Nitric Oxide Production and Thermoregulation in Paramecium caudatum

We investigated nitric oxide (NO) production in Paramecium caudatum, and the role of NO production in population growth in culture and thermoregulation. Nitrite (NO2 -) concentration in media containing P. caudatum [6350 ± 390 (SD) paramecia/ml] was 2.37 ± 0.53 μM after 6 h, compared to 0.16 ± 0.06 μM in media alone (p<0.005), and the NO synthase (NOS) inhibitor, Nω-nitro-L-arginine methylester (L-NAME), reduced [NO2 -] to 0.81 ± 0.24 μM (p<0.02). Media containing P. caudatum produced [3H]L-citrulline from [3H]Larginine, and the [3H]L-citrulline production was inhibited by L-NAME. Addition of A23187, a calcium ionophore, to the media resulted in greater [NO2 -] (1.49 ± 0.28 μM with no A23187, 2.51 ± 0.23 μM with 0.1 μM A23187 added, p<0.05). Western blot analysis revealed a 155 kDa protein that reacted with mouse NOS1 antibody. Paramecia concentration increased from 51 ± 9 per ml on day 0 to 943 ± 53 per ml on day 7. L-NAME decreased paramecia concentration at day 7 (0.1 mM, 720 ± 70 per ml; 1.0 mM, 761 ± 49 per ml; and 10 mM, 132 ± 32 per ml; p<0.05 compared to control for all 3 concentrations). In a thermal gradient, P. caudatum selected an environmental temperature (Ts) of 32.9 ± 0.3°C, addition of 10 mM L-NAME reduced Ts to 24.3 ± 0.3°C (p<0.05). These data suggest that P. caudatum produce NO via a calcium dependent NOS similar to mammalian NOS1, and inhibition of NO production reduced paramecia number in culture and decreased Ts.