In vivo effects of gamma-aminobutyric acid and beta-alanine on thyrotrophin secretion in the normal and hypothyroid rat

The effect of pharmacological doses of two amino acids neurotransmitters, gamma-aminobutyric acid (GABA) and beta-alanine (beta-Ala), on thyrotrophin (TSH) secretion was studied in normal and hypothyroid (PTU-treated) male rats. Inhibition of TSH secretion was observed in normal rats treated with the drugs, 30 min after their administration. Hypothyroid animals responded only to GABA administration, decreasing their serum TSH at 30 min. Response to thyrotrophin-releasing hormone (TRH) after 15 min of drug administration was blunted in GABA injected animals, as compared to saline-injected controls. When TRH was injected at the same time as GABA and beta-Ala, the response was significantly lower than in controls. It is suggested that beta-Ala and GABA act at the pituitary by impairing the TSH response to TRH. The possibility that beta-Ala actions may be due to decreased GABA catabolism is considered, since beta-Ala administration increased GABA synaptosomal levels.     

Gonadotropin secretion in azotemic male rats--effect of opioid blockade

The role of endogenous opioids in the control of gonadotropin secretion in uremic male rats was investigated using the narcotic antagonist, naloxone. In order to eliminate the effect of weight loss due to uremia-induced anorexia as a cause of previously described altered gonadotropin secretion in uremia, we also studied a group of normal pair-fed control animals who exhibited a weight loss comparable to that of the uremic animals. Naloxone administration had no effect on the basal or LRH-stimulated peak concentrations of LH and FSH in the normal or the uremic rats. Basal and LRH-stimulated gonadotropin responses in the pair-fed rats were comparable to those seen in the normal rats. Similarly, opioid blockade produced no change in the basal or LRH-stimulated gonadotropin responses in the pair-fed animals. Testosterone concentrations were significantly lower in the uremic and pair-fed animals compared to the normal rats. The data suggest that experimental renal failure is not associated with altered opioidergic tone, as it relates to gonadotropin secretion, or to diminished sensitivity of the gonadotroph to LRH stimulation. The decreased testosterone concentration seen in the uremic and pair-fed rats may reflect abnormalities in gonadal hormone secretion due to primary pathology occurring at the level of the gonad. These abnormalities may be reflected as diminished Leydig cell sensitivity to LH. The inappropriately low concentrations of LH in the presence of low testosterone together with normal gonadotropin response to exogenous LRH also suggest an abnormal secretion of endogenous LRH. It is not clear whether this presumed abnormality in LRH secretion is a primary event or is related to decreased testosterone production by the testes in the uremic and pair-fed rats.     

Potentiation by indomethacin of TRH-induced TSH secretion in the rat.

We have studied the effect of two inhibitors of prostaglandin synthesis on the basal and TRH-stimulated plasma TSH levels in the rat. Animals were injected sc daily with indomethacin 3 mg/0.5 ml) or aspirin (16--30 mg/0.5 ml) for 3 days. The plasma T4 and T3 were consistently lower in the indomethacin or aspirin groups than in the controls, while the basal TSH levels did not change. Indomethacin treatment significantly potentiated the TSH response to synthetic TRH (20 ng. iv) in intact and thyroidectomized rats. The pituitary TSH content was markedly increased by indomethacin, while hypothalamic TRH content did not change. In contrast, aspirin inhibited the TSH response to TRH in intact rats, when pituitary TSH content decreased significantly. No potentiation by aspirin of TRH-stimulated TSH response in the thyroidectomized rats was observed. The increased sensitivity of plasma TSH response to exogenous TRH in the indomethacin group is presumably due to higher pituitary TSH content than in the controls. The action of indomethacin appears to be mediated, at least in part, at the pituitary level. In addition, there is a dissociation between the action of indomethacin and the action of aspirin in the TSH response to TRH.     

Aging and gender affect the response of thyrotropin (TSH) to gastrin releasing peptide (GRP) in rats

We had previously shown that GRP acts directly at the pituitary gland inhibiting basal and TRH-stimulated TSH secretion in adult male rats. In this study we showed a gender dimorphism in this response of old animals pituitaries to GRP. In both female and male young adult animals, GRP-incubated pituitaries showed approximately 50% less basal and TRH-stimulated TSH secretion to the medium, without affecting the pituitary content of TSH. However, GRP did not have any significant effect upon TSH secretion in old male rats, but the old female showed the same degree of response to GRP as the young adult female rat, regarding basal and TRH-stimulated TSH secretion, while the TSH pituitary content after GRP incubation was higher than that of the young female group. Our data suggest a loss of thyrotrope responsiveness to GRP in aged male rats that could contribute to the decrease in TSH pituitary stores leading to lower basal and TRH-stimulated TSH secretion. Meanwhile, the preservation of GRP responsiveness could help in the relative maintenance of these parameters in the old female rat.     

The role of cholecystokinin octapeptide (CCK-8) in regulating thyrotropin secretion in rats. In vivo studies

The effect of cholecystokinin octapeptide (CCK-8) on basal and TRH-stimulated secretion of TSH was investigated in 67 male Sprague-Dawley rats. Blood for TSH determinations was sampled by cannulation of right heart auricle in urethane narcosis before and four times during 60 minutes following CCK-8 administration. It was found that CCK-8 administered to the lateral brain ventricle at a dose of 0.5 microgram per animal caused a decrease in blood serum TSH concentration but did not change the response of TSH to TRH. Intravenous administration of CCK-8 at doses of 2 and 20 micrograms/kg had no effect on blood serum concentration of TSH.     

Significance of serum thyrotropin and plasma dopamine concentration in the regulation of thyroid function in elderly subjects

In our previous study, we observed a tendency towards an age-related increase in the serum thyrotropin (TSH) concentration. Regulatory mechanisms of TSH secretion in elderly subjects were studied. In 43 elderly subjects, serum TSH did not correlate significantly with serum T4, T3 free T4 or rT3. Further, those with increased TSH (greater than 5 mU/l, 9 subjects) did not overlap with those with low T3 (less than 0.92 nmol/1, 8 subjects). Increases in serum TSH were not associated with the presence of circulating anti-thyroid autoantibodies. A TRH test using a 500 micrograms single bolus injection was performed in 15 subjects. TSH response (basal: 1.92 +/- 1.42 (s.d.) mU/1, peak: 11.25 +/- 5.33 mU/1, sigma: 26.74 +/- 12.89 mU/1, respectively) did not differ significantly from that of younger subjects. T3 response after TRH varied greatly and a close correlation was observed between basal T3 and peak T3 (r = 0.86), and also between peak T3 and delta T3 (r = 0.81). A significant correlation was observed between sigma TSH and basal T3 (r = 0.60). Neither plasma cortisol, epinephrine nor norepinephrine concentrations showed any significant correlation with basal and TRH-stimulated TSH or T3 concentrations. However, the plasma dopamine concentration correlated significantly with sigma TSH (r = 0.60) and basal T3 (r = 0.52), respectively. In conclusion, the increase in serum TSH observed in elderly subjects was felt to represent a physiological adaptation to maintain serum T3. Low T3 subjects appear to have a disturbance in this mechanism, with decreased TSH and T3 response to TRH stimulation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Trifluoperazine inhibits thyrotropin-releasing hormone-stimulated TSH secretion

In previous work changes of the thyrotropic secretion after administration of some substances affecting the calcium content in the cytosol were demonstrated. The object of the present investigation was to assess the hormonal response to the administration of trifluoperazine, a psychopharmaceutical preparation, the main mechanism of its action being the inactivation of the cytosol receptor for the calcium signal - calmodulin. The poor utilization of intracellular calcium of the secretory cell is then the factor which inhibits secretion proper. The thyrotropic secretory reserve (delta TSH) was assessed in the same subjects before and after trifluoperazine administration by the TRH test as the difference of values at rest and TRH-stimulated TSH levels during the 20th, 30th, 40th and 60th minute following intravenous administration of 200 micrograms TRH. It was revealed that this calmodulin antagonist administered for one week in amounts of 6-12 mg per day by mouth significantly inhibits the secretory response of TSH to TRH in healthy subjects during the 20th and 40th min. (P less than 0.05). The reproducibility of the TRH test repeated in a group of subjects not treated with trifluoperazine, however, under equal conditions and after the same time intervals as in the experiment with trifluoperazine was very satisfactory and thus physiological inhibition caused by repeated TRH administration could be ruled out. The inhibition of the secretory TSH response to TRH can be therefore considered the consequence of the direct effect of trifluoperazine on the thyrotropic secretory mechanism. Trifluoperazine significantly reduced serum calcium levels and raised phosphate levels, while it did not affect the blood levels of magnesium.(ABSTRACT TRUNCATED AT 250 WORDS)     

Abnormalities of the thyroid hormone negative feedback regulation of TSH secretion in spontaneously hypertensive rats.

Spontaneously hypertensive rats (SHR) are characterized by several neuroendocrine abnormalities including a chronic hypersecretion of thyrotropin (TSH) of unknown etiology. We hypothesized that the inappropriately high TSH secretion in SHR may be the result of an impaired thyroid hormone negative feedback regulation of hypothalamic thyrotropin-releasing hormone (TRH) and/or pituitary TSH production. To test this hypothesis, SHR or their normotensive Wistar-Kyoto (WKY) controls were treated with either methimazole (0.02% in drinking water) to induce hypothyroidism or administered L-thyroxine (T4) at a dose of 0.8 or 2.0 micrograms/100 g body weight/day to induce hyperthyroidism. All treatments were continued for 14 days after which animals were killed under low stress conditions. TSH concentrations in plasma and anterior pituitary tissue were 2-fold higher (P less than 0.01) in euthyroid SHR compared to WKY control rats while thyroid hormone (T3 and T4) levels were in the normal range. Hypothyroidism induced by either methimazole or thyroidectomy caused a significant (P less than 0.01) rise of plasma TSH levels in both WKY and SHR rats. However, relative to the TSH concentrations in control animals, the increase of plasma TSH in SHR was significantly blunted (P less than 0.01) in comparison to the WKY group. Hypothyroidism caused a significant depletion of TRH in stalk-median eminence (SME) tissue in both groups of rats. However, no differences between SHR and WKY rats were observed. The administration of thyroid hormone caused a dose dependent suppression of plasma TSH levels in both strains of rats. However, at both doses tested plasma TSH concentrations in SHR rats were significantly less suppressed (P less than 0.05) than those in WKY animals. Under in vitro conditions basal and potassium induced TRH release from SMEs derived from SHR was significantly (P less than 0.05) higher than that from WKY rats, whether expressed in absolute terms or as percent of content. These findings suggest that the thyroid hormone negative feedback regulation of TSH secretion may be impaired in SHR rats. Our data do not allow conclusions as to whether defects in the regulation of TSH production are located exclusively at the hypothalamic level. Since the overproduction of hypothalamic TRH and hypophysial TSH should lead to an increased thyroid hormone biosynthesis other defects in the hypothalamus-pituitary-thyroid-axis may contribute to the abnormal regulation of TSH secretion in SHR rats.     

The thyroid reserve in children with chronic lymphocytic thyroiditis revealed by the thyrotropin-releasing hormone and thyroid-stimulating hormone tests

Serum thyroid hormone and TSH concentrations were measured before and after the administration of TRH (10 micrograms/kg body weight) and bovine TSH (10 IU) in 14 children with chronic lymphocytic thyroiditis. The TRH test showed that the responsiveness of TSH was positively correlated with the basal TSH (P less than 0.001) and inversely with the increase in serum thyroid hormones, for delta T3 (P less than 0.05) and for delta T4 (P less than 0.001). Overall, the patients had significantly lower mean values for basal T4, but not for T3. The TSH test revealed that the delta T3 was positively correlated with delta T4 (P less than 0.05). delta T3 after TSH administration was positively correlated with it after TRH (P less than 0.05). The patients were divided into three groups on the basis of their peak TSH values after TRH administration. In Group 1 (peak value below 40 microU/ml; N = 5); T3 increased significantly after TRH and TSH administrations (P less than 0.05 and P less than 0.025, respectively). In addition, delta T4 was significant after TSH administration. In Group 2 (peak TSH above 40 and less than 100 microU/ml; N = 6); only delta T3 after TRH was significant (P less than 0.05). In Group 3 (peak TSH above 100 microU/ml; N = 3); the response of thyroid hormones was blunted. Thus, the thyroid hormone responses to endogenous TSH coincided with that to exogenous TSH, and the exaggerated TSH response to TRH indicates decreased thyroid reserve.     

Increase of thyrotropin response to thyrotropin-releasing hormone (TRH) and TRH release in rats during pregnancy

Regulation of thyrotropin (TSH) release by thyrotropin releasing hormone (TRH) in the anterior pituitary gland (AP) of pregnant rats was studied. The pregnant (day 7, 14, and 21) and diestrous rats were decapitated. AP was divided into 2 halves, and then incubated with Locke's solution at 37 degrees C for 30 min following a preincubation. After replacing with media, APs were incubated with Locke's solution containing 0, or 10 nM TRH for 30 min. Both basal and TRH-stimulated media were collected at the end of incubation. Medial basal hypothalamus (MBH) was incubated with Locke's medium at 37 degrees C for 30 min. Concentrations of TSH in medium and plasma samples as well as the cyclic 3':5' adenosine monophosphate (cAMP) content in APs and the levels of TRH in MBH medium were measured by radioimmunoassay. The levels of plasma TSH were higher in pregnant rats of day 21 than in diestrous rats. The spontaneous release of TSH in vitro was unaltered by pregnancy. TRH increased the release of TSH by AP, which was higher in pregnant than in diestrous rats. Maternal serum concentration of total T3 was decreased during the pregnancy. The basal release of hypothalamic TRH in vitro was greater in late pregnant rats than in diestrous rats. After TRH stimulation, the increase of the content of pituitary cAMP was greater in late pregnant rats than in diestrus animals. These results suggest that the greater secretion of TSH in pregnant rats is in part due to an increase of spontaneous release of TRH by MBH and a decrease of plasma thyroid hormones. Moreover, the higher level of plasma TSH in rats during late pregnancy is associated with the greater response of pituitary cAMP and TSH to TRH.     

Cyclosporine A inhibits the secretion of certain anterior pituitary hormones in patients with nephrotic syndrome.

To clarify the effects of cyclosporine A (CsA) on the secretion of serum thyrotropin (TSH), prolactin (PRL), luteinizing hormone (LH) and follicular stimulating hormone (FSH), we performed TRH and LH-RH testing in 4 patients with the nephrotic syndrome before and after the administration of CsA, 6 mg/kg/day for 4 to 12 weeks. Prior to CsA all patients responded normally to TRH with respect to TSH and PRL secretion. Two patients showed normal response of LH and FSH to LH-RH stimulation while the response in 2 other patients, who were both menopausal, was exaggerated. By the third or fourth week of CsA administration the basal and peak TSH and PRL values declined significantly in all patients in response to TRH stimulation while those of LH and FSH showed only a modest decrease in response to LH-RH stimulation. Two to 4 weeks after the cessation of CsA the response of TSH, PRL and FSH returned to the pretreatment level. These observations suggest that: 1) CsA exerts an inhibitory effect on the secretion of at least TSH and PRL in humans, and 2) the effect of CsA on the pituitary may be partially reversible after the cessation of the therapy.     

Influence of pimozide on a TRH induced TSH secretion in the rat during food deprivation.

Adult Wistar rats food deprived for 3 days had lower basal levels of TSH compared to normal fed animals. An increase of these lower levels to normal values was obtained following a prolonged (injections during 3 consecutive days) or acute treatment (single injection) with pimozide (1 mg/injection). Blood samples obtained after the last or an only injection of pimozide contained profound increased prolactin levels. Prolactin increase was more than 100-fold in fed and more than 30-fold in starved rats following prolonged pimozide treatment and more than 25-fold and 10-fold following a single injection of pimozide. An injection of 250 ng of TRH increased plasma concentrations of TSH in all groups, but this increase was more pronounced in fasted rats injected with pimozide during 3 consecutive days. It is concluded that fasting results in a dopaminergic inhibition of the sensitivity of the thyrotrophs to a TRH challenge.     

Dissociated response of thyrotropin and prolactin to dopamine receptor blockade with domperidone in hypothyroid subjects.

To investigate the hypothesis of an altered hypothalamic dopaminergic activity in primary hypothyroidism, eight patients with hypothyroidism and seven normal subjects, all female, were studied. All of them were submitted to two tests: TRH stimulation and after the administration of dopamine receptor-blocking drug, Domperidone. The hypothyroid patients with basal TSH values less than or equal to 60 mU/L (4 cases--group 1) had lower PRL levels than the remaining 4 subjects with TSH greater than 60 mU/L (group 2) (p less than 0.001), despite all patients presenting the PRL levels within the normal range. A significant increase occurred for both TSH and PRL after the administration of TRH and Domperidone in normal as well as in the hypothyroid subjects, except for TSH in group 1 after the administration of Domperidone. The area under the curve for PRL response to THR was not different between the normal subjects and both hypothyroid groups, while that under the curve for TSH was greater in the hypothyroidism as a whole than in the normal subjects (p = 0.006) and between the hypothyroid groups, being greater in group 2 than in 1 (p less than 0.009). In relation to Domperidone, the area under the curve for TSH was significantly higher in group 2 when compared to the normal controls (p less than 0.001), while for PRL it was not different between hypothyroid groups in relation to normal controls and when groups I and II were compared. These results suggest that the hypothalamic dopamine activity is not altered in primary hypothyroidism and favor the small relevance of dopamine on the control of TSH secretion.     

Characterization of an in vitro method for demonstrating thyrotropin-releasing hormone-stimulated prolactin secretion.

Anterior pituitaries of normal adult male rats were subjected to synthetic thyrotropin-releasing hormone (TRH) treatment in an acute incubation system which employed pretreatment of the glands with plasma obtained from the donor animals. Following a 60-min preincubation period in a 1:1 mixture of Krebs-Ringer bicarbonate buffer (KRB) and plasma, media and hemipituitary prolactin (PRL) concentrations were significantly (p less than 0.01) increased after a 40-min treatment with 500 pg TRH. The TRH effect was absent among hemipituitaries preincubated in KRB alone. Plasma obtained from older donors was more potent than was plasma from younger rats in this effect. TSH secretion was markedly increased by 500 pg TRH, whether or not plasma preincubation was employed. A dose response of PRL release to concentrations of TRH from 100 pg to 6.0 ng was observed. Crude extracts of median eminence also effected enhanced PRL release using the plasma preincubation technique. The results suggest that plasma preincubation of explanted pituitaries increases PRL cell sensitivity to TRH, perhaps by enzymatic inactivation of endogenous TRH bound to cellular membrane receptors.     

Effect of active and passive immunization with TRH on plasma TSH response to propylthiouracil.

The role of thyrotropin-releasing hormone (TRH) in the secretion of TSH from the anterior pituitary was investigated in rats by active and passive immunization with TRH. The plasma TSH response to propylthiouracil (PTU) in TRH-bovine serum albumin (BSA)-immunized rats was significantly lower than that of BSA-immunized or non-immunized rats. Similarly, the increased plasma TSH level following PTU treatment was significantly suppressed after iv injection of antiserum to TRH. However, the decline in plasma TSH levels was not complete. The results of the present study indicate, at least in part, the physiological significance of endogenous TRH in the regulation of pituitary TSH secretion.     

The effect of medial forebrain bundle lesion on thyrotropin secretion in the rat

The medial forebrain bundle (MFB) was partially lesioned with 6-hydroxydopamine (6-OHDA) in order to investigate the effect of deficient central noradrenergic regulation on thyrotropin (TSH) secretion in the rat. 6-OHDA injection into the MFB significantly reduced the noradrenaline (NA), dopamine (DA) and serotonin (5-HT) content of the whole hypothalamus. NA and 5-HT concentrations were also significantly decreased in the paraventricular nucleus (PVN). The MFB lesion did not affect the clonidine (250 g/kg, i.p.) induced stimulation of TSH release or the isoproterenol (1 mg/kg i.p.) induced decrease in TSH levels. Thyrotropin releasing hormone (TRH, 5 g/kg i.v.) caused a similar significant stimulation of TSH secretion in lesioned and non-lesioned rats. The present results do not support the hypothesis that the blunted TSH response to TRH observed in depressed patients results from a deficiency in noradrenergic neurotransmission.     

Influence of central dopaminergic activity on thyrotropin responsiveness to thyrotropin-releasing hormone in normal and thyrotoxic subjects

To study whether central dopaminergic activity influences TSH responsiveness to TRH in normal individuals and in patients with hyperthyroidism, three experiments (A, B and C) were carried out in 8 normal subjects, and two experiments (A and B) in 8 patients with untreated thyrotoxicosis. In experiment A oral placebo (PBO) preceded iv administration of 200 micrograms TRH by 90 min. In experiment B dopamine receptor blockade with 15 mg oral metoclopramide (MET) was given 90 min before iv administration of 200 micrograms TRH. In experiment C two oral doses (each dose 2.5 mg) of bromocriptine (BCT), known for dopamine agonistic properties, were given 9 and 1 hour before ingestion of 15 mg MET which, in turn, preceded iv injection of 200 micrograms TRH by 90 min. In the healthy subjects experiment A revealed a TSH responsiveness, as reflected by the TSH incremental area, which was 430 +/- 74. The corresponding TSH responsiveness was significantly larger in experiment B (661 +/- 138; P less than 0.02). In experiment C the TSH incremental area (332 +/- 102) did not differ significantly from the one obtained in experiment A. The thyrotrophs responded quite different to TRH in the group of thyrotoxic patients, where the TSH incremental area was zero regardless of whether PBO or MET were given as oral pretreatments. These results imply that central dopaminergic activity inhibits the pituitary thyrotrophs and modulates the TSH response to TRH in healthy subjects, but does not contribute significantly to the blocked TSH responsiveness in patients with untreated hyperthyroidism.     

Effect of prolonged intermittent thyrotropin-releasing hormone administration to fetal and neonatal rats.

Fetal and neonatal rats received daily subcutaneous injections of 10 microgram thyrotropin-releasing hormone (TRH) until 7 or 14 days postnatally. At 70 days the pups were challenged with 1 microgram TRH intravenously via an indwelling jugular cannula. Basal serum thyroxine, triiodothyronine, and thyroid-stimulating hormone (TSH) concentrations did not differ among the three groups. The mean TSH responses as determined by the mean peak TSH concentration and the total TSH response as determined by planimetry were not significantly different, and there was no significant difference in pituitary TSH content following the TRH challenge among the three groups. This study suggests that the integrity of the hypothalamo--pituitary axis in adult rats cannot be affected by the repeated administration of pharmacologic doses of TRH during the perinatal period.     

Physiological regulation of thyrotropin

The pattern of TSH secretion in man in pulsatile in addition to the well known circadian variation. The mechanism triggering TSH pulses remains unclear to date. Infusions of somatostatin or dopamine rapidly lowering basal TSH levels without suppressing the pulsatile pattern suggest that an episodic disinhibition exerted by a physiological inhibitor is not a likely cause. On the same basis, thyroid hormones do not appear to be candidates, since they similarly inhibit basal TSH levels after a time lag of several hours but again do not suppress pulsatile release of the hormone. In contrast, bolus injections of dexamethasone completely abolish pulsatile release of TSH for several hours despite a normal sensitivity of the pituitary to exogenous TRH, suggesting a hypothalamic action of the drug. The hypothesis that pulsatile TSH release might be governed by a pulsatile mode of a hypothalamic stimulator is supported by the observation that an infusion of nifedipine, a calcium channel blocker, which in vitro selectively inhibits the TRH effect on TSH but not prolactin secretion, exerts a comparable effect when it is infused in vivo.