Subthreshold depression in older subjects: an unmet therapeutic need.

Major depression, defined according to DSM IV TR criteria, is less common in older subjects, while other types of depression are two to three times more prevalent. This heterogeneous group of disturbances has received different names: depression not otherwise specified, minor depression, subthreshold or subsyndromal depression. Moreover, each condition has been defined using heterogeneous criteria by different authors. The term of subthreshold depression will be adopted in this position statement. Subthreshold depression has been associated with the same negative consequences of major depression, including reduced well being and quality of life, worsening health status, greater disability, increased morbidity and mortality. Nevertheless, there is a dearth of clinical trials in this area, and therefore older patients with subthreshold depression are either not treated or they are treated with the same non pharmacological and pharmacological therapies used for major depression, despite the lack of supporting scientific evidence. There is an urgent need to reach a consensus concerning the diagnostic criteria for subthreshold depression as well as to perform clinical trials to identify effective and safe therapies in this too long neglected patient group.

Chromaffin cells: the peripheral brain.

Chromaffin cells probably are the most intensively studied of the neural crest derivates. They are closely related to the nervous system, share with neurons some fundamental mechanisms and thus were the ideal model to study the basic mechanisms of neurobiology for many years. The lessons we have learned from chromaffin cell biology as a peripheral model for the brain and brain diseases pertain more than ever to the cutting edge research in neurobiology. Here, we highlight how studying this cell model can help unravel the basic mechanisms of cell renewal and regeneration both in the central nervous system (CNS) and neuroendocrine tissue and also can help in designing new strategies for regenerative therapies of the CNS.

Effects of TNF-α and IL-1 ß on the activation of genes related to inflammatory, immune responses and cell death in immortalized human HaCat keratinocytes.

The present experiments were designed to characterize by microarray analysis the transcriptional responses of human keratinocytes (HaCat) to TNF-α and IL-1 ß, given alone or in combination, in order to better understand the mechanisms underlying inflammatory, immune responses and cell death in which both cytokines play a pathophysiological role. Significant differences in the percentage and quality of genes dysregulated by TNF-α and IL-1 ß were shown. Both cytokines activated a series of genes involved in inflammatory, immune response as well as in cell death. In our experimental conditions, TNF-α, in contrast to IL-1 ß, did not induce a significant level of apoptosis in keratinocytes. However, given together both cytokines produced a significant decrease in apoptotic cells and synergistic transcriptional response which was due to the activation of several specific genes occurring after application of each cytokine. TNF-α and IL-1 ß evoked apoptotic effect and transcriptional responses were linked to the stimulation of their specific receptors since a pre-treatment with monoclonal antibodies vs TNF-α and/or IL-1 ß receptors was able to significantly reduce them.

Neuroprotective effect of hydrogen peroxide on an in vitro model of brain ischaemia.

BACKGROUND AND PURPOSE: Reactive oxygen species (ROS) have been postulated to play a crucial role in the pathogenesis of ischaemia-reperfusion injury. Among these, hydrogen peroxide (H(2)O(2)) is known to be a toxic compound responsible for free-radical-dependent neuronal damage. In recent years, however, the 'bad reputation' of H(2)O(2) and other ROS molecules has changed. The aim of this study was to assess the protective role of H(2)O(2) and modification in its endogenous production on the electrophysiological and morphological changes induced by oxygen/glucose deprivation (OGD) on CA1 hippocampal neurons. EXPERIMENTAL APPROACH: Neuroprotective effects of exogenous and endogenous H(2)O(2) were determined using extracellular electrophysiological recordings of field excitatory post synaptic potentials (fEPSPs) and morphological studies in a hippocampal slice preparation. In vitro OGD was delivered by switching to an artificial cerebrospinal fluid solution with no glucose and with oxygen replaced by nitrogen. KEY RESULTS: Neuroprotection against in vitro OGD was observed in slices treated with H(2)O(2) (3 mM). The rescuing action of H(2)O(2) was mediated by catalase as pre-treatment with the catalase inhibitor 3-amino-1,2,4-triazole blocked this effect. More interestingly, we showed that an increase of the endogenous levels of H(2)O(2), due to a combination of an inhibitor of the glutathione peroxidase enzyme and addition of Cu,Zn-superoxide dismutase in the tissue bath, prevented the OGD-induced irreversible depression of fEPSPs. CONCLUSIONS AND IMPLICATIONS: Taken together, our results suggest new possible strategies to lessen the damage produced by a transient brain ischaemia by increasing the endogenous tissue level of H(2)O(2).

Operative files of legal medical emergency in Calabria: study team.

In Calabria, a study team form legal medicine emergency (LME) formed to initiate a Crisis Unit (CU) able to manage and deal LME with complicated operations and roles, people and technical aspects involvement. First steps are planning, scene study and organization. Everything connecting the first and second emergency to have practicable application.

gp120 induces cell death in human neuroblastoma cells through the CXCR4 and CCR5 chemokine receptors.

To infect target cells, the human immunodeficiency virus (HIV) type I (HIV-1) must engage not only the well-known CD4 molecule, but it also requires one of several recently described coreceptors. In particular, the CXCR4 (LESTR/fusin) receptor allows fusion and entry of T-tropic strains of HIV, whereas CCR5 is the major coreceptor used by primary HIV-1 strains that infect macrophages and CD4(+) T-helper cells (M-tropic viruses). In addition, the alpha chemokine SDF1alpha and the beta chemokines MIP1alpha, MIP1beta, and RANTES, natural ligands of CXCR4 and CCR5, respectively, are potent soluble inhibitors of HIV infection by blocking the binding between the viral envelope glycoprotein gp120 and the coreceptors. Approximately two-thirds of individuals with acquired immunodeficiency syndrome (AIDS) show neurologic complications, which are referred to a syndrome called AIDS dementia complex or HIV-1-associated cognitive/motor complex. The HIV-1 coat glycoprotein gp120 has been proposed as the major etiologic agent for neuronal damage, mediating both direct and indirect effects on the CNS. Furthermore, recent findings showing the presence of chemokine receptors on the surface of different cell types resident in the CNS raise the possibility that the association of gp120 with these receptors may contribute to the pathogenesis of neurological dysfunction. Here, we address the possible role of alpha and beta chemokines in inhibiting gp120-mediated neurotoxicity using the human neuroblastoma CHP100 cell line as an experimental model. We have previously shown that, in CHP100 cells, picomolar concentrations of gp120 produce a significant increase in cell death, which seems to proceed through a Ca(2+) - and NMDA receptor-dependent cascade. In this study, we gained insight into the mechanism(s) of neurotoxicity elicited by the viral glycoprotein. We found that CHP100 cells constitutively express both CXCR4 and CCR5 receptors and that stimulation with phorbol 12-myristate 13-acetate down-regulates their expression, thus preventing gp120-induced cell death. Furthermore, all the natural ligands of these receptors exerted protective effects against gp120-mediated neuronal damage, although with different efficiencies. These findings, together with our previous reports, suggest that the neuronal injury observed in HIV-1 infection could be due to direct (or indirect) interactions between the viral protein gp120 and chemokine and/or NMDA receptors.

Cocaine induced T cell proliferation in the rat: role of amygdala dopamine D1 receptors.

The immunomodulatory effects of local administration of cocaine into the amygdala were studied in the rat. Intra-amygdala infusion of cocaine significantly and dose-dependently increased the proliferative response of splenocytes to concanavalin A (Con A). A similar effect on the immune response was also observed in rats, microinfused into the central amygdala with the selective D1 receptor agonist SKF 38393. The increase of the proliferative response of splenocytes to Con A was inhibited by coinfusion within the central amygdala of the dopamine D1 receptor antagonist SCH 23390, together with cocaine, but not by coinfusion of the dopamine D2 receptor antagonist eticlopride. These results suggest that cocaine may produce at least some of its effects on the immune system through the activation of brain dopamine neurotransmission and that the central amygdala may represent a critical structure mediating cocaine-induced T cell proliferation.

Paraquat: a useful tool for the in vivo study of mechanisms of neuronal cell death.

The present article reviews the results of experimental studies on paraquat neurotoxicity, started by our group several years ago--when clinical and experimental reports had increased the interest for the possibility that environmental chemicals, including paraquat, may be related to the development of Parkinson's disease-, and which are still continuing since paraquat appears to be a promising tool to study the mechanisms of neuronal cell death in vivo. Our observations have demonstrated that paraquat causes evident neurotoxic effects after intracerebroventricular or intracerebral injection in experimental animals; however, it seems that the herbicide does not exibit a selective neurotoxicity towards the dopaminergic nigro-striatal system since potent behavioural and electrocortical changes are induced by paraquat after injection in brain areas other than the substantia nigra and caudate nucleus. By studying the mechanisms through which paraquat induces neurotoxic effects in vivo, it was shown that either free radical production and activation of cholinergic and glutamatergic transmission may be regarded as related events which play a crucial role in paraquat-induced neurotoxicity. In addition, it was observed that in rats paraquat penetrates the blood-brain barrier following systemic administration to give rise to a differential brain regional distribution; the latter observation rises some concern over the hazard of paraquat as a potential environmental neurotoxin. Indeed, paraquat, administered systemically in rats produces behavioural excitation and brain damage. The brain damage appears to be selective for the pyriform cortex and this does not seem to be strictly related to the high concentrations reached by the herbicide in this area but to the higher vulnerability of this cortical area to the enhanced cholinergic transmission. The recent observation that paraquat, injected into the rat hippocampus, induces the expression of apoptotic neuronal cell death, appears of valuable interest also with a view to paraquat as an useful experimental model in the development of neuroprotective drugs able to block the molecular events which, once activated, are responsible for the induction of neuronal cell death.

The HIV envelope protein gp120 in the nervous system: interactions with nitric oxide, interleukin-1beta and nerve growth factor signalling, with pathological implications in vivo and in vitro.

The neuronal loss often described at post-mortem in the brain neocortex of patients suffering from AIDS has been proposed to be responsible for the development of the AIDS dementia complex. Neuroinvasive strains of the HIV virus infect macrophages, microglial cells, and multinucleated giant cells, but not neurones. Processing of the virus by cells of the myelomonocytic lineage yields viral products known to initiate a complex network of events that may lead to the death of neurones and to the development of AIDS-associated neurological syndrome. The HIV-1 coat protein gp120, in particular, has been proposed as a likely etiologic agent of the described neuronal loss because it causes the death of neurones in culture. More recently, it has been shown that brain cortical cell death caused in rats by intracerebroventricular injection of gp120 occurs via apoptosis. This observation broadens our knowledge of the pathophysiology of the reported neuronal cell loss and opens a new avenue of experimental research for the development of novel therapeutic strategies for the treatment of patients suffering from AIDS-associated neurological syndrome.

The effect of nitric oxide on cytokine-induced release of PGE2 by human cultured astroglial cells.

1. The role of the L-arginine-nitric oxide (NO) pathway on the formation of prostaglandin E2 (PGE2) by human cultured astroglial cells incubated with interleukin-1beta (IL-1beta) and tumour necrosis factor-alpha (TNF-alpha) was investigated. 2. Incubation of T 67 astroglial cell line with IL-beta (10 ng ml(-1)) and TNF-alpha (500 u ml(-1)) produced a significant (P<0.05) increase of both nitrite (the breakdown product of NO), cyclic GMP and PGE2 levels in cell supernatants. N omega-nitro-L-arginine methyl ester (L-NAME; 20-300 microM), an inhibitor of NO synthase (NOS), inhibited the increase of cyclic GMP and nitrite levels found in supernatants of cytokine-treated astroglial cells and reduced the release of PGE2. The latter effect showed that the enhanced arachidonic acid (AA) metabolism subsequent to stimulation of astroglial cells with IL-1beta and TNF-alpha was, at least in part, induced by NO. This occurred also when sodium nitroprusside (SNP; 120 microM), an NO donor, was incubated with astroglial cells, an effect antagonized by oxyhaemoglobin (OxyHb; 10 microM). 3. The inhibition elicited by L-NAME on PGE2-release by cytokine-treated astroglial cells was reversed by adding AA (40 microM), showing that the effect of NO on cytokine-dependent PGE2 release occurred at the cyclo-oxygenase (COX) level. Furthermore, the release of PGE2 in cytokine-treated astroglial cells was inhibited by indomethacin (10 microM), a COX inhibitor as well as by preincubating cells with dexamethasone (20 microM), an inhibitor of inducible enzymes, showing that the inducible isoform of COX (COX-2) was involved. 4. On the other hand, pretreating astroglial cells with methylene blue (MB; 10 microM), an inhibitor of NO biological activity acting at the guanylate cyclase level, failed to affect PGE2 release in cytokine-treated astroglial cells, leading to the conclusion that cyclic GMP changes related to NO formation are not involved in the generation of AA metabolites. 5. The present experiments demonstrated that the release of PGE2 by astroglial cells pretreated with IL-1beta and TNF-alpha is due to enhanced COX-2 activity via activation of the L-arginine-NO pathway, and this may be relevant to the understanding of the pathophysiological mechanisms underlying neuroimmune disorders.

Melatonin reduces paraquat-induced genotoxicity in mice.

The protection afforded by melatonin against paraquat-induced genotoxicity in both bone marrow and peripheral blood cells of mice was tested using micronuclei as an index of induced chromosomal damage. Melatonin (2 mg/kg) or an equal volume of saline was injected i.p. into mice 30 min prior to the i.p. administration of paraquat (two injections of 15 mg/kg; the paraquat injections were given with a 24 h interval) and thereafter at 6 h intervals to the conclusion of the study (72 h). Using fluorescence microscopy, the number of micronuclei in polychromatic erythrocytes (MN-PCE) per 2000 PCE (1000 PCE/slide) per mouse was counted both in blood and bone marrow, and the ratio of PCE to normochromatic erythrocytes (NCE) (PCE/NCE) was calculated. Paraquat treatment increased the number of MN-PCE at 24, 48, and 72 h, both in peripheral blood and bone marrow cells, while no differences were observed in the PCE/NCE ratio. Melatonin inhibited the paraquat-induced increase in MN-PCE by more than 50% at 48 and 72h. Paraquat toxicity is believed to be due to free radical generation. Since melatonin is known to be an efficient free radical scavenger, it is concluded that melatonin's protection against paraquat-induced genotoxicity is mediated, at least in part, by its free radical scavenging activity.

Cytotoxic effect of HIV-1 coat glycoprotein gp120 on human neuroblastoma CHP100 cells involves activation of the arachidonate cascade.

HIV type-1 (HIV-1) coat glycoprotein gp120 causes necrotic death in human neuroblastoma CHP100 cells. Here, we investigated the possible role of the arachidonate cascade and membrane peroxidation in gp120-induced cell necrosis. It is shown that gp120 increases the intracellular concentrations of prostaglandin E2 and leukotriene B4 by up-regulating the activity and expression of the arachidonate-metabolizing enzymes prostaglandin H synthase and 5-lipoxygenase respectively. Consistent with this observation, selective inhibitors of prostaglandin H synthase (i.e. indomethacin) and 5-lipoxygenase (i.e. MK886 and caffeic acid) protected CHP100 cells against gp120-induced necrosis. Treatment with gp120 also enhanced membrane lipid peroxidation and this may be implicated in the execution of cell damage. Interestingly, incubation with exogenous nitric oxide (NO) mimicked the effects of gp120 on necrotic death of CHP100 cells and activation of prostaglandin H synthase and 5-lipoxygenase. This suggests that NO might participate in the mechanism by which gp120 activates the arachidonate cascade.

Intrahippocampal injection of paraquat produces apoptotic cell death which is prevented by the lazaroid U74389G, in rats.

Injection of paraquat, a redox-cycling compound, into the rat hippocampus produces limbic seizures and hippocampal damage. Here we report that a proportion of the neuronal cell death caused by the herbicide occurs via an apoptotic mechanism which appears to be mediated by oxygen free radicals. Adult male Wistar rats (n=12) received a single dose of paraquat (25 nmol/0.5 microl; 0.5 microl/min rate) and were sacrificed 24 h later. Paraquat caused DNA fragmentation, nuclear chromatin marginalization and compaction in all hippocampal subsectors, 24 h after its injection, as revealed by both the TUNEL procedure and hematoxylin eosin staining of coronal brain tissue sections. Pre-treatment with the free radical scavenger lazaroid U74389G (30 mg/kg given i.p. 30 min beforehand) significantly reduced paraquat-induced apoptosis, but did not protect against non apoptotic neuronal cell loss caused by the herbicide.

Systemic administration of N omega-nitro-L-arginine methyl ester and indomethacin reduces the elevation of brain PGE2 content and prevents seizures and hippocampal damage evoked by LiCl and tacrine in rat.

Administration of tacrine (5 mg/kg i.p.), an anticholinesterase agent, in rats pretreated (24 h beforehand) with lithium chloride (LiCl; 12 mEq/kg i.p.) enhances the expression of neuronal nitric oxide (NO) synthase (NOS), increases NO, and causes seizures and hippocampal damage. Here we report immunohistochemistry evidence showing that in rat LiCl and tacrine enhance the expression of cyclooxygenase type 2 (COX-2) enzyme protein in the dorsal hippocampus and elevate brain PGE2 content during the preconvulsive period. The latter effect, but not enhanced COX-2 expression, is inhibited by previous (30 min before tacrine) administration of N omega-nitro-L-arginine-methyl ester (L-NAME; 10 mg/kg i.p.), an inhibitor of NO synthesis, thus implicating NO in the mechanism of stimulation of COX activity leading to elevation of brain PGE2 content. Indomethacin (10 mg/kg given i.p. 30 min before tacrine), an inhibitor of COX activity, prevented brain PGE2 elevation and abolished the expression of seizures and hippocampal damage thus supporting a role for this metabolite of the arachidonic acid cascade in the mechanisms of LiCl and tacrine-evoked neurotoxicity in rat.

Spontaneous induction of nitric oxide- and prostaglandin E2-release by hypoxic astroglial cells is modulated by interleukin 1 beta.

The effect of 0, 30, 60, 120, 240, 360 min hypoxia on the release of NO and PGE2 was investigated in human cultured astroglial cells. Exposure of astroglial cells to hypoxic injury produced a dose-dependent increase of the nitrite (the breakdown product of NO) level in the cell supernatant. In addition, a significant activation of the inducible isoform of NO synthase was seen, demonstrating that the enhancement on NO release produced by hypoxic injury was related to an increased biosynthesis of NO-generating enzyme(s). This effect was strongly antagonised by pretreating cells with dexamethasone (20 microM). The increase in NO release by hypoxic astroglial cells was accompanied by sustained release of PGE2, which was antagonised by the cyclooxygenase inhibitor indomethacin (10 microM), and partially attenuated by L-NAME (100 microM), a nitric oxide synthase inhibitor, showing that the release of PGE2 was driven by NO. Finally, inducible NOS activity elicited by hypoxic injury, was antagonised by incubating astroglial cells with antibodies directed against type 2 receptor for IL1 beta. In conclusion, hypoxia stimulates cytokine network in astroglial cells leading to enhanced release of NO and prostanoids and this may represent a key mechanism in cerebral blood flow disturbances.

Seizures and hippocampal damage produced by dendrotoxin-K in rats is prevented by the 21-aminosteroid U-74389G.

The epileptogenic and neurodegenerative effects of dendrotoxin K (DTx-K), from Dendroaspis polylepsis, a specific blocker of a noninactivating, voltage-sensitive K+ channel, were studied after focal injection into one dorsal hippocampus in rats pretreated with the 21-aminosteriod U-74389G, a scavenger of free oxygen radicals. Administration of 35 pmol DTx-K elicited in all of the treated animals (n = 6) motor seizures and bilateral electrocortical (ECoG) discharges after a latent period of approximately 5 min. At 24 h, histological examination of brain (n = 6) coronal sections (10 microns; n = 6 per brain) detected bilateral damage to the hippocampal formation. Quantitation of damage revealed significant bilateral neuronal cell loss in the CA1 and CA4 pyramidal cell layer and dentate gyrus granule cell layer relative to the corresponding brain regions of rats (n = 6) injected with bovine serum albumin (300 ng), which per se was ineffective in all respects. DTx-K (35 pmol) also caused a significant loss of CA3 pyramidal neurons ipsilateral to the site of toxin injection. Systemic (i.p.) administration of U-74389G (5 mg/kg given 30 min beforehand) delayed the onset of motor and ECoG seizures and reduced the number of epileptogenic discharges typically observed in rats receiving an injection of DTx-K (35 pmol) alone. Similarly, this treatment prevented the damage inflicted to the hippocampus by the toxin and in no instance was significant neuronal loss observed. At variance with these results, pretreatment with U-74389G (up to 10 mg/kg i.p.) failed to prevent seizures and CA1 hippocampal damage evoked by intra-hippocampal injection of alpha-DTx (35 pmol), a DTx-K homologue which preferentially inhibits a slowly inactivating, voltage-dependent K+ conductance in nerve cells. In conclusion, the present data support a role for free oxygen radicals in mediating hippocampal damage induced by DTx-K, but not alpha-DTx, and confirm the original deduction that these DTx homologues are complementary neurobiological tools to study mechanisms of seizures and neuronal death.

N-methyl-D-aspartate (NMDA) and non-NMDA glutamate receptor antagonists protect from apoptosis induced in the lateral geniculate nucleus of rabbits exposed to the dark.

Mono or binocular deprivation, during early postnatal development, produces dramatic effects on the anatomy and physiology of the visual system. Here we report that dark exposure induces apoptotic cell death in the lateral geniculate nucleus (LGN) of adult rabbit and this may be related to activation of N-methyl-D-aspartate (NMDA) and non-NMDA subtypes of glutamate receptors. In situ DNA fragmentation (TUNEL) was observed in the LGN of rabbits exposed to dark for 48 h. Morphological changes were confirmed on hematoxylin-eosin stained brain tissue coronal sections. Systemic treatment with CGP 040116 or MK 801, two NMDA receptor antagonists, and with GYKI 52466, a non-NMDA receptor antagonist, prevented in situ DNA fragmentation and nuclear chromatin marginalization and condensation. In no instance was apoptosis seen in rabbits kept under a normal light-dark cycle. Our findings indicate that glutamate, acting on NMDA and non-NMDA receptors, may be involved in the mechanisms of apoptotic cell death induced in the LGN of adult rabbits exposed to darkness.

Decrease of immunoreactive catalase protein in specific areas of ageing rat brain.

The activity of catalase, the main enzyme responsible for detoxification against hydrogen peroxide, decreases in specific brain areas of aged rats. The reduction of enzyme activity appears to be the consequence of a decreased protein expression rather than an impaired function of the native enzyme. In fact, diminution of the immunoreactive protein parallels enzyme activity decrease. Since the extent of decrease of both activity and protein content was observed to be area dependent, we hypothesise that this phenomenon may underlie, at least in part, the increased susceptibility of specific brain regions to oxidative insults observed in pathological situations related to ageing.