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. Author manuscript; available in PMC: 2019 Jun 14.
Published in final edited form as: Cell. 2018 May 24;173(7):1728–1741.e13. doi: 10.1016/j.cell.2018.04.027

Figure 7. The Ketogenic Diet and Bacterial Cross-Feeding Reduces Gamma Glutamyltranspeptidase (GGT) Activity, and GGT Inhibition Sufficiently Confers Seizure Protection.

Figure 7

(A) 6-Hz seizure thresholds in response to oral gavage with the GGT inhibitor, GGsTop, in SPF mice fed CD (left). n = 6, 9. Behavior in seizure-tested mice (right). Yellow line at y=10 seconds represents threshold for scoring seizures, and yellow triangle at 24 mA denotes starting current per experimental cohort. n = 16.

(B) 6-Hz seizure thresholds in response to supplementation with ketogenic amino acids in Abx-treated SPF mice enriched for A. muciniphila and Parabacteroides (left). n = 5, 6. Behavior in seizure-tested mice (right). n = 12.

(C) Total GGT activity per 100 mg feces from SPF CD, SPF KD, AkkPb KD, or AkkPb CD mice (left), and inhibition by GGsTop (right). n = 5.

(D) Total GGT activity per 100 mg feces from SPF CD animals treated with vehicle, A. muciniphila and Parabacteroides probiotic, or heat-killed bacteria for bi-daily for 28 days (left), and inhibition by GGsTop (right). n = 5.

(E) Levels of live A. muciniphila (Akk) after incubation in CD vs KD culture media or in CD or KD agar overlaid with M9 minimal media containing live P. merdae (PbM) or no bacteria (0). n = 3.

(F) Levels of live PbM after incubation in M9 minimal media overlaid on CD or KD agar containing Akk or no bacteria (0). n = 5.

(G) GGT activity in P. merdae grown in M9 media overlaid on CD agar containing A. muciniphila or no bacteria at t=24 hrs, and inhibition of GGT activity by GGsTop. n = 5.

(H) GGT activity in P. merdae grown in M9 media overlaid on KD agar containing A. muciniphila or no bacteria at t=24 hrs, and inhibition of GGT activity by GGsTop. n = 5.

Data are presented as mean ± SEM. Students t-test (A,B), Two-way ANOVA with Bonferroni (C, D), One-way ANOVA with Bonferroni (E-H): *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. SPF=specific pathogen-free, CD=control diet, KD=ketogenic diet, CC50=current intensity producing seizures in 50% of mice tested, AA=amino acids, veh=vehicle, Abx=pre-treated with antibiotics (ampicillin, vancomycin, neomycin, metronidazole), AkkPb=A. muciniphila, P. merdae and P. distasonis, GGsTop=GGT inhibitor, PbM=Parabacteroides merdae, Akk=Akkermansia muciniphila, M9=minimal media, GGT=gamma-glutamyltranspeptidase, AU=absorbance units. See also Figure S7.