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Amino acid uptake and incorporation into rat pituitary as affected by thyroidectomy and thyroxine administration
Vol.26, I%.$1967
BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS
AMINO ACID UPTAKE AND INCORPORATIONINTO RAT PITUITARY AS AFFECTEDBY THYROIDECTOMYAND THYROXINE ADMINISTRATION Teiichiro Department
Tonoue and Kiyoshi
of Physiology,
Yamamoto
Institute
Gunma University,
of Endocrinology,
Maebashi,
Japan
Received December 28, 1966 Since Hoskins proposed the 'feed-back' a homeostatic
control
been extensively
between the pituitary
investigated
have been many reports stimulating Suzuki,
concerning
Recently
TSH stimulates
protein
hand, the action studied
to confirm
pituitary
of action
19641,
There of thyroid 1964;
described
that
On the other has been
effect,
--e.g., thyroid changes of the
from or morphological
(see Purves,
has
(see Rail et al.,
hormones on the pituitary
only with the use of an over-all
changes, TSH release
the hypothesis.
Raghupathy --et al. (1963) synthesis of the thyroid.
of thyroid
in 1949,
and the thyroid
the sites
hormone (TSH) in the thyroid
1966).
hypothesis
and the sites
of the action
have
not been elucidated. Thyroid in the liver
hormone has been known to enhance protein
synthesis
(Sokoloff
1963;
Buchanan and Tapley, 1963)
and skeletal
and Kaufman, 1961.; Tata et al., 19661,
heart
and kidney
muscle (Brown, 1966).
(Michels
& &.,
In view of the peculiar
interrelationship between the thyroid and pituitary, it can be presumed that thyroid hormone depresses protein synthesis of the
315
Vd. 26, No. 3,1967
pituitary.
AND RJOMYSICAL RESEARCH COMMUNICATIONS
The present
is really into
-1
the case.
rat pituitary
stimulation
experiment
showed that the presumption
Both uptake and incorporation were stimulated
by.thyroidectomy
was depressed by thyroxine
Material
and Methods:
performed
surgically
administered
experiments.
2 to 3 weeks before
by a subcutaneous
100 g body weight. the pituitary
and the
administration.
Young Wistar rats
130 to 180 g were used for all
of amino acids
of body weights Thyroidectomy
experiments.
injection
was
T4 was
at a dose of 20 Dg per
Animals were sacrificed
by.decapitation
cleaned and incubated
was dissected,
of
and
with amino
acids. Experiments
on amino acid uptake were carried
method of Wool gt al. pituitary
was incubated
acid (AIB), Center,
(1965) with slight
accumulated
After
PC of AIB-1-14C
incubation
by the pituitary
water extract
for
counts in an unit
Experiments proteins
out after
modifications.
Amersham).
were extracted
protein
After
that is,
volume of tissue
for
incubation
the ratio
water to the pituitary
pituitary
was incubat-
m&uoles)(Radiochemical for
60 min at 37OC, proteins
was counted for radioactivity. was applied
selfabsorption
of the extract
into
anterior
'%(U)(5.88
25 Pg protein
corrections content
ratio,
were
the method of Wool & g&. (1965)
Sliced
and the extract
For the counting, therefore
The results
on amino acid incorporation
ed with 0.5 Ire of L-alanineCenter,
AIB
medium of the same volume.
were carried
with slight
(Radiochemical
with water and the
radioactivity.
expressed in terms of AIB distribution counts in incubation
The whole
60 min at 37'C,
was extracted
was counted for
of radioactivity
modifications.
the
with 0.25 Pmoles of o-aminoisobutyric
which contained'0.2
Amersham).
out after
was unnecessary.
was determined
316
to a planchet,
by measuring
The its
Vol. 26, No. $1967
turbidity
UOCHEMXL
after
concentration
AND MOPHYSlCAl
the addition
RESEARCH COMMUNICATIONS
of trichloroacetic
acid at a final
of 35%.
Results:
A representative
thyroidectomy
and thyroxine
shown in Table 1. This difference
result
of the effect
of
supplement on the uptake of AIB is
Thyro'idectomy
increased
was statistically
the uptake by 28%.
significant.
When T4 was
TABLE I JXf e cto
$m U
atake by rat nituitary
No. of animals
Treatment
'*lght Of pituitary
Distribution
ratio
of AIB-1-14C
Control
4
5.8
2.96 2 0.08*
Thyroidectomy
4
7.0
3.57
z 0.07
Thyroidectomy
+ T4 injection
3 hr
4
7.3
3.37
i 0.07*
12 hr
4
7.0
2.68
2 o&t*
*
Difference
administered
from thyroidectomy,
to the thyroidectomized
uptake by thyroidectomy injection
tion After
proteins
was stimulated
stimulation
AIB
3 hr after
than the level
significantly
T4
of the normal
on incorporation
The alanine
by thyroidectomy depressed 3 hr after
incorporation
These results
experiment
of the pituitary.
was significantly
12 hr, alanine
animals.
the stimulated
T4 injection.
Table 2 shows a representative into
rats,
decreased significantly
and the uptake was lower
rats 12 hr after of alanine
p
by 35%.
The
T4 injection.
was the same as in control
were confirmed
317
incorpora-
by repeated
experiments.
vol. 26,No.3, 1967
BIOCHEMICAL
AND BIOPHYSICAL RESEARCH COMMUNICATIONS
TABLE II Effect
of thyroidectomy
and TI, administration
No. of animals
Treatment
on alanine
L-Alanine-14C incorporation (cpm/mg protein)
Control
6
21960 t 195*
Thyroidectomy
6
29518
2 264
Thyroidectomy
*
+ T4 injection
3h
6
23210
: 264*
12 hr
6
22001
2 1048*
Difference
from thyroidectomy,
p
It should be noted here that both AIB uptake and alanine incorporation with other alanine
were very active tissues.
experimental
was about 70 times greater and diaphragm, The results
that thyroidectomy is
and
in the pituitary
when compared in the same
conditions.
Discussions tion
when compared
BIB uptake was about 5 times greater
incorporation
than in the liver
in the pituitary
described
above show clearly
causes a stimulation
of amino acids into
the pituitary
of uPtake and incorporaand that the stimulation
depressed by T4 supplement. It is well known that in hypothyroidism
TSH from the pituitary discharge
decreases
increases
the discharge
and in hyperthyroidism
(van Rees, 1966).
However,
suggest strongly
that the thyroid
the
the detailed
mechanisms do not appear to have been made clear. results
of
regulates
The present the pituitary
BIOCHEMICAL
Vol. 26, No. 3, 1967
function thyroid
through
a stimulatory
the effect
consistent thyroid effect
synthesis.
hormones on protein
pituitary, that
protein
is not clear its
in tissues
feed-back
other
is inhibitory relationship
High rates
of the pituitary
indicate
hormones of protein
of
than the
The fact is between the
though the mechanisms of this
at present.
trophic
the effect
has been reported.
of T4 upon the pituitary
and the pituitary,
synthesize
Concerning
synthesis
effect
with the peculiar
and incorporation
AN0 RIOPHYSICAL RESEARCH COMMUNICATIONS
reversed
of amino acid uptake the high potency to or polypeptide
nature.
References
Rail,
J. E., Robbins, J. and Lewallen, C. G. (1964) in Pincus, G., Thimann, K. V. and Astwood, E. B. (eds), The Hormones, Vol. V, Academic Press, New York 159. Sokoloff, L. and Kaufman, S. 11961) J. Biol. Chem.,&, 795. Suzuki M. (1966) Metabolism and Desease, , 796 (In Japanese). Tata, 5. R., Ernster L. and Lindberg, 0. Biophys Acta 51 61!ly63) Biochimm van Rees, G. 'P (l&A%*Endocr Wool, I. G., Cistels, J. J. and Moyer,'k. N: (1965) Biochim. Biophys. Acta, l.Q& 333.
319
BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS
AMINO ACID UPTAKE AND INCORPORATIONINTO RAT PITUITARY AS AFFECTEDBY THYROIDECTOMYAND THYROXINE ADMINISTRATION Teiichiro Department
Tonoue and Kiyoshi
of Physiology,
Yamamoto
Institute
Gunma University,
of Endocrinology,
Maebashi,
Japan
Received December 28, 1966 Since Hoskins proposed the 'feed-back' a homeostatic
control
been extensively
between the pituitary
investigated
have been many reports stimulating Suzuki,
concerning
Recently
TSH stimulates
protein
hand, the action studied
to confirm
pituitary
of action
19641,
There of thyroid 1964;
described
that
On the other has been
effect,
--e.g., thyroid changes of the
from or morphological
(see Purves,
has
(see Rail et al.,
hormones on the pituitary
only with the use of an over-all
changes, TSH release
the hypothesis.
Raghupathy --et al. (1963) synthesis of the thyroid.
of thyroid
in 1949,
and the thyroid
the sites
hormone (TSH) in the thyroid
1966).
hypothesis
and the sites
of the action
have
not been elucidated. Thyroid in the liver
hormone has been known to enhance protein
synthesis
(Sokoloff
1963;
Buchanan and Tapley, 1963)
and skeletal
and Kaufman, 1961.; Tata et al., 19661,
heart
and kidney
muscle (Brown, 1966).
(Michels
& &.,
In view of the peculiar
interrelationship between the thyroid and pituitary, it can be presumed that thyroid hormone depresses protein synthesis of the
315
Vd. 26, No. 3,1967
pituitary.
AND RJOMYSICAL RESEARCH COMMUNICATIONS
The present
is really into
-1
the case.
rat pituitary
stimulation
experiment
showed that the presumption
Both uptake and incorporation were stimulated
by.thyroidectomy
was depressed by thyroxine
Material
and Methods:
performed
surgically
administered
experiments.
2 to 3 weeks before
by a subcutaneous
100 g body weight. the pituitary
and the
Young Wistar rats
130 to 180 g were used for all
of amino acids
of body weights Thyroidectomy
experiments.
injection
was
T4 was
at a dose of 20 Dg per
Animals were sacrificed
by.decapitation
cleaned and incubated
was dissected,
of
and
with amino
acids. Experiments
on amino acid uptake were carried
method of Wool gt al. pituitary
was incubated
acid (AIB), Center,
(1965) with slight
accumulated
After
PC of AIB-1-14C
incubation
by the pituitary
water extract
for
counts in an unit
Experiments proteins
out after
modifications.
Amersham).
were extracted
protein
After
that is,
volume of tissue
for
incubation
the ratio
water to the pituitary
pituitary
was incubat-
m&uoles)(Radiochemical for
60 min at 37OC, proteins
was counted for radioactivity. was applied
selfabsorption
of the extract
into
anterior
'%(U)(5.88
25 Pg protein
corrections content
ratio,
were
the method of Wool & g&. (1965)
Sliced
and the extract
For the counting, therefore
The results
on amino acid incorporation
ed with 0.5 Ire of L-alanineCenter,
AIB
medium of the same volume.
were carried
with slight
(Radiochemical
with water and the
radioactivity.
expressed in terms of AIB distribution counts in incubation
The whole
60 min at 37'C,
was extracted
was counted for
of radioactivity
modifications.
the
with 0.25 Pmoles of o-aminoisobutyric
which contained'0.2
Amersham).
out after
was unnecessary.
was determined
316
to a planchet,
by measuring
The its
Vol. 26, No. $1967
turbidity
UOCHEMXL
after
concentration
AND MOPHYSlCAl
the addition
RESEARCH COMMUNICATIONS
of trichloroacetic
acid at a final
of 35%.
Results:
A representative
thyroidectomy
and thyroxine
shown in Table 1. This difference
result
of the effect
of
supplement on the uptake of AIB is
Thyro'idectomy
increased
was statistically
the uptake by 28%.
significant.
When T4 was
TABLE I JXf e cto
$m U
atake by rat nituitary
No. of animals
Treatment
'*lght Of pituitary
Distribution
ratio
of AIB-1-14C
Control
4
5.8
2.96 2 0.08*
Thyroidectomy
4
7.0
3.57
z 0.07
Thyroidectomy
+ T4 injection
3 hr
4
7.3
3.37
i 0.07*
12 hr
4
7.0
2.68
2 o&t*
*
Difference
administered
from thyroidectomy,
to the thyroidectomized
uptake by thyroidectomy injection
tion After
proteins
was stimulated
stimulation
AIB
3 hr after
than the level
significantly
T4
of the normal
on incorporation
The alanine
by thyroidectomy depressed 3 hr after
incorporation
These results
experiment
of the pituitary.
was significantly
12 hr, alanine
animals.
the stimulated
T4 injection.
Table 2 shows a representative into
rats,
decreased significantly
and the uptake was lower
rats 12 hr after of alanine
p
by 35%.
The
T4 injection.
was the same as in control
were confirmed
317
incorpora-
by repeated
experiments.
vol. 26,No.3, 1967
BIOCHEMICAL
AND BIOPHYSICAL RESEARCH COMMUNICATIONS
TABLE II Effect
of thyroidectomy
and TI, administration
No. of animals
Treatment
on alanine
L-Alanine-14C incorporation (cpm/mg protein)
Control
6
21960 t 195*
Thyroidectomy
6
29518
2 264
Thyroidectomy
*
+ T4 injection
3h
6
23210
: 264*
12 hr
6
22001
2 1048*
Difference
from thyroidectomy,
p
It should be noted here that both AIB uptake and alanine incorporation with other alanine
were very active tissues.
experimental
was about 70 times greater and diaphragm, The results
that thyroidectomy is
and
in the pituitary
when compared in the same
conditions.
Discussions tion
when compared
BIB uptake was about 5 times greater
incorporation
than in the liver
in the pituitary
described
above show clearly
causes a stimulation
of amino acids into
the pituitary
of uPtake and incorporaand that the stimulation
depressed by T4 supplement. It is well known that in hypothyroidism
TSH from the pituitary discharge
decreases
increases
the discharge
and in hyperthyroidism
(van Rees, 1966).
However,
suggest strongly
that the thyroid
the
the detailed
mechanisms do not appear to have been made clear. results
of
regulates
The present the pituitary
BIOCHEMICAL
Vol. 26, No. 3, 1967
function thyroid
through
a stimulatory
the effect
consistent thyroid effect
synthesis.
hormones on protein
pituitary, that
protein
is not clear its
in tissues
feed-back
other
is inhibitory relationship
High rates
of the pituitary
indicate
hormones of protein
of
than the
The fact is between the
though the mechanisms of this
at present.
trophic
the effect
has been reported.
of T4 upon the pituitary
and the pituitary,
synthesize
Concerning
synthesis
effect
with the peculiar
and incorporation
AN0 RIOPHYSICAL RESEARCH COMMUNICATIONS
reversed
of amino acid uptake the high potency to or polypeptide
nature.
References
Rail,
J. E., Robbins, J. and Lewallen, C. G. (1964) in Pincus, G., Thimann, K. V. and Astwood, E. B. (eds), The Hormones, Vol. V, Academic Press, New York 159. Sokoloff, L. and Kaufman, S. 11961) J. Biol. Chem.,&, 795. Suzuki M. (1966) Metabolism and Desease, , 796 (In Japanese). Tata, 5. R., Ernster L. and Lindberg, 0. Biophys Acta 51 61!ly63) Biochimm van Rees, G. 'P (l&A%*Endocr Wool, I. G., Cistels, J. J. and Moyer,'k. N: (1965) Biochim. Biophys. Acta, l.Q& 333.
319