Cell Death In Astrocytes Research Articles

1311 Striatal astrocytic cell death following systemic administration of mitochondrial toxin, 3-nitropropionic acid

1311 Striatal astrocytic cell death following systemic administration of mitochondrial toxin, 3-nitropropionic acid

AMPA receptor activation is rapidly toxic to cortical astrocytes when desensitization is blocked

Although cultured astrocytes express functional glutamate receptors, they are generally resistant to excitotoxic cell death. We explored the role of receptor desensitization in glutamate-mediated astrocyte injury. In cultures of type 1 astrocytes from mouse neocortex, brief application of AMPA evoked small, rapidly desensitizing inward currents, whereas kainate evoked small, sustained currents. Neither agonist increased cytosolic calcium, and astrocyte toxicity occurred only after 24 hr exposure to high (500-1000 microM) concentrations of kainate but not to AMPA or glutamate. Cyclothiazide, a drug that selectively blocks AMPA receptor desensitization, greatly potentiated AMPA- or kainate-gated currents and intracellular calcium elevation. Coapplication of 10-100 microM cyclothiazide with glutamate, AMPA, or kainate produced widespread astrocyte cell death within 2 hr or application. The enhancement of toxicity by cyclothiazide, which alone was not toxic, was concentration-dependent for each of the tested agonists (EC50 30-100 microM) and was blocked by further addition of the selective AMPA/kainate antagonist 2,3-dioxo-6-nitro-7-sulfamoylbenzo(f)quinoxaline (NBQX). NMDA caused no injury even in the presence of cyclothiazide. Cyclothiazide-enhanced injury varied with the age of astrocyte cultures; the maximal effect occurred at approximately 2 weeks in vitro, and little death was seen after 4 weeks. Type 1 astrocytes express AMPA-type glutamate receptors that are unmasked by reducing their desensitization with cyclothiazide. Although overactivation of AMPA receptors can be rapidly lethal to astrocytes, rapid desensitization normally limits this toxicity. The extent of AMPA receptor desensitization may be an important determinant of glial vulnerability to excitotoxic insults.

Astrocyte survival in the absence of exogenous substrate: Comparison of immature and mature cells

Astrocyte cultures prepared from newborn mouse neopallium were grown for either one or three weeks (representing, respectively, immature and mature astrocytes) and then exposed to deprivation of substrate (glucose and amino acids) for up to 48 hr. Cultures which had been deprived of metabolic substrates for either 24, 30, 36 or 48 hr were examined for lactate dehydrogenase efflux into the medium (an indicator of cell death) and ATP content. Significant cell death in mature astrocytes began after 30 hr of incubation in the substrate-deprived medium, a time when ATP had fallen to ∼10% of its initial value. Immature astrocytes survived on a substrate-free medium for 48 hr before there was any indication at all of cell death, and this corresponded to a time when ATP values had fallen to 5% of the initial values. These findings are compared to previous observations during simulated ischemia (substrate deprivation plus anoxia) when (1) there was a faster cell death and (2) cell death occurred at higher ATP levels.

Ischemia-induced death of astrocytes and neurons in primary culture: Pitfalls in quantifying neuronal cell death

We have demonstrated that both astrocytes and cerebellar granule cell neurons die during an ischemic insult only when there is complete loss of mitochondrial membrane potential. This was determined by comparing the ability of mitochondria to sequester rhodamine 123 with the ability of cells to exclude propidium iodide. We have also demonstrated that in astrocyte cultures cellular LDH loss correlates directly with propidium iodide uptake, i.e. cell death, and inversely with rhodamine 123 uptake. Thus, both LDH loss and rhodamine 123 uptake can be used to quantitatively measure astrocyte cell death in culture; however, this is not the case with neuronal cultures. It was demonstrated that even in highly enriched cerebellar granule cell neuron cultures where astrocytes comprise less than 10% of the total culture protein, approximately 40% of total culture LDH was in the astrocytes. This was also the case with rhodamine 123 sequestration. Caution must therefore be used when using the LDH technique to determine the proportion of neurons that have died in such enriched neuronal cultures.