Studies in non-neural cells have suggested that a fall in mitochondrial membrane potential (DeltaPsiM) is one of the earliest events in apoptosis. It is not known whether neural apoptosis caused by nerve growth factor (NGF) and serum withdrawal involves a decrease in DeltaPsiM. We used epifluorescence and laser confocal microscopy with the mitochondrial potentiometric dyes chloromethyl-tetramethylrosamine methyl ester and 5,5',6, 6'-tetrachloro-1,1',3,3'-tetraethybenzimidazol carbocyanine iodide to estimate DeltaPsiM. PC12 cells were differentiated in media containing serum and NGF for 6 d before withdrawal of trophic support. After washing, the cells were incubated with media containing serum and NGF (M/S+N), media without serum and NGF, or media with the "trophic-like" monoamine oxidase B inhibitor, (-)-deprenyl. Mitochondria in cells without trophic support underwent a progressive shift to lower DeltaPsiM values that was significant by 3 hr after washing. The percentages of cells with nuclear chromatin condensation or nuclear DNA fragmentation were not significantly increased above those for cells in M/S+N until 6 hr after washing. Replacement of cells into M/S+N or treatment with (-)-deprenyl markedly reduced the proportion of mitochondria with decreased DeltaPsiM. Measurements of cytoplasmic peroxyl radical levels with 2',7'-dihydrodichlorofluorescein fluorescence and intramitochondrial Ca2+ with dihydro-rhodamine-2-acetylmethyl ester indicated that cytoplasmic peroxyl radical levels were not increased until after 6 hr, whereas increases in intramitochondrial Ca2+ paralleled the decreases in DeltaPsiM. (-)-Deprenyl appeared to alter the relationship between intramitochondrial Ca2+ levels and DeltaPsiM, possibly through its reported capacity to increase the synthesis of proteins such as BCL-2.

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SUMMARY BackgroundAssessment of symptoms should be the primary outcome measure in dyspepsia clinical trials. This requires a reliable, valid and responsive questionnaire that measures the frequency and severity of dyspepsia. The Leeds Dyspepsia Questionnaire fulfils these characteristics, but is long and was not designed for self-completion, so a shorter questionnaire was developed (the Short-Form Leeds Dyspepsia Questionnaire).

Using S. pombe as a model system in which to assay CED-4 function, we have identified a potential direct role for CED-4 in chromatin condensation. Chromatin condensation is a ubiquitous feature of metazoan apoptosis that has yet to be linked to an effector. The CED-9-mediated rescue of CED-4-induced lethality in this system and the interaction of the two proteins in the yeast two-hybrid analysis suggest that CED-9 inhibits CED-4 action by direct physical association.