The foundation of our understanding of metabolic physiology is built on discoveries in fundamental, but isolated model systems.

Results from genes to organelles and cells may belie the physiognome.

A mechanism is only as important as its functional impact in the whole organism.

Well-Controlled Animal Models Bridge Cell Biology to

the Physiology of Exercise

The Study of Glucoregulation

Provocative or Sensitizing Tests

• Physical Exercise

• Hormone and Metabolic Challenges (e.g. hyperinsulinemic, euglycemic glucose clamps)

• Etcetera

Four Grams of GlucoseFour Grams of Glucose

Maintaining 4 Grams of Glucose in the Blood

Sedentary, Postabsorptive

Brain

Muscle

Liver

Fat

Liver

Blood

Glucose~4 grams

Maintaining 4 Grams of Glucose in the Blood

Feeding

Brain

Muscle

Liver

Fat

Liver

Blood

Glucose~4 grams

GI Tract

Stimulus(Insulin)

Suppression(Insulin)

Maintaining 4 Grams of Glucose in the Blood

Exercise

Brain

Muscle

Stimulus

Liver

Fat

Liver

Blood

Glucose~4 grams

Stimulus

Why don’t we get hypoglycemic when we exercise?

Exercise

Time (minutes)

-30

6

06

0100

0

GlucoseUtilization

Hepatic GlucoseProduction

Arterial PlasmaGlucose

mg·dl-1

mg ･ kg-1 ･min-1

mg ･ kg-1 ･min-1

If the liver does not release more glucose

during exercise

. . . Hypoglycemiarapidly ensues

0 60

Five Guiding Principles to Study of Metabolism in vivo

• Glucose metabolism is all about flux control.

• Glucose flux control is distributed amongst distinct systems that require an in vivo model to be fully understood.

• Glucose fluxes are most sensitively regulated and therefore best studied in the conscious state.

• Novel animal models can be used to bridge basic and clinical research.

• Provocative tests are often necessary to precipitate phenotypes and reveal functional limitations.

Endocrine and Sympathetic Nerve Response to Exercise

ArterialGlucagon

pg·ml-1

ArterialCatecholamines

pg·ml-1

Arterial Insulin

µU·ml-1

0

40

80

120

0

100

200

300

-60 -30 0 30 60 90 120 150

Exercise

Time (min)

Glucagon

Insulin

Norepinephrine

Epinephrine

0

8

12

16

A Minimal Overview of the Circulation

trunkand

lowerextremities

venous

arterial

portal vein

pancreasgutliver

Investigator sees…

Liver sees…

headand

upper extremities

heartandlungs

Chronically-Catheterized Conscious Dog Model

-50 0 50 100 150

PlasmaEpinephrine

(pg·ml-1)

Time (min)

ExerciseBasal

0

100

200

300

Arterial

Portal Vein

Hepatic Vein

-50 0 50 100 1500

100

200

300

Portal Vein

Hepatic Vein

Artery

Time (min)

ExerciseBasal

PlasmaGlucagon(pg·ml-1)

trunkand

lowerextremities

venous

arterial

portal vein

pancreasgutliver

Investigator sees…

Liver sees…

headand

upper extremities

heartandlungs

Protocols: Role of Glucagon

Equilibration Basal Moderate Treadmill Exercise

Basal Intraportal InsulinExercise-Simulated Intraportal Insulin

Saline Variable Glucose

Basal Intraportal Glucagon

Basal Intraportal GlucagonExercise-Simulated Intraportal Glucagon

-120 min -40 0 150

Protocol A

Protocol B

Somatostatin + [3-3H]glucose + [U-14C]alanine

0

50

100

150

Arterial Glucagon

pg/ml

Simulated Glucagon

Basal Glucagon

0

5

10

15

-60 -30 0 30 60 90 120 150

Time (min)

Arterial Insulin

µU/ml

Exercise

Exercise as a model to study glucagon action

Basal Glucagon

Simulated Glucagon

-40 0 30 60 90 120 1500

2

4

6

8

10

40

80

120

Arterial Plasma Glucose

mg·dl-1

Exercise

Basal Glucagon

SimulatedGlucagon

Basal Glucagon

Simulated Glucagon

Hepatic Glucose

Production

mg·kg-1·min-1

0

Exercise-induced Increment in Glucagon Stimulates Hepatic Glucose Production

Simulated Glucagon

Basal Glucagon

Exercise-induced Increment in Glucagon Stimulates Gluconeogenesis from Alanine

0

100

200

300

400

0

100

200

300

400

Gluconeogenesisfrom Alanine

(% Basal)

IntrahepaticGluconeogenicEfficiency

from Alanine

(% Basal)

Simulated Glucagon

Basal Glucagon

-60 -30 0 30 60 90 120 150

Time (min)

Exercise

0

1

2

3

4

5

6

Rest

Increase in Endogenous

GlucoseProduction

(mg·kg-1·min-1)

Exercise

Comparison of the Effects of Similar

Increases in Glucagon at Rest

and during Exercise

Why is Glucagon so Effective during Exercise?

Liver

Insulin

Glucagon

Adipose

WorkingMuscleWorkingMuscle

GlycerolNEFA

LactateAmino Acids

GlyGly

GNGGNG

Autonomic Nerve Activity

Epi

?

AminoAcids

Substrates

Signals

PancreasPancreas

IntestineIntestine

AdrenalAdrenal

Glucose4 gramsGlucose4 grams

BrainBrain

IL6

RBP4

Why is Glucagon so Effective during Exercise?

Liver

Insulin

Glucagon

Adipose

WorkingMuscleWorkingMuscle

GlycerolNEFA

LactateAmino Acids

GlyGly

GNGGNG

Autonomic Nerve Activity

Epi

?

AminoAcids

Substrates

Signals

PancreasPancreas

IntestineIntestine

AdrenalAdrenal

Glucose4 gramsGlucose4 grams

BrainBrain

IL6

RBP4

Body is in a ‘Gluconeogenic Mode’

Why is Glucagon so Effective during Exercise?

Liver

Insulin

Glucagon

Adipose

WorkingMuscleWorkingMuscle

GlycerolNEFA

LactateAmino Acids

GlyGly

GNGGNG

Autonomic Nerve Activity

Epi

?

AminoAcids

Substrates

Signals

PancreasPancreas

IntestineIntestine

AdrenalAdrenal

Glucose4 gramsGlucose4 grams

BrainBrain

IL6

RBP4

Body is in a ‘Gluconeogenic Mode’

Effects are Potentiated by the Fall in Insulin

Why is Glucagon so Effective during Exercise?

Liver

Insulin

Glucagon

Adipose

WorkingMuscleWorkingMuscle

GlycerolNEFA

LactateAmino Acids

GlyGly

GNGGNG

Autonomic Nerve Activity

Epi

?

AminoAcids

Substrates

Signals

PancreasPancreas

IntestineIntestine

AdrenalAdrenal

Glucose4 gramsGlucose4 grams

BrainBrain

IL6

RBP4

Body is in a ‘Gluconeogenic Mode’

Effects are Potentiated by the Fall in Insulin

Glucose Uptake Prevents Hyperglycemia

Protocol: Study of Splanchnic Amino Acid Metabolism during Exercise

Equilibration Basal Treadmill Exercise

-120 min -30 0 150

[5-15N]Glutamine + [1-13C]Leucine

The Exercise-induced Glucagon Response is Essential to the Increment in Hepatic Glutamine Extraction

0.00

0.20

0.40

0.60

25-50 75-100 125-150

Exercise Duration(min)

Basal 25-50 75-100 125-150

Exercise Duration(min)

Basal

HepaticFractionalGlutamineExtraction

Simulated Glucagon Basal Glucagon

**

*

††

The Exercise-induced Glucagon Response Drives Urea Formation in the Liver

Simulated Glucagon

0

5

10

15

20

25-50 75-100 125-150

Exercise Duration (min)

Basal 25-50 75-100 125-150

Exercise Duration (min)

Basal

Net HepaticUrea Output(mol·kg-1･ min-1)

**

*

 

Basal Glucagon

The Exercise-induced Glucagon Response is Required for the Accelerated transfer of Glutamine Amide Nitrogen to Urea in the Liver

Formation of Urea fromGlutamine Amide Nitrogen

during Exercise

(mol·kg-1･ min-1)

SimulatedGlucagon

BasalGlucagon

0.0

1.0

2.0

3.0

 

Energy State and the Liver

Energy State in the Liver is Controlled by Glucagon

*

Studies using the Phloridzin-Euglycemic Clamp further Illustrate the Role of Glucagon in Liver Energy Balance

Blood is Regulated like a Homeostat

Liver is the Battery (rechargeable)

RBP4

Substrates and Signals Implicated in Control of Glucose Fluxes to Working Muscle during Exercise

Liver

Adipose

GlycerolNEFA

LactateAmino Acids

GlyGly

GNGGNG

Epi

AminoAcids

Substrates

Signals

PancreasPancreas

IntestineIntestine

Glucose4 gramsGlucose4 grams

BrainBrain

IL6

Sensors Carotid Sinus Liver/Portal Vein Working Muscle

Feedforward

Feedback Chemical Mechanical

WorkingMuscleWorkingMuscle

Autonomic Nerve Activity

AdrenalAdrenal

Insulin

Glucagon

IL6

What about the Famous Catecholamine Response to Exercise?

• Epinephrine plays little to no role in control of glucose production during exercise. Moates et al Am J Physiol 255: E428-E436, 1988.

• Hepatic nerves are not necessary for the exercise-induced rise in glucose production. Wasserman et al Am J Physiol 259: E195-E203, 1990.

• Liver specific blockade of both - and -adrenergic receptors do not attenuate the increase in glucose production during exercise. Coker et al Am J Physiol 273: E831-E838, 1997. Coker et al Am J Physiol 278: 444-451, 2000.

Catecholamines

Essential, in association with the fall in insulin, for extrahepatic substrate mobilization during exercise.

Muscle glycogenolysis

Adipose tissue lipolysis

Glycerol

NE

TG

NEFA Flux is Accelerated during Moderate Exercise by Increased Lipolysis and Decreased Re-esterification

Glucose

FFA

TG

Glycerol

TG

FFA

FFA

TG

Glycerol

G3P

ATP

Glycerol

NE

TG

NEFA Flux is Accelerated during Moderate Exercise by Increased Lipolysis and Decreased Re-esterification

Glucose

FFA

TG

Glycerol

TG

FFA

FFA

TG

Glycerol

G3P

ATP

Four Grams of GlucoseControlling Rate of Removal

• blood flow• capillary recruitment• spatial barriers

Extracellular Membrane

• hexokinase #• hexokinase compartmentation• spatial barriers

Intracellular

glucose 6-phosphate

glucose

• transporter #• transporter activity

Strategy

Selectively remove sites of resistance to MGU in conscious mice by using transgenic mice or pharmacological methods.

Ohm’s Law Applied to Glucose Influx

V1 V2 V3 V4Resistor1 Resistor3Resistor2

Current (I)

V1 = I · Resistor1

Gextracell= Ig · Rextracell

Ga Ge Gi 0RExtracell RTransport RPhosp

Glucose Influx (Ig)

Gtransport= Ig · Rtransport Gphos = Ig · RPhosp

V2 = I · Resistor2 V3 = I · Resistor3

Ohm’s Law to Determine Sites of Resistance to Muscle Glucose Uptake

Ga Ge Gi 0

Glucose Influx

WT

GLUT4Tg

HKTg

GLUT4Tg

HKTg

Transgenics

ptf 2002/jea 2005

Chronically Catheterized, Conscious Unstressed Mouse

[2-14C]DG

Vein Artery

Sample

Insulin

Glucose

Blood

[3-3H]Glc

From: Glucose Clamping the Conscious Mouse by Vanderbilt MMPC 2005

Metabolic Control Analysis of MGU

• Control Coefficient ( C ) = lnRg/ln[E]

• Sum of Control Coefficients in a Defined Pathway is 1

i.e. Cd + Ct + Cp = 1

Control Coefficients for MGU by Mouse Muscle Comprised of Type II Fibers

Rest

Insulin(~80 µU/ml)

Delivery Transport Phosphorylation

0.1 0.9 0.0

0.5 0.1 0.4

Exercise Protocol

Acclimation Sedentary or Exercise

-90 30 min50

[2-3H]DG Bolus

Excise Tissues

Sedentary and Exercising Mice

WTHKTg

GLUT4Tg

HKTg + GLUT4Tg

0

50

100

150

200

250

0 5 10 15 20 25 30

Time (min)

Sedentary

0 5 10 15 20 25 30

Time (min)

Exercise

* * * ** *

* *

**

Blood Glucose (mg·dl-1)

Fueger et al. Am J Physiol; 286: E77-84, 2004

Sedentary and Exercising Mice

0

20

40

0

10

20Muscle Glucose Uptake

(mol·100g-1·min-1)

Sedentary

Exercise

SVL

Gastrocnemius

0

50

100Soleus

GLUT4Tg

†

†

WT

†

†

†

HKTg

+ GLUT4Tg

HKTG

†

†

†

Fueger et al. Am J Physiol; 286: E77-84, 2004

Control Coefficients for MGU by Mouse Muscle Comprised of Type II Fibers

Exercise

Delivery Transport Phosphorylation

0.1 0.9 0.0

0.5 0.1 0.4

0.2 0.0 0.8

Rest

Insulin(~80 µU/ml)

Distributed Control of Muscle Glucose Uptake

• Transport is clearly the primary barrier to muscle glucose uptake in the fasted, sedentary state.

• Transport is so effectively regulated by exercise and insulin that the membrane is no longer the primary barrier to muscle glucose uptake.

• The resistance to insulin-stimulated muscle glucose uptake with high fat feeding is due, in large part, to defects in the delivery of glucose to the muscle.

The vast majority of the literature on the regulation of glucose uptake is comprised of studies in isolated muscle tissue or cells that are blind to fundamental control mechanisms involved in muscle glucose uptake.

Four Grams of Glucose

Extracellular Membrane IntracellularLivergluconeogenic precursorsglycogen

glucose 6-phosphate

The distributed control of blood glucose allows for more precise control of glucose homeostasis, multiple mechanisms of glucose flux control, and multiple targets to correct dysregulation of metabolism such as is seen in diabetes

Carefully conducted studies in the whole animal are necessary to

ascribe function to putative controllers of glucose homeostasis.

Extracellular Membrane Intracellular

glucose 6-phosphate

glucose