Pier Luigi Canonico

Mitotic signaling by β-amyloid causes neuronal death

Aggregates of β‐amyloid peptide (βAP), the main constituent of amyloid plaques in Alzheimers brain, kill neurons by a not yet defined mechanism, leading to apoptotic death. Here, we report that both full‐length βAP(j_40) or (1–42) and its active fragment βAP(25_35) act as proliferative signals for differentiated cortical neurons, driving them into the cell cycle. The cycle followed some of the steps observed in proliferating cells, including induction of cyclin Di, phosphorylation of retinoblastoma, and induction of cyclin E and A, but did not progress beyond S phase. Inactivation of cyclin‐dependent protein kinase‐4 or ‐2 prevented both the entry into S phase and the development of apoptosis in βAP(25_35)‐treated neurons. We conclude that neurons must cross the G1/S transition before succumbing to βAP signaling, and therefore multiple steps within this pathway may be targets for neuroprotective agents.—Copani, A., Condorelli, F., Caruso, A., Vancheri, C., Sala, A., Giuffrida Stella, A. M., Canonico, P. L., Nicoletti, F., Sortino, M. A. Mitotic signaling by β ‐amyloid causes neuronal death. FASEB J. 13, 2225–2234 (1999)

Impaired Adult Neurogenesis Associated with Short-Term Memory Defects in NF-κB p50-Deficient Mice

Neurogenesis proceeds throughout adulthood in the brain of most mammalian species, but the molecular mechanisms underlying the regulation of stem/progenitor cell proliferation, survival, maturation, and differentiation have not been completely unraveled. We have studied hippocampal neurogenesis in NF-κB p50-deficient mice. Here we demonstrate that in absence of p50, the net rate of neural precursor proliferation does not change, but some of the steps leading to the final neuron differentiation status are hampered, resulting in ∼50% reduction in the number of newly born neurons in the adult mutant hippocampus. Additionally, in p50−/− mice, we observed a selective defect in short-term spatial memory performance without impairment of hippocampal-dependent spatial long-term memory and learning. Our results highlight the role of NF-κB p50 in hippocampal neurogenesis and in short-term spatial memory.

Comparative Effects of Estradiol Benzoate, the Antiestrogen Clomiphene Citrate, and the Progestin Medroxyprogesterone Acetate on Kainic Acid‐Induced Seizures in Male and Female Rats

Summary: We have investigated the comparative effects of estradiol benzoate (EB), the antiestrogenclomiphene citrate (CC), and the progesting medroxyprogesterone acetate (MPA) on seizures induced by systemic injection of kainic acid (15 mg/kg i.p.) in male and female rats. Subcutaneous administration for 10 days of EB (10 μg/kg) or high doses of CC (50 mg/kg) significantly potentiated kainate‐induced seizures, with the effect being more pronounced in male animals. Doses of 2.5 mg/kg of CC potentiated kainate‐induced seizures in male rats but were ineffective in female rats. Low doses of CC (0.5 mg/kg) exhibited a mild anticonvulsant effect in both sexes. repeated administration of MPA (2.5 mg/kg) partially protected female animals against kainate‐induced seizures; in male animls, MPA induced a 30% increase in the seizure severity score, although the difference fro the score of control male rats was not significant. These data suggest hat sex steroids influence kainate‐induced seizures in a sex‐dependent manner and that the effects of the antiestrogen CC are dose dependent. This should be taken into account in view of a possible use of CC and MPA in hormonal therapy for seizure disorders.

Characterization of ionotropic glutamate receptors in human lymphocytes.

The effect of L‐glutamate (Glu) on human lymphocyte function was studied by measuring anti‐CD3 monoclonal antibody (mAb) or phytohaemagglutinin (PHA)‐induced intracellular Ca2+ ([Ca2+]i) rise (Fura‐2 method), and cell proliferation (MTT assay). Glu (0.001 – 100 μM) did not modify basal lymphocyte [Ca2+]i, but significantly potentiated the effects of anti‐CD3 mAb or PHA. Maximal [Ca2+]i rises over resting cells were: 165±8 and 247±10 nM at 3.0×10−2 mg ml−1 anti‐CD3 mAb; 201±4 and 266±9 nM at 5.0×10−2 mg ml−1 PHA, in the absence or presence of 1 μM Glu, respectively. The Glu effect showed a bell‐shape concentration‐dependent relationship, with a maximum (+90±3% for anti‐CD3 mAb and +57±2% for PHA over Glu‐untreated cells) at 1 μM. Non‐NMDA receptor agonists (1 μM) showed a greater efficacy (+76±2% for (S)‐AMPA; +78±4% for KA), if compared to NMDA (+46±2%), or Glu itself. Ionotropic Glu receptor antagonists completely inhibited the effects of the corresponding specific receptor agonists (1 μM). The IC50 values calculated were: 0.9 μM for D‐AP5; 0.6 μM for (+)‐MK801; 0.3 μM for NBQX. Both NBQX and KYNA were able to abolish Glu effect. The IC50s calculated were: 3.4 μM for NBQX; 0.4 μM for KYNA. Glu (0.1 – 1 mM) did not change the resting cell proliferation, whereas Glu (1 mM) significant inhibited (−27±4%) PHA (1.0×10−2 mg ml−1)‐induced lymphocyte proliferation at 72 h. In conclusion, human lymphocytes express ionotropic Glu receptors functionally operating as modulators of cell activation.

Desensitization of metabotropic glutamate receptors in neuronal cultures

Abstract: Preexposure of cultured cerebellar neurons to glutamate reduced the stimulation of polyphosphoinositide (PPI) hydrolysis induced by subsequent addition of glutamate without affecting the response to the muscarinic receptor agonist carbamylcholine. Desensitization of glutamate‐stimulated PPI hydrolysis developed rapidly and persisted up to 48 h after removal of glutamate from the incubation medium. Stimulation of PPI hydrolysis by quisqualate was abolished in cultures pretreated with quisqualate or glutamate, but not with N‐methyl‐D‐aspartate (NMDA). In contrast, pretreatment with NMDA reduced the stimulation of PPI hydrolysis induced by a subsequent addition of NMDA, leaving the action of quisqualate intact. The lack of cross‐desensitization between NMDA and quisqualate supports the existence of two distinct subtypes of glutamate receptors coupled to PPI hydrolysis. Desensitization induced by a 30‐min (but not by a 6‐h) exposure to glutamate was attenuated or prevented by putative protein kinase C inhibitors, including mono‐ and trisialogangliosides, sphingosine, and polymyxin B, but not by inhibitors of arachidonic acid metabolism, nor by the nonselective calpain inhibitor leupeptin, nor by the lectin concanavalin A. These results suggest that desensitization of metabotropic glutamate receptors involves, at least in its rapid component, activation of protein kinase C.

Inhibitors of histone deacetylase (HDAC) restore the p53 pathway in neuroblastoma cells

Inhibitors of histone deacetylase (HDAC) are emerging as a promising class of anti‐cancer drugs, but a generic deregulation of transcription in neoplastic cells cannot fully explain their therapeutic effects. In this study we evaluated alternative molecular mechanisms by which HDAC inhibitors could affect neuroblastoma viability.

Nootropic Drugs Positively Modulate α-Amino-3-Hydroxy-5-Methyl-4-Isoxazolepropionic Acid-Sensitive Glutamate Receptors in Neuronal Cultures

Abstract: Micromolar concentrations of piracetam, aniracetam, and oxiracetam enhanced α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionic acid (AMPA)‐stimulated 45Ca2+ influx in primary cultures of cerebellar granule cells. Nootropic drugs increased the efficacy but not the potency of AMPA and their action persisted in the presence of the voltage‐sensitive calcium channel blocker nifedipine. Potentiation by oxiracetam was specific for AMPA receptor‐mediated signal transduction, as the drug changed neither the stimulation of 45Ca2+ influx by kainate or N‐methyl‐d‐aspartate nor the activation of inositol phospholipid hydrolysis elicited by quisqualate or (±)‐1‐aminocyclopentane‐trans‐1,3‐dicarboxylic acid. Piracetam, aniracetam, and oxiracetam increased the maximal density of the specific binding sites for [3H]AMPA in synaptic membranes from rat cerebral cortex. Taken collectively, these results support the view that nootropic drugs act as positive modulators of AMPA‐sensitive glutamate receptors in neurons.

Ubiquinone protects cultured neurons against spontaneous and excitotoxin-induced degeneration.

Ubiquinone is an endogenous quinone with pharmacological actions mainly related to its antioxidant properties. Here we report that ubiquinone protects cultured cerebellar granule cells against glutamate-induced neurotoxicity. In control cultures at 9 days of maturation in vitro (DIV), a 30-min exposure to 100 μM glutamate induced neuronal degeneration, as reflected by the great percentage (>90%) of cells labeled with propidium iodide 24 h after the exposure. Glutamate-induced neuronal death was dramatically reduced in cultures treated daily with ubiquinone since the second DIV. In these cultures, glutamate failed to induce a “delayed” increase in the influx of 45Ca2+, an established parameter of excitotoxicity. Similarly, repeated addition of ubiquinone attenuated in a concentration-dependent manner the age-dependent degeneration of granule cells that is due to the toxic action of the endogenous glutamate progressively released into the medium. These results suggest that ubiquinone may be a useful drug in the therapy of acute and chronic neurodegenerative diseases related to hyperactivity of excitatory amino acid neurotransmission.

Prolactin induces grooming in the rat: Possible involvement of nigrostriatal dopaminergic system

The possibility that Prolactin-induced grooming involves the nigrostriatal dopaminergic system was studied. Intracerebroventricular injection of rat Prolactin (PRL) in an amount of 10 micrograms induced grooming in male rats, and neostriatal injection of haloperidol (1 microgram/1 microliter) markedly suppressed this effect. Local administration of 6-OHDA in the substantia nigra also abolished the influence of intracerebroventricularly administered PRL. Bilateral injections of PRL (10 micrograms/l microliters) in the neostriatum failed to induce grooming, whereas bilateral injections of peptide into the substantia nigra (1 microgram/0.5 micrograms) elicited the behavioral response. It is probable that PRL induces grooming in the rat by interacting with the nigrostriatal dopaminergic transmission through an action on the cell bodies rather than in presynaptic terminals or at the postsynaptic level of this system.

Behavioral effects of phosphatidylserine in aged rats

The behavioral effects of 5 days of administration of phosphatidylserine (PS) was studied in aged rats. The intraperitoneal (5, 10, and 20 mg/kg) or intracerebroventricular (5, 10, and 20 micrograms/2 microliters) injection of PS liposomes caused a facilitated acquisition of active avoidance behavior as studied in shuttle-box and pole jumping test situations. The retention of active and passive avoidance responses was also improved. No substantial difference between PS-treated and control animals was apparent either in the responsiveness to electrical footshock or in the motor activity tested in an open field. Grooming behavior appeared to be enhanced in rats treated with the highest dose of the substance. Since PS affects both central catecholaminergic and cholinergic transmission, which is known to be impaired in old animals, the possibility that the behavioral effects of PS involve brain dopamine and/or acetylcholine systems is discussed.