Endocrinology Vol. 138, No. 12 5231-5237
Copyright © 1997 by The Endocrine Society
Thyroid Hormones Inhibit Type 2 Iodothyronine Deiodinase in the Rat Cerebral Cortex by Both Pre- and Posttranslational Mechanisms1
Lynn A. Burmeister,
John Pachucki and
Donald L. St. Germain
Division of Endocrinology and Metabolism, Department of Medicine,
University of Pittsburgh School of Medicine (L.A.B., J.P.), Pittsburgh,
Pennsylvania 15261; and the Department of Medicine and Physiology,
Dartmouth Medical School (D.L.S.G.), Lebanon, New Hampshire 03756
Address all correspondence and requests for reprints to: Dr. Lynn A. Burmeister, Division of Endocrinology and Metabolism, Department of Medicine, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15261
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Abstract
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The type 2 5'-deiodinase (D2) appears to play an important role in
maintaining the intracerebral T3 content relatively
constant during changes in thyroidal state. Previous studies have
demonstrated that the regulation of this enzyme by thyroid hormone and
its analogs occurs at a posttranslational level. The availability of
the rat D2 complementary DNA now allows an assessment of whether
pretranslational regulation of this enzyme also occurs in the cerebral
cortex. In rats rendered hypothyroid by the addition of methimazole to
the drinking water, D2 messenger RNA (mRNA) is increased 70%
(P = 0.03). Treatment with
L-T3 (50 µg/100 g BW) for 4 days results in
an 80% decrease in D2 mRNA compared with that in euthyroid controls
(P < 0.001). Administration of lower doses of
L-T3 (0.253 µg/100 g BW·day) is
associated with a dose-dependent decrease in cortical D2 mRNA, but
little or no change in D2 activity. The decrease in D2 mRNA in response
to T3 treatment can be demonstrated within 4 h.
Treatment of hypothyroid rats for 2 weeks with graded doses of
L-T4 (0.11.5 µg/100 g BW·day) results in
a significant decrease in cortical D2 activity, but not mRNA. The
association between D2 activity and D2 mRNA in euthyroid, hypothyroid,
and hormone-treated rats across a full range of thyroidal states
suggests that L-T4 treatment is associated with
greater changes in cortical D2 activity (via posttranslational effects)
than mRNA, whereas L-T3 treatment has a greater
effect on decreasing D2 mRNA (i.e. pretranslational
effects). In conclusion, these studies demonstrate both pre- and
posttranslational regulation of cortical D2 expression. The relative
contribution of each mechanism depends on the ambient thyroid hormone
concentration.
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Introduction
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IN THE hypo- or hyperthyroid state, whole
brain T3 content is altered to a considerably
lesser extent than observed in other organs, such as the liver (1, 2).
This relative stability of brain T3 level appears
to be due at least in part to autoregulatory mechanisms within the
brain itself that regulate both T4 to
T3 conversion (3, 4, 5) and the metabolism of
T4 and T3 to inactive
compounds (6). For example, the fractional conversion of
T4 to T3 in brain is
increased in the hypothyroid state, whereas it is decreased by this
condition in the liver (1, 2, 5, 7).
The type 2 iodothyronine 5'-deiodinase (D2) appears to be an important
factor in the ability of the central nervous system to adapt to
alterations in thyroid hormone status. This enzyme, which is present in
the anterior pituitary gland and brown adipose tissue as well as in
several regions of the brain (8, 9, 10), deiodinates
T4 at the 5'-position to form
T3 and appears to be the principal source of
intracellular T3 in these tissues (3, 4, 5, 7, 11).
D2 activity is rapidly and markedly altered by changes in thyroid
hormone levels; hypothyroidism results in enhanced activity, whereas
the opposite effect occurs in hyperthyroidism (9, 12, 13). Furthermore,
it has recently been reported that 90% of T3
production in hypothyroid rats occurs in slowly exchanging tissues,
such as brain and brown adipose tissue (BAT), where D2 predominates as
the enzyme catalyzing T4 to
T3 conversion (14).
Several lines of evidence strongly suggest that posttranslational
mechanisms are involved in the regulation of D2 activity by thyroid
hormone (15, 16, 17, 18, 19, 20, 21, 22). Specifically, thyroid hormones have been observed
both in vivo in rodents and in several cell culture systems
to induce a decrease in D2 activity that is more rapid than that
observed when gene transcription or protein synthesis is blocked with
actinomycin D or cycloheximide, respectively (15, 18, 19). In addition,
T4 and rT3 are considerably
more potent that T3 in inducing inactivation of
D2 when administered acutely to hypothyroid animals (16, 18, 23, 24).
Given that transcriptional regulation by thyroid hormone is primarily,
if not exclusively, mediated by T3 binding to its
nuclear receptors (25), the greater potency of T4
and rT3 in acutely inhibiting D2 activity is
further evidence that this process involves extranuclear metabolic
events such as alterations in the rate of D2 protein turnover or
translocation of the enzyme between various cellular compartments
(19, 20, 21, 26).
In the present studies, we sought to determine whether the regulation
of D2 in cerebral cortex might involve pretranslational as well as
posttranslational mechanisms. A direct assessment of this possibility
has only recently become feasible with the cloning of complementary
DNAs (cDNAs) for this enzyme from several species (13, 27). Indeed,
initial studies in rats have demonstrated significant changes in D2
messenger RNA (mRNA) levels in BAT and the anterior pituitary gland in
response to alterations in thyroid hormone status (13). To assess the
pretranslational regulation of D2 in the cerebral cortex by thyroid
hormone, D2 mRNA and activity levels were compared in euthyroid and
hypothyroid rats and in animals treated with graded doses of
L-T3 or
L-T4.
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Materials and Methods
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Materials
[
-32P]deoxy-CTP,
[125I]rT3, and Gene Screen
membranes were purchased from New England Nuclear (Boston, MA).
Decaprime II oligolabeling kits were purchased from Ambion (Austin,
TX). L-T4,
L-T3, and methimazole (MMI) were
purchased from Sigma Chemical Co. (St. Louis, MO). Reagents for
determining protein concentration were obtained from Bio-Rad
Laboratories (Richmond, CA).
Animals and treatments
All treatments were approved and in compliance with the
requirements of the animal care and use committee of the University of
Pittsburgh or Dartmouth Medical School. Male Sprague-Dawley rats,
weighing 175200 g, were purchased from Zivic Miller (Zelienople, PA),
Harlan Sprague-Dawley (Indianapolis, IN), or Charles River Laboratories
(Burlington, MA). Rats had free access to water and food at all times
during the study. Hypothyroidism was induced by the addition of MMI
(0.025% or 0.05%) to the drinking water. Hypothyroidism was assessed
clinically by failure to gain weight at the expected rate and could be
observed within 2 weeks of beginning MMI treatment.
In an initial study, the experimental groups (n = 6 rats/group)
included euthyroid control animals, animals receiving MMI for 4 weeks,
and euthyroid animals treated with
L-T3 (50 µg/100 g BW, sc, daily)
for 4 days before death to induce a hyperthyroid state.
A second series of experiments was conducted using a longer period of
thyroid hormone treatment. Rats taking MMI for 2 weeks were continued
on this agent for an additional 2 weeks, during which time they were
administered once daily ip injections of
L-T4,
L-T3, or 10 mM sodium
hydroxide vehicle. Rats treated with
L-T4 received daily ip doses of 0.1
(n = 6), 0.2 (n = 6), 0.5 (n = 6), 1.5 (n = 10), 3
(n = 5), 4.5 (n = 5), or 10 µg/100 g BW (n = 4).
Treatment with L-T3 involved
administering doses of 0.25 (n = 10), 1 (n = 6), 1.7 (n
= 4), 3 (n = 4), or 5 (n = 4) µg/100 g BW. Sixteen rats
receiving MMI, treated only with vehicle, served as hypothyroid
controls. Six euthyroid rats served as additional controls. The chosen
doses of administered thyroid hormone were based on previous reports
suggesting that L-T4 at 510
µg/100 g BW·day induces thyrotoxicosis (28, 29). Furthermore,
T3 nuclear receptor saturation with an
accompanying alteration in the transcriptional activity of specific
liver genes is induced with L-T3
treatment in a dose of 1550 µg/day (30) or with a single dose of
200 µg T3 (31). Serum T4,
serum T3, T3 uptake, and
cortical D2 mRNA concentrations were measured at the time of death in
all chronically treated animals. Hepatic type I iodothyronine
deiodinase (D1) mRNA and activity and cerebral cortex D2 activity were
assayed in selected samples, including euthyroid rats and those
receiving MMI receiving vehicle or low dose
L-T3 or
L-T4 treatment (dosage range, 0.13
µg/100 g BW).
The time course of thyroid hormone effects on D2 mRNA and activity was
determined using rats rendered hypothyroid by the inclusion of MMI in
their drinking water for 50 days. At the end of this period, animals
were administered vehicle, 50 µg
L-T4 (n = 5), or 200 µg
L-T3 (n = 5) as a single ip dose
24 h before death. This experiment was repeated in MMI-treated
rats who received vehicle or 200 µg
L-T3 4, 12, or 24 h before
death, and results were compared with those from a euthyroid control
group (n = 4/group). Serum T4, serum
T3, T3 uptake, and cortical
D2 mRNA and activity were measured in all animals.
Rats were killed in the morning (24 h after the last dose of
L-T3 or
L-T4, or as otherwise noted) under
Nembutal anesthesia by exsanguination through the carotid arteries.
Brains were quickly removed, and the cortex was dissected out on ice
for storage at -80 C until RNA extraction or homogenization for
activity.
Isolation and analysis of RNA
In the initial experiment, RNA was isolated, and Northern
analysis was performed according to previously described procedures
(13) using 20 µg total RNA from the cerebral cortex of individual
rats. Final wash conditions used 0.1 x SSPE (sodium chloride,
sodium phosphate, EDTA)-0.1% SDS at 60 C for 1 h. Quantitation of
the hybridization signal was performed by phosphorimaging. The signal
was corrected for differences in loading and transfer by reprobing the
blot with a rat ß-actin probe and determining the ratio of the D2
band to the ß-actin band for each lane.
In other experiments, RNA extraction was performed by a modification of
the guanidine thiocyanate method (32) and was quantitated by reading
the optical density at 260
. Using 20 µg total RNA/lane, agarose
gel electrophoresis was performed under denaturing conditions (33). The
RNA was transferred overnight via capillary action to a nylon membrane
and UV cross-linked (34). An XbaI fragment of the rat D2
(13) or a PvuII fragment of the rat D1 cDNA (35) was labeled
to high specific activity using the random primer technique and then
hybridized to the membrane at 43 C for 2448 h (36, 37). Membranes
were washed at 63 C (for D2) or at 50 C (for D1) for a minimum of 40
min each in 2 x SSC (standard saline citrate)-0.1% SDS, 25
mM NaHPO4 (pH 7.2)-1 mM EDTA-0.1%
SDS, and 25 mM NaHPO4 (pH 7.2)-1 mM
EDTA-1% SDS. Quantitation of mRNA density was performed by
phosphorimaging the specific band with normalization to the 28S
ribosomal RNA band to correct for variations in loading. The 28S band
was visualized by ethidium bromide staining and photographed with type
665 positive-negative Polaroid film, and the density was quantitated by
densitometry of the negative (38, 39). The data are reported in
relative units corrected for the amount of 28S RNA present in each
lane. The relative level of D2 mRNA between gels and experiments was
standardized to either a rat brain or liver standard placed on all
gels.
Measurement of 5'-deiodinase activity
A portion of the cerebral cortex from individual animals was
homogenized (1:5, wt/vol) in 0.25 M sucrose and 0.02
M Tris-HCl, pH 7.4, and assayed for D2 activity using 1
nM radiolabeled rT3 as substrate and
in the presence of 20 mM dithiothreitol, 1 mM
EDTA, and 0.1 mM 6n-propyl-2-thiouracil, as
previously described (13). D2 activity is expressed as
femtomoles/h·mg protein. Livers were homogenized at 1:10 in the same
buffer, diluted 1:100, and assayed for D1 activity as described above,
except that no 6n-propyl-2-thiouracil was included in the
assay mixture. For a given experiment, all determinations were made in
a single assay, with duplicate determinations for each sample. D1
activity is expressed as picomoles/h·mg protein. Protein
concentrations were determined by the method of Bradford (40).
Hormone measurements
Serum total T4, total
T3, and T3 uptake were
measured by RIA (Coat-A-Count, Diagnostic Products Corp., Los Angeles,
CA). A rat thyroid hormone binding ratio was obtained by normalizing
the T3 uptake determination to a rat euthyroid
standard. Free T4 index and free
T3 index were calculated by multiplying the serum
total T4 or total T3,
respectively, by the thyroid hormone binding ratio. The
T4 and T3 concentrations
obtained in the study are expressed as micrograms per dl and nanogram
per ml, respectively. The free T4 index and free
T3 index are without units.
Statistical analysis
Statistical analysis was performed in some cases using
log-transformed data. Where appropriate, linear regression was
performed or comparisons between groups were assessed by ANOVA, with
appropriate post-hoc mean comparisons by the method of Tukey
(41) (Systat Program, Evanston, IL) or Dunnetts test for multiple
comparisons with a control group. P < 0.05 is
considered significant.
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Results
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Using a rat D2 cDNA probe, Northern analysis of total RNA from rat
cerebral cortex identified a predominant and prominent band
approximately 7 kilobases in length (Fig. 1a
). This is the same size RNA species
identified previously on Northern blots of BAT, anterior pituitary, and
cerebral cortex using polyadenylated RNA (13). Quantitation of this
7-kilobase band by phosphorimager analysis permitted a comparison of
the effects of hormone treatments on D2 mRNA levels. Representative
blots of cerebral cortical RNA samples from hypothyroid rats and
animals treated with L-T3 and
L-T4 are shown in Fig. 1
, b and
c.

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Figure 1. Expression of D2 mRNA in rat cerebral cortex. a,
Lane 1, One hundred and fifty nanograms of polyadenylated RNA from
hypothyroid rat cerebral cortex. Lane 2, Twenty micrograms of total RNA
from hypothyroid rat cortex. Molecular size markers and the 28S and 18S
ribosomal RNA bands are indicated. b, Relative levels of D2 mRNA in
cerebral cortex. Lanes 1 and 2, L-T3 (5
µg/100 g BW·day)-treated rats (5237 ± 800 relative units);
lanes 3 and 4, MMI-treated rats (9665 ± 2621 relative units);
lanes 5 and 6, L-T4 (10 µg/100
g·day)-treated rats (4744 ± 303 relative units). The data are
corrected for density of the 28S ribosomal band by ethidium bromide
staining. Shown are the mean ± SEM. c, Ethidium
bromide staining of 28S and 18S RNA from the samples shown in b.
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In the initial experiment, D2 mRNA levels were compared in euthyroid
rats, hypothyroid rats, and rats rendered hyperthyroid by the
administration of high doses of L-T3
(50 µg/100 g BW·day) for 4 days. As shown in Fig. 2
, hypothyroidism was associated with a
70% increase in cortical D2 mRNA levels compared with those in
euthyroid control animals (P = 0.03). In contrast,
T3 administration for 4 days resulted in an 80%
decrease in mRNA levels compared with levels in this control group
(P < 0.001). Likewise, as shown in Table 1
, administration of
T3 (5 µg/100 g BW·day) or
T4 (10 µg/100 g BW·day) for 14 days to
MMI-treated rats resulted in 59% and 71% decreases, respectively, in
the cortical D2 mRNA level compared with that in the 28-day MMI-treated
hypothyroid group.

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Figure 2. Relative D2 mRNA level in cerebral cortex of
euthyroid, hypothyroid, and 4-day L-T3-treated
(50 µg/100 g BW·day) rats. *, P < 0.001
compared with hypothyroid. Shown are the mean ± SEM
(n = 6/group).
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To examine the effects of physiological levels of thyroid hormone on
the regulation of cortical D2 mRNA and activity, hypothyroid rats were
treated for 2 weeks with daily doses of vehicle or
L-T3 ranging from 0.253 µg/100 g
BW. A group of euthyroid animals served as an additional control group.
The administration of increasing doses of
L-T3 induced a significant increase
in the serum free T3 index (P <
0.001; Fig. 3a
). This was associated with
a dose-dependent decrease in cerebral cortex D2 mRNA (P
= 0.006; Fig. 3b
). However, L-T3
administration did not statistically alter the high levels of D2
activity observed in the hypothyroid state (P = 0.31;
Fig. 3c
). For comparison, hepatic D1 mRNA and activity were determined
in the same animals (Fig. 3
, d and e). As expected from prior studies
(35, 42, 43), D1 mRNA and activity progressively increased with
increasing doses of T3 (P <
0.001). Of note, significant increases in these parameters were noted
at the lowest dose of T3 administered (0.25
µg/100 g BW·day), whereas higher doses (3 µg/100 g BW·day) were
required to effect inhibition of D2 mRNA in the cerebral cortex.

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Figure 3. Effects of L-T3
administration on cortical and hepatic deiodinase expression.
Hypothyroidism was induced with MMI treatment for 28 days. Animals were
administered L-T3 (0.253 µg/100 g BW·day)
or vehicle by ip injection once daily during the last 2 weeks of
treatment. They were killed in the morning, 24 h after the last
dose of L-T3. Six euthyroid rats served as a
separate control group. a, Free T3 index; b, cerebral
cortex D2 mRNA; c, cerebral cortex D2 activity; d, hepatic D1 mRNA; e,
hepatic D1 activity. *, P < 0.001; ,
P < 0.05 (relative to zero dose hypothyroid
vehicle in post-hoc analysis). Values are the mean
± SEM (n = 6/group).
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T3 treatment rapidly decreased cerebral cortical
D2 mRNA levels. In an initial experiment, a single 200-µg dose of
L-T3 was administered ip to
hypothyroid rats 24 h before death. The D2 mRNA level decreased by
90% in the L-T3-treated animals
(hypothyroid, 2.76 ± 0.15; T3-treated,
0.27 ± 0.18; P = 0.003; n = 5/group). In a
second experiment, D2 mRNA and activity levels were determined 4, 12,
and 24 h after L-T3
administration. As shown in Fig. 4a
, the
free T3 index was increased markedly by 4 h
after L-T3 administration and
remained elevated for 24 h (P < 0.001).
Associated with this, the D2 mRNA level was significantly decreased at
4 h (P < 0.01; Fig. 4b
) and declined further at
1224 h (P < 0.001). This was associated with a
progressive decrease in D2 activity (P = 0.02; Fig. 4c
)
that appeared to be somewhat delayed and of lesser magnitude than the
changes in the D2 mRNA level.

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Figure 4. Time course of response of hypothyroid rats to
T3 treatment. Rats were treated with MMI for 42 days and
then a single ip dose of 200 µg L-T3 or
vehicle was administered. All rats were killed between 08001200. a,
Free T3 index; b, cortex D2 mRNA; c, cortex D2 activity. *,
P < 0.001; , P < 0.01
(compared with hypothyroid vehicle-treated group in
post-hoc analysis). Values are the mean ±
SEM (n = 4/group).
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The effects of chronic L-T4 treatment
on cortical D2 and hepatic D1 mRNA and activity were also examined. As
in the L-T3 experiment, hypothyroid
rats were administered vehicle or
L-T4 daily in doses ranging from
0.11.5 µg/100 g BW·day for 2 weeks. Animals were killed 24 h
after the last dose. Significant increases in free
T4 index and free T3 index
were observed in a dose-dependent fashion, with the largest dose
administered resulting in values at 24 h comparable to those
observed in the euthyroid control group (P < 0.001;
Fig. 5
, a and b). Although cerebral
cortical D2 activity significantly decreased (P <
0.001), cortical D2 mRNA remained relatively stable, with the exception
of an apparent increase in mRNA associated with the 0.1 µg/100 g
BW·day L-T4 dose that proved not to
be statistically significant (P = 0.13 vs.
hypothyroid rats). Again, as expected, hepatic D1 mRNA and activity
could be readily induced by L-T4
treatment (P < 0.001, Fig. 5
, a and c).

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Figure 5. Effects of L-T4
administration on cortical and hepatic deiodinase expression.
Hypothyroidism was induced with MMI treatment for 28 days. Animals were
administered L-T4 (0.13 µg/100 g BW·day)
or vehicle by ip injection once daily during the last 2 weeks of
treatment. They were killed in the morning, 24 h after the last
dose of L-T4. Six euthyroid rats served as a
separate control group. a, Free T4 index and free
T3 index; b, cerebral cortex D2 mRNA and activity; c,
hepatic D1 mRNA and activity. *, P < 0.001; ,
P < 0.05; ¥, P = 0.053
(compared with hypothyroid vehicle-treated rats). Values are the
mean ± SEM (n = 6/group).
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The relationship between cortical D2 mRNA and activity in euthyroid,
hypothyroid, and L-T4- and
L-T3-treated rats is illustrated in
Fig. 6
. These data represent the values
for the individual animals from the experiments depicted in Figs. 3
and 5
as well as other confirmatory experiments not shown. Here, it can be
seen that the transition from the euthyroid to the hypothyroid state is
associated with a relatively larger increase in D2 activity than in
mRNA concentration. When comparing the effects of
T4 vs. T3
treatment on these parameters, it is apparent that
T4 treatment is associated with large changes in
D2 activity relative to alterations in mRNA levels, whereas the
opposite occurs with T3 treatment, in that mRNA
levels are primarily affected.
The relationships between serum T3 levels and
cortical D2 mRNA and activity are shown in Fig. 7
. A highly significant inverse linear
relationship is observed between log D2 mRNA and the log serum free
T3 index (Fig. 7a
; r = 0.520 and
P < 0.001; r = 0.61 if we exclude the outlier),
with overlap in the values regardless of whether the animals were
treated with L-T4 or
L-T3. No such relationship was found
between log D2 mRNA and log serum free T4 index
(r = 0.115; P = 0.213). In contrast to Fig. 7a
, a
marked difference was noted in T4- and
T3-injected animals in the relationship between
the log D2 activity and the log free T3 index
(Fig. 7b
; T4-treatment: r = 0.74;
P < 0.001; T3 treatment: r
= 0.65; P < 0.001). Thus, for any given level of free
T3 index, D2 activity was significantly lower in
the T4-treated animals than in those given
T3. Of importance, the euthyroid control group
(indicated by the box in Fig. 7b
) falls along the line for
the T4-injected animals, suggesting that
T4 exerts the dominant controlling effect on D2
activity in the euthyroid animal.
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Discussion
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Previous studies using both in vivo and in
vitro model systems have clearly demonstrated the importance of
posttranslational mechanisms in the regulation of D2 activity by
thyroid hormones (15, 16, 17, 18, 19, 20, 21, 22). Significant effects of thyroid status on D2
mRNA levels in the anterior pituitary gland and BAT have also recently
been reported (13). The present studies extend these observations by
demonstrating that pretranslational regulation of D2 also occurs in the
rat cerebral cortex in response to altered thyroid status. Thus,
hypothyroidism resulted in a significant increase in D2 mRNA levels
compared with those in euthyroid control animals, whereas the
administration of either T3 or
T4 rapidly decreased transcript levels. These
changes in thyroid status were associated with alterations in D2 mRNA
levels over an approximately 10-fold range of values.
In confirmation of previous studies (16, 44), treatment of hypothyroid
rats with L-T4 resulted in a greater
degree of inhibition of D2 activity than did administration of similar
doses of L-T3. In addition,
L-T4 administration resulted in
greater proportional changes in D2 activity relative to alterations in
mRNA levels. For example, chronic
L-T4 administered to hypothyroid
animals at a dose of 0.2 µg/100 g BW·day resulted in a significant
inhibition of D2 activity, yet no change in D2 mRNA levels occurred.
Indeed, treatment with L-T4 at doses
as high as 1.5 µg/100 g BW·day did not significantly decrease
cortical D2 mRNA, whereas it had a profound effect on inhibiting D2
activity. Such observations are compatible with a predominantly
posttranslational mechanism of D2 inhibition by
L-T4. In contrast, treatment with
L-T3 resulted in larger changes in D2
mRNA levels than in activity. Thus,
L-T3 treatment at 3 µg/100 g
BW·day did not significantly decrease cortical activity, but it was
associated with a significant decrease in cortical D2 mRNA, suggesting
a primary effect of this thyroid hormone on pretranslational D2
regulation. However, at high doses of administered hormone [chronic
L-T3 (5 µg/100 g BW·day),
L-T4 (10 µg/100 g BW·day), or
L-T3 (200 µg), or
L-T4 (50 µg) as a single ip dose
24 h before death], the level of cerebral cortex D2 mRNA
suppression was the same with either
L-T4 or
L-T3 (data not shown).
The predominant effect of thyroidal status on regulating D2 activity
occurs in the transition from the euthyroid to the hypothyroid state,
whereas D2 mRNA levels appear to vary across the entire range of
thyroid hormone levels. Thus, the relative importance of the
pretranslational and posttranslational mechanisms in regulating D2
expression probably depends on the intracellular concentrations of both
L-T3 and
L-T4. These dual mechanisms of
regulating D2 expression may work in concert to ensure that D2
activity, and thus T4 to T3
conversion, is appropriate to maintain the cortex in a euthyroid state.
Of note, the posttranslational effects of
L-T4 on D2 activity have been
demonstrated to occur rapidly (in <2 h) (12, 15), whereas the effects
of L-T3 in inhibiting D2 mRNA appear
to take somewhat longer. Thus, the pretranslational mechanism of D2
regulation may assume greater importance over a more chronic time
frame. It is also interesting to speculate that a decrease in D2 mRNA
in the hyperthyroid state may, by decreasing translation of the D2
protein, allow for the conservation of selenocysteine so that it can be
directed into the synthesis of other essential selenoproteins, such as
D3, whose activity is increased in this condition.
Differences were noted in these studies in the apparent sensitivity of
cortical D2 and hepatic D1 to regulation by thyroid hormones. For
example, low doses of L-T3 that did
not affect cortical D2 activity or mRNA levels did induce significant
increases in hepatic D1 mRNA and activity. This finding may reflect the
observations of others that tissue concentrations of
T3 are higher in the liver than in the cerebral
cortex over a range of thyroid states (45, 46). Alternatively, these
differences may indicate an inherently greater sensitivity of hepatic
D1, compared with cortical D2, to pretranslational regulation by
L-T3. In contrast, the exquisite
sensitivity of D2 in the brain to posttranslational regulation by
L-T4 is evident by the marked
inhibition of D2 activity by administered doses of this hormone that
have little or no effect on hepatic D1 levels.
In conclusion, we have demonstrated that D2 in the rat cerebral cortex
is regulated by thyroid hormones at a pretranslational as well as a
posttranslational level. This provides an additional mechanism of
autoregulatory control over the activity of this important enzyme.
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Acknowledgments
|
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The authors thank Theresa Toczek, Walburga Croteau, and Elena
Martinez for excellent technical assistance; Mark OMalley and Dr. Jim
Goss for assistance with brain dissections; and Dr. Sidney Morris, Jr.,
for encouragement.
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Footnotes
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1 This work was supported by NIDDK Grant DK-0214703 (to L.A.B.) and
DK-42271 (to D.L.S.). 
Received July 2, 1997.
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