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[A1CR]

We CRONies often consider our glucose levels to be an
indicator of the
beneficial effects of CR, and often take pride in their low
levels, relative
to those who do not CR. Well, it appears that several types of
http://en.wikipedia.org/wiki/Glucose_transporter (GLUT) are
required for the
effectiveness of CR. "Glycerol 3-phosphate dehydrogenase:
GPDH ... is an
enzyme which catalyzes the reversible reaction between
dihydroxyacetone
phosphate and glycerol 3-phosphate with NAD as coenzyme. It
is known that
GPDH activity increases significantly during differentiation
of progenitor
cells into adipocytes. This enzyme activity has been used as
an index for
monitoring the fat synthesis."

A new paper appears to have examined GLUTs in CRed
codfish. They
apparently found that starvation led to heart tissue GLUT4
and GPDH
decreasing. Maybe this is far-fetched, but was their
relevance in GLUT
being proposed to be involved in the action of resveratrol,
which acts via
NAD coenzyme by MR in http://tinyurl.com/ynxebv and CR
countering
one of the GLUTs in the heart in the paper below?

Apparently, the hepatosomatic index (HSI) is defined as:
"Liver weight as a
percentage of the whole body weight". For the condition
factor, the
explanation in http://tinyurl.com/wjk72 appears to indicate
it is an
indication of the well being of a fish.

AP estimates the following values. For Fig. 5, relative to
the 2-month fed
fish, the 2-month starved and refed fish values in % were
for: body mass, 65
and 71; length, 95 and 92; condition factor, 80 and 106; and
HSI, 75 and 85.
Comparable values for Fig. 6 were: blood glucose, 23 and 68;
and liver
glycogen, 12 and 175. The blood glucose levels at liver
glycogen levels
were 4 units at 0 units and 8 units at 175 units,
respectively, with much
variation among individual codfish. For Fig. 8, starved and
re-fed % values
relative to fed values were, respectively: heart GLUT4, 53
and 108; heart
GPDH, about the same as for GLUT4, with somewhat less
reduction for starved
and a small reduction for re-fed codfish; white muscle
GLUT4, 225 and 125;
liver GLUT2, 70 and 94; and live GLUT, somewhat reduced
changes for both
starved and re-fed codfish.

Hall JR, Short CE, Driedzic WR.
Sequence of Atlantic cod (Gadus morhua) GLUT4, GLUT2 and
GPDH: developmental
stage expression, tissue expression and relationship to
starvation-induced
changes in blood glucose.
J Exp Biol. 2006 Nov 15;209(Pt 22):4490-502.
PMID: 17079719


... Food deprivation for 2 months was used as a vehicle to
monitor GLUT
expression at different blood glucose levels. Starvation
resulted in a
decrease in blood glucose and liver glycogen that recovered
following 20
days of re-feeding. GLUT4 expression in heart was decreased
with starvation
and increased with re-feeding. GLUT4 mRNA level in heart
correlated with
blood glucose. It is suggested that this relationship is
related to insulin
responsiveness. GLUT4 expression in white muscle increased
with starvation
and decreased with re-feeding. It is proposed that this is
due to the
necessity to maintain high levels of the glucose transporter
protein in the
face of starvation-associated proteolysis. GLUT2 expression
in liver
correlated with blood glucose, consistent with higher rates
of glucose
transport from liver to blood in the fed state than in the
food-deprived
state. Glycerol-3-phosphate dehydrogenase (GPDH) ... is
ubiquitously
expressed. Expression in heart decreased with starvation and
increased with
refeeding, whereas expression in liver did not change with
starvation. ...

... Here we report ... GLUT4, GLUT2 and GPDH ... response to
starvation. The
most important finding is that heart GLUT4 and liver GLUT2
expression
correlate with plasma glucose, whereas, white muscle GLUT4
does not.

... Fasting/re-feeding was used as a means to alter blood
glucose levels.
Atlantic cod, less than 1 year old ... One group was fed a
commercial diet
... while the other was deprived of food. ... Fed and
starved fish were
sampled after 1 and 2 months. The food-deprived fish were
then re-fed and
sampled after 20 days. ...

... The effects of fasting/re-feeding
Atlantic cod were deprived of food for 1 or 2 months and
thereafter re-fed
for 20 days. Control (i.e. fed) fish were sampled at the
same time as the 1
or 2 month food-deprived fish. There was no significant
difference in fish
length amongst the five groups (Fig. 5). After 2 months the
starved fish
weighed significantly less than the time matched controls
(P=0.014).
Re-feeding, following 2 months of starvation, did not lead
to a change in
body mass. There was no significant difference in condition
factor amongst
the five groups. Overall, the hepatosomatic index tended to
change during
the course of the study (P=0.06) with the starved groups
having lower mean
values than the time matched fed fish.

As there were no significant differences in blood glucose,
liver glycogen or
liver lipids between the two groups of fed fish these values
were pooled in
the following analysis. Blood glucose levels decreased from
9.4 µmol ml-1 to
approximately 2.3 µmol ml-1 following 1 month starvation
(Fig. 6). There was
no further change during the second month of food
deprivation. With
re-feeding, blood glucose levels increased to 6.7 µmol ml-1,
a value not
significantly different from the control group. Liver
glycogen levels
decreased from 100 µmol glucosyl units g-1 to 38 µmol
glucosyl units g-1
after 1 month and to 5.1 µmol glucosyl units g-1 following 2
months of
starvation. Twenty days of re-feeding led to increases in
glycogen levels to
172 µmol glucosyl units g-1, a value significantly higher
than all the other
groups. When values for blood glucose and liver glycogen
from all
individuals were considered, there was a strong correlation
between the two
variables. There were two fish in the control group with
blood glucose
levels of 239 and 202 µmol ml-1. As these values were so far
removed from
the norm they were not included in the analysis.

Neither total lipid nor triglyceride level in liver changed
even with 2
months of starvation (Fig. 7). Total lipid level was
approximately 500 mg
g-1, equivalent to 50% of the liver mass. Triglyceride
accounted for
approximately 90% of the total lipid pool.

Expression of GLUT4, GLUT2 and GPDH was assessed by qRT-PCR
in fish that had
been continuously fed, starved for 2 months, and re-fed for
20 days (Fig.
8). GLUT4 expression was analyzed in two tissues that highly
express GLUT4,
heart and white muscle. In heart, GLUT4 expression changed
significantly
(P=0.016) and was twofold higher in the fed fish than in the
starved fish.
When the food-deprived fish were re-fed, GLUT4 levels
doubled with respect
to starved levels returning to those seen in the control fed
fish. In white
muscle, GLUT4 expression also showed a significant change
(P=0.021) but in
an opposite pattern to that seen in heart. GLUT4 levels were
2.5-fold higher
in white muscle of starved fish than in control fed fish.
When food-deprived
fish were re-fed, GLUT4 levels dropped 1.8-fold, with
respect to levels seen
in the starved fish, returning to values that were not
significantly
different from those in the control fed fish. GPDH
expression was analyzed
in heart and liver and GLUT2 in liver only. The pattern of
GPDH expression
in heart showed significant change, similar to that of
GLUT4, in that GPDH
level was 1.7-fold lower in starved than in control fed
fish. GPDH
expression in heart then increased upon re-feeding to a
value not
significantly different from the control fed fish. In liver,
there was no
statistically significant difference in GLUT2 and GPDH
expression amongst
fed, food-deprived and re-fed groups.

When values for GLUT expression, from all individuals, were
plotted against
blood glucose, there was a significant correlation between
heart GLUT4 and
blood glucose and between liver GLUT2 and blood glucose
(Fig. 9). There was
no correlation between white muscle GLUT4 and blood glucose.

Discussion

... Impact of starvation on metabolic fuels and GLUT expression
The period of food deprivation was well within the tolerance
limits of
Atlantic cod as evidenced by maintenance of condition factor
and levels of
lipids in liver. However, the challenge resulted in
decreases in blood
glucose and liver glycogen, as previously reported (Black
and Love, 1986;
Hemre et al., 1990). Such starvation-induced decreases in
blood glucose are
associated with parallel decreases in plasma insulin (Hemre
et al., 1990;
Sunby et al., 1991). Although insulin was not measured in
the current
experiments, we assume that it decreased with starvation,
given that the
water temperature (5-8°C in previous studies; 8°C current
experiment) and
the length of starvation (3-4 weeks in previous studies; 4-8
weeks in
current experiment) were in the same range as in the earlier
work of Hemre
et al. (Hemre et al., 1990) and Sunby et al. (Sunby et al.,
1991).
Re-feeding led to a recovery of blood glucose levels and an
overshoot in
liver glycogen, again as reported by Black and Love (Black
and Love, 1986).
We note here for the first time a correlation between
glycogen content in
liver and blood glucose, suggesting that the former sets the
glucose level
available to other tissues.

Expression of GLUT4 in heart decreased during starvation,
increased with
re-feeding and correlated with blood glucose. The most
plausible explanation
for this is that high levels of blood glucose are associated
with elevated
levels of insulin and this in turn results in activation of
GLUT4
transcription. The scenario depicted for heart of Atlantic
cod matches that
in red muscle of brown trout, in which starvation is
associated with
decreases in blood insulin and glucose in association with
decreases in red
muscle GLUT4 (Capilla et al., 2002).

Expression of GLUT4 in white muscle was the mirror image of
that in heart.
Food deprivation resulted in an increase in mRNA levels,
which returned to
control values with re-feeding. GLUT4 expression in white
muscle did not
correlate with blood glucose. Differences between red muscle
and white
muscle, with respect to alterations in GLUT4 levels, have
been previously
noted in trout although the significance of it may have gone
unrecognized
(Capilla et al., 2002). GLUT4 mRNA levels decreased in red
muscle of starved
brown trout but in white muscle, although the there was no
significant
difference in expression level, the average value increased
by about 25%. In
rainbow trout injected with porcine insulin, there was an
increase in GLUT4
mRNA in red muscle but no change in white muscle. It appears
that the GLUT4
responsiveness to insulin and/or high blood glucose levels
noted in heart of
Atlantic cod and red muscle of trout does not carry over to
white muscle.
Indeed starvation is associated with increases in GLUT4
mRNA. An explanation
to account for this is that during starvation there is
proteolysis in white
muscle of Atlantic cod (Black and Love, 1986) that probably
includes GLUT4.
The data presented here are for GLUT 4 mRNA and not
transporter protein. In
order to maintain glucose transport there may need to be an
increase in
synthesis of this protein and this is reflected in increased
GLUT4
expression. A response similar to that reported here for
Atlantic cod has
been observed in rats, in which a 3-day fast led to a two-
to threefold
increase in GLUT4 transcription and GLUT4 mRNA in white
muscle but no change
in red muscle (Neufer et al., 1993).

GLUT2 mRNA levels correlated with blood glucose. In mammals,
GLUT2 serves to
facilitate glucose transport either into or out of liver
cells, dependent on
dietary and hormonal status. One scenario to account for the
data is that as
starvation proceeds, liver glycogen is depleted and delivery
of glucose from
liver to blood is diminished therefore reducing the need to
maintain high
levels of GLUT2 protein. In the only other study that we are
aware of,
regarding GLUT2 in fish, the transporter was highly
expressed in liver of
rainbow trout and the level was not influenced by 4 days of
food deprivation
(Panserat et al., 2001).

Glycerol-3-phosphate dehydrogenase
GPDH from Atlantic cod encodes a deduced amino acid sequence
that is very
similar to that found in a number of other fish species. The
enzyme GPDH
plays a pivotal role in the synthesis of glycerol
3-phosphate required for
triglyceride/phospholipid synthesis and in the conversion of
glycerol
3-phosphate to dihydroxyacetone in the breakdown of these
components. GPDH
expression was apparent at fertilization, increased by about
threefold by
halfway to hatching, and remained elevated throughout the
remainder of the
developmental period. In juvenile fish, GPDH was detected in
all tissues
sampled with the highest levels, typically occurring in
those tissues with
high lipid turnover, such as brain, intestine and liver. The
starvation
challenge was not substantive enough to induce changes in
liver triglyceride
levels. In this context it is not surprising that GPDH mRNA
levels did not
change in liver. GPDH expression in heart decreased with
starvation and
recovered during re-feeding. The simplest explanation for
this is a decrease
in phospholipid/triglyceride synthesis in heart during
starvation associated
with decreased synthesis of glycerol 3-phosphate.

Conclusion
In summary, GLUT4, GLUT2 and GPDH cDNAs from Atlantic cod
were cloned and
sequenced. In accordance with the mammalian model GLUT4 is
expressed
primarily in heart and muscle, whereas GLUT2 is expressed at
the highest
levels in liver. GPDH is expressed in all tissues assessed,
with highest
levels in tissues that have high rates of lipid turnover.
Starvation was
associated with decreases in blood glucose, liver glycogen
and heart GLUT4
and GPDH. These parameters recovered with re-feeding. By
contrast, white
muscle GLUT4 increased with starvation and returned to
pre-starved levels
with re-feeding. The physiological significance of this is
yet to be
resolved. GLUT2 expression in liver correlated with blood
glucose levels,
probably reflecting glycogen depletion in liver during
starvation with
reduced movement of glucose from liver to blood.