Quantification of secreted Paneth cell granules in response to butyric acid and leucine: (a) Enteroids at days 5 to 7 were injected with sodium butyrate (right bar, 17.3 ± 4.2) or PBS (left bar, 3.7 ± 3.8). Secreted granules were counted for 30 min after injection of reagents. Data are expressed as the total number of secreted granules from five Paneth cells ± SEM (n = 3). * p < 0.05 by Student’s t-test. (b) Secreted granules after the introduction of L-threonine (middle bar, 9.0 ± 9.6) and L-leucine (right bar, 34.0 ± 13.1) were compared to that after PBS (left bar, 2.7 ± 2.1). Data are expressed as the total number of secreted granules from five Paneth cells ± SEM (n = 3). * p < 0.05 by Tukey’s multiple comparisons test. (c) For the Gpr41 inhibition assay, sodium butyrate (50 mM) and beta-hydroxybutyrate (BHB) (1 mM) were co-injected. BHB significantly inhibited butyrate granules secretion. (d) For the Slc7a8 inhibition assay, L-leucine (1 mM leucine) and 2-amino-2-norbornane-carboxylic acid (BCH) (6 mM) were co-injected. BCH showed a tendency to inhibit granule secretion induced by leucine.