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Paradigms of sleep deprivation (SD) and memory testing in rodents (laboratory rats and mice) are here reviewed. The vast majority of these studies have been aimed at understanding the contribution of sleep to cognition, and in particular to memory. Relatively little attention, instead, has been devoted to SD as a challenge to induce a transient memory impairment, and therefore as a tool to test cognitive enhancers in drug discovery. The purpose of this article is to provide an overview of the studies that have accurately described methodological aspects of the SD protocol and behavioral paradigm in order to critically assess them and propose SD protocols that could be employed as cognitive challenge. Total SD, partial or state-selective SD (rapid eye movement SD) procedures are first reviewed, followed by procedures to investigate SD-induced impairment of learning and memory consolidation. Thus, a platform of knowledge is here provided for laboratory protocols that could be used to assess the efficacy of drugs designed to improve memory performance in rodents, including rodent models of neurodegenerative diseases that cause cognitive deficits, and Alzheimer’s disease in particular. Issues in the interpretation of such preclinical data and their predictive value for clinical translation are also discussed.

Intramuscular injections of botulinum toxin A were made into the snout of 3-month- and 3-week-old rats, resulting in transient paralysis of the facial muscles. Nicotinamide adenine dinucleotide phosphate (NADPH)-diaphorase histochemistry, which is a marker of nitric oxide synthase activity in fixed tissue and, in particular, in injured motoneurons, was studied in the facial nucleus. At variance with control injections of saline, the histochemical staining was found to be induced in facial motoneurons after botulinum toxin injection. The occurrence and persistence of the histochemical positivity in facial motoneurons paralleled that of muscle paralysis. These findings indicate that the enzyme of synthesis of the free radical nitric oxide can be induced in motoneurons after a functional disconnection from the target, which spares the axon and is associated with cell survival.

Induction of NADPH-diaphorase (NDP) following ischemic infarction was studied by means of histochemistry in the rat cerebral cortex 1, 2, 7, and 14 days after distal occlusion of the right middle cerebral artery (MCA). The fine structure of cells in the penumbra region of the necrotic center was also investigated. MCA distal occlusion resulted in ischemic lesion of the frontoparietal cortex of variable extent; NDP induction was detected in neurons, astrocytes, macrophages, and endothelial cells, with a regional specificity and a temporal gradient. One, two, and seven days after MCA occlusion, weak NDP positivity was consistently induced in some pyramidal neurons in cortical areas neighboring the necrotic area; NDP induction was also seen in pyramidal neurons of the ipsilateral anterior cingulate and infralimbic cortices and in the tenia tecta. In addition, numerous NDP-positive pyramidal neurons were detected in the contralateral frontoparietal cortex after relatively large ischemic lesions. Two weeks after MCA occlusion, NDP induction in neurons was only evident in the deep cortical layers near the lesion. NDP histochemistry combined with glial fibrillary acidic protein immunofluorescence, performed 7 days after MCA occlusion, indicated that the astrocytes at the periphery of the necrotic area were hypertrophic and some of them were also NDP-positive. One and two days after MCA occlusion, numerous macrophages displaying NDP positivity of variable intensity were seen at the periphery of the necrotic area and in the external capsule of the ischemic cerebral hemisphere. Many endothelial cells in the cortex and subcortical white matter were consistently NDP-positive in all rats. Electron microscopic study indicated that the area adjacent to the necrotic center was composed of fibrous astrocytes, with the morphological characteristics of proliferation, and numerous lysosome-filled macrophages. Altogether the present results suggest that focal cerebral ischemia may induce in different cell types nitric oxide synthase, which is equivalent to NDP in fixed tissue. The induction of nitric oxide synthase may be related to (1) blood-flow regulation at relatively early postischemic stages, which may decline when collateral circulation is established, and/or (2) cytotoxic or neuroprotective mechanisms.

Neuronal cell populations giving origin to bifurcating projections to the septum and the entorhinal cortex were studied in the rat by means of double retrograde labeling using the fluorescent tracers Fast Blue and Diamidino Yellow. Double labeled pyramidal neurons were consistently detected in the temporal level of the CA1 area and subiculum of the hippocampal formation, where they represented at least 50% of the cells retrogradely labeled from the entorhinal injections. Double labeled neurons were also detected in the amygdala, where they prevailed in the basal complex. Scattered double labeled neurons were observed in a number of hypothalamic nuclei, with a slight predominance in the preoptic region. Finally, a few double labeled cells were detected in the midline thalamus, and especially in the thalamic paraventricular nucleus. In all these structures, double labeled neurons were located ispilaterally to the injection sites. The present data indicate that the septum and entorhinal cortex are tightly interconnected by axonal bifurcations deriving from a variety of telencephalic and diencephalic sources.

The term “axon” was coined by Koelliker in 1896. To celebrate the occasion of this centennial, the contributions made in the 19th century by Remak and Deiters to the discovery of the structural uniqueness of the axon, and its continuity with the nerve cell body are highlighted briefly, and an overview is provided of the pioneering studies performed in the 18th century. Leeuwenhoek, Dutch pioneer of microscopic investigations, first observed nerve fibers in 1717 and described them as “very minute vessels.” Fontana, rigorous and ingenious investigator primarily interested in the biological effects of the viper venom, is credited with the first report of nerve fibers, of which he provided in 1781 a remarkable description as “transparent, uniform and simple cylinders,” “primitive organic elements” that can form a nerve. Copyright © 1996 Elsevier Science Inc.

The experimental infection with the parasite Trypanosoma brucei in the rat provides a unique model of dysfunction of the sleep regulatory mechanisms, because the length of synchronized sleep episodes is selectively and dramatically reduced in the advanced stages of the disease. In the present study, melatonin was acutely administered (3 mg/kg SC) to trypanosome-infected rats, before the sleep onset. This treatment resulted in a significant increase of the length of synchronized sleep episodes in respect to the infected animals and to those that had received only the vehicle. Thus, melatonin restored a normal sleep pattern during the infection. Similar findings were obtained with the new melatonin agonist S-20098. The sleep parameters were not significantly modified by either melatonin or S-20098 acute administration to noninfected animals. These findings indicate that exogenous melatonin and S-20098 exert a selective regulatory action on sleep fragmentation during experimental trypanosomiasis.

Structural asymmetry in diencephalic regions has been reported in a number of studies since the pioneering observations by Kemali and Braitenberg, Atlas of the frog's brain. Springer Verlag: 1969. Anatomical differences between the left and right habenulae have been identified in many lower vertebrate species. While there are few reports of structural asymmetry in the dorsal thalamus, there is evidence that asymmetrical thalamofugal projections can be induced in the visual system of chicks by lateralized sensory stimulation prior to hatching. Finally, there have been consistent reports of differences between and right sides of the hypothalamus in their sensitivity to the effects of circulating gonadal hormones in rats. In most cases, these asymmetries are sex-linked and correspond to a lateralization of function. Although the significance of these diencephalic asymmetries is still enigmatic, their existence indicates that asymmetry is not a phylogenetically recent feature of the brain, and the left-right differences in the brain may be mediated by a common ontogenetic mechanism and may underlie the development of highly specialized functions.

Volkensin and ricin, either free or conjugated with colloidal gold, were injected into the cerebellar cortex of rats. The inferior olive and pontine nuclei were examined to verify the retrograde axonal transport of these two toxins, and the consequent neuronal damage. No evidence was obtained of a retrograde axonal transport of ricin in these pathways. Injection of gold-conjugated volkensin in the cerebellar cortex resulted in retrogradely labelled neurones in the inferior olive after 3 h, and in the pontine nuclei after 6 h. Degenerative changes were very severe in the retrogradely labelled neurones 48 h after the gold-conjugated volkensin injection. In the Nissl-stained material, neuronal degeneration started to be evident in the inferior olive 12 h, and in pontine nuclei 6 h, after volkensin injection. The neuronal degeneration in both the inferior olive and pons increased up to 4 days after the injection. These findings provide direct evidence of the retrograde axonal transport of volkensin in the central nervous system, and of the time course of the consequent degenerative changes in the afferents to the cerebellar cortex.

Information processing streams that proceed through the sensory unimodal and multimodal areas of the cerebral cortex, and hence the limbic system, bring from perception to memory. This itinerary through the neural networks underlying perception and cognition is briefly outlined through famous passages from Marcel Proust's "Swann's Way", in which the flavour of the "petites madeleines" triggers vivid recollection.

Background and aim In the framework of IMI PharmaCog project (Grant Agreement n°115009, www.pharmacog.org ), this study evaluated whether cortical sources of resting state EEG rhythms were related to cerebrospinal fluid A®42 level in amnesic mild cognitive impairment (MCI) subjects and have translational value in mouse model of AD. Methods The research data (including human biological samples) were sourced ethically and used in line with international ethical standards. EEG rhythms were recorded in 127 aMCI subjects. Cortical sources of global delta (2–4 Hz) and low-frequency alpha (8–10.5 Hz) EEG rhythms were estimated by LORETA package. Back translation was tested on on-going EEG rhythms in wild type and transgenic mouse models of AD developing accumulation of A®42 in the brain (i.e. PDAPP, TASTPM). Results and conclusions (1) delta (

Objectives In the framework of IMI PharmaCog project (Grant Agreement n°115009), we evaluated whether spectral electroencephalographic (EEG) markers of motor activity change across aging in wild type (WT) mice and in transgenic models of AD. Methods All animal studies were carried out ethically (86/609/EEC). EEG data were recorded from bipolar fronto-parietal electrode in 85 WT mice, 22 PDAPP, and 33 TASTPM mice. Artifact-free EEG segments during wake active and passive state (no sleep) were used as input for EEG power density analysis. Results Statistical results (p Conclusions EEG markers of motor activity are useful to unveil neurophysiological mechanisms of cortical neural synchronization characterizing the physiological and pathological aging in mice.

Aging-related changes in the peripheral immune response are well documented, but less is known about changes of the immune response in the central nervous system. Reactivity of microglia, effectors of the brain innate immunity, is known to increase in the aged brain, but little attention has been hitherto devoted to T cell recruitment. Data in rodents point to a gradual enhancement of T cell homing to the brain in the steady state since the middle age. Experimental findings also point to enhanced transmigration of lymphocytes as part of an amplified response of the aging brain to acute exogenous inflammatory insults. Thus, available data support the capacity of the aged brain to mount a robust immune response, in contrast with peripheral immunity decline, and indicate that such central response involves recruitment of lymphocytes. These findings open many questions, including blood-brain barrier molecular regulation and infiltrated T cell subtypes during normal aging. The crosstalk between T cells, glia and neurons also remains to be clarified in the aged brain parenchyma. This intercellular dialogue and related signaling could be relevant for both protection of the aged brain and its vulnerability to neurological disease.

Background Human African trypanosomiasis (HAT) or sleeping sickness leads to a complex neuropsychiatric syndrome with characteristic sleep alterations. Current division into a first, hemolymphatic stage and second, meningoencephalitic stage is primarily based on the detection of white blood cells and/or trypanosomes in the cerebrospinal fluid. The validity of this criterion is, however, debated, and novel laboratory biomarkers are under study. Objective clinical HAT evaluation and monitoring is therefore needed. Polysomnography has effectively documented sleep-wake disturbances during HAT, but could be difficult to apply as routine technology in field work. The non-invasive, cost-effective technique of actigraphy has been widely validated as a tool for the ambulatory evaluation of sleep disturbances. In this pilot study, actigraphy was applied to the clinical assessment of HAT patients. Methods/Principal Findings Actigraphy was recorded in patients infected by Trypanosoma brucei gambiense, and age- and sex-matched control subjects. Simultaneous nocturnal polysomnography was also performed in the patients. Nine patients, including one child, were analyzed at admission and two of them also during specific treatment. Parameters, analyzed with user-friendly software, included sleep time evaluated from rest-activity signals, rest-activity rhythm waveform and characteristics. The findings showed sleep-wake alterations of various degrees of severity, which in some patients did not parallel white blood cell counts in the cerebrospinal fluid. Actigraphic recording also showed improvement of the analyzed parameters after treatment initiation. Nocturnal polysomnography showed alterations of sleep time closely corresponding to those derived from actigraphy. Conclusions/Significance The data indicate that actigraphy can be an interesting tool for HAT evaluation, providing valuable clinical information through simple technology, well suited also for long-term follow-up. Actigraphy could therefore objectively contribute to the clinical assessment of HAT patients. This method could be incorporated into a clinical scoring system adapted to HAT to be used in the evaluation of novel treatments and laboratory biomarkers., Author Summary The clinical picture of the parasitic disease human African trypanosomiasis (HAT, also called sleeping sickness) is dominated by sleep alterations. We here used actigraphy to evaluate patients affected by the Gambiense form of HAT. Actigraphy is based on the use of battery-run, wrist-worn devices similar to watches, widely used in middle-high income countries for ambulatory monitoring of sleep disturbances. This pilot study was motivated by the fact that the use of polysomnography, which is the gold standard technology for the evaluation of sleep disorders and has greatly contributed to the objective identification of signs of disease in HAT, faces tangible challenges in resource-limited countries where the disease is endemic. We here show that actigraphy provides objective data on the severity of sleep-wake disturbances that characterize HAT. This technique, which does not disturb the patient's routine activities and can be applied at home, could therefore represent an interesting, non-invasive tool for objective HAT clinical assessment and long-term monitoring under field conditions. The use of this method could provide an adjunct marker of HAT severity and for treatment follow-up, or be evaluated in combination with other disease biomarkers in body fluids that are currently under investigation in many laboratories.

Modafinil (MOD) is a wake-promoting drug with pro-cognitive properties. Despite its increasing use, the neuronal substrates of MOD action remain elusive. In particular, animal studies have highlighted a putative role of diencephalic areas as primary neuronal substrate of MOD action, with inconsistent evidence of recruitment of fronto-cortical areas despite the established pro-cognitive effects of the drug. Moreover, most animal studies have employed doses of MOD of limited clinical relevance. We used pharmacological magnetic resonance imaging (phMRI) in the anesthetized rat to map the circuitry activated by a MOD dose producing clinically relevant plasma exposure, as here ascertained by pharmacokinetic measurements. We observed prominent and sustained activation of the prefrontal and cingulate cortex, together with weaker but significant activation of the somatosensory cortex, medial thalamic domains, hippocampus, ventral striatum and dorsal raphe. Correlation analysis of phMRI data highlighted enhanced connectivity within a neural network including dopamine projections from the ventral tegmental area to the nucleus accumbens. The pro-arousing effect of MOD was assessed using electroencephalographic recording under anesthetic conditions comparable to those used for phMRI, together with the corresponding Fos immunoreactivity distribution. MOD produced electroencephalogram desynchronization, resulting in reduced delta and increased theta frequency bands, and a pattern of Fos induction largely consistent with the phMRI study. Altogether, these findings show that clinically relevant MOD doses can robustly activate fronto-cortical areas involved in higher cognitive functions and a network of pro-arousing areas, which provide a plausible substrate for the wake-promoting and pro-cognitive effects of the drug.

Summary Background In the AVERROES study, apixaban, a novel factor Xa inhibitor, reduced the risk of stroke or systemic embolism in patients with atrial fibrillation who were at high risk of stroke but unsuitable for vitamin K antagonist therapy. We aimed to investigate whether the subgroup of patients with previous stroke or transient ischaemic attack (TIA) would show a greater benefit from apixaban compared with aspirin than would patients without previous cerebrovascular events. Methods In AVERROES, 5599 patients (mean age 70 years) with atrial fibrillation who were at increased risk of stroke and unsuitable for vitamin K antagonist therapy were randomly assigned to receive apixaban (5 mg twice daily) or aspirin (81–324 mg per day). The mean follow-up was 1·1 years. The primary efficacy outcome was stroke or systemic embolism; the primary safety outcome was major bleeding. Patients and investigators were masked to study treatment. In this prespecified subgroup analysis, we used Kaplan-Meier estimates of 1-year event risk and Cox proportional hazards regression models to compare the effects of apixaban in patients with and without previous stroke or TIA. AVERROES is registered at ClinicalTrials.gov, number NCT00496769. Findings In patients with previous stroke or TIA, ten events of stroke or systemic embolism occurred in the apixaban group (n=390, cumulative hazard 2·39% per year) compared with 33 in the aspirin group (n=374, 9·16% per year; hazard ratio [HR] 0·29, 95% CI 0·15–0·60). In those without previous stroke or TIA, 41 events occurred in the apixaban group (n=2417, 1·68% per year) compared with 80 in the aspirin group (n=2415, 3·06% per year; HR 0·51, 95% CI 0·35–0·74). The p value for interaction of the effects of aspirin and apixaban with previous cerebrovascular events was 0·17. Major bleeding was more frequent in patients with history of stroke or TIA than in patients without (HR 2·88, 95% CI 1·77–4·55) but risk of this event did not differ between treatment groups. Interpretation In patients with atrial fibrillation, apixaban is similarly effective whether or not patients have had a previous stroke or TIA. Given that those with previous stroke or TIA have a higher risk of stroke, the absolute benefits might be greater in these patients. Funding Bristol-Myers Squibb and Pfizer.

Human African trypanosomiasis (HAT), or sleeping sickness, is a severe disease caused by Trypanosoma brucei (T.b.). The disease hallmark is sleep alterations. Brain involvement in HAT is a crucial pathogenetic step for disease diagnosis and therapy. In this study, a rat model of African trypanosomiasis was used to assess changes of sleep-wake, rest-activity, and body temperature rhythms in the time window previously shown as crucial for brain parenchyma invasion by T.b. to determine potential biomarkers of this event. Chronic radiotelemetric monitoring in Sprague-Dawley rats was used to continuously record electroencephalogram, electromyogram, rest-activity, and body temperature in the same animals before (baseline recording) and after infection. Rats were infected with T.b. brucei. Data were acquired from 1 to 20 d after infection (parasite neuroinvasion initiates at 11-13 d post-infection in this model), and were compared to baseline values. Sleep parameters were manually scored from electroencephalographic-electromyographic tracings. Circadian rhythms of sleep time, slow-wave activity, rest-activity, and body temperature were studied using cosinor rhythmometry. Results revealed alterations of most of the analyzed parameters. In particular, sleep pattern and sleep-wake organization plus rest-activity and body temperature rhythms exhibited early quantitative and qualitative alterations, which became marked around the time interval crucial for parasite neuroinvasion or shortly after. Data derived from actigrams showed close correspondence with those from hypnograms, suggesting that rest-activity could be useful to monitor sleep-wake alterations in African trypanosomiasis.

Induction of an interferon-gamma-like molecule, previously isolated from neurons (N-IFN-gamma), and of the neuronal isoform I of the synthetic enzyme of the free radical nitric oxide, nitric oxide synthase I, as well as of NADPH-diaphorase, were examined in axotomized dorsal motor vagal and hypoglossal neurons. Unilateral transection of the vagal and hypoglossal nerves was performed in the same rat and an induction of N-IFN-gamma and nitric oxide synthase I immunostaining as well as NADPH-diaphorase histochemical positivity was observed in the ipsilateral motoneurons after 2-4 days. The immuno- and enzyme-histochemical positivities were much stronger in the dorsal motor vagal neurons than in hypoglossal neurons. Two and 4 weeks after axotomy N-IFN-gamma immunoreactivity and NADPH-diaphorase positivity persisted in the former, but started to decrease in the latter neurons. Previous data have shown that 23 weeks after nerve transection the majority of the dorsal motor vagal neurons are lost, while the majority of the hypoglossal neurons survive. The high and persistent expression of N-IFN-gamma and nitric oxide synthase I after axotomy in the dorsal motor vagal neurons, that are largely destined to die, indicates that the co-induction of these two molecules may be implicated in the pathogenesis of neuronal degeneration.

Several lines of evidence indicate that neuromuscular junction (NMJ) destruction and disassembly is an early phenomenon in amyotrophic lateral sclerosis (ALS). Here we analyzed by confocal and electron microscopy the NMJ structure in the diaphragm of SOD1G93A mice at symptom onset. In these mice, which provide a model for familial ALS, diaphragm denervation (~50%) as well as gastrocnemius denervation (~40%) was found. In addition, the size of the synaptic vesicle pool was reduced and alterations of mitochondria were observed in approximately 40% of the remaining presynaptic terminals. Chronic treatment of SOD1G93A mice with the anabolic steroid nandrolone during the presymptomatic stage preserved the diaphragm muscle mass and features indicative of synaptic activity. These features were represented by the number of vesicles docked within 200 nm from the presynaptic membrane and area of acetylcholine receptor clusters. Structural preservation of mitochondria was documented in presynaptic terminals. However, innervation of diaphragm muscle fibers was only slightly increased in nandrolone-treated SOD1-mutant mice. Altogether the results point out and define fine structural alterations of diaphragm NMJs in the murine model of familial ALS at symptom onset, and indicate that nandrolone may prevent or delay structural alterations in NMJ mitochondria and stimulate presynaptic activity but does not prevent muscle denervation during the disease.

Konzo is a self-limiting central motor-system disease associated with food dependency on cassava and low dietary intake of sulfur amino acids (SAA). Under conditions of SAA-deficiency, ingested cassava cyanogens yield metabolites that include thiocyanate and cyanate, a protein carbamoylating agent. We studied the physical and biochemical modifications of rat serum and spinal cord proteins arising from intoxication of young adult rats with 50-200 mg/kg linamarin, or 200 mg/kg sodium cyanate (NaOCN), or vehicle (saline) and fed either a normal amino acid- or SAA-deficient diet for up to two weeks. Animals under SAA-deficient diet and treatment with linamarin or NaOCN developed hind limb tremors or motor weakness, respectively. LC/MS-MS analysis revealed differential albumin carbamoylation in animals treated with NaOCN, vs. linamarin/SAA-deficient diet, or vehicle. 2D-DIGE and MALDITOF/MS-MS analysis of the spinal cord proteome showed differential expression of proteins involved in oxidative mechanisms (e.g. peroxiredoxin 6), endocytic vesicular trafficking (e.g. dynamin 1), protein folding (e.g. protein disulfide isomerase), and maintenance of the cytoskeleton integrity (e.g. α-spectrin). Studies are needed to elucidate the role of the aformentioned modifications in the pathogenesis of cassava-associated motor system disease.

Aging is often accompanied by increased levels of inflammatory molecules in the organism, but age-related changes in the brain response to inflammatory challenges still require clarification. We here investigated in mice whether cytokine signaling and T-cell neuroinvasion undergo age-related changes. We first analyzed the expression of molecules involved in T-cell infiltration and cytokine signaling regulation in the septum and hippocampus of 2-3 months and 20- to 24-month-old mice at 4h after intracerebroventricular injections of tumor necrosis factor (TNF)-alpha or interferon-gammaversus saline injections. Transcripts of the chemokine CXCL9, intercellular adhesion molecule (ICAM)-1 and suppressor of cytokine signaling molecules (SOCS) 1 and 3 were increased in both age groups after cytokine injection; microglia-derived matrix metalloproteinase (MMP) 12 mRNA was induced in old mice also after control saline injections. Age-related changes in ICAM-1 protein expression and T-cell infiltration were then analyzed in mice of 3-4, 8-9 and 15-16 months at 48h after TNF-alpha injections. ICAM-1 immunoreactivity, and Western blotting in striatum, septum, hippocampus and hypothalamus showed progressive age-related enhancement of TNF-alpha-elicited ICAM-1 upregulation. Double immunofluorescence revealed ICAM-1 expression in microglia and astrocytic processes. CD3(+), CD4(+) and CD8(+) T-cells exhibited progressive age-related increases in brain parenchyma and choroid plexus after cytokine exposure. The findings indicate that the brain responses to inflammatory challenges are not only preserved with advancing age, but also include gradual amplification of ICAM-1 expression and T-cell recruitment. The data highlight molecular and cellular correlates of age-related increase of brain sensitivity to inflammatory stimuli, which could be involved in altered brain vulnerability during aging.

In the rat thalamus, immunoreactivity for the calcium binding protein calbindin (Cb) is mostly confined to neuronal cell bodies, sometimes revealing proximal dendrites, of the midline, intralaminar and posterior regions. Substance P (SP)-, cholecystokinin (CCK)- and Leu-enkephalin (L-ENK)-immunoreactive (ir) elements in the thalamus are fibre-like structures, intermingled with punctate elements probably representing axonal arborizations and their synaptic boutons. These peptidergic fibres are unevenly distributed in several thalamic domains, including the areas that contain Cb-ir neurons. The relationship between Cb-ir cell bodies and these three different peptidergic systems of thalamic innervation was studied with immunohistochemistry. Single-labelling experiments on adjacent sections and double immunostaining on the same section were performed. A considerable overlap between Cb-ir perikarya and SP-ir fibres was found in most thalamic nuclei. In particular, in the intralaminar nuclei and posterior complex. SP-ir punctate elements were frequently observed in close proximity to Cb-ir cell bodies and dendrites. On the other hand, no consistent topographical correspondence between Cb-ir perikarya and CCK- or L-ENK-ir fibres was evident. Altogether, the present data suggest a selective anatomical and, possibly, functional relationship between SP and Cb in at least a subpopulation of rat thalamic neurons.

Ultrastructural visualization of non-electron-dense fluorescent retrograde neuronal labeling was attempted by means of photo-oxidation. This procedure was used to convert the fluorescence of neurons labeled by the tracers propidium iodide, rhodamine latex microspheres and fluorogold into a stable diaminobenzidine reaction product. The ultrastructural study revealed an accumulation of electron-dense material in these cells both within lysosomes and scattered in the cytoplasmic matrix. Comparison with several different sets of control samples indicated that this material, on the basis of its amount, electron density and appearance, specifically represents the photoconversion reaction product. The effects of the intensity of the fluorescent labeling and of a prolonged photoconversion on the fine structural features of the reaction product are also described and discussed. The present findings indicate that photoconversion can be effectively applied to ultrastructural study of fluorescent retrogradely labeled neurons. The specificity of the photoconversion reaction product should be tested routinely for each fluorochrome and tissue sample.

The chapter provides an itinerary of knowledge on nervous system anatomy as one of the pillars of clinical neurology. The journey starts from the Renaissance explosion on the approach to the human body, its functions and its diseases, dealing with the seminal contributions of Leonardo da Vinci and Vesalius. The itinerary proceeds through the contributions of the 17th century, especially by Thomas Willis and the pioneering investigations of Marcello Malpighi and Antony van Leeuwenhoek, and onto the 18th century. The itinerary thus leads to the progress from gross anatomy to the microscopic investigation of the nervous system in the 19th century: the reticular theories, the revolution of the neural doctrine and their protagonists (Camillo Golgi and Santiago Ramón y Cajal), which initiated the modern era of the neurosciences. The chapter also includes sections on the contributions of developmental neuroanatomy to neurology, on the history of tract tracing, and on the cytoarchitecture of the cerebral cortex. The never-ending story of the anatomical foundations of clinical neurology continues to evolve at the dawn of the 21st century, including knowledge that guides deep brain stimulation, and novel approaches to the anatomy of the living brain based on rapidly developing neuroimaging technology.

The chapter starts from the Renaissance (although the origins of Italian neurology can be traced back to the Middle Ages), when treatises of nervous system physiopathology still followed Hippocratic and Galenic "humoral" theories. In Italy, as elsewhere in Europe, the concepts of humoral pathology were abandoned in the 18th century, when neurology was influenced by novel trends. Neurology acquired the status of clinical discipline (as "clinic of mental diseases") after national reunification (declared in 1861 but completed much later). At the end of the 19th and first decades of the 20th century, eminent Italian "neuropsychiatrists" (including, among many others, Ugo Cerletti, who introduced electroconvulsive shock therapy in 1938) stimulated novel knowledge and approaches, "centers of excellence" flourished, and "Neurological Institutes" were founded. In the first half of the 20th century, the history of Italian neurology was dominated by World Wars I and II (which stimulated studies on the wounded) and the fascist regime in-between the Wars (when the flow of information was instead very limited). Italy became a republic in 1946, and modern neurology and its distinction from psychiatry were finally promoted. The chapter also provides detailed accounts of scientific societies and journals dedicated to the neurological sciences in Italy.

The chapter provides an itinerary of knowledge on nervous system anatomy as one of the pillars of clinical neurology. The journey starts from the Renaissance explosion on the approach to the human body, its functions and its diseases, dealing with the seminal contributions of Leonardo da Vinci and Vesalius. The itinerary proceeds through the contributions of the 17th century, especially by Thomas Willis and the pioneering investigations of Marcello Malpighi and Antony van Leeuwenhoek, and onto the 18th century. The itinerary thus leads to the progress from gross anatomy to the microscopic investigation of the nervous system in the 19th century: the reticular theories, the revolution of the neural doctrine and their protagonists (Camillo Golgi and Santiago Ramon y Cajal), which initiated the modern era of the neurosciences. The chapter also includes sections on the contributions of developmental neuroanatomy to neurology, on the history of tract tracing, and on the cytoarchitecture of the cerebral cortex. The never-ending story of the anatomical foundations of clinical neurology continues to evolve at the dawn of the 21st century, including knowledge that guides deep brain stimulation, and novel approaches to the anatomy of the living brain based on rapidly developing neuroimaging technology.

The chapter starts from the Renaissance (although the origins of Italian neurology can be traced back to the Middle Ages), when treatises of nervous system physiopathology still followed Hippocratic and Galenic “humoral” theories. In Italy, as elsewhere in Europe, the concepts of humoral pathology were abandoned in the 18th century, when neurology was influenced by novel trends. Neurology acquired the status of clinical discipline (as “clinic of mental diseases”) after national reunification (declared in 1861 but completed much later). At the end of the 19th and first decades of the 20th century, eminent Italian “neuropsychiatrists” (including, among many others, Ugo Cerletti, who introduced electroconvulsive shock therapy in 1938) stimulated novel knowledge and approaches, “centers of excellence” flourished, and “Neurological Institutes” were founded. In the first half of the 20th century, the history of Italian neurology was dominated by World Wars I and II (which stimulated studies on the wounded) and the fascist regime in-between the Wars (when the flow of information was instead very limited). Italy became a republic in 1946, and modern neurology and its distinction from psychiatry were finally promoted. The chapter also provides detailed accounts of scientific societies and journals dedicated to the neurological sciences in Italy.

Perturbations of neurovascular integrity and breakdown of the blood–brain barrier may lead to neuronal hypersynchronization and epileptiform activity. However, the mechanisms underlying these processes are still unclear. Furthermore, inflammatory processes related to the activation of brain vessel endothelia and/or BBB leakage have been reported in epilepsy, but no explanations have yet been provided for the relevant mechanisms of action. Our main goal is to adopt an integrated and multidisciplinary approach to this issue, taking into account different factors in the in vivo study of alterations of the ‘whole brain’ in epilepsy. Toward that end, we combine electrophysiological, neuropathological, and neuroimmunological data, in order to consider the multiple players involved in vivo in the pathogenesis of epilepsy. We report that vascular alterations and BBB leakage induced by protracted seizures are not merely epiphenomena detected in animal models of chemically induced epilepsy, but may represent crucial steps of a cascade of events responsible for epileptogenic mechanisms. Furthermore, we suggest that such events can implicate leukocyte recruitment and cytokine and chemokine release.

The olfactory system has been implicated in the pathogenesis of transmissible spongiform encephalopathies (TSEs). To examine this issue and identify the pattern of TSE agent spread after intranasal administration, we inoculated a high-infectious dose of neurotropic scrapie strain 263K into the nasal cavity of Syrian hamsters. All animals allowed to survive became symptomatic with a mean incubation period of 162.4 days. Analysis at different time points revealed deposition of the pathological prion protein (PrP(TSE)) in nasal-associated lymphoid tissues in the absence of brain involvement from 80 days post-infection (50% of the incubation period). Olfactory-related structures and brainstem nuclei were involved from 100 days post-inoculation (62% of the incubation period) when animals were still asymptomatic. Intriguingly, vagal or trigeminal nuclei were identified as early sites of PrP(TSE) deposition in some pre-symptomatic animals. These findings indicate that the 263K scrapie agent is unable to effectively spread from the olfactory neuroepithelium to the olfactory-related structures and that, after intranasal inoculation, neuroinvasion occurs through olfactory-unrelated pathways.

Terminal arborizations expressing nerve growth factor receptor (NGF-R) have been detected with immunohistochemistry in the reticular thalamic nucleus of neonate, adult and aged rats. Intracerebroventricular administration of nerve growth factor (NGF) resulted in a dramatic increase in NGF-R immunoreactivity throughout the lifespan. This effect was paralleled by a concomitant increase in NGF-R immunopositivity in the neurons of the basal forebrain, which was here demonstrated also in aged animals, thus indicating that the NGF-R immunoreactivity within the reticular thalamic nucleus derives in all likelihood from cholinergic neuronal cell bodies of the basal forebrain. Our results demonstrate a prominent ability of NGF to up-regulate its receptors within fibres innervating the reticular thalamic nucleus, and show that this up-regulation of NGF-R is maintained throughout the lifetime. Altogether this indicates that the reticular thalamic nucleus may represent a new, important site of action of endogenous NGF or NGF-like molecules within the brain. In view of the crucial role played by the reticular thalamic nucleus in gating thalamocortical information, the autoregulation of NGF-R in this structure may have important concomitants in both physiological and pathological conditions.

Biological rhythms, and especially the sleep/wake cycle, are frequently disrupted during senescence. This draws attention to the study of aging-related changes in the hypothalamic suprachiasmatic nucleus (SCN), the master circadian pacemaker. The authors here compared the SCN of young and old mice, analyzing presynaptic terminals, including the gamma-aminobutyric acid (GABA)ergic network, and molecules related to the regulation of GABA, the main neurotransmitter of SCN neurons. Transcripts of the α3 subunit of the GABAA receptor and the GABA-synthesizing enzyme glutamic acid decarboxylase isoform 67 (GAD67) were analyzed with real-time RT-PCR and GAD67 protein with Western blotting. These parameters did not show significant changes between the 2 age groups. Presynaptic terminals were identified in confocal microscopy with synaptophysin immunofluorescence, and the GABAergic subset of those terminals was revealed by the colocalization of GAD67 and synaptophysin. Quantitative analysis of labeled synaptic endings performed in 2 SCN subregions, where retinal afferents are known to be, respectively, very dense or very sparse, revealed marked aging-related changes. In both subregions, the evaluated parameters (the number of and the area covered by presynaptic terminals and by their GABAergic subset) were significantly decreased in old versus young mice. No significant differences were found between SCN tissue samples from animals sacrificed at different times of day, in either age group. Altogether, the data point out marked reduction in the synaptic network of the aging biological clock, which also affects GABAergic terminals. Such alterations could underlie aging-related SCN dysfunction, including low-amplitude output during senescence.

The existence of Golgi type II neurons was verified in the anterior intralaminar central lateral (CL) nucleus of the cat thalamus, and its projection cell types were identified, by means of Golgi impregnation. CL principal neurons were found to display a large- or medium-sized cell body and a radiate dendritic pattern. Their primary dendrites were limited in number, and had a rather long course; they were poorly ramified. The axons of principal neurons were impregnated only occasionally and for a short distance. Projection neurons of the 'bushy' or tufted type, described in the main thalamic sensory nuclei, were not identified in the CL in the present study. Typical Golgi type II neurons were found throughout CL. They were mainly small-sized, and displayed a rich dendritic arborization characterized by dendritic appendages. The axons of Golgi type II neurons were seen to give rise to extensive local arborizations. The present findings indicate that in the cat CL, principal cells are mainly represented by radiate neurons. Typical local circuit neurons also are evident in CL, suggesting that the activity of anterior intralaminar structures is regulated by intrinsic mechanisms similar to those operating in the main thalamic relay nuclei.

The organization of the thalamic midline efferents to the amygdaloid complex, hippocampal formation, and nucleus accumbens was investigated in the rat by means of multiple retrograde fluorescent tracing. The present findings indicate that these connections derive from separate cell populations of the thalamic midline, with a low degree of divergent collateralization upon more than one of the targets examined. The neural populations projecting to the amygdala, hippocampus, or accumbens are highly intermingled throughout the thalamic midline, but display some topographical prevalence. Midline thalamo-hippocampal cells are concentrated in the nucleus reuniens; thalamo-accumbens neurons prevail in the ventral portion of the paraventricular nucleus, and in the central medial nucleus. Thalamo-amygdaloid cells display a topographical prevalence in the rostral third of the thalamic midline and are concentrated in the dorsal part of the paraventricular nucleus and in the medial part of the nucleus reuniens. Both dorsally in the paraventricular nucleus and ventrally in the nucleus reuniens, thalamo-amygdaloid cells are located closer to the ependymal lining than the neurons projecting to the hippocampus or nucleus accumbens. Further, thalamo-amygdaloid cells, especially in the paraventricular nucleus, extend their dendritic processes in the vicinity of the ependymal lining, where they arborize profusely. These features indicate a close topographical relationship of neurons projecting to the amygdala with ependymal cells. The fairly discrete origin of midline outputs to the amygdala, hippocampus, and accumbens indicates that the flow of information is conveyed through separate channels from the thalamic midline to limbic and limbic-related targets. Together with the literature on the limbic afferents to the thalamus, these findings emphasize the relationships between the thalamus and the limbic system subserved by parallel input-output routes. However, because of the overlap of the projection cell populations, the thalamic midline may represent a locus of interaction among neurons connected with different parts of the limbic system. The functional implications of these findings are discussed in relation to the "nonspecific" thalamic system, as well as to the circuits involved in memory formation.

Tract tracing techniques provide tools for the study of the termination or origin of central neural pathways or peripheral nerves. The understanding of the organization of neural circuits has represented one of the major goals in neuroscience since its birth. In the second half of the 19th century, the pioneers in tract tracing discovered that retrograde degeneration of neuronal cell bodies and anterograde degeneration of fibers could be used to trace pathways in the nervous system. Thus, “the earliest way to identify the neurons sending their axons to a given neural structure was to destroy the structure” (20). In addition, the Golgi method, which impregnates random subsets of neuronal cell bodies and processes in their entirety, played a crucial role not only in unraveling the basic structure of the nervous system, but also in pioneering investigations on its connectivity. Cajal’s seminal studies on the wiring of the nervous system were in fact based on Golgi-impregnated material (5). It is very hard, however, to effectively impregnate the axons and to reconstruct their trajectory over long distances with the Golgi staining. This techconnections, i.e., at a limited distance from the cell body.

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The word inflammation derives from Latin inflammare, meaning to set in flame, and signifies the heat and redness related to an increased blood flow and vasodilation in the affected tissue. Although vasodilation, edema, and extravasation of white blood cells are the hallmarks of inflammation, its definition has become broader and has been more loosely applied to tissue reactions to injuries. With the discovery of molecules that mediate inflammation and, in particular, the extensive number of cytokines that regulate the cellular response to inflammation, an inflammatory component is now frequently ascribed to neurodegenerative diseases that were traditionally considered as noninflammatory. This is due to the production of proinflammatory cytokines in activated microglial cells and astrocytes in such diseases. The classical hallmarks of inflammation are, however, lacking. On the other hand, events considered as components of systemic inflammation occur also in normal brain function, e.g., increase in local blood flow related to increased brain activities, and release of certain proinflammatory cytokines, including those that may be involved in mediating sleep–wakefulness. In this overview we will focus on recent observations on how mediators of vascular responses in inflammation outside the nervous system (e.g., histamine) and proinflammatory cytokines may affect the biological clock in the brain and be involved in the regulation of the switch between sleep and wakefulness, and hence cause disturbances in the function of this switch during diseases.

Apoptosis plays a key role in normal aging and neurodegeneration. It is now known that normal aging implies low-grade inflammation and increases susceptibility to neurodegenerative diseases, which, in turn, include a neuroinflammatory component. We here investigated, using mice of 2–3 months, 10–11 months, or 18–21 months of age, the expression of apoptosis-regulatory proteins in cortical brain regions in response to intracerebroventricular administration of pro-inflammatory cytokines. A mixture of interferon-γ and tumor necrosis factor-α was injected, using vehicle (phosphate-buffered saline) as control. At 4 days, levels of the anti-apoptotic Bcl-2 and pro-apoptotic Bax proteins in the cerebral cortex and hippocampus, examined with Western blotting, were markedly upregulated by cytokine exposure in mice of all age groups with respect to controls. Interestingly, cytokine-elicited Bcl-2 upregulation was aging-dependent, with significant enhancement paralleling the animals’ age. Cytokine-elicited Bax expression did not exhibit instead significant aging-related variation. Using the same paradigm and 1 or 2 day survival, Bcl-2 immunoreactivity was observed mainly in neurons of cortex and hippocampus of both control and cytokine-treated mice of all age groups. Furthermore, immunohistochemistry confirmed the enhancement of cytokine-elicited Bcl-2 expression in the cerebral cortex and hippocampus of old mice, and showed that this finding was already evident in the second day after cytokine exposure. The data point out the novel finding that Bcl-2 and Bax expression in cortical brain regions is differentially regulated during senescence in response to an acute inflammatory challenge. Aging-related Bcl-2 increases in neurons after cytokine exposure could contribute to amplify neuroprotective mechanisms in the old brain.

Objective To describe a novel molecular and pathological phenotype of Creutzfeldt-Jakob disease. Patient A 69-year-old woman with behavioral and personality changes followed by rapidly evolving dementia. Results Postmortem examination of the brain showed intracellular prion protein deposition and axonal swellings filled with amyloid fibrils. Biochemical analysis of the pathological prion protein disclosed a previously unrecognized PrP Sc tertiary structure lacking diglycosylated species. Genetic analysis revealed a wild-type prion protein gene. The prion agent responsible for this atypical phenotype was successfully passaged to bank voles. Conclusion To our knowledge, our results define a new human prion disorder characterized by intracellular accumulation of a novel type of pathological prion protein.

International audience; In humans, activity rhythms become fragmented and attenuated in the elderly. This suggests an alteration of the circadian system per se that could in turn affect the expression of biological rhythms. In primates, very few studies have analyzed the effect of aging on the circadian system. The mouse lemur provides a unique model of aging in non-human primates. To assess the effect of aging on the circadian system of this primate, we recorded the circadian and daily rhythms of locomotor activity of mouse lemurs of various ages. We also examined age-related changes in the daily rhythm of immunoreactivities for vasoactive intestinal polypeptide (VIP) and arginine-vasopressin (AVP) in suprachiasmatic nucleus neurons (SCN), two major peptides of the biological clock. Compared to adult animals, aged mouse lemurs showed a significant increase in daytime activity and an advanced activity onset. Moreover, when maintained in constant dim red light, aged animals exhibited a shortening of the free-running period compared to adult animals. In adults, AVP immunoreactivity (ir) peaked during the second part of the day, and VIP ir peaked during the night. In aged mouse lemurs, the peaks of AVP ir and VIP ir were significantly shifted with no change in amplitude. AVP ir was most intense at the beginning of the night; whereas, VIP ir peaked at the beginning of the daytime. A weakened oscillator could account for the rhythmic disorders often observed in the elderly. Changes in the daily rhythms of AVP ir and VIP ir may affect the ability of the SCN to transmit rhythmic information to other neural target sites, and thereby modify the expression of some biological rhythms.

The possibility that genius, special talents, and skills can be explained by an analysis of brain structure has fascinated people for centuries. Scientists from various disciplines and backgrounds have investigated this intriguing concept in many different ways. This exploration is the subject of the book Postcards from the brain museum. The author, Brian Burrell, defines this quest as the “anatomy of genius” and explains that it encroaches upon key issues of past and contemporary neuroscience. These issues are highlighted by the long search for localization of function in the brain, including the nineteenth century debate over whether specific functions are precisely localized within the brain. The issue of the anatomy of genius also touches upon the major philosophical question of the relationship between the brain and the mind or how the invisible products of the brain relate to the physical structure of the organ itself. Burrell, determined to be a “neuro-detective” in the hunt for brains and heads preserved in jars, brain casts, and face masks kept in museums or forgotten on shelves, presents the results of his search in his new book. He traces the story of the anatomy of genius through accounts of unique individuals after whom the chapters are named. Burrell discusses philosopher Rene Descartes and his doctrine of ventricular localization, which sets the pineal gland within the skull as a site of convergence of sensory stimuli. He also considers anatomist Franz Joseph Gall and his doctrine of organology, i.e., the idea that certain skills and talents can be defined based on the bumps on the skull, reflecting — incorrectly as it turns out, but with a theoretically innovative approach — the development of underlying cerebral organs devoted to given faculties. This doctrine was soon defined as phrenology and was later discredited. This information is followed by tales of Lord Byron’s huge brain and by the story of mathematician Carl Friedrich Gauss’s brain and other brains studied at the University of Gottingen. The chapter about neurologist Paul Broca nicely reconstructs his discovery of a brain center for language. A chapter about neuropsychiatrist Cesare Lombroso, the father of criminal anthropology, and his writings on the “criminal man” and “man of genius” is followed by a chapter about Edouard Seguin, who introduced special education for the handicapped, less relevant in the context of this book. The next part of the book recounts the story of Charles Guiteau, who murdered US President James A. Garfield, and provides biographical sketches of and anedoctes regarding neurologist Edward C. Spitzka and anatomist Burt Wilder (who started the Wilder Brain Collection at Cornell University). The end of the book covers Walt Whitman’s encounter with phrenology and more recent stories on the brains of Russian revolutionary Vladimir Ilyich Lenin and physicist Albert Einstein. The story of Lenin’s brain is reconstructed, as it should be, through its political impact. The story on Einstein’s brain is probably too recent and controversial for an objective evaluation. The book is interesting and entertaining in most chapters but also insists sometimes on macabre details and off-focus anecdotes. The main problem is that it pretends to give an account of the history of the neurosciences by basing it on the narratives of the so-called “elite brains.” The book is informative for scientists especially interested in the brains of individuals with peculiar skills. However, the history of the neurosciences is obviously much more complex than this book would lead one to believe and is based on data totally unrelated to the history of brains of famous individuals. The book has other weaknesses, such as the correlation between the anatomy of genius and racism or “racial hygiene.” If one believes in a superior brain, then an inferior brain must also exist. This may be historically important, but it is not politically correct to present the two issues of genius and prejudice as interconnected, and this problem is dealt with rather superficially by Burrell. The brain of a genius is not superior, but it’s a brain that has developed certain skills. There are also several mistakes that make the book seem not as scholarly as it should be. For example, the theory of physiognomics is much older than the 1770s, as claimed in the book. In addition, it was Adolf Hitler, who is not mentioned in the book, and not Cesare Lombroso, who inspired Mussolini’s racial laws. Furthermore, Burrell’s view is that the study of brain anatomy based on the architecture of the cerebral cortex is a subjective exercise because interindividual variations make it totally unreliable, but this is not the case. In fact, the cortex has been subdivided by Korbinian Brodmann into numbered areas, and it is not the phrenological map that Burrell proposes. Burrell confuses the discovery of cortical organization and compartmentalization — one of the pillars of modern neuroscience — with a subjective exercise of anatomists. Finally, a more comprehensive and accurate bibliography would have enriched a book that seems to be the result of a passionate search but seems also to have been written in haste. Altogether, the book provides Burrell’s view of the localization of talents in the brain, and his cruise among documents and accounts is certainly informative. However, the book makes the same historical mistake it criticizes, leaving anecdotes as such without providing deeper insight into the history of neuroscience .

The organization of the main dopaminergic cell groups in the brain, located in the ventral mesencephalic tegmentum, and the circuits in which they are inserted are reviewed here, with emphasis on rodents. Subdivisions based on cytoarchitecture (substantia nigra, ventral tegmental area and related nuclei, retrorubral field), dopaminergic phenotype (A8, A9 and A10 cell groups) and organization in dorsal and ventral tiers are discussed and compared. Dendritic release and gap junctional protein expression, interactions with glial cells, molecular and cellular features of the chemical repertoire of midbrain dopaminergic neurons and their main inputs are also reviewed. An account is given on basal ganglia circuits, including the organization of the direct, indirect and hyperdirect pathways of information processing and dopamine modulation of these pathways. Data on the dopaminergic innervation of limbic structures, including the extended amygdala, and the distribution and laminar organization of dopaminergic fibers in the cerebral cortex are summarized. The last part of the chapter focuses on the distribution of dopamine receptor subtypes and their relative densities in different brain structures. For each of the D 1 , D 2 , D 3 , D 4 and D 1B/5 receptors, an overview and distributional maps are provided, followed by data on their localization in the rat basal ganglia, cerebral cortex and limbic system, and a comparison with findings obtained in the human and nonhuman primate brain. This chapter thus presents an overview, at the molecular, cellular and systems levels, of central dopaminergic circuits involved in state-setting modulatory systems, generation and integration of motor behavior, cognitive functions and reward mechanisms.

Previous studies demonstrated that the spiny rat Proechimys guyannensis exhibits resistance to experimental epilepsy. Neural activation was studied in the Proechimys hippocampus, using Fos induction, within 24 h after pilocarpine-induced seizures; neurodegenerative events were investigated in parallel, using FluoroJade B histochemistry. These parameters were selected since pilocarpine-induced limbic epilepsy is known to elicit immediate early gene expression and cell loss in the hippocampus of seizure-prone laboratory rodents. At variance with matched experiments in Wistar rats, pilocarpine injection resulted in Proechimys in seizure episodes that, as previously reported, did not develop into status epilepticus. At 3 h and 8 h after seizure onset, Fos immunoreactivity filled the dentate gyrus of both rat species, and was quite marked in pyramidal cells of the Proechimys Ammon's horn. At 24 h, Fos immunoreactivity dropped in the Wistar hippocampus and persisted in Proechimys. At 8 h and 24 h, FluoroJade-stained neurons were very few in the Proechimys hippocampus, whereas they were abundant in that of Wistar rats. Double immunohistochemistry for Fos and parvalbumin, the protein expressed by fast-spiking hippocampal interneurons, indicated that Fos was induced up to 24 h in the vast majority of parvalbumin-containing cells of the Proechimys hippocampus, and in a minority of these cells in the Wistar hippocampus. The findings demonstrate that early postepileptic neurodegeneration is very limited in the Proechimys hippocampus, in which sustained Fos induction persists for several hours. The findings also indicate that Fos induction and persistence may not correlate with seizure intensity and may not be associated with neuronal death. Finally, the data implicate differential mechanisms of interneuron activity in anti-convulsant and pro-convulsant phenomena.

An Italian collaborative group on motor neuron disorders, including amyotrophic lateral sclerosis (ALS) and its variants, has been recently created, combining various academic groups and laboratories involved in basic and clinical research. The aim is to exploit all the specific expertise and combine efforts at a national level to better understand and fight these fatal diseases. This review summarizes the achievements of the different groups and outlines prospects for future research. Basic research deals with the etiopathogenesis of motor neuron diseases. In vitro and in vivo models of superoxide dismutase 1 (SOD1) mutations are used to investigate the mechanisms of motor neuron death associated with this gene defect. The role of excitotoxicity, immune response, intracellular aggregates and mitochondrial alterations is studied with an integrated approach, at the molecular and cellular levels. Transgenic mice carrying the human mutated SOD1, and the wobbler mouse, a spontaneous model for motor neuron degeneration, offer unique opportunities for testing new therapies in vivo related or not to SOD1 mutations. Clinical research has focused mostly on the incidence and determinants of ALS in several areas of Italy. The incidence of the disease is now among the highest according to the results of population-based regional registries. Compared to earlier studies, more recent Italian investigations show an increase in the incidence and mortality related to ALS. Findings on the role of environmental risk factors are inconsistent. Methodological issues have also been raised by Italian groups regarding the diagnosis and treatment. The validity of the El Escorial diagnostic classification has been questioned where investigators and carers have not received formal training. Pitfalls and methodological drawbacks of randomized clinical trials have been highlighted based on the results of collaborative trials by Italian investigators. Information is now available on non-pharmacological treatments and palliative care, and the economic aspects and quality of life of ALS patients are being investigated.

The relationship between efferents of the hypothalamic suprachiasmatic nucleus (SCN) and neurons of the thalamic paraventricular nucleus (PVT) projecting to the amygdala was investigated in the rat using tract tracing in light and electron microscopy. Biotinylated dextran amine was used to label anterogradely SCN efferents. These fibers were found to reach the thalamic midline, terminating in PVT, through three pathways: anterodorsally through the preoptic region, dorsally through the periventricular hypothalamus, and through the contralateral medial hypothalamic and preoptic areas after crossing the midline in the optic chiasm. Preterminal and terminal-like elements labeled from the SCN were distributed throughout the rostrocaudal extent of PVT, with an anteroposterior gradient of density. Labeled terminal elements were densest in the dorsal portion of PVT beneath the ependymal lining and some of them entered the ependyma. Anterograde tracing of SCN fibers was combined with injections of retrograde tracers in the amygdala. Numerous retrogradely labeled cell bodies were seen throughout PVT, with a prevalence in its anterodorsal portion. Overlap was detected between puncta labeled from the SCN and retrogradely labeled neurons, especially in the anterodorsal sector of PVT, where numerous puncta were in close apposition to thalamo-amygdaloid cells. Electron microscopy revealed that boutons labeled from the SCN established synaptic contacts with dendritic profiles of PVT neurons labeled from the amygdala. The findings demonstrate that information processed in the biological clock is conveyed to the amygdala through PVT, indicating that this nucleus plays a role in the transfer of circadian timing information to the limbic system.

Expression of the calcium binding protein (CaBP) calretinin (CR) was studied with immunohistochemistry in the pineal complex and habenular nuclei (HN) of the developing and adult frog Rana esculenta. The frog pineal complex is a medial structure formed by two interconnected components, the frontal organ and the pineal organ or epiphysis; the habenular nuclei are bilateral and are asymmetric due to subdivision of the left dorsal nucleus into medial and lateral components. In the pineal complex, calretinin immunostaining of cells and fibers was consistently observed in developing and adult frogs. In the habenulae, calretinin immunoreactivity exhibited instead marked variations during development, and was expressed only in cells of the medial subnucleus of the left dorsal habenula. In particular, calretinin was detected at larval stages, peaked during metamorphosis, was markedly downregulated at the end of metamorphosis, and was evident again in adulthood. This sequence of calretinin expression was confirmed by quantitative analysis of immunoreactive cells in the left habenula. In tadpoles, calretinin-positive cells exhibited a dorsoventral gradient of density, while in adulthood, they were distributed throughout the dorsoventral extent of the medial subnucleus. The study demonstrates a peculiar developmental pattern, with transient downregulation, of asymmetric calretinin expression in the frog epithalamus. The findings indicate that calcium and calcium buffering systems may play critical roles in neurogenetic and neuronal migration processes implicated in the formation of the asymmetric habenular portion in amphibians. In addition, the reappearance of calretinin expression in the adult frog supports a distinct functional role of the asymmetric habenular component in amphibians.

Olfactory cortexes and the olfactory tracts are involved in sporadic Creutzfeldt-Jakob disease. We examined peripheral regions of the olfactory sensory pathway, including the olfactory mucosa, to assess whether pathologic infectious prion protein (PrPSc) is deposited in the epithelium lining the nasal cavity.We studied nine patients with neuropathologically confirmed sporadic Creutzfeldt-Jakob disease. We obtained the brain, the cribriform plate with the attached olfactory mucosa, and the surrounding respiratory epithelium at autopsy. Control samples of nasal mucosa were obtained post mortem or at biopsy from age-matched control subjects and from control patients with other neurodegenerative diseases. The olfactory and respiratory mucosa and the intracranial olfactory system were analyzed by light microscopy, immunohistochemistry, and Western blotting for pathological changes and for deposition of PrPSc.In all nine patients with sporadic Creutzfeldt-Jakob disease, PrPSc was found in the olfactory cilia and central olfactory pathway but not in the respiratory mucosa. No PrPSc was detected in any of the tissue samples from the 11 controls.Our pathological and biochemical studies show that PrPSc is deposited in the neuroepithelium of the olfactory mucosa in patients with sporadic Creutzfeldt-Jakob disease, indicating that olfactory biopsy may provide diagnostic information in living patients. The olfactory pathway may represent a route of infection and a means of spreading prions.

A brief survey of cortical development is presented, focusing on neuronal migration and its alterations. Corticogenesis is achieved through ordered temporospatial steps, via the formation of transient structures, and successive waves of cell proliferation and migration (followed by cell differentiation and maturation), and apoptotic cell death. The appearance of the proliferative ventricular zone and marginal zone, and of the superficial primordial plexiform layer, is followed by the formation of the prospective layer I, of the subplate, whose neurons are destined to die, and of the cortical plate that will give rise to layers II-VI. Cells arising in the ventricular zone migrate radially using radial glia as a scaffold, and are destined to form pyramidal cells. Cortical interneurons are mainly generated in the ganglionic eminence and migrate along axonal substrates following tangential routes. Disorders of this complex process lead to a wide range of alterations, and focal derangements of cortical organization have been grouped under the term focal cortical dysplasia (FCD). As the result of a neuropathological revision of FCD cases with intractable epilepsy, a novel classification comprising three subgroups of FCD has been introduced, and is supported by electroclinical and neuroimaging data, as well as by the postsurgical outcome of patients: i). architectural dysplasia, characterized by altered cortical lamination; ii). cytoarchitectural dysplasia, with the occurrence of giant neurons besides cortical dyslamination; iii). Taylor-type cortical dysplasia, in which altered cortical lamination is consistently associated with the occurrence of giant, dysmorphic and ectopic neurons, and frequently with the so-called balloon cells.