Figure 2

Enzymatic methylation of tryptamine does not differ between WT and INMT-KO rats (a) The gene structure of rat INMT with exons represented in black boxes. The ___location and sequence of the INMT guide RNA (gRNA) is shown with a red triangle. (b) The 2-base pair deletion responsible for generating the KO is shown below the wild type (WT) sequence with the ___location of the gRNA underlined. (c) Western blot showing the presence of INMT (green bands) in WT, but not KO rat lung tissues. The original image including all lanes in the molecular weight marker is included as Supplementary Figure S1. (d) Radiometric enzyme assays with tissue extracts from rabbit lung, as well as rat brain and rat lung from both WT and INMT-KO rats. Each point represents the average of 3 triplicate experiments from an individual animal for rabbits (n = 3), and rats (n = 6) for each genotype. Plots show averages with error bars indicating standard deviations. CPM: counts per minute. Rabbit lung tissue extracts (n = 3) showed tryptamine-dependent activity > 20-fold higher than rat tissues (average specific activity [SA] = 322.2 ± 8.81, 95% confidence interval [CI] of mean = 300.3–344.0). Tryptamine-dependent activity did not differ between WT and KO rats in brain (t = 0.30, mean difference = − 0.36, 95% CI = − 3.05–2.33, p = 0.77) or lung tissues (t = 0.52, mean difference = − 0.25, 95% CI = − 1.33–0.84, p = 0.62). In brain, average SA was 14.68 ± 2.34, 95% CI of mean = 12.22–17.13 in WT, and 14.32 ± 1.76, 95% CI of mean = 12.47–16.16 in KO. In lung, average SA was 6.36 ± 0.66, 95% CI of mean = 5.66–7.05 in WT, and 6.11 ± 0.96, 95% CI of mean = 5.10–7.12 in KO. Tryptamine-dependent activity was significantly higher in brain tissues relative to lung (t = 8.38, mean difference = 8.32, 95% CI = 5.87–10.77, p = 0.0002 for WT and t = 10.04, mean difference = 8.21, 95% CI = 6.31–10.11, p < 0.0001 for KO).