Silvia Piccirilli

Evidence that the HIV-1 coat protein gp120 causes neuronal apoptosis in the neocortex of rat via a mechanism involving CXCR4 chemokine receptor.

The HIV-1 coat protein, gp120 (100 ng given intracerebroventricularly (i.c.v.) daily for seven consecutive days) causes DNA fragmentation in the brain neocortex of rat. In neocortical cells bearing ultrastructural features typical of apoptosis, electron microscopy revealed specific immunopositivity for neurofilament cytoskeletal proteins, suggesting the neuronal nature of dying cells. Neuronal apoptosis by gp120 implicates CXCR4 chemokine receptors; in fact, in rats receiving a single daily, non-neurotoxic, dose of SDF-1alpha (0.25 pmoles given i.c.v. for 7 days before gp120), the natural ligand of CXCR4 receptor, apoptosis was significantly hindered. The mechanism of SDF-1alpha protection involves inhibition of gp120-enhanced expression of IL-1beta, a cytokine implicated in the mechanisms of apoptosis induced by the viral protein in the neocortex of rat.

Paraquat-and Rotenone-Induced Models of Parkinson's Disease

Parkinsons disease (PD) is a neurodegenerative disorder mainly characterized by a loss of dopaminergic (DA) neurons in the substantia nigra pars compacta. In recent years, several new genes and environmental factors have been implicated in PD, and their impact on DA neuronal cell death is slowly emerging. However, PD etiology remains unknown, whereas its pathogenesis begins to be clarified as a multifactorial cascade of deleterious factors. Recent epidemiological studies have linked exposure to environmental agents, including pesticides, with an increased risk of developing the disease. As a result, over the last two decades the ‘environmental hypothesis’ of PD has gained considerable interest. This speculates that agricultural chemicals in the environment, by producing selective dopaminergic cell death, can contribute to the development of the disease. However, a causal role for pesticides in the etiology of PD has yet to be definitively established. Importantly, most insights into PD pathogenesis came from investigations performed in experimental models of PD, especially those produced by neurotoxins. This review presents data obtained in our laboratories along with current views on the neurotoxic actions induced by the two most popular parkinsonian pesticide neurotoxins, namely paraquat and rotenone. Although confined to these two chemicals, mechanistic studies underlying dopaminergic cell death are of the utmost importance to identify new drug targets for the treatment of PD.

Systemic Administration of Nω-Nitro-l-Arginine Methyl Ester and Indomethacin Reduces the Elevation of Brain PGE2Content 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.

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

Reactive oxygen species (ROS) have been postulated to play a crucial role in the pathogenesis of ischaemia‐reperfusion injury. Among these, hydrogen peroxide (H2O2) is known to be a toxic compound responsible for free‐radical‐dependent neuronal damage. In recent years, however, the ‘bad reputation’ of H2O2 and other ROS molecules has changed. The aim of this study was to assess the protective role of H2O2 and modification in its endogenous production on the electrophysiological and morphological changes induced by oxygen/glucose deprivation (OGD) on CA1 hippocampal neurons.

Cellular localization of TRPC3 channel in rat brain: preferential distribution to oligodendrocytes

In the present work we describe the cellular localization of TRPC3 in non-excitable cells as compared to the neurons in normal rat brain. We performed a double labeling study for TRPC3 and one of the following cell-specific markers: mouse anti-glial fibrillary acidic protein (GFAP) for astrocytes; mouse anti-RIP for oligodendrocytes, or mouse anti-OX42 for microglia, or mouse anti-NeUN for neuronal nuclei or mouse anti-tyrosine hydroxylase (TH) for detection of dopaminergic neurons of the substantia nigra. Our double label immunofluorescence study showed that that TRPC3 is mainly localized in oligodendrocytes. These result were confirmed by the electron microscopy study, which showed TRPC3 immunoreactivity in oligodendrocytes. Consistent with the evidence that calcium homeostasis is important to oligodendrocytes for development, myelination, and demyelination [Microsc. Res. Tech. 52 (2001) 672], we can speculate that the distribution of TRPC3 in oligodendrocytes plays a role in myelination and or demyelination processes.

Inducible nitric oxide synthase is involved in the mechanisms of cocaine enhanced neuronal apoptosis induced by HIV-1 gp120 in theneocortex of rat

Cocaine, often abused by human immunodeficiency virus (HIV) infected patients, has been suggested to worsen the HIV associated dementia via unknown mechanisms. Here we report that subchronic treatment with a dose of cocaine (30 mg/kg i.p.), unable per se to cause neuronal death, increases the number of apoptotic cells typically observed in the neocortex of rats treated with HIV-1 gp120 (100 ng given i.c.v.). A pre-treatment with MK801 (0.3 mg/kg i.p.), a NMDA receptor antagonist, L-NAME (10 mg/kg i.p.) and 7-nitroindazole (50 mg/kg i.p.), two specific inhibitors of NOS, or with 1400 W (1 mg/kg s.c.), a selective inhibitor of inducible NOS (iNOS), minimized neurotoxicity by combined administration of cocaine and gp120 thus implicating iNOS. This conclusion is supported by the evidence that cocaine increases brain neocortical citrulline, the co-product of NO synthesis.

The Blockade of K+-ATP channels has neuroprotective effects in an in vitro model of brain ischemia

There is a common belief that the opening of K(+)-ATP channels during an ischemic episode has protective effects on neuronal functions by inducing a reduction in energy consumption. However, recent studies have also proposed that activation of these channels might have deleterious effects on cells survival possibly after a stroke or during long-lasting neurodegenerative processes. Considering these contrasting results, we have used a hippocampal in vitro slice preparation in order to investigate the possible effects of K(+)-ATP channel blockers on the electrophysiological and morphological changes induced by a transient episode of ischemia (oxygen and glucose deprivation) on CA1 pyramidal neurons. Therefore, we found that tolbutamide and glibenclamide, both nonselective K(+)-ATP channel blockers, produce neuroprotective effects against in vitro ischemia. Interestingly, the mitochondrial K(+)-ATP channel blocker 5-hydroxydecanoate and various K(+) channel blockers did not exert neuroprotection. Our results are consistent with the concept that a decreased activity of the plasmalemmal K(+)-ATP conductances may have a protective effect during episodes of transient cerebral ischemia.

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.

Multifaceted roles of nitric oxide in the lateral geniculate nucleus: from visual signal transduction to neuronal apoptosis

The lateral geniculate nucleus (LGN) is the thalamic relay of retinal inputs to the visual cortex. It contains a rich array of brain terminals, which modulate the visual signals to the cortex. Several data have documented that beside cholinergic, GABA-nergic, istaminergic, serotoninergic, and glutamatergic signals, the LGN contains also fibers and interneurons expressing the enzyme that produces nitric oxide (NO). Here, we review the documented physiological roles of NO in the transmission of visual inputs to the cortex and in the processes of activity-dependent refinement of LGN connections. Moreover we focus on the recently suggested role of NO in processes of neurotoxicity in the LGN. Particular relevance is given to studies documenting that, through an excitotoxic cascade, NO triggers apoptosis in the LGN of new-born rats deprived of vision in one eye. Data are also discussed on a possible role of NO in the mechanisms of LGN neuronal loss induced by glaucoma. We believe that a better understanding of the role of NO in the LGN may contribute to discover new experimental strategies for the treatment of degenerative ophthalmic diseases.

Enhanced anandamide degradation is associated with neuronal apoptosis induced by the HIV‐1 coat glycoprotein gp120 in the rat neocortex

Human immunodeficiency virus type‐1 coat glycoprotein gp120 causes delayed apoptosis in rat brain neocortex. Here, we investigated the possible role of the endocannabinoid system in this process. It is shown that gp120 causes a time‐dependent increase in the activity and immunoreactivity of the anandamide (AEA)‐hydrolyzing enzyme fatty acid amide hydrolase (FAAH), paralleled by increased activity of the AEA membrane transporter and decreased endogenous levels of AEA. The AEA‐synthesizing phospholipase D and the AEA‐binding receptors were not affected by gp120. None of the changes induced by gp120 in the cortex were induced by bovine serum albumin, nor were they observed in the hippocampus of the same animals. Also, the activity of 5‐lipoxygenase, which generates AEA derivatives able to inhibit FAAH, decreased down to approximately 25% of the control activity upon gp120 treatment, due to reduced protein level (∼45%). In addition, the FAAH inhibitor methyl‐arachidonoyl fluorophosphonate significantly reduced gp120‐induced apoptosis in rat brain neocortex, whereas selective blockers of AEA membrane transporter or of AEA‐binding receptors were ineffective. Taken together, these results suggest that gp120, by activating FAAH, decreases endogenous levels of AEA, and the latter effect seems instrumental in the execution of delayed neuronal apoptosis in the brain neocortex of rats.