The glial‐derived neurotrophic protein S100β has been implicated in the development and maintenance of the nervous system. S100β has also been postulated to play a role in mechanisms of neuropathology because of its specific localization and selective overexpression in Alzheimer's disease. However, the exact relationship between S100β overexpression and neurodegeneration is unclear. Recent data have demonstrated that treatment of cultured rat astrocytes with high concentrations of S100β results in a potent activation of inducible nitric oxide synthase (iNOS) and a subsequent generation of nitric oxide (NO), which can lead to astrocytic cell death. To investigate whether S100β‐induced NO release from astrocytes might influence neurons, we studied S100β effects on neuroblastoma B104 cells or primary hippocampal neurons co‐cultured with astrocytes. We found that S100β treatment of astrocyte‐neuron co‐cultures resulted in neuronal cell death by both necrosis and apoptosis. Neuronal cell death induced by S100β required the presence of astrocytes and depended on activation of iNOS. Cell death correlated with the levels of NO and was blocked by a specific NOS inhibitor. Our data support the idea that overexpression of S100β may be an exacerbating factor in the neurodegeneration of Alzheimer's disease.

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The glia-derived, neurotrophic protein S100␤ has been implicated in development and maintenance of the nervous system. However, S100␤ has also been postulated to play a role in mechanisms of neuropathology, because of its specific localization and selective overexpression in Alzheimer's disease. To begin to address the question of whether S100␤ can induce potentially toxic signaling pathways, we examined the effects of the protein on nitric oxide synthase (NOS) activity in cultures of rat cortical astrocytes. S100␤ treatment of astrocytes induced a time-and dose-dependent increase in accumulation of the NO metabolite, nitrite, in the conditioned medium. The S100␤-stimulated nitrite production was blocked by cycloheximide and by the NOS inhibitor N-nitro-L-arginine methylester, but not by the inactive D-isomer of the inhibitor. Direct measurement of NOS enzymatic activity in cell extracts and analysis of NOS mRNA levels showed that the NOS activated by S100␤ addition is the calcium-independent, inducible isoform. Furthermore, the specificity of the effects of S100␤ on activation of NOS was demonstrated by the inability of S100␣ and calmodulin to induce an increase in nitrite levels. Our data indicate that S100␤ can induce a potent activation of inducible NOS in astrocytes, an observation that might have relevance to the role of S100␤ in neuropathology.The normal development and maintenance of the brain involves the temporal and spatial coordination and proper functioning of a number of intracellular and cell-cell signaling events, and the contribution of glial cells to these signaling processes is becoming more widely appreciated. The classical concept of the role of glia in brain function is rapidly changing with newer evidence of the crucial nature of these cells in controlling neurotransmitter levels, maintaining calcium homeostasis, and synthesizing and releasing neurotrophic and growth factors (for review, see Ref. 1). One such glia-derived factor is S100␤, a protein that promotes neuritic outgrowth of specific neuronal populations (e.g. cortical (2, 3), dorsal root ganglia (4), serotonergic (5, 6), and motoneurons (7)) and enhances survival of neurons during development (7,8) and after insult (9). S100␤ is also a glial mitogen, inducing phosphoinositide hydrolysis, increases in intracellular calcium, and protooncogene expression (10, 11). These trophic functions require nanomolar concentrations of a disulfide-linked S100␤ dimer (see Ref. 12). Thus, S100␤ may be beneficial during development of the nervous system, and increased S100␤ expression and secretion following acute glial activation in response to central nervous system injury may be one mechanism the brain uses in attempts to repair injured neurons.However, S100␤ may also reach concentrations that are deleterious, e.g. in neurodegenerative diseases like Alzheimer's disease and Down syndrome where chronic glial activation occurs (13). It has been found that S100␤ levels in severely affected brain regions of Alzheimer's disease patients are severalfol...

S100 beta is a calcium binding protein expressed primarily by astrocytes in the brain. In initiating studies of the toxic signalling pathways activated by high concentrations of S100 beta, we previously demonstrated that treatment of astrocytes with microM S100 beta results in a potent stimulation of the mRNA level and enzyme activity of inducible nitric oxide (NO) synthase, an enzyme previously implicated in glial pathology. We provide evidence here that NO formation stimulated by S100 beta can lead to cell death in astrocytes, with characteristics defined for apoptosis. Incubation of astrocytes with S100 beta for 48 h results in an increased percentage of astrocytes undergoing apoptotic cell death, as determined with the TUNEL technique, assays of DNA fragmentation and lactate dehydrogenase release. The cell death induced in responses to S100 beta addition correlates with the levels of NO formation, and an inhibitor of nitric oxide synthase attenuates the NO formation elicited by S100 beta, as well as the cell death. Therefore, we propose that S100 beta has the potential to be trophic or toxic. Although S100 beta may be involved in development, homeostasis and repair, chronic overexpression of the protein may mediate toxic responses or even cell death.

We have previously shown that ␤-amyloid (A␤) induces astrocyte activation in vitro and that this reaction is attenuated by the addition of exogenous apolipoprotein E (apoE)-containing particles. However, the effects of A␤ on endogenous apoE and apoJ levels and the potential role of apoE receptors in astrocyte activation have not been addressed. Three activating stimuli (lipopolysaccharide, dibutyryl cAMP, and aged A␤ 1-42) were used to induce activation of rat astrocyte cultures, as assessed by changes in morphology and an increase in interleukin-1␤. However, only A␤ also induced ϳ50% reduction in the amount of released apoE and apoJ and an 8-fold increase in the levels of cell-associated apoE and apoJ. Experiments using two concentrations of receptor-associated protein, an inhibitor of apoE receptors with a differential affinity for the low density lipoprotein receptor (LDLR) and the LDLR-related protein (LRP), suggest that LRP mediates A␤-induced astrocyte activation, whereas LDLR mediates the A␤-induced changes in apoE levels. Receptor-associated protein had no effect on apoJ levels or on activation by either dibutyryl cAMP or lipopolysaccharide. These data suggest that apoE receptors translate the presence of extracellular A␤ into cellular responses, both initiating and modulating the inflammatory response induced by A␤.