Importance of the hepatic arterial glucose level in generation
of the portal signal in conscious dogs
Po-Shiuan
Hsieh1,
Mary
Courtney
Moore1,
Doss W.
Neal2, and
Alan D.
Cherrington1,2
1 Department of Molecular Physiology and Biophysics,
2 Diabetes Research and Training Center, Vanderbilt University
School of Medicine, Nashville, Tennessee 37232
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ABSTRACT |
The aim of this study was to
determine whether the elimination of the hepatic arterial-portal (A-P)
venous glucose gradient would alter the effects of portal glucose
delivery on hepatic or peripheral glucose uptake. Three groups of
42-h-fasted conscious dogs (n = 7/group) were studied.
After a 40-min basal period, somatostatin was infused peripherally
along with intraportal insulin (7.2 pmol·kg
1·min
1) and glucagon (0.65 ng·kg
1·min
1). In test
period 1 (90 min), glucose was infused into a peripheral vein to
double the hepatic glucose load (HGL) in all groups. In test
period 2 (90 min) of the control group (CONT), saline was infused
intraportally; in the other two groups, glucose was infused intraportally (22.2 µmol·kg
1·min
1). In the
second group (PD), saline was simultaneously infused into the
hepatic artery; in the third group (PD+HAD), glucose was infused into
the hepatic artery to eliminate the negative hepatic A-P glucose
gradient. HGL was twofold basal in each test period. Net hepatic
glucose uptake (NHGU) was 10.1 ± 2.2 and 12.8 ± 2.1 vs.
11.5 ± 1.6 and 23.8 ± 3.3* vs. 9.0 ± 2.4 and
13.8 ± 4.2 µmol · kg
1·min
1 in the two periods of CONT,
PD, and PD+HAD, respectively (* P < 0.05 vs. same
test period in PD and PD+HAD). NHGU was 28.9 ± 1.2 and 39.5 ± 4.3 vs. 26.3 ± 3.7 and 24.5 ± 3.7* vs. 36.1 ± 3.8 and 53.3 ± 8.5 µmol·kg
1·min
1 in the first
and second periods of CONT, PD, and PD+HAD, respectively (* P < 0.05 vs. same test period in PD and PD+HAD).
Thus the increment in NHGU and decrement in extrahepatic glucose uptake
caused by the portal signal were significantly reduced by hepatic
arterial glucose infusion. These results suggest that the hepatic
arterial glucose level plays an important role in generation of the
effect of portal glucose delivery on glucose uptake by liver and muscle.
liver; liver nerve; hepatic glucose uptake
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INTRODUCTION |
THE ROUTE OF GLUCOSE
DELIVERY is one of the key determinants of net hepatic glucose
uptake (NHGU), with NHGU being enhanced two- to threefold during
infusion of glucose into the portal vein vs. a peripheral vein
(11). The ability of intraportal glucose delivery to
increase NHGU is not dictated by the absolute glucose concentration in
the portal vein but is instead a function of the magnitude of the
negative arterial-portal (A-P) glucose gradient. It seems likely,
therefore, that after portal glucose delivery, the portal glucose level
is sensed and compared with the arterial glucose level, which is
simultaneously monitored at some as yet undetermined site. Three
possible arterial reference sites have been suggested: the hepatic
artery (6, 24), the arterial blood supply of
the hypothalamus (4), and the arterial blood reaching the
carotid bodies (2, 3).
Using isolated perfused rat liver, Gardemann et al. (6)
and Stumpel and Jungermann (24) showed that a negative
glucose gradient between the hepatic artery and the portal vein could be transformed into a metabolic signal locally within the liver and
that the action of acetylcholine on the hepatocyte seemed to be
involved in this signal transduction. Recently, Horikawa et al.
(7) reported that both portal vein and hepatic arterial glucose infusion stimulated NHGU in the conscious dog. These authors went on to suggest that glucose sensors within the liver, rather than
the portal vein, are involved in the augmentation of NHGU.
Data also exist to support the contention that the portal glucose
level is compared with an arterial glucose level sensed outside the
liver. Results from Matsuhisa et al. (9) suggested that
minimizing the glucose gradient between the portal vein and the central
nervous system diminished hepatic glucose uptake. Our previous study
(8) overcame many of the problems associated with their
study design, however, and failed to confirm their results. In our
study, there was no diminution of NHGU during portal glucose delivery
when glucose was infused simultaneously into the carotid and vertebral
arteries to maintain the brain isoglycemic with the portal vein
(8). Other studies (2, 3) have
shown that changes in the blood glucose concentration within the
carotid body can affect glucose homeostasis. This implies that the
carotid bodies provide another potential reference site for arterial
glucose sensing. However, infusion of glucose into the carotid and
vertebral arteries, as in our earlier study (8), would
also alter the glucose level in the carotid bodies. Our earlier data,
therefore, do not support carotid body involvement in the operation of
the portal signal.
The aim of the present study was to clarify this issue by determining
whether elimination of the hepatic A-P venous glucose gradient within
the liver would alter the changes in net hepatic and peripheral glucose
uptake induced by portal glucose delivery in conscious dogs.
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METHODS |
Animals and surgical procedures.
Studies were carried out on twenty-one 42-h-fasted conscious mongrel
dogs of either sex weighing 21-28 kg. All animals were maintained
on a diet of meat (Kal-Kan, Vernon, CA) and chow (Purina Lab Canine
Diet no. 5006; Purina Mills, St. Louis, MO) composed of 34% protein,
14.5% fat, 46% carbohydrate, and 5.5% fiber based on dry weight. The
protocol was approved by the Vanderbilt University Medical Center
Animal Care Committee, and the animals were housed according to the
guidelines of the American Association for the Accreditation of
Laboratory Animal Care International. Approximately 16 days before the
study, each dog underwent surgery under general anesthesia for
placement of sampling catheters into a hepatic vein, the portal vein,
and a femoral artery and infusion catheters in a splenic and a jejunal
vein (11). A Silastic catheter was inserted into the
gastroduodenal artery, and its tip was advanced 3-4 cm retrograde
into the common hepatic artery. The distal end of the gastroduodenal
artery was then ligated to ensure total delivery of glucose to the
hepatic bed. Ultrasonic flow probes (Transonic Systems, Ithaca, NY)
were positioned around the portal vein and hepatic artery. The proximal
ends of the probes and catheters were placed in subcutaneous pockets
(11).
The hematocrit and leukocyte count were measured 1-2 days before
the study. A dog was studied only if it exhibited a hematocrit of
>36%, leukocyte count of <18,000/mm3, consumption of
more than three-fourths of the daily ration, and normal stools. On the
morning of the study, the proximal ends of the flow probes and
surgically implanted catheters were exteriorized, the catheters were
cleared, the dog was placed in a Pavlov harness, and intravenous access
was established in three peripheral veins.
Experimental design.
At
120 min, a primed (36 µCi) continuous (0.3 µCi/min) peripheral
infusion of D-[3-3H]glucose and a continuous
peripheral infusion of indocyanine green dye [(ICG) Sigma Chemicals,
St. Louis, MO, 4 µg·kg
1·min
1] were begun in
all three protocols. The ICG infusion allowed confirmation of hepatic
vein catheter placement and provided a second measurement of hepatic
blood flow. After 80 min (from
120 to
40 min) of dye equilibration,
there was a 40-min (from
40 min to time 0) basal period
followed by two 90-min experimental periods. At time 0,
constant infusions of several solutions were begun. Somatostatin (0.8 µg·kg
1·min
1; Bachem,
Torrance, CA) was infused to suppress endogenous insulin and glucagon
secretion. Insulin (7.2 pmol·kg
1·min
1) and glucagon
(0.65 ng·kg
1·min
1; both from
Eli Lilly, Indianapolis, IN) were infused intraportally at rates
designed to elevate insulin three- to fourfold and to keep glucagon
basal. In addition, a primed continuous peripheral infusion of 50%
dextrose was begun so that the arterial blood glucose level could be
quickly clamped at a value approximately equal to twofold
basal. In the first experimental period, glucose was infused only
through a peripheral vein to double the glucose load to the liver in
all groups. In the control study (CONT, n = 7) the
conditions established in the first test period were continued
throughout the second experimental period. During the second
experimental period of the other two groups, glucose was infused
intraportally (22.2 µmol·kg
1·min
1). In one
group, saline was concurrently infused into the hepatic artery
(PD, n = 7), but in the other group, glucose
was concurrently infused into the hepatic artery (8.0 ± 0.5 µmol·kg
1·min
1) to eliminate
the glucose gradient between the portal vein and the hepatic artery
(PD+HAD, n = 7). The peripheral glucose infusion rate
was modified as required to maintain the hepatic glucose load equal to
twofold basal in each protocol. Dextrose, 20 and 5%, was used for the
portal and hepatic arterial glucose infusions, respectively, and
p-aminohippuric acid (PAH; Sigma Chemicals, St.
Louis, MO) was added to the infusates to assess mixing of the infused
glucose with blood in the portal and hepatic veins, as described
previously (11, 18). Blood sampling
was performed as previously described (8).
Processing and analysis of samples.
Plasma glucose was assayed by the glucose oxidase method with a
Beckman glucose analyzer (Fullerton, CA). Plasma insulin and glucagon
concentrations were determined by RIA, as previously described
(8). Blood glucose and blood lactate levels were determined by fluorometric enzymatic assays on perchloric acid-treated samples, as previously described (8). PAH was also
measured in perchloric acid-deproteinized blood (11,
18).
Calculations.
When substrates are infused into the portal vein, the possibility of
poor mixing with the blood in the laminar flow of the portal
circulation is of concern. In addition, multiple branching patterns of
the hepatic artery in the dog (21) raise concern about the
mixing of infusates with the blood of the hepatic artery. In both PD
and PD+HAD, the mixing of the infusate with portal vein blood was
assessed by comparing the portal PAH infusion rate with the calculated
appearance rate of PAH in the portal vein [the difference between the
rate of PAH exiting in portal blood and the rate of PAH entering the
splanchnic bed through the arterial system (11,
18)], as shown in the equation below
where A and P represent the femoral arterial and portal venous
blood concentrations, and PBF represents portal blood flow.
The mixing of the infusate with blood by the time it reached the
hepatic vein was assessed in a similar fashion. In the PD group, we
used the equation
whereas in the PD+HAD group, we used the equation
where HBF represents hepatic blood flow.
In the PD+HAD group, we used an indirect (I) approach to assessing the
mixing in the hepatic artery, because we could not sample hepatic
arterial blood. To increase our ability to assess the mixing of
arterially delivered PAH by the time the arterial blood reached the
hepatic vein, we infused PAH at a higher rate into the hepatic artery
than into the portal vein (0.37 ± 0.03 vs. 0.23 ± 0.01 mg·kg
1·min
1). Mixing of the
hepatic arterial infusate within the liver in the PD+HAD group was
assessed using the equation
This assumes that mixing of portally delivered PAH is similar in
the portal and hepatic veins, which is usually the case.
Samples were considered statistically unmixed (95% confidence that
mixing did not occur) if hepatic or portal vein recovery of PAH was
40% greater or less than the actual amount of PAH infused (11, 18). An experiment was excluded from the
database if poor mixing was observed at more than one of the four time
points in the portal glucose infusion period or if the average mixing in a dog was <80% or >120% of the expected value. In PD, 5 of 12 dogs studied were excluded because of poor mixing; in PD+HAD, 6 of 13 dogs studied were excluded because of poor mixing in the portal vein
and/or the hepatic artery. In the dogs retained in the database, the
average ratio of PAH recovery in the portal vein to the intraportal PAH
infusion rate was 0.9 ± 0.1 in both PD and PD+HAD, with a ratio
of 1.0 representing perfect mixing. The ratio of PAH recovery in the
hepatic vein to the PAH infusion rate was 0.8 ± 0.1 and 0.9 ± 0.1 in PD and PD+HAD, respectively. The recovery of PAH infused in
the hepatic artery was 1.0 ± 0.1 during the hepatic arterial
glucose infusion period in the PD+HAD group. When a dog was retained in
the database, all of the data points were used, whether they were mixed
or not, because mixing errors occur randomly.
The mean results for HBF obtained with ultrasonic flow probes and ICG,
respectively, in the two study periods for each group were not
significantly different. The data shown in the figures are those
obtained with the flow probes, because their use did not require an
assumption regarding the distribution of arterial and portal
contribution to hepatic blood flow. Plasma glucose values were
converted to whole blood glucose values by use of a correction factor,
as previously described (18).
The rate of substrate delivery to the liver, or hepatic substrate load,
was calculated by a direct (D) method as
where [S] is the substrate concentration, and ABF refers to
blood flow through the hepatic artery. A similar method was used to
calculate the hepatic sinusoidal insulin and glucagon concentrations
where [H] is the hormone concentration, and HS refers to
hepatic sinusoid. To minimize any potential errors arising from either
incomplete mixing of glucose during intraportal glucose infusion or a
lack of precise measurement of the distribution of hepatic blood flow,
the hepatic glucose load was also calculated by an indirect method
where G is the blood glucose concentration, GIRPO is
the intraportal glucose infusion rate, GIRHA is the hepatic
arterial glucose infusion rate, and GUG is the uptake of glucose from
the blood by the gastrointestinal tract, calculated on the basis of the
previously described relationship between the arterial blood glucose
concentration and GUG (11, 18).
The load of a substrate exiting the liver was calculated as
Direct and indirect methods were used in calculation of net
hepatic balance (NHB). The direct calculation was:
NHBD = loadout
loadin
(D). The indirect calculation was: NHBI = loadout
loadin (I). The data in the
figures represent those calculated by the indirect calculation of net
hepatic glucose balance (NHGB) during the portal glucose infusion
periods so that we would be consistent with our previous publications;
nevertheless, the mean values were not significantly different,
regardless of the method used in calculation. Lactate balance was
calculated by the direct method. Net fractional substrate extraction by
the liver was calculated as the ratio of NHB (I) to loadin
(I). Nonhepatic glucose uptake (non-HGU) was calculated by subtracting
the rate of NHGU (I) from the total glucose infusion rate. The net
hepatic balance of glucose equivalents was calculated as the sum of the
balances of NHGB (I) and lactate when the latter had been converted to
glucose equivalents. This calculation serves as an indicator of the
carbon used for glycogen deposition; however, it ignores carbon derived from gluconeogenic precursor uptake (
3
µmol·kg
1·min
1) and glucose
used for oxidation (
1.5
µmol·kg
1·min
1), which tend
to offset each other.
The calculation of hepatic arterial glucose infusion rate was
based on our desire to maintain the hepatic arterial glucose level at a
value slightly above the portal glucose level
In the calculation of the hepatic arterial glucose infusion
rate, the hepatic arterial and portal blood flows were measured by
Transonic flow probes. Our goal was to maintain the positive A-P
glucose gradient that exists in the presence of peripheral glucose delivery.
Data are presented as means ± SE. SYSTAT (SYSTAT, Evanston, IL)
was used for statistical analysis. The time course data were analyzed
by repeated-measures ANOVA with post hoc analysis and univariate
F tests. Results were considered statistically significant at P < 0.05.
 |
RESULTS |
Plasma insulin and glucagon concentrations.
Arterial (data not shown) and liver sinusoidal insulin concentrations
rose nearly three- to fourfold in all groups. Glucagon levels, on the
other hand, remained basal. Neither hormone differed between groups
(Fig. 1).

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Fig. 1.
Calculated plasma insulin (top) and glucagon
(bottom) levels in the hepatic sinusoid in 42-h-fasted
conscious dogs during the basal and two experimental periods in PD
(portal vein glucose delivery), PD+HAD (portal vein and hepatic artery
glucose delivery), and CONT (control) groups (n = 7/group). SRIF, somatostatin; PO+HA Glucose, intraportal and hepatic
arterial glucose infusion. There are no significant differences between
groups.
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Blood glucose levels, the A-P glucose gradient and hepatic blood
flow.
Peripheral glucose infusion in the first test period doubled the blood
glucose levels (Fig. 2) such that the
arterial, portal venous, and hepatic venous glucose levels were not
significantly different (NS) among the three groups. In the second test
period, the arterial blood glucose level was slightly higher in CONT
than in the other two groups (9.3 ± 0.2* vs. 8.6 ± 0.2 and
8.3 ± 0.2 mM, * P < 0.05 vs. PD and PD+HAD,
respectively). The portal and hepatic glucose levels, on the other
hand, were indistinguishable in the three groups. In PD, intraportal
glucose infusion switched the A-P blood glucose gradient from 0.10 ± 0.04 (period 1) to
0.75 ± 0.08 mM and
thereby presented the liver with a portal signal (Fig.
3B). In PD+HAD, the
hepatic arterial glucose infusion offset the portal glucose infusion so
that a positive glucose gradient was maintained between the hepatic
artery and the portal vein, even in the latter experimental period
(0.22 ± 0.03 vs. 0.55 ± 0.08 mM; Fig. 3A).
However, there was still a negative glucose gradient between the
femoral artery and the portal vein (
0.57 ± 0.10 mM). A positive
A-P glucose gradient was maintained throughout the experiment in CONT
(0.11 ± 0.03 and 0.14 ± 0.07 mM in the two test periods,
respectively).

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Fig. 2.
Blood glucose levels in the femoral artery, portal vein,
and hepatic vein in 42-h-fasted conscious dogs during the basal and two
experimental periods in PD, PD+HAD, and CONT groups (n = 7/group). * P < 0.05 vs. CONT at this time
point.
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Fig. 3.
Blood glucose levels in the hepatic arteries
(calculated), portal vein, and femoral artery in 42-h-fasted conscious
dogs during the 4 sampling points in the second experimental period in
PD+HAD (A) and PD (B); n = 7/group.
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The HBF was not different over time or between groups except in the
latter test period when HBF was slightly lower in CONT than in the
other two groups (26 ± 1* vs. 32 ± 2 vs. 31 ± 3 ml·kg
1·min
1;
* P < 0.05 vs. the other groups).
Hepatic glucose load, net hepatic glucose balance, and net hepatic
fractional extraction of glucose.
The hepatic glucose loads (HGL) did not differ (268 ± 29 and
264 ± 24 µmol·kg
1·min
1) between the
two test periods in CONT. The HGLs in the other two groups were
similar, although the HGL rose slightly in period 2 in these
groups (252 ± 19 and 281 ± 18 vs. 233 ± 19 and
278 ± 21*µmol·kg
1·min
1 in PD and
PD+HAD; * P < 0.05 vs. period 1; Fig.
4, top). Net hepatic glucose
balance (NHGB, i.e., net hepatic glucose output and NHGU) is shown in
Fig. 4 (bottom). In the basal period, net hepatic glucose
output did not differ between PD, PD+HAD, and CONT (9.7 ± 1.1, 9.7 ± 1.6, and 9.3 ± 1.7 µmol·kg
1·min
1,
respectively). Peripheral glucose infusion in the presence of hyperinsulinemia resulted in similar rates of NHGU in all three groups
(11.5 ± 1.6, 9.0 ± 2.3, and 10.1 ± 2.2 µmol·kg
1·min
1
in PD, PD+HAD, and CONT, respectively). During the latter experimental period, NHGU increased to 23.8 ± 3.3 µmol·kg
1·min
1 in PD
(
12.4 ± 3.2 µmol·kg
1·min
1,
P < 0.05), to 13.8 ± 4.2 µmol·kg
1·min
1 in
PD+HAD (
4.9 ± 2.4 µmol·kg
1·min
1, NS), and to
12.8 ± 2.1 µmol·kg
1·min
1 in CONT (
2.7 ± 1.5 µmol·kg
1·min
1, NS). NHGU
did not differ between PD+HAD and CONT at any time, indicating that
elimination of the glucose difference between the hepatic artery and
the portal vein markedly reduced the effect of portal glucose delivery
on NHGU (Fig. 4, bottom). When the data were analyzed with D
(rather than I), NHGU increased to 20.5 ± 2.9 µmol·kg
1·min
1 in PD
(
9.1 ± 2.4 µmol·kg
1·min
1,
P < 0.05), to 8.4 ± 2.7 µmol·kg
1·min
1 in PD+HAD
(
0.3 ± 0.7 µmol·kg
1·min
1, NS), and
to 12.8 ± 2.1 µmol·kg
1·min
1 in CONT (
2.7 ± 1.5 µmol·kg
1·min
1, NS).

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Fig. 4.
Hepatic glucose load (top) and net hepatic
glucose balance (bottom) in 42-h-fasted conscious dogs
during the basal and two experimental periods in PD, PD+HAD, and CONT
(n = 7/group). P < 0.05, CONT
vs. PD at this time point; * P < 0.05, PD+HAD vs.
PD at this time point.
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Net hepatic fractional extraction of glucose (NHFEG; Fig.
5) showed a similar trend. NHFEG values
were 0.047 ± 0.008 and 0.092 ± 0.016* vs. 0.045 ± 0.012 and 0.051 ± 0.016 vs. 0.039 ± 0.007 and 0.051 ± 0.009 in the two test periods of PD, PD+HAD, and CONT, respectively
(* P < 0.05 vs. period 1). In the latter
experimental period, the increment of NHFEG induced by the portal
signal was completely suppressed by hepatic arterial glucose infusion.
NHFEG did not differ between PD+HAD and CONT.

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Fig. 5.
Net hepatic fractional extraction in 42-h-fasted
conscious dogs during the basal and two experimental periods in PD,
PD+HAD, and CONT (n = 7/group). P < 0.05, CONT vs. PD at this time point; * P < 0.05, PD+HAD vs. PD at this time point.
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Nonhepatic glucose uptake.
Peripheral glucose infusion during period 1 resulted in
nonhepatic glucose uptake (non-HGU; Fig.
6) of 26.3 ± 3.7, 36.1 ± 3.8, and 28.9 ± 1.2 µmol·kg
1·min
1 in PD,
PD+HAD, and CONT, respectively. In period 2, non-HGU was 24.5 ± 3.7, 53.3 ± 8.5, and 39.5 ± 4.3 µmol·kg
1·min
1 in PD,
PD+HAD, and CONT, respectively. Non-HGU rose by 10.5 ± 4.2 µmol·kg
1·min
1 in
period 2 of CONT. In PD, non-HGU fell by 1.7 ± 4.7 µmol·kg
1·min
1 in
period 2, and thus the net effect of the portal signal
was to decrease non-HGU by 12.4 ± 4.7 µmol·kg
1·min
1. In PD+HAD,
non-HGU rose by 17.5 ± 5.9 µmol·kg
1·min
1 in
period 2. The change in non-HGU did not differ between
PD+HAD and CONT. Thus hepatic arterial glucose delivery completely
blocked the suppressive effect of portal glucose delivery on non-HGU.

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Fig. 6.
Nonhepatic glucose uptake in 42-h-fasted conscious dogs
during the basal and two experimental periods in PD, PD+HAD, and CONT
(n = 7/group). P < 0.05, CONT
vs. PD at this time point; * P < 0.05, PD+HAD vs.
PD at this time point.
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The total glucose infusion rate increased moderately (20-50%)
between periods 1 and 2 in each protocol
(37.7 ± 3.5 and 48.4 ± 3.1 vs. 44.7 ± 4.3 and
67.4 ± 6.8 vs. 39.0 ± 2.2 and 52.3 ± 3.7 µmol·kg
1·min
1 in PD,
PD+HAD, and CONT, respectively; data not shown).
Net hepatic lactate balance.
In response to peripheral glucose infusion, net hepatic lactate balance
switched from uptake (10.8 ± 1.7, 8.7 ± 1.3 and 5.9 ± 2.2 µmol·kg
1·min
1) to
output (4.4 ± 1.3, 3.6 ± 2.4 and 5.8 ± 1.8 µmol·kg
1·min
1) in the first
test period of PD, PD+HAD, and CONT, respectively. Net hepatic lactate
output fell slightly in the latter experimental period but was not
significantly different among the three groups (Table
1). Blood lactate levels in the femoral
artery, portal vein, and hepatic vein were also not different among the
three groups (data not shown).
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Table 1.
Net hepatic lactate balance during basal and two experimental periods
in PD+HAD, PD and CONT groups of 42-h-fasted conscious dogs
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Net hepatic balance of glucose equivalents.
The net balance of glucose equivalents across the liver represents the
combination of glucose and lactate balance (after the latter is
converted to glucose equivalents) and serves as an index of glycogen
deposition. The net balance of glucose equivalents across the liver
switched from output (4.0 ± 1.3, 5.4 ± 1.7, and 9.0 ± 1.7 µmol·kg
1·min
1)
to uptake (9.2 ± 1.9, 6.9 ± 1.8, and 7.2 ± 1.8 µmol·kg
1·min
1) in PD,
PD+HAD, and CONT, respectively, in response to peripheral glucose
infusion (Fig. 7). In the latter
experimental period, the uptakes of glucose equivalents in the three
groups were 22.1 ± 3.2,* 13.8 ± 4.0, and 10.9 ± 1.6 µmol·kg
1·min
1,
respectively. Hepatic arterial glucose infusion markedly reduced the
stimulatory effect of the portal signal on glycogen deposition (* P < 0.05 vs. the other two groups).

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Fig. 7.
Net hepatic balance of glucose equivalents in 42-h-fasted
conscious dogs during the basal and two experimental periods in PD,
PD+HAD, and CONT (n = 7/group). The net balance of
glucose equivalents is calculated as the sum of the net balance of
glucose and lactate, the latter converted to glucose equivalents.
P < 0.05, CONT vs. PD at this time point;
* P < 0.05, PD+HAD vs. PD at this time point.
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DISCUSSION |
Previous studies have demonstrated that the portal signal is an
important component of the metabolic response to feeding
(4, 11, 12, 18). We
have established that a negative A-P gradient is necessary for the
initiation of the portal signal (19). However, the
reference glucose concentration against which the portal glucose concentration is compared is still not clear. This study sought to
determine whether elimination of a negative glucose gradient between
the hepatic artery and the portal vein would alter the effects of the
portal signal on glucose uptake by the liver and/or peripheral tissues.
The present data demonstrate that the effects of the portal signal on
hepatic and peripheral glucose uptake and on the stimulation of hepatic
glycogen storage were markedly reduced by eliminating the hepatic A-P
glucose gradient within the liver in conscious dogs.
The results from the present study suggest that the portal signal is
generated within the liver itself. They are consistent with the work of
Gardemann et al. (6) and Stumpel and Jungermann (24) in the perfused liver. However, the intrahepatic
mechanism by which the portal signal is generated and the way in which
it is transduced into an effect on hepatic glucose uptake are still unknown. Stumpel et al. suggested that the parasympathetic nervous system may be involved. In their studies with isolated perfused rat
livers, the increment in insulin-stimulated glucose uptake induced by a
negative A-P glucose gradient was markedly reduced by either portal or
arterial delivery of atropine. In addition, they showed that the effect
of a negative A-P glucose gradient on NHGU could be mimicked by
addition of acetylcholine to either the portal or arterial perfusate.
The combination of acetylcholine and adrenergic blockers infused
intraportally in the conscious dog rapidly stimulated NHGU in the
presence of hyperinsulinemia and hyperglycemia, but adrenergic blockade
alone did not alter NHGU (23). The above change in NHGU
was consistent with those induced by the portal signal in terms of both
time course and magnitude. These observations suggest that the
muscarinic nervous system within the liver is somehow involved in the
generation of the hepatic effect of the portal signal and/or in the
transduction of its effect into a biological action.
Neurophysiological data indicate that glucose-responsive neurons in
specific hypothalamic regions and in the dorsomedial medulla oblongata
modulate glucose metabolism in some organs (liver and pancreas) through
autonomic efferent nerves (16, 17). However, only one study (9) has suggested that the head glucose
level is involved in the generation of the portal signal. Furthermore, the quantitative accuracy of that study has been questioned on several
grounds, including the mixing of their infusate in portal blood
(8). In a previous study designed to examine the same question, we infused glucose (22.2 µmol·kg
1·min
1)
intraportally at a rate known to increase NHGU maximally in the
presence of a fourfold rise in insulin. Unlike Matsuhisa et al.
(9), we used a rate that minimized potential problems of portal glucose mixing (a favorable noise-to-signal ratio). In addition,
we used an independent method (PAH assay) to assess the mixing of the
infused glucose in the portal vein and the hepatic vein, and this in
turn allowed evaluation of the potential effects of imperfect mixing in
the two individual protocols. Also, we infused glucose into the carotid
and vertebral arteries bilaterally instead of unilaterally, as had
Matsuhisa et al., to ensure a uniform elimination of the difference
between brain and portal vein glucose. Our results clearly demonstrated
that, under hyperglycemic hyperinsulinemic conditions, raising the head
arterial glucose level did not modify the increase in NHGU seen in
response to portal glucose delivery. Thus the brain arterial glucose
level does not appear to provide the reference information used in
generation of the portal signal in conscious dogs. To date, our data
suggest that the liver, not the brain, is the organ critical for
orchestration of the distribution of dietary glucose after a
carbohydrate meal.
A recent report by Horikawa et al. (7) suggested that
augmentation of hepatic glucose uptake was not dependent on the sign of
the A-P venous glucose gradient and that a difference in either direction (A > P or P > A) was effective. They
used conscious dogs in an experiment consisting of a 30-min control and
three 90-min test periods. After the control period, glucose (55.6 µmol·kg
1·min
1) was first
infused via the superior mesenteric vein; it was then infused into both
the superior mesenteric vein and the gastroduodenal artery (27.8 µmol·kg
1·min
1 in each
vessel); and finally, it was infused solely into the gastroduodenal
artery (55.6 µmol·kg
1·min
1).
Unfortunately, the data from that study must be questioned on several
grounds. First, the authors did not assess the mixing of their infusate
with blood in any vessel, and poor portal glucose mixing was likely,
given the extremely high glucose infusion rates used. This is further
evident from the inconsistency between NHGU calculated by the direct
and indirect methods in the second test period. When the indirect
method of glucose balance calculation was used, it appeared that NHGU
decreased 50% during concurrent infusion of glucose into the portal
and arterial systems, suggesting an inhibition of NHGU by hepatic
arterial glucose infusion. On the other hand, no decrease in NHGU was
evident when the direct method of calculation was used, making it
difficult to draw a conclusion. Second, the pancreatic hormones were
not clamped, and as a result they varied somewhat over the course of
the study. Third, the authors failed to include a control group that
would account for changes over time during the study. Finally, the
plasma glucose values were not in steady state, thus limiting the
accuracy of the arteriovenous difference calculation.
In the present study, we made several improvements over the study
design of Horikawa et al. (7) to minimize the above
errors. First, the hepatic arterial glucose infusion (8.0 ± 0.5 µmol·kg
1·min
1) was kept
small to minimize the impact of any mixing problems (i.e., to improve
the signal-to-noise ratio). Even under our carefully controlled
conditions, mixing of the infusate in the hepatic artery was probably
not perfect. This is evidenced by the fact that the positive gradient
between the hepatic artery and the portal vein in PD+HAD was slightly
greater during the latter experimental period than during the first.
Second, we used PAH to assess mixing of both the portal and hepatic
arterial infusates and only "mixed" dogs were utilized for data
analysis. Third, we used two control groups so that we could isolate
the effects of hyperglycemia, hyperinsulinemia, and the portal signal
over time. Fourth, we clamped insulin, glucagon, and the HGL so that
they did not differ among the protocols. Finally, we made our
measurements in a steady-state period. The present results thus clearly
demonstrate that a gradient between the hepatic arterial and portal
vein glucose levels is critical for the generation of the hepatic and
extrahepatic effects of the portal signal.
It is also possible, however, that the portal signal is sensed within
the liver and that the transduction of its effect is mediated by the
autonomic nervous system outside the liver. This is made more likely by
the need to explain the coordinate but discrete responses of the liver
and nonhepatic tissues (primarily muscle). It is clear that the glucose
portal signal not only enhances hepatic glucose uptake but also
suppresses nonhepatic glucose uptake (Fig. 6). If this were not true,
the glucose infusion rate would have been the same in PD and PD+HAD.
Earlier reports have described the existence of neural pathways that
link the liver to the brain and the brain to the liver and various
endocrine organs (5, 20). Afferent fibers in
the hepatic branch of the vagus nerve (14) and neurons in
the lateral hypothalamus (22) can respond to the presence
of glucose in the portal vein. Functional studies also have
demonstrated that an intact nerve supply to the liver appears to be
vital for a normal hepatic response to intraduodenal or intraportal
glucose delivery (1, 10). Niijima and
colleagues (13, 15) reported that the
stimulation of hepatic afferents can alter the efferent activity of the
adrenal and vagal pancreatic nerves in the rabbit and rat. There is no doubt that an autonomic link between the liver and peripheral organs
exists, but the extent to which it is involved in the initiation of the
effects of the portal signal remains to be determined.
Recently, several reports from Xie and Lautt (26,
27) focused on the relationship between peripheral insulin
resistance and the activity of hepatic parasympathetic nervous system.
They reported that surgical denervation of the hepatic anterior plexus or intraportal atropine infusion reduced the magnitude of insulin's effectiveness in skeletal muscle but had no effect on the liver or gut
(25, 26). Complete hepatic denervation plus
vagotomy did not cause further impairment, thus indicating that all of the relevant nerves reached the liver via the anterior plexus. Futhermore, intraportal, but not peripheral, acetylcholine infusion reversed insulin resistance produced by liver denervation
(27). These studies suggest that the hepatic
parasympathetic nerves regulate release of a liver-generated factor
that selectively controls insulin effectiveness in skeletal muscle.
Because the intrahepatic parasympathetic system has been suggested to
be a key determinant in generation and/or transduction of the effect of
the portal signal (24), the results from Xie and Lautt
(25-27) further imply that the anterior plexus around
the hepatic artery might be a potential reference site for generation
of the suppressive effect of the portal signal on nonhepatic glucose
uptake. This possibility needs to be investigated further.
In summary, the elimination of the negative glucose gradient between
hepatic artery and portal vein markedly reduced the effects of the
portal signal on hepatic and peripheral glucose uptake. This suggests
that the liver plays a primary role in the regulation of postprandial
glucose distribution, not only by augmenting its own uptake of glucose
but by preventing glucose uptake by skeletal muscle.
 |
ACKNOWLEDGEMENTS |
The authors would like to acknowledge the technical assistance of
Wanda Snead and Pam Venson in the hormone core laboratory of the
Vanderbilt University Medical Center Diabetes Research and Training Center.
 |
FOOTNOTES |
The work was supported by National Institute of Diabetes and Digestive
and Kidney Diseases Grant R-01 DK-43706 and Diabetes Research and
Training Center Grant SP-60 AM-20593.
Address for reprint requests and other correspondence:
M. C. Moore, 702 Light Hall, Dept. of Molecular Physiology and
Biophysics, Vanderbilt University School of Medicine, Nashville, TN
37232-0615 (E-mail: genie.moore{at}mcmail.vanderbilt.edu).
Present affiliation of P.-S. Hsieh: Department of Biology and Anatomy,
National Defense Medical Center, Taipei 114, Taiwan.
The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement"
in accordance with 18 U.S.C. §1734 solely to indicate this fact.
Received 28 December 1999; accepted in final form 7 March 2000.
 |
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