We have created early-onset transgenic (Tg) models by exploiting the synergistic effects of familial Alzheimer's disease mutations on amyloid ␤-peptide (A␤) biogenesis. TgCRND8 mice encode a double mutant form of amyloid precursor protein 695 (KM670/671NL؉V717F) under the control of the PrP gene promoter. Thioflavine S-positive A␤ amyloid deposits are present at 3 months, with dense-cored plaques and neuritic pathology evident from 5 months of age. TgCRND8 mice exhibit 3,200 -4,600 pmol of A␤42 per g brain at age 6 months, with an excess of A␤42 over A␤40. High level production of the pathogenic A␤42 form of A␤ peptide was associated with an early impairment in TgCRND8 mice in acquisition and learning reversal in the reference memory version of the Morris water maze, present by 3 months of age. Notably, learning impairment in young mice was offset by immunization against A␤42

Interleukin-10 (IL-10) is a powerful suppressor of cellular immune responses, with a postulated role in brain inflammation. First, we have evaluated the role of this cytokine in ischaemic brain damage using IL-10 knockout (IL-10-/-) mice. The middle cerebral artery (MCA) was occluded in either IL-10-/- or wild-type animals of corresponding strain (C57Bl/6) and age. Infarct volume was assessed 24 h later in serial brain sections. Brain infarct produced by MCA occlusion was 30% larger in the IL-10-/- than in wild-type mice (21. 8 +/- 1.2 vs. 16.9 +/- 1.0 mm3, respectively; P < 0.01; Student's t-test). To further characterize these findings, studies were extended to in vitro models. Primary neuronal cortical cultures derived from IL-10-/- animals were more susceptible to both excitotoxicity and combined oxygen-glucose deprivation compared with cell cultures from wild-type mice. Moreover, when added to the culture medium, recombinant murine IL-10 (0.1-100 ng/mL) exerted a concentration-dependent prevention of neuronal damage induced by excitotoxicity in both cortical and cerebellar granule cell cultures taken from either strain. The accordance of in vivo and in vitro data allows us to suggest a potential neuroprotective role of IL-10 against cerebral ischaemia when administered exogenously or made available from endogenous sources.

Blockade of adenosine receptors can reduce cerebral infarct size in the model of global ischaemia. Using the potent and selective A2A adenosine receptor antagonist, SCH 58261, we assessed whether A2A receptors are involved in the neuronal damage following focal cerebral ischaemia as induced by occluding the left middle cerebral artery. SCH 58261 (0.01 mg/kg either i.p. or i.v.) administered to normotensive rats 10 min after ischaemia markedly reduced cortical infarct volume as measured 24 h later (30% vs controls, p < 0.05). Similar effects were observed when SCH 58261 (0.01 mg/kg, i.p.) was administered to hypertensive rats (28% infarct volume reduction vs controls, p < 0.05). Neuroprotective properties of SCH 58261 administered after ischaemia indicate that blockade of A2A adenosine receptors is a potentially useful biological target for the reduction of brain injury.

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Recently, new technologies based on biosensors and called label free have been developed. These technologies eliminate the need for using markers and dyes. The authors applied one of these technologies, based on measurement of cell impedance variation, to study the pharmacological profiles of ligands for the cannabinoid receptor 2 (CB2), a Gi-coupled receptor, and for the metabopotropic glutamate receptor 1 (mGluR1), a Gq-coupled receptor. Reference agonists and antagonists/inverse agonists for the 2 receptors were applied to recombinant cell lines and impedance monitored over time. Agonists (JWH133 and CP55940 for CB2; quisqualate, glutamate, 1S-3R-ACPD, and S-3,5-DHPG for mGluR1) triggered a variation of impedance consistent in both potency and efficacy with data obtained using classical assays measuring cAMP or Ca2+ levels. This effect was not present in the parental nontransfected cell line, confirming specific receptor-mediated response. Application of antagonists (AM630 for CB2; YM298198, SCH1014222, J&J16259685, and CPCCOEt for mGluR1) reduced agonist-induced impedance changes. The only exception was the mGluR1 antagonist BAY367620 that, while active in the Ca2+ assay, was inactive in the impedance assay. Overall, these results confirm the possibility of using cell impedance–based technology to study the pharmacological profile of ligands acting at G-protein-coupled receptors coupled to different downstream signaling pathways.

A direct pathophysiological role of Familial Alzheimer's Disease (FAD)-associated Presenilin 1 (PS1) mutations in neuronal vulnerability remains a controversial matter. We evaluated the relationship between PS1 and excitotoxicity in four different experimental models of neurotoxicity by using primary neurons from (i) transgenic (tg) mice overexpressing a human FAD-linked PS1 variant (L286V mutation), (ii) tg mice overexpressing human wild-type (wt) PS1, (iii) PS1 knockout mice, and (iv) wt mice in which PS1 gene expression was knocked down by antisense treatment. We found that primary neurons overexpressing mutated PS1 showed an increased vulnerability to both excitotoxic and hypoxic-hypoglycemic damage when compared with neurons obtained from either mice overexpressing human wt PS1 or in wt mice. In addition, reduced excitotoxic damage was obtained in neurons in which PS1 expression was absent or diminished. Data obtained in in vivo experimental models of excitotoxicity partially supported the in vitro observations. Accelerated neuronal death was demonstrated in the hippocampus of mice overexpressing mutated PS1 after peripheral administration of kainic acid in comparison with wt animals. However, measurement of the infarct volume after middle cerebral artery occlusion did not show significant difference between the two animal groups. The results altogether suggest that expression of FAD-linked PS1 variants increases the vulnerability of neurons to a specific type of damage in which excitotoxicity plays a relevant role. In addition, they support the view that reduction of endogenous PS1 expression results in neuroprotection.