Dr. Diego Bohórquez and Dr. Zhen Yan present new technologies for obesity and metabolic research. Dr. Bohórquez discusses gut-brain interactions and the role of enteroendocrine cells, presenting case studies using optogenetics and automated home cage phenotyping. Dr. Yan introduces a novel voluntary weightlifting model for mice that does not require human handling. Long-term weightlifting training in mice leads to muscle hypertrophy, improved contractile function, and enhanced glucose metabolism and insulin sensitivity. The new technologies explore gut-brain connections and models of resistance exercise.

1. Obesity Series 2020
Diego Bohórquez, PhD
Assistant Professor of Medicine
Duke University School of Medicine
Durham, NC, USA
Cravings and Weightlifting Squats:
Technologies that Explore New
Metabolic and Behavioral Research
Zhen Yan, PhD
Professor of Medicine,
University of Virginia
Charlottesville, VA, USA

2. Dr. Zhen Yan and Dr. Diego Bohórquez present case
studies demonstrating the use of automated home
cage phenotyping for preclinical obesity research.
Welcome to Obesity 2020
Cravings and Weightlifting Squats:
Technologies that Explore New
Metabolic and Behavioral Research

3. The Bohórquez
Laboratory
Duke University
A gut brain atelier
A
gut
choice

4. No other competing interests
are declared.
Dr. Bohórquez is a member of the
scientific board of Holobiome Inc.

5. Miller and Kessen. J Comp Physiol Psychol. 1952; 45(6): 555–564

6. de Araujo. Neuron. 2008; 57: 930–941

7. Photo:©DiegoV.Bohórquez

8. enteroendocrine cell:
Specialized cells of the GI tract and pancreas with
endocrine function. They produce hormones or
peptides in response to various stimuli and
release them into the bloodstream for systemic
effect, diffuse them as local messengers,
or transmit them to the enteric nervous system to
activate nervous responses.
Wikipedia, 2020

9. neuropod cell:
A gut sensory epithelial cell that
synapses with nerves to
transduce stimuli in
milliseconds.
Kaelberer et al. Science. 2018

10. 10 µm
Bohórquez et al. JCI. 2015; 125 (2): 782

11. Bohórquez et al. JCI. 2015; 125 (2): 782

12. Bellono et al. Cell. 2017; 170 (1): 185–198

13. Bohórquez et al. JCI. 2015; 125 (2): 782
Rabies infects
a type of
epithelial cell

14. @gutbrains
∆G-rabies-GFP
Nucleus tractus Solitarius |
CCKCRE_rabGTvA
Kaelberer et al. and Bohórquez Science. 2018; 361 (6408): eaat5236

15. Kaelberer et al. and Bohórquez Science. 2018; 361 (6408): eaat5236

16. Han et al. Cell. 2018; 75 (3): 665-678
“…fast optogenetic activation of R-
Nodose Ganglia terminals [to NTS]
sustained self-stimulation behavior…”

17. Vagal cuff
Kaelberer et al. and Bohórquez Science. 2018; 361 (6408): eaat5236

18. sucrose
532nm laser
EECs are
necessary for
sugar sensing
Kaelberer et al. and Bohórquez Science. 2018; 361 (6408): eaat5236

19. sucrose
Sugar and
the vagus
+devazepide
Kaelberer et al. and Bohórquez Science. 2018; 361 (6408): eaat5236

20. 34,860 Images
4.3 nm/pxl
500 microns
In collaboration with David GC Hildebrand and Wei-Chung Lee | Harvard University

21. Glutamate
Vesicular glutamate expression
Kaelberer et al. and Bohórquez Science. 2018; 361 (6408): eaat5236

22. food 1 food 2
body
weight
activity
Automated
behavior
phenotyping
The Bohórquez laboratory

23. Preference for
caloric sugar
The Bohórquez laboratory

24. Preference for
caloric sugar
The Bohórquez laboratory

25. Gut optogenetics
The Bohórquez laboratory

26. The Bohórquez laboratory
Gut optogenetics
The Bohórquez laboratory

27. UNPUBLISHED DATA
The Bohórquez Laboratory. Unpublished.

28. UNPUBLISHED DATA
The Bohórquez Laboratory. Unpublished.
Wild Type

29. - The Bohórquez Laboratory
“Behind every bright brain
is a wise gut.”

30. May 11-13, 2021

31. Our Team

32. THANK YOU.

33. follow us
@gutbrains
The Bohórquez
Laboratory
Duke University
To treat the brain from the gut.

34. A novel voluntary weightlifting
model in mice for resistance
exercise research
Zhen Yan, PhD
Professor of Medicine: Cardiovascular Medicine
The Robert M. Berne Cardiovascular Research Center
University of Virginia
Obesity Series 2020
Copyright 2020 Z. Yan, APS and InsideScientific. All Rights Reserved.

35. Exercise promotes health and prevents diseases
Exercise capacity
Cachexia
Obesity & T2D

36. Petriz et al. 2016, J Cell Physiol
Aerobic (Endurance) Exercise vs. Strength
(Resistance) Training
Strength Training

37. Akimoto et al, AJP, 2005;
Choi et al. JAP, 2005
Voluntary wheel running is a physiological model
of endurance exercise

38. Discrepancy of publications with “voluntary wheel running”
vs. “ladder climbing” in rodents
0
50
100
150
Ladder Climbing
Voluntary Wheel Running
Year
NumberofPublications

39. Cholewa et al, 2014, J Cell Physiol
(A) Surgical ablation
(B) Tenotomy
(C) voluntary plantar extension
(D) 85° weighted ladder climb
(E) non-voluntary hind-limb
extension
(F) passive stretch
(G) 90° weighted ladder climb
(H) electric stimulated squat
Existing animal models of resistance exercise
(I) modified non-voluntary hind-
limb extension
(J) modified flywheel with hind-
limb suspension
(K) operantly conditioned squat
(L) modified operantly
conditioned squat
(M) jumping submersed in water
with overload

40. Tamaki et al, 2009, Hist & Cell Biol
50–100% hypertrophy 2 wks of SA in mice
8–12% hypertrophy 10 wks WL in humans
Hornberger et al,
2004, Can J Appl
Physiol Goodman et al, 2011, J Physiol; Lessard et al, 2018, Nat Commun
Invasive Labor intense Non-physiological
Blazevich et al, 2007, J Appl Physiol
Limitations of the existing animal models of weight
(resistance) training
A B

41. 11
LiftingActivity(pushes/min)
Time (h)
The prototype of voluntary weightlifting cage top
Unpublished Data

42. W
ater bottle
Weight
Adjustable
ramp
Lever
Magnet
Sensor
Food
container
Hinge
Nose to heal
length – 0.7 cm
Collar
Di et al. FASEB J, 2020
The design of the novel voluntary weightlifting cage top (1)

43. 3D printing of weightlifting cage top
Di et al. FASEB J, 2020

44. CON
Fn14
18S
WL
Dscr1
Nr4a3
Cytb
Tweak
D
scr1
Fn14N
r4a3Tw
eak
C
ytb
0
1
2
3
4
Foldchange
CON
WL
**
*
**
0 2 4 6 8 10 12
0
10
20
30
40
Activity(Lifts/5min)
Time (h)
Total: 419 ± 82
Q
C
TA
H
eart
Liver
0
1
2
3
Fn14(Foldchange)
**
**
Q
C
TA
H
eart
Liver
0
1
2
3
4
Dscr1(Foldchange)
**
Squat-like activity validated by transcriptional analysis
Di et al. FASEB J, 2020

45. Con
Row Z-Score
-1 0 1
CON1
WL1
CON2
CON3
WL2
WL3
341 up-regulated
149 down-regulated
Transcriptional response to acute weightlifting
Di et al. FASEB J, 2020

46. Acclimatization:
Day 1: Level up and ramp up (Up-up)
Day 2: Level down and ramp up (Up-down)
Day 3: Level down and ramp down (Down-down)
Staging:
5 days/week, starting at 100% body weight load
and 20% increment each training day until 240%
body weight load
Training:
5 days/week at 240% body weight load 0 20 40 60
0
500
1000
1500
0
100
200
300
Time (days)
Weightlifting(times)
Load(%bodyweight)
W
ater bottle
Weight
Adjustable
ramp
Lever
Magnet
Sensor
Food
container
Hinge
Weightlifting training protocol

47. Voluntary weightlifting training promotes
muscle hypertrophy
Con
WL
C
on
W
L
0
2
4
6
SUnSET-Puromycin(A.U.)
*
C
O
N
W
L
Puromycin
Gapdh
PonceauS
C
O
N
W
L
C
O
N
W
L
C
O
N
W
L
C
O
N
W
L
C
O
N
W
L
PonceauSPuromycin
QC PL Heart Liver
0
3
6
9
Musclemass(mg/mm)
CON
WL
***
0
500
1000
1500
CSA(mm2/gBW)
**
0
1
2
Puromycin(Foldchange)
CON
WL
*
**
**
0 20 40 60
0
500
1000
1500
0
100
200
300
Time (days)
Weightlifting(times)
Load(%bodyweight)
0
2
4
6
Puromycin(Foldchange)
**
Weightlifting(times)
Di et al. FASEB J, 2020

48. Di et al. FASEB J, 2020
0
2
4
6
10
12
14
Foldchange
CON
WL
*
***
***p=0.05
***
50 kDa
Raptor
p-p70S6K(T389)
p70S6K
p-4e-bp1(S65)
4e-bp1
Gapdh
150 kDa
50 kDa
50 kDa
20 kDa
20 kDa
37 kDa
CON WL
50 kDa p-Akt(S473)
Akt50 kDa
Raptor
p-p70S6K(T389)
p70S6K
p-4e-bp1(S65)
4e-bp1
Gapdh
150 kDa
50 kDa
50 kDa
20 kDa
20 kDa
37 kDa
CON WL
50 kDa p-Akt(S473)
Akt
Voluntary weightlifting training promotes signaling
pathways of protein synthesis
Perez-Schindler et al. 2013, PNAS

49. Electrodes
Footplate &
Force Transducer
C
O
N
W
L
0
500
1000
1500
Runingdistance(m)
C
O
N
W
L
0
4
8
12
Bloodlactate(mmol/L)
***
Pre
Post
***
0 200 400 600
0
10
20
30
40
50
Torque(mN*m)
CON
WL
Time (ms)
C
O
N
W
L
0.00
0.05
0.10
0.15
0.20
0.25
Tetanictorque(mN*m/mg)
**
C
O
N
W
L
0
400
800
1200
Tetanicmaxdx/dt
***
10
20
30
40
60
80100125150200
0.0
0.5
1.0
1.5
Torque(mN*m/g)
Con
WL
Frequency (Hz)
**
***
20
40
60
80
100
120
0.0
0.2
0.4
0.6
0.8
Torque(mN*m/g)
Con
WL
Time (sec)
Gapdh
CON WL
CV
15 kDa
37 kDa
50 kDa
37 kDa
25 kDa
15 kDa
20 kDa
200 kDa
200 kDa
200 kDa
Cox4
CIII
CIV
CII
CI
Myh2 (IIa)
Myh4 (IIb)
Myh7 (I)
Di et al.
FASEB J, 2020
Voluntary weightlifting training promotes contractile
function, but not endurance capacity

50. p-Akt (S473)
Akt
Gapdh
50 kDa
35 kDa
CON WL CON WL
50 kDa
Pre insulin Post insulin
Pre
Post
0
5
10
15
20
p-Akt/Akt(Foldchange)
CON
WL
*
**
**
**
C
O
N
W
L
0
5000
10000
15000
20000
25000
AUC(mg*min/dL)
***
0 1 2 3 4 5 6 7 8
20
25
30
35
40
Bodyweight(g)
Time (weeks)
*
CON
WL
**
*
*
**
** ***
Fatm
ass
Lean
body
m
ass
Free
w
ater
Totalw
ater
0.00
0.02
0.04
20
40
70
100
Bodycomposition(%)
CON
WL
C
O
N
W
L
0
10
20
30
Epididymalfat(mg/mm)
*
Voluntary weightlifting enhances insulin sensitivity
Di et al. FASEB J, 2020

51. CON WL
Glut4
p62
LC3-I
LC3-II
Gapdh
15 kDa
37 kDa
75 kDa
50 kDa
37 kDa
10 kDa
Atg775 kDa
Atg650 kDa
G
lut4
p62
LC
3LC
3-II/I
Atg7
Atg6
0.0
0.5
1.0
1.5
2.0
Foldchange
CON
WL
*
*
**
Voluntary weightlifting promotes autophagy
Di et al. FASEB J, 2020

52. • There is an urgent need of developing a physiological resistance exercise model
for large-scale animal studies.
• We developed a novel voluntary weightlifting cage top for mice that does not
require human handling during the experiment.
• An acute bout of weightlifting activities elicit robust transcriptional responses
only in recruited skeletal muscles.
• Long-term weightlifting training (8 wks) promotes skeletal muscle hypertrophy,
contractile function along with enhanced expression of the mTOR pathway and
protein synthesis.
• Long-term weightlifting training (8 wks) promotes whole-body glucose
metabolism with enhanced skeletal muscle insulin sensitivity and autophagy.
Take-home message

53. Z line
Mito
Sarcomere
Endurance and resistance exercise models in research

54. Di Cui (Hunan Univ.)
The Yan Lab
Laboratory of Exercise Physiology
Becky Wilson (Duke)

55. Acknowledgments
Yan Lab
Di Cui
Josh Drake
Becky Wilson
RJ Shute
Beven Lewellen
Mei Zhang
Henan Zhao
UVA Collaborators
Silvia Sabik
Suna Onengut
Stwart Berr
Steve Rich
Charles Farber
R01 AR050429

56. Thank you for
participating!
CLICK HERE to learn
more and watch the
webinar
Diego Bohórquez, PhD
Assistant Professor of Medicine
Duke University School of Medicine
Durham, NC, USA
Zhen Yan, PhD
Professor of Medicine,
University of Virginia
Charlottesville, VA, USA