Anna Teresa Palamara

Nerve growth factor inhibits apoptosis in memory B lymphocytes via inactivation of p38 MAPK, prevention of Bcl-2 phosphorylation, and cytochrome c release

Survival of memory B lymphocytes is tightly linked to the integrity of the Bcl-2 protein and is regulated by a nerve growth factor (NGF) autocrine circuit. In factor-starved memory B cells, the addition of exogenous NGF promptly induced p38 mitogen-activated protein kinase (MAPK), but not c-Jun N-terminal kinase (JNK), dephosphorylation. Conversely, withdrawal of endogenous NGF was followed by p38 MAPK activation and translocation onto mitochondria, whereby it combined with and phosphorylated Bcl-2, as assessed by co-immunoprecipitation and kinase assays in vivo and in vitro. Mitochondria isolated from human memory B cells, then exposed to recombinant p38 MAPK, released cytochrome c, as did mitochondria from Bcl-2-negative MDCK cells loaded with recombinant Bcl-2. Apoptosis induced by NGF neutralization could be blocked by the specific p38 MAPK inhibitor SB203580 or by Bcl-2 mutations in Ser-87 or Thr-56. These data demonstrate that the molecular mechanisms underlying the survival factor function of NGF critically rely upon the continuous inactivation of p38 MAPK, a Bcl-2-modifying enzyme.

Low molecular weight, non-peptidic agonists of TrkA receptor with NGF-mimetic activity

Exploitation of the biologic activity of neurotrophins is desirable for medical purposes, but their protein nature intrinsically bears adverse pharmacokinetic properties. Here, we report synthesis and biologic characterization of a novel class of low molecular weight, non-peptidic compounds with NGF (nerve growth factor)-mimetic properties. MT2, a representative compound, bound to Trk (tropomyosin kinase receptor)A chain on NGF-sensitive cells, as well as in cell-free assays, at nanomolar concentrations and induced TrkA autophosphorylation and receptor-mediated internalization. MT2 binding involved at least two amino-acid residues within TrkA molecule. Like NGF, MT2 increased phosphorylation of extracellular signal-regulated kinase1/2 and Akt proteins and production of MKP-1 phosphatase (dual specificity phosphatase 1), modulated p38 mitogen-activated protein kinase activation, sustained survival of serum-starved PC12 or RDG cells, and promoted their differentiation. However, the intensity of such responses was heterogenous, as the ability of maintaining survival was equally possessed by NGF and MT2, whereas the induction of differentiation was expressed at definitely lower levels by the mimetic. Analysis of TrkA autophosphorylation patterns induced by MT2 revealed a strong tyrosine (Tyr)490 and a limited Tyr785 and Tyr674/675 activation, findings coherent with the observed functional divarication. Consistently, in an NGF-deprived rat hippocampal neuronal model of Alzheimer Disease, MT2 could correct the biochemical abnormalities and sustain cell survival. Thus, NGF mimetics may reveal interesting investigational tools in neurobiology, as well as promising drug candidates.

P10.8 Herpes simplex virus type 1 (HSV-1) increases the excitability of rat neocortical neurons and triggers amyloid precursor protein (APP) processing

caudate-putamen (CPU), subthalamic nucleus (STN) and susbstantia nigra pars reticulate (SNr) in 15 awake rats before and after the administration of apomorphine (5 mg/kg) and haloperidol (1 mg/kg). In basal condition, the phase of the delta band (2.5 Hz) entrained the amplitudes of the gamma (50 & 80 Hz) and high frequency oscillations (HFO, 150 Hz) bands. In the motor cortex, the high gamma (80 Hz) and the HFO bands were significantly modulated by the delta activity. In contrast, in the basal ganglia nuclei the most strongly modulated band was the low gamma band (50 Hz), while the high gamma showed a lesser modulation and the HFO band was only modulated in the CPU. Apomorphine injection induced a frequency increase in the modulating activity into the theta range (7.5 Hz) with the modulated amplitude mainly in the high gamma band for the four nuclei. On the other hand, the effect of haloperidol over the frequency of the modulating activity was the opposite, a reduction of the phase frequency (1.75 Hz), with a strengthening of the modulation in the high gamma band. Delta-theta to gamma-HFO cross-frequency coupling occurs in the rat motor circuit. The frequency and magnitude of these phase-to-amplitude interactions are modulated by dopamine agonist and antagonist drugs. This work was supported by the UTE proyect CIMA and by grants from the Fondo de Investigaciones Sanitarias (PI 07/0034) and the Ministerio de Ciencia e Innovación (BFU201