A physiological dose of triiodothyronine normalizes cardiac myosin adenosine triphosphatase activity and changes myosin isoenzyme distribution in semistarved rats

The possibility that the lowering of thyroid hormone levels which occurs in the nonthyroidal illness syndrome results in a hypothyroid state at the cardiac tissue level was examined in semistarved rats. Rats were fed 50% of their normal food intake in the form of a regular diet (R. diet) or low carbohydrate diet (L.C. diet) for 8 weeks. Animals semistarved for 8 weeks on the R. diet lost 42% of their body weight, while plasma T3 and T4 levels decreased by 45-50%. Semistarvation on the L.C. diet resulted in a 19% weight loss and a similar 46-49% decrease in plasma T3 and T4 levels. Ca++-activated myosin ATPase activity declined by 28% and 48% with the R. and L.C. diets, respectively [normal rats myosin ATPase, 1.30 +/- 0.18 mumol Pi/(mg protein . min) (mean +/- SD); semistarvation R diet, 0.93 +/- 0.15; semistarvation L.C. diet, 0.67 +/- 0.15]. The administration of physiological amounts of T3 (0.3 micrograms T3/100 g BW daily) restored the cardiac myosin ATPase activity in both groups. To confirm that the T3 effect was due to a normalization of the thyroid status at the tissue level, hypothyroid animals on a normal diet were injected with 0.3 micrograms T3 for 4 weeks, which resulted in normalization of myosin ATPase activity levels. Thyroidectomized rats receiving daily T3 injections, and when placed on a 50% reduction of food intake for 4 weeks still maintained normal myosin ATPase activity even though they lost 36% of their body weight. Distribution of cardiac myosin isoenzymes was determined by pyrophosphate polyacrylamide gel electrophoresis. In normal cardiac ventricles, myosin isoenzyme V1 predominates and represents 68 +/- 7% (+/- SD) of the total myosin. Semistarvation resulted in a redistribution of myosin isoenzymes so that V3 myosin was the predominant species (53 +/- 3% of the total myosin). The administration of 0.3 microgram T3/100 g BW daily for 4 weeks to semistarved rats reverted myosin isoenzyme distribution to V1 predominance (V1 myosin, 54 +/- 3% of the total myosin). These results indicate that the semistarvation-induced lowering plasma T3 and T4 levels is an important determinant of myosin ATPase activity and myosin isoenzyme distribution. Restoration of myosin ATPase activity to its normal level and return to myosin V1 predominance after T3 administration make it likely that these changes are related to the lowering of thyroid hormone levels.     

Myosin isoenzyme distribution and Ca+2-activated myosin ATPase activity in the rat heart is influenced by fructose feeding and triiodothyronine

Studies were conducted to determine if the level of cardiac Ca+2-activated myosin ATPase activity and ventricular myosin isoenzyme distribution are influenced by both T3 administration and fructose feeding. Previous studies have shown that in the cardiac ventricle of hypothyroid rats, only myosin V3 is present, and the Ca+2-activated myosin ATPase activity is markedly decreased. Hypothyroid [thyroidectomized (Tx)] rats were fed a diet containing 60% fructose or a regular diet (47% complex carbohydrates) for 4 weeks. Fructose feeding of hypothyroid rats led to a significant increase in Ca+2-activated myosin ATPase activity (Tx regular diet, 0.33 +/- 0.02 mumol Pi/mg protein X min; Tx fructose diet, 0.54 +/- 0.04 mumol Pi/mg protein X min). In addition, myosin V1 was detectable in the heart of fructose-fed Tx rats, but was absent in Tx rats on the regular diet. To determine if fructose had an effect of similar magnitude in animals of different thyroid states, Tx rats were injected with 0.075, 0.150, 0.225, and 0.300 micrograms T3/100 g BW daily and placed on fructose or regular diets. The fructose-induced increase in Ca+2-myosin ATPase activity was between 24-27% in Tx rats receiving 0-0.15 micrograms T3/100 g BW daily. In animals receiving 0.225 and 0.300 micrograms T3/100 g BW daily, fructose feeding did not induce a significant increase in myosin ATPase activity. This is due to the fact that the Ca+2-activated myosin ATPase activities of euthyroid and hyperthyroid animals are not significantly different from each other. In hypothyroid rats receiving a 60% glucose diet, Ca+2-myosin ATPase activity showed a significant 20% increase above the value in regular diet-fed Tx rats. Fructose- and glucose-induced changes in Ca+2-myosin ATPase activity occurred in the absence of changes in thyroid hormone or insulin levels. Our findings may indicate that cardiac carbohydrate consumption influences the predominance of ventricular myosin isoenzymes in the rat heart.     

Age dependent changes in myosin ATPase activity in the myocardium of spontaneously hypertensive rats

Male spontaneously hypertensive rats (SHR) and age matched Wistar Kyoto normotensive (WKY) rats of 5 weeks, 16 weeks, and 52 weeks of age were used to determine whether duration of hypertension has any effect on contractile protein ATPase and myosin isoenzyme distribution. Myofibrils, actomyosin, and myosin were isolated from the left ventricles of WKY rats and SHR and assayed for myosin ATPase activity and myosin isoenzyme distribution. Myofibrillar ATPase activity was assayed at various free [Ca++] ranging from 10(-7) to 10(-5) mol X litre-1. Ca++ stimulated actomyosin ATPase activity was determined at several Ca++ concentrations both at low ionic strength, which favours actin-myosin interaction, and at high ionic strength, which diminishes actin interaction with myosin. Purified myosin ATPase activity was assayed in the presence of K+-EDTA and in the presence of several concentrations of Ca++. Actin activated myosin ATPase activity was assayed using 26 mumol X litre-1 skeletal muscle actin. Under all these assay conditions no differences were observed in the contractile protein ATPase activity between SHR and WKY rats in any age group. On the other hand, in both SHR and WKY rats the contractile protein ATPase activity under all assay conditions was significantly decreased in 52 week old rats compared with 5 week old rats. The predominant myosin isoenzyme was Vi in 5 week and 16 week old WKY rats and SHR. In 52 week old WKY rats and SHR, however, significant amounts of isoenzymes V2 and V3 were present along with V1. Percentage distribution of V1, V2, V3 isoenzymes calculated from densitometric scans of gels did not show any differences between WKY rats and SHR in any age group. These results suggest that neither myosin ATPase activity nor myosin isoenzyme distribution is altered in the moderately hypertrophied left ventricles of SHR. Moreover, the data indicate that the myocardium of SHR, despite the persistence of pressure overload, undergoes a similar decrease in myosin ATPase activity and an increase in myosin isoenzyme V3 to age matched normotensive WKY rats.     

Changes in myosin isoenzymes, ATPase activity, and contraction duration in rat cardiac muscle with aging can be modulated by thyroxine

To determine whether the relative decline in cardiac myosin isoenzyme V1 with maturation continues progressively into senescence and whether thyroxine could reverse age-associated changes in the myosin isoenzyme profile and contraction, rats 2, 8, and 24 months old were treated with thyroxine, 6.4 mg/kg, for 7 days. Myosin isoenzymes, Ca2+-myosin ATPase activities, and isometric contractile function were measured in cardiac preparations from thyroxine-treated animals and age-matched controls. Right ventricular hypertrophy did not occur with aging in controls. Thyroxine increased right ventricular weight in each age group compared to the control group. Body weight decreased by 10% in all thyroxine-treated rats. The relative right ventricular V1 isoenzyme content progressively decreased from 75 +/- 1% to 54 +/- 1% and 14 +/- 1% in controls at 2, 8, and 24 months, respectively, and was associated with a reciprocal increase in V3 myosin isoenzyme. Ca2+-myosin ATPase activity also progressively declined monotonically with age in the control rats from 854 +/- 28 nmol Pi/mg prot/min at 2 months to 529 +/- 28 nmol Pi/mg prot/min at 24 months. Thyroxine administration increased right ventricular V1 at each age to 97 +/- 2%, 73 +/- 2%, and 59 +/- 2% at 2, 8, and 24 months, respectively. A thyroxine induced increase in the Ca2+-myosin ATPase activity could be detected only in the 24-month-old animals.(ABSTRACT TRUNCATED AT 250 WORDS)     

Alterations in rat cardiac myosin isozymes induced by whole-body irradiation are prevented by 3,5,3'-L-triiodothyronine

Changes in cardiac myosin isozymes and serum thyroid hormone levels were investigated in rats following 10 Gy whole-body gamma irradiation. The percent beta-myosin heavy chain increased from 21.3 +/- 1.8 to 28.1 +/- 6.8 (NS) at 3-day postirradiation, 37.7 +/- 1.9 (P less than .001) at 6-day postirradiation, and 43.8 +/- 3.3 (P less than .001) at 9-day postirradiation. Along with the change in myosin isozymes was a significant 53% decrease (P less than .001) in the serum thyroxine (T4) level by day 3 postirradiation, remaining depressed through day 9 postirradiation. The serum 3,5,3'-triiodothyronine (T3) level, however, was normal until day 9, when significant depression was also observed. In contrast, the thyroid-stimulating hormone (TSH) level was significantly increased by fourfold at day 3, returning to near normal values by day 9 postirradiation. Daily injections of physiological doses of T3 (0.3 microgram/100 g body weight) prevented the change in the myosin isozymes following whole-body irradiation. Daily pharmacological injections of T3 (3.0 micrograms/100 g body weight) to the irradiated rats produced a further decrease in the percent beta-myosin heavy chain (below control values) indicating tissue hyperthyroidism. Thus, this study suggests that the change in myosin isozymes following whole-body irradiation is caused by an alteration in thyroid hormone activity.     

The effects of amiodarone on serum thyroid hormones and hepatic thyroxine 5'-monodeiodination in rats

Amiodarone (2-n-butyl-3,4'-diethylaminoethoxy-3', 5'-diiodobenzoyl-benzofurane) is an antiarrhythmic drug which increases serum T4 and rT3 levels in patients and lowers serum T3 levels. To investigate its effects on T4 metabolism and its cardiac action, we fed amiodarone to male Fisher rats at doses of 5, 15, and 45 mg/kg BW X day; controls received potassium iodide for 4-7 weeks, and another group received sodium ipodate. At 4 weeks, amiodarone caused a dose-dependent increase in the serum T4 concentration and a slight reduction of serum TSH without a change in the serum T3 concentration. These changes were not present at 7 weeks. Sodium ipodate raised serum T4 concentrations at both times. Rats treated with T4 (150 micrograms/kg BW X day) to suppress thyroidal secretion of hormone and with amiodarone (15 mg/kg) had marked reduction of serum T3 concentrations compared with controls receiving T4 without amiodarone. Liver homogenates from rats treated with amiodarone showed marked reduction on T4 5'-monodeiodinase activity in a dose-related manner. Amiodarone added to liver homogenates in vitro at concentrations of 0.001-1 mM did not inhibit T3 production from T4, whereas ipodate added in vitro (0.01-1 mM) did inhibit T3 production. Rats treated with amiodarone showed a lowering of the resting heart rate and a reduction of the increment in heart rate after iv isoproterenol administration. The cardiac Ca++ myosin ATPase activity was reduced in rats receiving amiodarone (45 mg/kg) compared with that in controls. The data indicate that rats treated with amiodarone have reduced peripheral conversion of T4 to T3 owing to impaired hepatic T4 5'-monodeiodinase activity. In addition, these rats have slowing of heart rate and reduction of cardiac Ca++ myosin ATPase activity. These findings are consistent with the hypothesis that amiodarone blocks some effects of thyroid hormone on the heart, but additional studies are needed to test this hypothesis.     

Alterations in cardiac myosin isoenzymes distribution as an adaptation to chronic environmental heat stress in the rat

The distribution of cardiac myosin isoenzymes, using gel pyrophosphate electrophoresis, was studied in laboratory rats during acclimation to heat (34 degrees C, 0-2 month) with and without daily administration of triiodotyronin (0.3 micrograms/100 g b.wt) Thyroxin (T4) and triiodotyronin (T3) concentrations during that period were measured in the acclimated rats as well. Control rats exhibited only the V1 myosin form during the entire experimental period. In the heat acclimated rats, after three weeks of acclimation, in addition to the V1 band, bands of V2 and V3 myosin forms appeared. After the fourth week of acclimation V3 became the dominant myosin. Changes in the cardiac isoenzymes distribution occurred 1 week after a significant decrease in T3 was recorded. Administration of additional thyroxin dose didn't allow the development of V3 band in the heat acclimated rats. It was concluded that the changes observed in cardiac isoenzymes distribution could be related to an alteration in thyroid activity and that these changes could play a role in the adaptation of the cardiac muscle to chronic heat.     

Hormonal influences on cardiac myosin ATPase activity and myosin isoenzyme distribution

It has been recognized for a long time that changes in hormone secretion can influence cardiac function; however, the biochemical basis for these changes has only recently been clarified. In this review the influences of hormonal status on the contractile protein myosin is discussed. Myosin has a rod-like portion and a globular head and consists of two myosin heavy chains (MHC) and four light chains (LC), two of which are identical. The globular head is the site of an ATP-splitting enzyme, the myosin ATPase, and increases in myosin ATPase activity are closely related to an increased velocity of contraction of the heart. Myosin ATPase activity shows marked response to alterations in thyroid hormone, insulin, glucocorticoid, testosterone and catecholamine levels, but marked animal species differences in this response occur. Thyroid hormone administration to normal rabbits, for example, increases myosin ATPase activity markedly, but the myosin ATPase activity of hyperthyroid rats remains unchanged. In contrast, in hypothyroid rats myosin ATPase activity is markedly decreased but the hypothyroid rabbit shows no such response. These species-related differences in the hormonal response of myosin ATPase activity result from the predominance pattern of specific myosin isoenzymes. In the normal rat heart three myosin isoenzymes, v₁, V₂ and V₃, can be separated electrophoretically. Myosin V₁ predominates (70% of total myosin), and has the highest myosin ATPase activity, whereas in rabbits myosin v₃, which has a lower myosin ATPase activity, is the predominant isomyosin. Thyroid hormone administration to rabbits induces myosin V₁ predominance and therefore increases myosin ATPase activity, whereas in hyperthyroid rats only a small further increase in V₁ predominance can occur. The alterations in myosin isoenzyme predominance and myosin ATPase activity are closely correlated to changes in cardiac contractility. Hormone-induced alterations in myosin isoenzyme predominance are mediated through changes in the formation of two isoforms of myosin heavy chain. Changes in the expression of different myosin heavy chain genes are most likely responsible for the thyroid hormone and insulin-induced alterations in myosin isoenzyme predominance. Investigation of the control of myosin heavy chain formation can provide further insights into the hormonal control of a multigene family as well as broaden our understanding of the molecular events which result in altered cardiac contractility. It is currently unclear if androgens, glucocorticoids and catecholamines influence myosin ATPase activity through changes in myosin isoenzyme predominance resulting from alterations in myosin heavy chain gene expression. Post-translational modifications of myosin heavy chain and light chain polypeptides have also to be considered.     

Diurnal variations of plasma growth hormone, thyrotropin, thyroxine, and triiodothyronine in streptozotocin-diabetic and food-restricted rats

The pattern of spontaneous GH, TSH, T4, and T3 secretion has been studied in male rats in response to a 15-day period of streptozotocin diabetes or food restriction. Beginning at 0900 h, groups of control (C), food-restricted (FR), diabetic (D), and insulin-treated D rats were killed every 60-90 min for a 8-h period. Food restriction resulted in a significant depression of the GH, TSH, T4, and T3 peaks, whereas diabetes caused complete suppression of episodic secretion of each hormone. Insulin (6 U/100 g BW X day for 12 days) administration to D rats restored the normal pattern of secretion. In D and FR rats, pituitary GH concentrations were lower than in C rats, whereas pituitary TSH concentrations were similar to those in controls. Thus, as compared to C rats, FR and D rats showed an inhibition in GH, TSH, T4, and T3 secretion, most marked in D animals. Since diabetes is associated with a deficiency of circulating thyroid hormones, the potential roles of T4 and T3 on pituitary GH concentration and secretion in D rats were evaluated. Treatment of D rats with insulin (3 U/100 g BW X day), T4 (1.8 micrograms/100 g BW X day), or T3 (0.30 microgram/100 g BW X day) for 12 days resulted in a significant but limited increase in pituitary GH content. When administered together with insulin, the net effects of T4 or T3 with insulin appeared additive. T4 administration to D rats produced a significant though limited increase in plasma GH concentrations and weight gain, whereas both values were unaffected by T3. Simultaneous administration of T4 and insulin resulted in significant increased plasma GH concentration to levels greater than those in C rats. However, plasma GH levels in rats treated with T3 plus insulin were greater than those in D rats, but lower than in C animals. The results indicate that the decreased pituitary GH content of D rats can be corrected, at least in part, by T4 and T3.     

Effect of amiodarone on rat heart myosin isoenzymes

The effects of amiodarone on heart weight, production of 14C-CO2 from labelled glucose, myosin ATPase activity, and myosin isoenzyme patterns were determined by comparing control and amiodarone-treated male Wistar rats. Since it has been suggested that amiodarone may interfere with thyroid hormone action on the heart, similar experiments were also carried out in hypothyroid and amiodarone-plus-triiodothyronine(T3)-treated rats, and the data were compared to those obtained in amiodarone-treated rats. Amiodarone treatment for 6 weeks resulted in lower heart weight, decreased atrial production of 14C-CO2 from labelled glucose, decreased myosin Ca-ATPase activity, and preferential synthesis of V3 isomyosin. These effects were similar to those observed in hypothyroid rats but were lesser in magnitude. T3 treatment of amiodarone-treated rats reversed all the changes induced by amiodarone. Serum thyroxine (T4) was higher in amiodarone-treated than in control rats, while serum T3 was similar. Serum T3 was higher in the amiodarone-plus-T3 than in the amiodarone-treated group. These results show that 1) amiodarone-induced changes resemble hypothyroidism with respect to cardiac myosin expression and atrial CO2 production, 2) amiodarone causes hypothyroid-like changes despite normal serum T3 and increased serum T4, and 3) T3 reverses the effects of amiodarone. These data support the hypothesis that amiodarone inhibits the action of thyroid hormone on the heart.     

The effect of altered thyroid status on pituitary hormone messenger ribonucleic acid concentrations in the rat

To study the effects of altered thyroid status on pretranslational control of pituitary hormones, adult male rats were given propylthiouracil for 6 weeks and underwent the following studies. 1) Rats were injected with T3 at 10 micrograms/100 g BW daily for 10 days. 2) Rats were given T3 injections at 0, 0.01, 0.1, 1.0, or 10 micrograms/100 g BW for 10 days. 3) Rats were killed 0, 1, 6, or 24 h after a single injection of T3 at 10 micrograms/100 g BW or after 5 or 10 days of daily T3 injections. Pituitary mRNA concentrations of TSH beta, alpha-subunit, PRL, GH, POMC, FSH beta, and LH beta were determined for individual animals. Marked increases in TSH beta and alpha-subunit mRNAs occurred after PTU treatment, and these changes were reversed by 1.0 microgram/100 g BW T3 and within 24 h of a single T3 injection of 10 micrograms/100 g BW. Further increases in the dose or time course of T3 administration led to a relatively greater suppression of TSH beta mRNA levels than alpha-subunit mRNA levels. In contrast, GH and PRL mRNA levels were low in hypothyroid animals, and both rose toward control levels with 0.1 microgram/100 g BW T3 and by 24 h after a single T3 dose. Induction of hyperthyroidism did not further increase GH mRNA levels above control, but increased PRL mRNA levels 2-fold over control. No changes were seen in FSH beta, LH beta, or POMC mRNA levels with any treatment. Thus, studies of altered thyroid status in the rat reveal dose-response and time-course variability in the pretranslational control of TSH beta, alpha-subunit, GH, and PRL by thyroid hormone.     

In vitro effects of thyroid hormones on red blood cell Ca++-dependent ATPase activity

Thyroid hormones (TH) have been shown to exert a direct stimulatory effect on the Ca++-dependent ATPase from human and other mammalian erythrocytes. In this in vitro system, T4 has been shown to be more effective than T3. In the present study, TH effects on Ca++-dependent ATPase were investigated, using rabbit and human erythrocyte membranes, after preincubation with 10(-10) M T4, in the presence or in the absence of exogenous calmodulin (CaM) (5.10(-12) M to 5.10(-9) M). Ca++-dependent ATPase activity was measured as inorganic phosphate (Pi) release from 1 mM ATP. The results showed that basal Ca++-dependent ATPase activity in rabbits was moderately increased by T4 (1.44 +/- 0.05 vs 1.32 +/- 0.04 mumol Pi/mg protein/90 min, mean +/- SE; p less than 0.05). The time course of Pi release did not show any stimulatory effect of T4 during the first hour of incubation. The effect of T4 became apparent, however, 1 h after the addition of ATP (delta T4: 15%). With human membranes, T4 induced a relative stimulation of the Ca++-dependent ATPase of 8-10% (p less than 0.05) in experimental conditions where the enzyme was not maximally stimulated by CaM (delta CaM over basal activity: 5-40%). In conditions of high CaM stimulation (delta CaM: 50-320%), T4 had no effect. These results confirm that Ca++-dependent ATPase activity is increased by T4. The effect of T4 is small, and appears as a late event during incubation with ATP. Stimulation by T4 is expressed in states of low enzyme activation by CaM.(ABSTRACT TRUNCATED AT 250 WORDS)     

Myocardial mechanical and myosin isoenzyme alterations in streptozotocin-diabetic rats

Fifteen week old male Wistar rats (n = 7) were made diabetic by intravenous injection of streptozotocin (50 mg/kg). Age-matched, untreated male Wistar rats (n = 9) served as controls. Hearts were removed after 5-6 weeks of diabetes, and the isometric developed tension (T) of isolated left ventricular papillary muscles and its first derivative (dT/dt) were measured at a frequency of 0.2 Hz. During testing, the muscles were perfused with Tyrode's solution (Ca2+ concentration was half of normal Tyrode's solution, pH 7.4, 32 degrees C, bubbled with 95% O2 and 5% CO2). In addition, the left ventricular isoenzyme pattern, which is related to myocardial energetics, was determined by pyrophosphate gel electrophoresis. There was no significant difference in isometric developed tension between diabetic and control rats (DM: 2.90 +/- 0.89 vs controls: 2.87 +/- 0.85 g/mm2, mean +/- SD), but in diabetic rats, dT/dtmax decreased significantly as compared with controls (DM: 23.5 +/- 4.2 vs controls: 31.9 +/- 7.9 g/mm2.s, p less than 0.05). Myocardial mechanical responses to isoproterenol (10(-7)M) and dibutyryl cyclic AMP (10(-5)M) also decreased in diabetic rats. The left ventricular myosin isoenzyme pattern shifted toward VM-3 in diabetic rats (VM-3: DM: 74.9 +/- 10.7 vs controls: 9.5 +/- 4.1%, p less than 0.001). These results indicate that diabetes influences myocardial contractility and changes cardiac energetics. Post-receptor processes may play a role in myocardial mechanical responses to catecholamines in streptozotocin-diabetic rats.     

Thyroid hormone and growth hormone interact to regulate insulin-like growth factor-I messenger ribonucleic acid and circulating levels in the rat

Thyroid hormones influence growth in part by altering the secretion and effects of GH. GH, in turn, mediates its effects by regulating the synthesis and secretion of insulin-like growth factor-I (IGF-I). IGF-I is a pleiotropic growth factor that is synthesized by many tissues and acts on many tissues to regulate both cellular replication and differentiated function. We have studied the direct effects of thyroid hormones and the combined effects of thyroid hormones and GH on the regulation of IGF-I synthesis and secretion in hypophysectomized (hypox) rats in vivo. All rats, except normal littermates and a hypox control group, received 100 micrograms hydrocortisone/100 g BW for 10 days. Circulating IGF-I was measured by specific RIA (normal rats, 1 U/ml), and hepatic IGF-I mRNA was measured by Northern blot hybridization with an antisense cRNA probe. 1) Hypox rats treated with hGH (75 micrograms, ip, twice daily) for 10 days gained 17 g BW vs. 70 g for normal littermates. GH markedly increased hepatic IGF-I mRNA and circulating IGF-I (0.52 +/- 0.14 U/ml 12 h after the last GH injection vs. 0.03 +/- 0.02 for hypox controls). 2) T4 (1 micrograms/100 g BW, ip) for 10 days increased neither weight, hepatic IGF-I mRNA, nor circulating IGF-I. 3) Rats treated with T4 for 10 days followed by a single injection of 1 mg GH, ip, increased hepatic IGF-I mRNA and circulating IGF-I levels comparably as in rats receiving acute GH alone (IGF-I, 12 h, 0.31 +/- 0.09 vs. 0.36 +/- 0.06 U/ml). 4) Hypox rats treated with a single injection of T3 (1.5 micrograms/100 g BW, ip) had slightly increased hepatic IGF-I mRNA, but showed no significant change in circulating IGF-I levels. 5) A single injection of T3 plus GH to hypox rats increased IGF-I mRNA levels above those in rats injected with GH alone and increased serum IGF-I levels to 0.48 +/- 0.12 U/ml compared to 0.36 +/- 0.06 U/ml for GH alone. 6) After 10 days of GH treatment, a single injection of T3 lowered both hepatic IGF-I mRNA and circulating IGF-I (0.52 +/- 0.14 to 0.16 +/- 0.06 U/ml, 6 h after T3). These studies demonstrate that thyroid hormones have relatively little direct effect on IGF-I synthesis but can have major effects on GH-stimulated IGF-I synthesis and secretion. The pattern of these effects depends on the integrity of the pituitary gland, prior exposure of the liver to GH and/or thyroid hormones, and the temporal relationship between GH and thyroid hormone administration.     

Thyroid vascular conductance: differential effects of elevated plasma thyrotropin (TSH) induced by treatment with thioamides or TSH-releasing hormone

We have reported previously that thyroid gland blood flow, expressed as vascular conductance (C) per mass, is decreased at very low and increased at very high chronic plasma TSH concentrations, but is apparently unchanged over a broad range of plasma TSH concentrations encompassing normal levels. The aim of the present study was to examine the apparently very steep dose-response relationship between elevated plasma TSH and thyroid vascular C/mass. In the first series of experiments, endogenous plasma TSH concentrations were manipulated by treating male Sprague-Dawley rats (250-280 g) for 6 days as follows: 1) controls (0.5 ml saline/day, ip), 2) propylthiouracil injections (2.0 mg PTU/day, ip), 3) PTU plus partial thyroid hormone replacement (2.0 mg PTU/day and 0.3-0.9 microgram T4 plus 0.075-0.225 microgram T3/100 g.day via continuous sc infusion), or 4) TRH (9-1200 micrograms TRH/100 g.day via continuous iv infusion). The vascular C values of the thyroid gland, salivary gland, kidney, and pancreas were determined using the reference sample version of the radioactive microsphere technique. PTU treatment led to the expected hypothyroidism, increased plasma TSH concentrations (959 +/- 66 vs. 154 +/- 22 ng/dl), increased thyroid weight (9.19 +/- 0.36 vs. 4.60 +/- 0.15 mg/100 g), and increased thyroid vascular C/mass (495 +/- 51 vs. 127 +/- 20 microliters/mm Hg.g/min). PTU-treated rats receiving partial thyroid hormone replacement demonstrated a dose-related suppression of plasma TSH, thyroid weight, and thyroid vascular C. Although, TRH treatments resulted in increased plasma TSH concentrations (e.g. 1200 micrograms TRH, 706 +/- 46 ng/dl) and thyroid weight (e.g. 1200 micrograms TRH, 7.45 +/- 0.41 mg/100 g), thyroid vascular C per tissue mass was not significantly increased after any TRH treatment (e.g. 1200 micrograms TRH, 166 +/- 19 microliters/mm Hg.g/min). Thus, at similarly elevated plasma TSH concentrations, the thyroid vascular C/mass of PTU- and TRH-treated rats constituted separate populations. Both PTU- and TRH-induced thyroid growth were accompanied by similar alterations in thyroid gland morphology (i.e. increased cellular mass with little change in the total amount of colloid). To investigate the mechanisms involved, groups of rats were treated for 6 days as follows: 1) control, 2) PTU or methimazole (25 mg MMI/day, ip), 3) PTU or MMI plus thyroid hormone replacement (1.2 micrograms T4 plus 0.3 microgram T3/d.100 g), 4) TRH (12 micrograms/100 g.day), and 5) PTU or MMI, thyroid hormones, and TRH.(ABSTRACT TRUNCATED AT 400 WORDS)     

Effect of administration of growth hormone on plasma and intracellular levels of thyroxine and tri-iodothyronine in thyroidectomized thyroxine-treated rats.

We have studied the effects of the administration of GH on plasma levels and peripheral production of tri-iodothyronine (T3) from thyroxine (T4) in thyroidectomized male Wistar rats given a continuous i.v. infusion of T4 (1 microgram/100 g body weight per day) and GH (120 micrograms per day) for 3 weeks. Tracer doses of 131I-labelled T3 and 125I-labelled T4 were added to the infusion. At isotopic equilibrium (10 days after the addition of 125I-labelled T4) the rats were bled and perfused. The plasma appearance rate for T3 was higher (10.6 +/- 1.3 vs 8.4 +/- 2.8 pmol/h per 100 g body weight, P = 0.05) and plasma TSH was lower (246 +/- 24 vs 470 +/- 135 pmol/l, P less than 0.01) in GH-treated rats. The amount of T3 in liver (12.3 +/- 2.8 vs 5.5 +/- 1.7 pmol/g wet weight, P less than 0.01), kidney (11.5 +/- 1.4 vs 6.5 +/- 1.4 pmol/g wet weight, P less than 0.01) and pituitary (8.8 +/- 2.7 vs 4.8 +/- 0.5 pmol/g wet weight, P less than 0.01) was higher than in controls, mainly as a result of an increased local production of T3 from T4, but plasma-derived T3 was also higher in most organs. We found an increased intracellular T3 concentration in the pituitary which may be responsible for the lower plasma TSH concentration in the GH-treated rats. Since the increase in locally produced T3 is found particularly in liver, kidney and pituitary, typical organs that express 5'-deiodinase activity, we suggest that GH acts on thyroid hormone metabolism by stimulating type-I deiodinase activity.     

Thyroid hormone regulation of hypothalamic immunoreactive thyrotropin

Immunoreactive TSH (IR-TSH) has been identified in the hypothalamus and other brain areas in intact and hypophysectomized rats. To determine if thyroid hormones regulate IR-TSH in the rat brain, the effects of T3 and T4 on pituitary and hypothalamic IR-TSH content were studied in intact and hypophysectomized rats. Female rats received daily injections of T3 (0.1, 1.0, or 10 micrograms/100 g BW) or T4 (0.5, 2.5, or 5.0 micrograms/100 g BW) for 7 days. The chronic administration of T3 and T4 decreased the plasma concentration and pituitary content of IR-TSH in a dose-dependent manner. In intact rats, the administration of T3 did not affect the content of IR-TSH in the median eminence, ventral hypothalamus, or dorsal hypothalamus. In contrast, administration of T4 significantly increased the content of IR-TSH in the median eminence and ventral hypothalamus in a dose-dependent manner. Similarly, in hypophysectomized rats daily administration of T4 resulted in a dose-dependent increase in the concentration of IR-TSH in the median eminence and ventral hypothalamus. In hypophysectomized rats infusion of colchicine into the lateral ventricle, which blocks axonal transport, decreased the content of IR-TSH in the median eminence (631 +/- 59 vs. 439 +/- 36 microU/mg), increased the content of IR-TSH in the ventral hypothalamus (45 +/- 5 vs. 87 +/- 10 microU/mg), and blocked the T4-induced increase in IR-TSH in the median eminence. In intact rats, administration of iopanoic acid, which blocks the conversion of T4 to T3, decreased the content of IR-TSH in the median eminence (688 +/- 44 vs. 450 +/- 38 microU/mg) and ventral hypothalamus (103 +/- 16 vs. 70 +/- 5 microU/mg) and blocked the T4-induced increase in IR-TSH in the median eminence (1025 +/- 82 vs. 644 +/- 62 microU/mg) and ventral hypothalamus (229 +/- 23 vs. 117 +/- 11 microU/mg). These data indicate that TSH content in the median eminence and ventral hypothalamus is regulated by a thyroid hormone-sensitive mechanism which requires the local monodeiodination of T4 to T3. In addition, the effect of colchicine on the IR-TSH content in the median eminence and the T4-induced increase in brain IR-TSH are consistent with previous reports demonstrating that hypothalamic IR-TSH is regulated independently of pituitary and serum TSH.     

Isolation and characterization of myosin heavy chain isozymes of the bovine conduction system

To determine the characteristics of cardiac myosin in the conduction system, a pure Purkinje fiber preparation, consisting of atrioventricular nodes and the ventricular conduction system, was obtained from bovine hearts. Two types of myosin heavy chain isozymes, alpha-type and beta-type, were fractionated by affinity chromotography using monoclonal antibodies CMA19 and HMC50, which are specific for the alpha-type heavy chain and beta-type heavy chain, respectively. Competitive enzyme-linked immunosorbent assay demonstrated that the content of beta-type in the atrioventricular node (30-40%) was higher than that in atrial ordinary myocardium (10-20%) and that of the alpha-type was 30-40% in the ventricular conduction system, which was much higher than that in the ventricular ordinary myocardium (less than 10%). By one- and two-dimensional electrophoresis of the peptides produced by partial and complete digestion, the peptide compositions of alpha-type and beta-type in the conduction system were shown to be very similar to those of alpha-type and beta-type in ordinary myocardium, respectively. The CA2+-activated ATPase activity of myosin of the atrioventricular nodes was lower than that of ordinary atrial myosin (0.46 +/- 0.03 versus 0.58 +/- 0.02 mumol Pi/mg/min, mean +/- SEM, p less than 0.05) and in contrast, that of ventricular specialized myocardium was higher than that of myosin in the ventricular ordinary working myocardium (0.32 +/- 0.03 versus 0.22 +/- 0.01 mumol Pi/mg/min, p less than 0.05). This was in good agreement with the relative proportion of myosin isozymes.(ABSTRACT TRUNCATED AT 250 WORDS)     

Force transient time course in heart muscle with high and low V1 to V3 myosin isoenzyme ratio

Values of mechanical and biochemical indicators of muscle performance measured in heart muscle having mostly the V1 isoenzyme of myosin are different than measurements of the same indicators made in muscle having mostly the V3 isoenzyme of myosin. It has been suggested that these differences in performance indicators might be attributable to subtle differences in myosin-actin crossbridge cycling kinetics between the V1 and the V3 isoforms of myosin. To investigate this, we derived information about myosin-actin cycling kinetics from the time course of force transients following rapid small amplitude length releases applied to chemically "skinned", isometrically contracting trabeculae from the hearts of normal (greater than = 90% V1) and propylthiouracil treated (greater than = 90% V3) rats. The rate constant for rapid force recovery measured in trabeculae from normal rats was twice that measured in trabeculae from treated rats (88.8 +/- 18.8 (n = 12) vs. 43.7 +/- 6.5 (n = 10)/s, mean +/- S.D.). We interpret this difference in rate constants as evidence that the kinetics of at least one step in the interaction of myosin with actin depends on the isoenzyme of myosin present in the heart.     

Peripheral metabolism of homologous thyrotropin in euthyroid and hypothyroid rats: acute effects of thyrotropin-releasing hormone, triiodothyronine, and thyroxine.

The peripheral metabolism and metabolic clearance rate (MCR) of homologous TSH was studied in euthyroid and hypothyroid rats. Incubation of freshly labeled [125I]iodo-TSH with rat serum revealed a labeled nonimmunoreactive protein in the void volume of a Sephadex G-100 column which could not be detected by conventional chromatographic purification. Removal of this contaminant from the tracer reduced the nonspecific binding in the absence of serum and increased the binding of tracer in the absence of added exogenous TSH. Injection of [125I]iodo-TSH into rats was followed within 15 min by the appearance of at least three labeled protein components. Gel filtration showed that these peaks were trichloroacetic acid (TCA)-precipitable proteins of larger molecular weight than TSH, but not all were precipitable by antibody to rat TSH. The disappearance rate of TCA-precipitable 125I (t1/2 = 28 min) was significantly longer than the disappearance rate of immunoprecipitable 125I (t1/2 = 22 min). The disappearance rate of immunoprecipitable [125I]iodo-TSH was identical to that of injected purified rat TSH and of the TRH-induced TSH increment in euthyroid rats. The disappearance rate os suppressible TSH (after 100 microgram T3) in hypothyroid animals was only slightly longer than the rate of disappearance of immunoprecipitable [125I]iodo-TSH (40 vs. 36 min) in the same rats. The calculated MCR of TSH was slightly lower (P less than 0.05) in hypothyroid rats (18.3 +/- 3.0 ml/h/100 g BW, mean +/- SD) than it was in euthyroid rats (22.6 +/- 2.1). The pituitary TSH concentration in hypothyroid rats was 29 mU/mg wet wt, similar to that of euthyroid animals. These results indicate that the turnover rate of pituitary TSH in hypothyroid rats with serum TSH concentrations of 1400-3000 microunit/ml is 7-14 times/day. Therefore, the significant increase we observed in pituitary TSH concentration 1 h after T4 (1.5 microgram/100 g BW) or T3 (0.15 microgram/100 g BW) administration indicates that the 35% decrease in plasma TSH at this interval is due to inhibition of TSH release, not to inhibition of TSH synthesis.