Impact of gut microbiome on sleep and glymphatic system functioning

1. Induction of intestinal dysbiosis through broad-spectrum antibiotic gavage, high-fat feeding: effects
on the microbiota-gut-brain axis and sleep behavior in mice
Jonathan Lendrum1
, Bradley Seebach1
, Barrett Klein1
, Andrew Berns2
, Sumei Liu1
Department of Biology1
, Department of Computer Science2
, University of Wisconsin La Crosse, 1725
State St La Crosse, WI 54601
Objective: The glymphatic system, a perivascular circulatory system active during sleep phases and the
recently uncovered meningeal lymphatic vasculature have brought into question the long-standing
notion of immune privilege of the central nervous system (CNS). Furthermore, within the last decade
there has been a growing appreciation for the importance of coevolved, host-microbial interactions in
shaping brain development and behavior. The aim of this study is to investigate relationships between
altered compositions of intestinal microbiota and impaired sleep function in mice by assessment of
brain-gut axis pathways.
Methods: We individually housed three groups of five C57BL/6 mice in cages custom-fit with an infrared
security camera system used to observe and record sleep behavior. Five of the mice were gavaged with
a broad-spectrum antibiotic cocktail consisting of ampicillin, neomycin, metronidazole, and vancomycin
to perturb intestinal microbiota and induce dysbiosis. To a second group of mice we used high fat (60%
kcal) diet feeding, which has been shown to significantly alter microbial compositions in the gut. The
final five mice served as the control and received the same low fat (10% kcal) diet as the antibiotic-
treated group. We used Ussing chamber techniques to assess mucosal barrier function, whole blood
parameters were measured using a HemaVet FS Automated Hematology Analyzer, and energy harvest
was determined using bomb calorimetry of fecal samples (IKA C 200 Calorimeter).
Interpretation: We found that antibiotics and high fat diet caused a dramatic increase in intestinal
permeability. Spleen size and white blood cell count was significantly reduced in antibiotic treated mice,
resembling germ free phenotypes. Additionally, antibiotics treatment decreased the capacity to harvest
energy from the diet. The preliminary results of this study strongly suggest that different altered
compositions of gut microbial populations impairs mucosal barrier function similarly, the consequence
of which may lead to glymphatic system disruption and subsequent sleep dysfunction. Our results
indicate that the microbiome of antibiotic treated mice has a decreased capacity to harvest energy from
the diet. A detailed post analysis of the mouse fecal content and recorded video data through next
generation sequencing and high-throughput video tracking software that applies the 40 Second Rule (an
algorithm based on extended body immobility used to accurately predict sleep behavior in mice) will
hopefully give us insight into how and why microbial populations may regulate sleep phases, and why
reduction in the diversity of bacteria in the gut (as a result of poor nutrition, or antibiotic exposure) may
be associated with the pathophysiology of neurodegenerative and metabolic diseases commonly seen
with congruent brain and gut dysfunction. The results of which will be included in the presentation upon
completion.
Keywords: Antibiotics, High Fat Diet, Sleep Behavior, Obesity, Dysbiosis, Intestinal Permeability, Gut-
Brain Axis, Immune System
Acknowledgement: NIH R15 DK097460-01A1 (SL) and UW-L undergraduate research grant (JL).