzed with TPF microscopy GFP-tagged brain DCs in healthy thy1GFP-M mice and during infection with Trypanosoma brucei (Tb). This parasite is the etiological agent of human African trypanosomiasis or sleeping sickness, whose encephalitic stage is fatal if untreated, and in which pathogenetic mechanisms of the neuroimmune response remain to be clarified. Our in vivo observations showed, in normal conditions, GFP-DCs in the subarachnoid space and meninges, where these cells were mainly static and occasionally in a probing-like motion. A motile behavior of GFP-DCs was also observed in the upper cortical layers, supporting a role of immunosurveillance of DCs in the healthy brain. Striking changes of motility and quantity of GFP-DCs were observed in the brain of Tb-infected thyGFP-M mice. During the early meningoencephalitic stage, GFP-DCs invaded the parenchyma with rapid and wide displacements, and also occurred in static adhesion to or crawling on the inflamed endothelium. With disease progression, a drastic decrease in the number of GFP-DCs was observed at the brain surface, and GFP-DCs appeared arranged in static clusters of cells exhibiting numerous processes, likely to increase the cell membrane surface on which antigens are exposed. Preliminary in vivo observations of thy1GFP-M mice infected with transgenic fluorescent Tb have shown direct interactions between GFP-DCs and the parasites; further analyses are ongoing. Taken together, the present in vivo investigations not only reveal a motile behavior of DCs at the brain surface and in the upper cortical layers, but also suggest a relevant role of brain DCs in African trypanosome infection. In particular, these cells could play a role in the transition from immune resistance to immune tolerance during this severe brain infection
Human African trypanosomiasis, also known as sleeping sickness, is a severe disease caused by the... more Human African trypanosomiasis, also known as sleeping sickness, is a severe disease caused by the parasite Trypanosoma brucei (T.b.), in which systemic infection evolves into meningoencephalitis1. One of the most effective strategies adopted by several parasites to attack the host immune system is to interfere with dendritic cells (DCs)2, which play a key role of immune surveillance. In African trypanosomiasis, DCs have been analyzed in peripheral organs3,4 but have not been hitherto examined in the brain. We recently demonstrated that thy1GFP-M transgenic mice represent a novel tool for the study of brain DCs because they express green fluorescent protein (GFP) not only in neurons but also in DCs5. Here, we investigated in vivo, by two-photon microscopy, DCs and their interaction with T.b. in infected thy1GFP-M mice at different time points during the meningoencephalitic stage of the disease
Dendritic cells (DCs) are a subset of leukocytes which promote the immune response by antigenpres... more Dendritic cells (DCs) are a subset of leukocytes which promote the immune response by antigenpresentation to T cells. In the central nervous system (CNS), DCs are poorly explored during inflammation and infection. Both DCs and brain\u2013derived antigens are drained by cerebrospinal fluid in the afferent lymphatic vessels of cervical lymph nodes, where antigen presentation mainly occurs. We recently demonstrated that in the thy1GFP-M transgenic mice DCs express green fluorescent protein ( GFP ) . These mice represent, therefore, a novel animal model for the study of DCs in vivo by two-photon microscopy ( TPM) . Here, we investigated DCs in an experimental model of CNS infection represented by African trypanosomiasis, also known as sleeping sickness. The causative agent of this disease, which is deadly if untreated, is the parasite Trypanosoma brucei ( Tb ) . After infection, the disease evolves from a first, hemolymphatic stage, to a second, menigoencephalitic stage, in which both T cells and parasites cross the blood-brain barrier and enter the brain parenchyma by mechanisms that still require full clarification. Our aim was understand whether DCs lead upstream events for T cells/Tb neuroinvasion. In vivo transcranial acquisition by TPM in thy1GFP-M mice infected with fluorescent trypanosomes ( DsRed Tb ) showed, during the hemolymphatic stage, direct interactions in the blood vessels between DCs and Ds-Red Tb, possibly representing antigen capture events. Moreover, during the early meningoencephalitic stage, we found that DCs are recruited from the blood and invade the brain parenchyma, where they exhibit random motions. At an advanced phase of the disease, DCs are mainly arranged in static clusters which incorporate the parasite. Confocal analysis showed that the sub-capsular zone of cervical lymph nodes, where DCs are rare or absent in basal conditions, is invaded by migratory DCs during the late meningoencephalitic stage; at the same time point, both migratory and resident DCs preferentially contact CD8+ T cells, probably to present Tb antigens and to prime the immune-response. The present findings provide for the first time evidence of a dynamic role of DCs during CNS infection and point to a role of DCs in mechanisms of pathogen neuroinvasion
Dendritic cells (DCs) present self or no-self antigens to T cells, to modulate the immune respons... more Dendritic cells (DCs) present self or no-self antigens to T cells, to modulate the immune response. DCs are widely studied in peripheral organs, but little is known about their function in the brain. We recently demonstrated (Laperchia et al., 2013) that DCs can be monitored in vivo by two-photon microscopy in thy1GFP-M transgenic mice, in which a subset of DCs is tagged by green fluorescent protein (GFP-DCs). Here we investigated by two-photon microscopy the migratory pattern of brain GDP-DCs both in basal condition and during the meningo-encephalitic stage of an experimental model of African trypanosomiasis, also known as sleeping sickness, induced by the infection with parasites Trypanosome brucei brucei. Trough a chronically implanted brain window in thy1GFP-M mice we found, in basal conditions, GFP-DCs floating in the cerebrospinal fluid or static at the pia mater/parenchyma interface. At an early stage of the meningoencephalitis, circulating GFP-DCs were in contact with the parasites, maybe representing an antigen capture process. Subsequently, DCs roll and crawl on the inflamed endothelium and were massively recruited from the blood stream to the brain parenchyma, where exhibited rapid and wide displacements. At a late stage, the number of motile GFP-DCs was significantly reduced and, interestingly, GFP-DCs were mainly arranged in static clusters that incorporate the parasite. Our results show for the first time the migratory pattern of DCs during invasion of the inflamed brain and suggest a role of brain DCs in the passage from brain immune-resistance to immune-tolerance during a parasitic infection
The perirhinal cortex (PRh) is strategically located between the neocortex and memory-related str... more The perirhinal cortex (PRh) is strategically located between the neocortex and memory-related structures such as the entorhinal cortex and the hippocampal formation. The pattern of strong reciprocal connections between these areas, together with experimental evidence that PRh damage induces specific memory deficits, has placed this cortical region at the center of a growing interest for its role in learning and memory mechanisms. The aim of the present study is to clarify the involvement of PRh in learning and retention in a novel experimental model of spatial working memory, the water T-maze. The data show that pre-acquisition neurotoxic PRh lesions caused task-learning deficits. This impairment was observed during the acquisition phase as well as the retrieval phase. On the other hand, a post-acquisition PRh neurotoxic lesion failed to impair the acquisition and the retrieval of the water T-maze task performed 32 day after lesion. These results suggest a possible key role of PRh in the acquisition but not in the retention of a working memory task. Furthermore, these results show that the water T-maze may be a suitable learning paradigm to study different components of learning and memory.
Astrocytic networks and gap junctional communication mediated by connexins (Cxs) have been repeat... more Astrocytic networks and gap junctional communication mediated by connexins (Cxs) have been repeatedly implicated in seizures, epileptogenesis, and epilepsy. However, the effect of seizures on Cx expression is controversial. The present study focused on the response of Cxs to status epilepticus (SE), which is in turn an epileptogenic insult. The expression of neuronal Cx36 and astrocytic Cx30 and Cx43 mRNAs was investigated in the brain of rats in the first day after pilocarpine-induced SE. In situ hybridization revealed a progressive decrease in Cx43 and Cx30 mRNA levels, significantly marked 24 h after SE onset in neocortical areas and the hippocampus, and in most thalamic domains, whereas Cx36 mRNA did not exhibit obvious changes. Regional evaluation with quantitative real-time-RT-PCR confirmed Cx43 and Cx30 mRNA downregulation 24 h after SE, when ongoing neuronal cell death was found in the same brain regions. Immunolabeling showed at the same time point marked a decrease in Cx43...
Dendritic cells (DCs) are a subset of leukocytes which promote the immune response by antigenpres... more Dendritic cells (DCs) are a subset of leukocytes which promote the immune response by antigenpresentation to T cells. In the central nervous system (CNS), DCs are poorly explored during inflammation and infection. Both DCs and brain\u2013derived antigens are drained by cerebrospinal fluid in the afferent lymphatic vessels of cervical lymph nodes, where antigen presentation mainly occurs. We recently demonstrated that in the thy1GFP-M transgenic mice DCs express green fluorescent protein ( GFP ) . These mice represent, therefore, a novel animal model for the study of DCs in vivo by two-photon microscopy ( TPM) . Here, we investigated DCs in an experimental model of CNS infection represented by African trypanosomiasis, also known as sleeping sickness. The causative agent of this disease, which is deadly if untreated, is the parasite Trypanosoma brucei ( Tb ) . After infection, the disease evolves from a first, hemolymphatic stage, to a second, menigoencephalitic stage, in which both T...
The perirhinal cortex (PRh) is strategically located between the neocortex and memory-related str... more The perirhinal cortex (PRh) is strategically located between the neocortex and memory-related structures such as the entorhinal cortex and the hippocampal formation. The pattern of strong reciprocal connections between these areas, together with experimental evidence that PRh damage induces specific memory deficits, has placed this cortical region at the center of a growing interest for its role in learning and memory mechanisms. The aim of the present study is to clarify the involvement of PRh in learning and retention in a novel experimental model of spatial working memory, the water T-maze. The data show that pre-acquisition neurotoxic PRh lesions caused task-learning deficits. This impairment was observed during the acquisition phase as well as the retrieval phase. On the other hand, a post-acquisition PRh neurotoxic lesion failed to impair the acquisition and the retrieval of the water T-maze task performed 32 day after lesion. These results suggest a possible key role of PRh in the acquisition but not in the retention of a working memory task. Furthermore, these results show that the water T-maze may be a suitable learning paradigm to study different components of learning and memory.
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