Figure 4

Putative nitric oxide synthase immunoreactivity (NOS-IR, green) in the brain of G. insensibilis. (a) Entire brain showing the bilateral NOS-IR distribution. The signal is concentrated at the level of the cell clusters pmptc, alvc, ofc, vpolc, and vpmc (arrows, see Fig. 2 for abbreviations). (b) and (c) NOS IR in somata and neuropils. NOS-IR is localized in the somata of neuronal cell clusters [arrows in (b) and (c)]. NOS-IR is also found in neuropil regions as a punctuate signal [asterisks in (c)]. (d to f) Metacercariae in the protocerebrum of MAD gammarids surrounded by intense punctate NOS-IR. (g) and (h) One juvenile (asterisk) and two mature metacercariae in the protocerebrum of a MAD gammarid. The NOS label is shown in (g) whereas the propidium iodide signal is seen in (h); the white arrows indicate NOS-IR tissues present around a mature metacercaria but not around a young larva (asterisk). The black arrow points to the invagination of the cyst wall displayed in pictures (i) an (j). (i) and (j) Confocal sections at different levels of an invagination of the cyst wall in a metacercaria. The sections are tangential to the cyst; the arrows point to NOS-IR particles. In (b), (e), (g), (h), (i), (j), single confocal sections; in (a), (c), (d), (f), stacks of confocal sections. For clarity the propidium iodide counterstain has been omitted in (a) and (d). Anterior is up; cw, cyst wall; h, host; p, parasite. Scale bars: (a), (d), (g), (h), 300 μm; (e), 100 μm; (c), 30 m; (b), (f), (i), (j), 20 μm;.