Mitochondrial mass and membrane potential in coelomocytes from the earthworm Eisenia foetida: studies with fluorescent probes in single intact cells.
Earthworm coelomocytes exist in two forms, i.e., small (SC) and large (LC) cells, as demonstrated by velocity sedimentation, electron microscopy, and FCM. However, we know little concerning the functional activities of various, important organelles, such as mitochondria. In comparison with SC, LC from Eisenia foetida have a higher number of mitochondria, and, accordingly, showed a greater fluorescence intensity when mitochondrial mass was measured by nonyl acridine orange and FCM. To measure MMP we used both the lipophilic cationic probe JC-1 and Rh123. The intracellular localization of JC-1 in SC and LC was observed by fluorescence microscopy. Using JC-1, MMP was analyzed separately on SC and LC by FCM, and significant percentages of coelomocytes (> 95% of SC and about 90% of LC) displayed a high MMP. Adding 0.1 microM VAL caused most SC to depolarize, while this occurred in only a few LC. Rh123 gave different results: no effects of VAL were observed either in SC or in LC. In coelomocytes there may be several energy-independent Rh123-binding sites whose role must still be elucidated. On the whole, these data indicate that it is possible to analyze mitochondrial parameters by FCM in intact invertebrate coelomocytes, and that the type of cell and the probe used have a critical importance.
Cossarizza
A
A
Department of Biomedical Sciences, University of Modena, Italy.
Cooper
E L
EL
Quaglino
D
D
Salvioli
S
S
Kalachnikova
G
G
Franceschi
C
C
D000818:Animals; D002469:Cell Separation; D002470:Cell Survival; D005456:Fluorescent Dyes; D007107:Immune System / Q000166:cytology / Q000502:physiology* / Q000648:ultrastructure; D008564:Membrane Potentials / Q000187:drug effects; D008928:Mitochondria / Q000502:physiology* / Q000648:ultrastructure*; D009835:Oligochaeta; D013050:Spectrometry, Fluorescence; D014634:Valinomycin / Q000494:pharmacology
D005456:Fluorescent Dyes; D014634:Valinomycin