Glucocorticoid Withdrawal Research Articles

Temporally sensitive trophic responsiveness of the adrenalectomized rat anterior pituitary to dexamethasone challenge: relationship between mitotic activity and apoptotic sensitivity.

Depending on timing and dose, exogenous glucocorticoids induce a wave of apoptosis in the adult rat anterior pituitary, a response that is enhanced by adrenalectomy. In this study, we show that the size of the glucocorticoid-sensitive apoptotic population progressively increases during the week following surgical adrenalectomy, plateaus for a further week, then spontaneously declines to levels seen in intact animals by 4 wk. Mitotic activity, in contrast, rises rapidly post adrenalectomy but returns to baseline within 2 wk. Increased mitotic activity precedes the increase in the population of cells that undergo glucocorticoid-induced apoptosis and the subsequent decline in mitotic activity precedes the decline in apoptotic sensitivity despite persistent elevation of hypothalamic CRH and pituitary proopiomelanocortin transcripts. If glucocorticoid exposure is delayed until 4 wk post adrenalectomy when the apoptotic response has returned to baseline, glucocorticoid withdrawal, by transiently increasing mitotic activity, again primes the formation of an expanded glucocorticoid-sensitive apoptotic cell population. These data suggest that apoptotic sensitivity is largely confined to cells that have recently entered the cell cycle. This observation is further corroborated by demonstrating an abrupt glucocorticoid-induced step-down in the bromodeoxyuridine-labeling index to basal levels in rats given daily injections of bromodeoxyuridine during the week following adrenalectomy.

Anterior pituitary trophic responses to dexamethasone withdrawal and repeated dexamethasone exposures.

Glucocorticoid withdrawal, depending on the dose and duration of treatment, results in a transient but sometimes prolonged reduction in hypothalamo-pituitary-adrenal (HPA) axis secretory responsiveness. As the anatomic basis of HPA axis suppression remains uncertain, we have directly examined changes in trophic activity within the rat anterior pituitary gland following dexamethasone withdrawal and re-treatment. Treatment of adrenalectomised, male Wistar rats with dexamethasone results in a discrete, highly significant burst of apoptosis in the anterior pituitary with concurrent suppression of mitosis. Despite a surge in mitotic activity immediately after dexamethasone withdrawal, calculated total anterior pituitary cell populations remain below that seen in untreated adrenalectomised controls. Repeated exposures to dexamethasone show that the dexamethasone-sensitive cell population that is deleted by apoptosis is partially but not completely restored. As the amplitude of apoptotic bursts induced by second and third dexamethasone exposures are similar but smaller than that induced by initial exposure, it appears that the very first exposure to dexamethasone deletes a subset of anterior pituitary cells that are either not restored at all, or are only replaced very slowly. The reduced proportion of corticotrophs contributing to the increase in mitotic index after dexamethasone withdrawal corroborates this. Although continued cell turnover within the pituitary predicts that the absolute cellular deficit would diminish with time, the effects seen may contribute to the delayed recovery of pituitary axis function following cessation of glucocorticoid treatment.

The physiology of corticotropin-releasing hormone deficiency in mice

A review of the generation and characterization of corticotropin-releasing hormone (CRH)-deficient mice is presented. The studies summarized demonstrate the central role of CRH in the pituitary-adrenal axis response to stress, circadian stimulation, and glucocorticoid withdrawal. Additionally, pro-inflammatory actions of CRH at sites of local inflammation are given further support. In contrast, behavioral effects during stress that had been ascribed to CRH action are not altered in CRH-deficient mice. The normal behavioral response to stress in CRH-deficient mice strongly suggests the importance of other, possibly as yet undiscovered, CRH-like molecules.

Pituitary-Adrenal Axis Regulation in Crh-Deficient Mice

Corticotropin-releasing hormone (CRH)-deficient (knockout (KO)) mice demonstrate severely impaired adrenal responses to restraint, ether, and fasting, and lack the normal diurnal glucocorticoid (GC) rhythm. Here, we summarize recent studies determining the role of CRH in augmenting plasma adrenocorticotrophic hormone (ACTH) concentration after glucocorticoid withdrawal and pituitary-adrenal axis stimulation in the context of inflammation. Even though GC insufficient, basal pituitary proopiomelanocortin (POMC) mRNA, ACTH peptide content within the pituitary, and plasma ACTH concentrations are not elevated in CRH KO mice. POMC mRNA content in CRH KO mice increases following adrenalectomy, and this increase is reversed by GC, but not aldosterone, replacement. In marked contrast to the increase in POMC mRNA, plasma ACTH does not increase in the CRH KO mice following adrenalectomy. Administration of CRH to adrenalectomized CRH KO mice results in acute, robust ACTH secretion. Thus, loss of GC feedback can increase POMC gene expression in the pituitary, but CRH action is essential for increased secretion of ACTH into the circulation. While GC secretion is impaired in CRH KO mice after most stimuli, we have found near-normal GC responses to inflammation and systemic immune challenge. Studies in mice with CRH and IL-6 deficiency reveal that IL-6 is essential for activation of the pituitary-adrenal axis during inflammatory and other stressors in the absence of CRH.

Pretreatment with glucocorticoids enhances T-cell effector function: possible implication for immune rebound accompanying glucocorticoid withdrawal.

Glucocorticoids (GCs) exert their immunosuppressive/antiproliferative effects largely through inhibition of cytokine expression, and paradoxically upregulate the expression of (proinflammatory) cytokine receptors on select nonlymphoid cells. Clinically, withdrawal of GCs was frequently associated with worsening of the outcome of heightened immunity disorders, thereby implicating enhanced cytokine and cytokine receptor expression as a possible consequence of acute/short-term GCs withdrawal. In view of the significance of this complication of GC therapy, we addressed the effect of GC withdrawal on cytokine receptor expression and subsequent T-cell effector function, using the proliferation of human T cells as biological readout. To mimic GC withdrawal, T cells were treated with GCs or controls, stimulated, and incubated for 16-20 h at 37 degrees C, washed, and reactivated for a further 4-48 h. Surface marker expression was assessed by FACS analysis, and proliferation was determined by measuring the cellular uptake of tritiated thymidine. Dexamethasone (DEX) and prednisolone (PRED), in a concentration-dependent manner, inhibited T-cell proliferation induced by anti-CD28 Ab + PMA. However, pretreatment of T cells activated with mitogens, cross-linking antibodies, or PMA + ionomycin ("CD3-bypass" stimulation regimen), but not resting T cells, with DEX or PRED resulted in a marked increase in IL-IR, IL-2R alpha, and IL-6R expression, which was accompanied by a significant enhancement in T-cell proliferation. This effect of GCs was neither stimulus specific nor did it result from alteration in cell viability, and was paralleled by augmentation in cytokine (rIL-2) effects on DEX-pretreated and preactivated T cells. Taken together, our results underline the dual effects of GCs in regulating T-cell activation and cytokine expression. In essence, GCs directly inhibited T-cell proliferation by suppressing cytokine production, and, by enhancing cytokine receptor expression, pretreatment with GCs augmented T-cell proliferation.

Regulation of pituitary corticotropin releasing hormone (CRH) receptor mRNA and CRH binding during adrenalectomy: role of glucocorticoids and hypothalamic factors.

The role of glucocorticoids and hypothalamic factors on CRH receptor expression in the pituitary were studied by analysis of the effects of adrenalectomy and suppression of CRH and VP secretion by hypothalamic lesions in the rat. Consistent with previous in situ hybridization studies, Northern blots showed that pituitary CRH receptor mRNA decreased only transiently after adrenalectomy, falling to 51% of the control levels after 18 h, and returning to control values after 6 days (112%). The early decrease was prevented by dexamethasone injection, 100 micrograms, s.c. The role of increased levels of CRH and VP in the pituitary portal circulation on the transient decrease in CRH receptor mRNA levels after adrenalectomy were studied by in situ hybridization in rats subjected in PVN lesions or median eminence deafferentation by hypothalamic anterolateral cuts (ALC). PVN lesion (12 days) or ALC (8 days) resulted in undetectable irCRH and VP in the external zone of the median eminence and had no effect on basal levels of pituitary POMC mRNA, CRH binding and CRH receptor mRNA. In sham lesioned rats, adrenalectomy for 18 h or 4 days caused the expected increases in pituitary POMC hnRNA and mRNA, and decreases in CRH binding. CRH-R mRNA levels decreased by about 50% after 18 h adrenalectomy but returned to basal by 4 days. PVN lesion or ALC fully prevented the fall in CRH binding after 18 h or 4 days adrenalectomy and the increase in POMC mRNA after 4 days adrenalectomy, whereas only attenuated the decrease in CRH receptor mRNA and increase in POMC mRNA levels after 18 h adrenalectomy. Administration of a CRH antagonist did not affect CRH receptor mRNA and POMC hnRNA and mRNA indicating that residual CRH in the median eminence after hypothalamic surgery is not responsible for the effect of adrenalectomy. These studies confirm previous in situ hybridization studies showing that adrenalectomy causes transient decreases in pituitary CRH receptor mRNA levels. The data indicate that while increases in hypothalamic CRH secretion following glucocorticoid withdrawal mediate pituitary CRH receptor binding loss and the increase in POMC expression after long-term adrenalectomy, CRH only partially accounts for the early changes in CRH receptor mRNA and POMC mRNA.

Antiglucocorticoids as Treatments for Depression : Rationale for Use and Therapeutic Potential.

There is considerable interest in the possible antidepressant properties of antiglucocorticoids. Studies of the hypothalamic-pituitary-adrenal (HPA) axis in depression, coupled with increasing recognition of the importance of glucocorticoids in brain function, strongly support such interest.Psychiatric symptomatology in Cushing's syndrome (a disorder involving hypersecretion of cortisol), the adverse cognitive effects of glucocorticoids, and the suppressive effects of conventional antidepressants on HPA function all suggest that HPA abnormalities could cause or contribute to depression. However, the psychiatrie effects of glucocortieoids and glucocorticoid withdrawal suggest that HPA hyperactivity compensates for some other defect. While preliminary clinical studies suggest that both glucocorticoids and antiglucocorticoids may have efficacy in depression, the toxicity and loss of efficacy of available drugs during long term administration will probably necessitate the development of new agents to advance this field.

Altered body composition and fuel metabolism in stable kidney transplant patients on immuno-suppressive monotherapy with cyclosporine A.

Glucocorticoid associated altered body fat distribution and muscle wasting are well known following kidney grafting. Whether an immunosuppressive regimen after glucocorticoid withdrawal (i.e. monotherapy with cyclosporine A (CsA)) is associated with normalization of altered body fat distribution and muscle mass remains to be determined. Therefore 18 renal transplant patients (nine males and nine females, 64 +/- 5 (mean +/- SEM) months since transplantation; CsA-monotherapy: 38 +/- 7 months) and 18 age, sex and body mass index matched healthy volunteers were investigated using indirect calorimetry and dual energy X-ray absorptiometry. Lean body mass (LBM) was decreased in patients mostly due to loss of striated muscle in the legs (P < 0.01). Compared to healthy controls, fat mass was increased in head and trunk (P < 0.01) and similar in extremities. Resting energy expenditure expressed per kg LBM was increased by more than 10% (P < 0.05) in patients vs. controls. Plasma insulin and glucose concentrations, total serum cholesterol (C), triglyceride levels and the ratio of LDL-C to HDL-C were all elevated (P < 0.01) in patients as compared with controls. In summary, renal transplant patients on immunosuppressive monotherapy with CsA demonstrate decreased muscle mass despite discontinuation of prednisone therapy. The increased upper body fat might account, at least in part, for peripheral hyperinsulinaemia and dyslipidaemia observed in kidney transplant patients even years after successful transplantation.

Possible involvement of glucocorticoids in the reduction of serum insulin levels by interleukin-1 in the rat.

Evidence is accumulating that adrenal steroids may be involved in the metabolic effects of cytokines. We evaluated the possible involvement of glucocorticoids in the inhibition of pancreatic insulin secretion by interleukin-1 beta (IL-1 beta), one of the cytokines produced by inflammatory cells. In the first group of experiments, adrenalectomized rats showed a significant reduction in basal and glucose (0.5 g/kg, i.v.)-stimulated immunoreactive insulin (IRI) levels after injection of IL-1 beta (1.0 microgram/kg), but intact rats did not. Pretreatment with IL-1 beta increased plasma glucose levels 2 and 15 min after an i.v. bolus of glucose in adrenalectomized rats. In the second group of experiments, dexamethasone supplement (0.1 mg/kg) given to adrenalectomized rats cancelled the reduction in plasma glucose levels by IL-1 beta, and rats treated with 1.0 mg dexamethasone/kg showed a significant increase in basal IRI levels and enhanced serum IRI levels after IL-1 beta injection. However, 1.0 mg deoxycorticosterone/kg given daily for 7 days failed to cancel the effect of IL-1 beta on the reduction of serum IRI levels, although it attenuated the weight loss after adrenalectomy. The data suggested that withdrawal of glucocorticoids after adrenalectomy potentiates the effect of IL-1 beta on the reduction of serum IRI levels. Glucocorticoids may have a protective action against the reduction of serum IRI levels by IL-1 beta.

Corticotrophin-releasing factor and arginine vasopressin in the hypothalamo-hypophyseal portal blood of rats following high-dose glucocorticoid treatment and withdrawal

Hypothalamo-hypophyseal portal blood was obtained from rats treated with chronic, high-dose prednisolone in the drinking water and after subsequent withdrawal of the steroid for up to 7 days. Corticotrophin-releasing factor (CRF) immunoreactivity in portal blood was reduced by treatment with prednisolone but not completely abolished. There was a rapid recovery of CRF to control values within 3 days of withdrawal of prednisolone. There was no significant change in arginine vasopressin (AVP) in portal blood over the time-course of the experiment.

Stress responsiveness of hypothalamic corticotrophin-releasing factor and pituitary pro-opiomelanocortin mRNAs following high-dose glucocorticoid treatment and withdrawal in the rat

In-situ hybridization with synthetic oligonucleotide probes was used to determine the mRNA content of corticotrophin-releasing factor (CRF) and proenkephalin A mRNA in the paraventricular nucleus, and of pro-opiomelanocortin (POMC) mRNA in the anterior pituitary gland of male rats immediately after, and during recovery from, chronic high-dose prednisolone treatment. Levels of transcripts for mRNA for both CRF and POMC were markedly reduced after the treatment, but there was a rapid return to control values for CRF mRNA within 18 h of steroid withdrawal. In untreated animals, the stressful stimulus of i.p. hypertonic saline increased CRF and proenkephalin A mRNA within 4 h with no significant difference in response seen whether the tissues were removed at 13.00 or 20.00 h. The increase in POMC mRNA did not reach statistical significance in these animals. Although prednisolone resulted in a marked reduction of basal CRF mRNA, the stress-induced increment of CRF mRNA remained comparable with that found in untreated animals. On the day following cessation of prednisolone treatment at 09.00 h, basal and stress levels of CRF mRNA were significantly higher in rats killed at 20.00 h than at 13.00 h. Proenkephalin A mRNA transcripts were below quantifiable levels of detection in control or non-stressed prednisolone-treated animals at all the time-points studied. Stress, however, resulted in the accumulation of proenkephalin A mRNA in control animals. This response was inhibited by prednisolone treatment and only returned 18 h after withdrawal. Prednisolone treatment reduced POMC mRNA below the levels detected in untreated animals, with no detectable response to stress.(ABSTRACT TRUNCATED AT 250 WORDS)

Regulation of liver angiotensinogen mRNA by glucocorticoids and thyroxine

This study examines the immediate and longer-term changes in angiotensinogen mRNA and in plasma angiotensinogen which follow the withdrawal and replacement of glucocorticoids or thyroxine. RNA from rat liver was analysed by Northern blot and dot-blot hybridization with a 40-mer oligodeoxynucleotide probe specific for angiotensinogen mRNA. Adrenalectomy decreased plasma angiotensinogen and angiotensinogen mRNA to 55 and 50% of control values respectively over a period of 16 days. Similar decreases were obtained after propylthiouracil (1 mg/kg for 16 days) treatment, except when injected simultaneously with T 3 (3 μg/kg/day). Over the same period plasma renin activity increased in adrenalectomized rats from 4.1 ± 0.8 to 7.0 ± 1.5 pmol angiotensin I (AI)/ml/h, and decreased in propylthiouracil-treated rats from 3.8 ± 0.4 to 1.6 ± 0.4 pmol AI/ml/h. Approximate half-times of 2–3 days were calculated for both plasma angiotensinogen and angiotensinogen mRNA post-adrenalectomy or after propylthiouracil treatment. Dexamethasone (400 μg/kg i.m.) given to intact rats rapidly increased angiotensinogen mRNA to a maximum of 250% of control by 8 h and with a half-maximal response of 2.8 h. Plasma angiotensinogen responded similarly, apart from an initial delay of 2 h. Treatment with different doses of propylthiouracil and dexamethasone showed that responses were dose-related. We conclude that changes in plasma angiotensinogen and angiotensinogen mRNA are closely correlated, and that under various physiological circumstances angiotensinogen mRNA has a rapid rate of accumulation but a slow rate of decay.

Regulation of plasma fibronectin biosynthesis by glucocorticoids in chick hepatocyte cultures.

Plasma fibronectin is an acute-phase reactant synthesized by hepatocytes. Glucocorticoids are one of the major factors implicated in controlling the hepatic acute-phase response. To study the regulatory effects of glucocorticoids on plasma fibronectin biosynthesis, a model chick hepatocyte culture system under serum- and hormone-free conditions was used. In the presence of either dexamethasone or corticosterone, secreted plasma fibronectin increased maximally to 2.8-fold above basal levels. The stimulatory effect of the hormones was maintained only in their continuous presence, since plasma fibronectin production dropped to near basal levels within 16 h of glucocorticoid withdrawal. Pulse-chase studies indicated that pretreatment of cells with dexamethasone stimulated the level of secreted plasma fibronectin but had no effect on its rate of secretion. The increase in plasma fibronectin production by dexamethasone was abolished in a dose-dependent manner by the addition of progestin, an antagonist of dexamethasone known to compete specifically for the liver glucocorticoid receptor. Actinomycin D and alpha-amanitin, which are inhibitors of transcription, also blocked the early dexamethasone effect on plasma fibronectin synthesis. Slot blot hybridization of total RNA samples from dexamethasone-treated cultures revealed a 6-fold stimulatory rise in fibronectin mRNA during the first 6 h of treatment, which later declined and was no longer evident at 48 and 72 h. However, fibronectin mRNA levels were elevated to about the same extent in control and dexamethasone-treated cells at the later time points. During the same time period (0 to 72 h), plasma fibronectin protein levels rose and remained elevated. Evaluation of pulse-chase experiments following pretreatment with hormone for 48 h demonstrated that equal amounts of plasma fibronectin were translated by dexamethasone-treated and control cells, but only 42% of labeled plasma fibronectin was secreted by control cells compared with 93% for dexamethasone-treated cells. These findings suggest that the early phase of glucocorticoid regulation of plasma fibronectin biosynthesis occurs at the transcriptional level and is mediated through the specific action of the glucocorticoid receptor. A later phase of glucocorticoid-stimulated plasma fibronectin biosynthesis results from modulation of post-translational processing events leading to secretion of an increased amount of newly translated plasma fibronectin polypeptides.

Inhibition by glucocorticoids of phosphate transport in primary cultured renal cells.

The regulation by glucocorticoids of phosphate transport in primary cultured chick renal cells was examined. Dexamethasone inhibited the Na+-dependent phosphate uptake system. Na+-independent phosphate uptake and Na+-dependent uptakes of alpha-methylglucoside and L-proline were unaffected. The mineralocorticoid aldosterone did not alter phosphate uptake. The inhibition of Na+-dependent phosphate uptake by dexamethasone was concentration-dependent, exhibited an induction period, was blocked by inhibitors of RNA and protein synthesis, and was rapidly reversed when the steroid was removed. Following reversal, the cells could respond a second time to the glucocorticoid. However, this time the response was rapid, could be evoked at least for 24 h after glucocorticoid withdrawal, and might be prevented by actinomycin D and cycloheximide. These findings demonstrate that glucocorticoids act on renal cells to modulate phosphate transport and suggest that the renal cell system provides an attractive model to examine the mechanism by which glucocorticoids control gene expression and regulate plasma membrane transport function.

Thyroid function in adrenocortical insufficiency during withdrawal and re-administration of glucocorticoid substitution.

The regulatory function of glucocorticoids on thyroid hormone concentrations was studied in patients with adrenocortical insufficiency (ACI, n = 8) and in healthy subjects (n = 6). In patients with ACI withdrawal of glucocorticoid substitution for 84 h led to an increase in serum concentrations of total triiodothyronine (TT3) from 110 +/- 20 to 133 +/- 22 ng/dl and a decrease of serum reverse-T3 (rT3) from 23 +/- 6 to 18 +/- 6 ng/dl, whereas subsequent administration of dexamethasone (0.5 mg po q.i.d. for 3 days) induced a fall in TT3 (129 +/- 22 to 88 +/- 16 ng/dl) and a rise in rT3 (17 +/- 5 to 37 +/- 11 ng/dl) concentrations. Serum levels of total thyroxine (TT4) were unchanged by either withdrawal or re-administration of glucocorticoids. Basal plasma thyrotrophin (TSH) concentrations were unchanged by glucocorticoid withdrawal and fell from 2.2 +/- 1.5 to 1.1 +/- 0.8 mU/l during subsequent dexamethasone therapy. In healthy subjects a decrease of TT3 (89 +/- 8 to 69 +/- 8 ng/dl) and an increase in rT3 (19 +/- 5 to 32 +/- 8 ng/dl) concentrations were seen after stimulation of endogenous cortisol production by prolonged infusion of ACTH1-24 (0.5 mg, t = 8 h on 2 consecutive days), whereas concentrations of TT4 remained unchanged. During subsequent administration of dexamethasone serum level of both. TT3 and rT3 returned to basal levels. Thus, changes in thyroid hormone concentrations are induced by alterations of substitution therapy in patients with ACI. In healthy subjects the application of 2 mg dexamethasone/day following prolonged maximal stimulation of endogenous cortisol by iv ACTH may represent a state of relative glucocorticoid deficiency, thus explaining the observed hormonal changes, which are inverse to those generally induced in healthy man by endogenous or exogenous glucocorticoids.

Triphasic changes in plasma ACTH concentration and brain serotonin synthesis rate following adrenalectomy in rats.

Following bilateral adrenalectomy in adult male rats, there occurs a pattern of triphasic change in basal plasma concentration of radioimmunoassayable ACTH. Plasma ACTH is markedly elevated at 2 h, has returned down almost to normal at 20 h and is again markedly elevated 96 h after adrenalectomy. We have examined serotonin (5HT) synthesis rats in several brain regions, anterior hypothalamus, posterior hypothalamus, and brain stem, at these times after adrenalectomy using the accumulation of 5-hydroxytryptophan (5HTP) after inhibition of aromatic L-amino acid decarboxylase with m-hydroxybenzylhydrazine. In both anterior hypothalamus and brain stem, decrease 5HT synthesis rates were observed at 2 and 96 h after adrenalectomy, but at 20 h 5HT synthesis rates were normal. This pattern was not observed in the posterior hypothalamus. Thus, we demonstrated inverse correlations between 5HT synthesis rates in anterior hypothalamus and brain stem, but not posterior hypothalamus, and basal plasma ACTH concentration throughout the period of triphasic change following adrenalectomy in adult male rats. Both the adrenalectomy-induced increases in plasma ACTH concentration and the adrenalectomy-induced decreases in brain 5HT synthesis rates were inhibited by treatment with dexamethasone, suggesting that the changes resulted from glucocorticoid withdrawal. The data are consistent with a role of brain 5HT neurons with cell bodies in brain stem and nerve endings in anterior hypothalamus in the regulation of the triphasic changes in plasma ACTH concentration following adrenalectomy in rats.

Antibody to 1-24 ACTH and Hypofunction of the Adrenal Cortex in a Patient who was Unilaterally Adrenalectomized for Cushing's Syndrome

We report here on a patient who was unilaterally adrenalectomized for Cushing's syndrome, and who developed antibodies to 1-24 ACTH. A 49-year-old nurse had undergone right adrenalectomy for adrenal adenoma. After surgery, she was treated with 0.5 mg of 1-24 ACTH-Z together with glucocorticoid replacement therapy for 40 days. Thereafter she was given 0.25-0.5 mg of ACTH-Z every other day for 4 months. ACTH-Z was then stopped for a year but glucocorticoid therapy was continued. About one year prior to this admission, 1 mg of ACTH-Z was again initiated 1 to 2 times a week. Glucocorticoid therapy was not withdrawn during the four years after adrenalectomy. She was admitted for the purpose of withdrawal of glucocorticoids. Her serum was found to bind labeled ACTH. This labeled ACTH was competitively displaced from binding by unlabeled hormones. Finally, reaction with specific antihuman Ig demonstrated an antibody of the IgG class. The titer of the antibodies gradually decreased after the discontinuation of ACTH-Z, but it is still present in measurable quantity in her serum. The clinical significance of the circulating anti-ACTH antibody in her serum is discussed.

The recovery of the hypothalamo-pituitary-adrenal axis after transsphenoidal operation in three patients with Cushing's disease. The effect of prior external pituitary irradiation.

Abstract. The recovery of the hypothalamo-pituitary-adrenal axis after selective transsphenoidal adenomectomy was studied in 3 patients with Cushing's disease by measuring basal plasma ACTH and cortisol concentrations, cortisol secretion rate, the diurnal rhythm of cortisol, and the reaction of cortisol to lysine vasopressin (LVP), of compound S to metyrapone and of cortisol and growth hormone to an insulin-induced hypoglycaemia. The third patient had been treated previously by external pituitary irradiation. In 2 patients basal plasma ACTH levels returned within normal values before plasma cortisol, but no supra-physiological plasma concentrations of ACTH were seen as has been observed after withdrawal of exogenous glucocorticoids. With regard to the different stimulation tests: at first the normal reaction of plasma cortisol to LVP returned after 3 months, at the same time as the restoration of growth hormone secretion in response to hypoglycaemia. A normalization of the reaction to metyrapone was seen thereafter while finally the reaction of cortisol to an insulin-induced hypoglycaemia and the diurnal rhythm of plasma cortisol returned 15 to 18 months after operation in the first patient and after 12 months in the second patient. Selective adenomectomy had also been carried out in the third patient, as evidenced by normal TSH, LH and FSH secretion. Hypocortisolism, and a deficient ACTH and growth hormone secretion in response to the stimuli mentioned, however, did not normalize up till 22 months after operation. The restoration of the hypothalamo-pituitary-adrenal axis after selective pituitary adenomectomy in Cushing's disease was prevented in this patient by prior external pituitary irradiation.

Brain serotonin turnover correlates inversely with plasma adrenocorticotropin during the triphasic response to adrenalectomy in rats.

After bilateral adrenalectomy in adult male rats a triphasic change occurs in the plasma concentration of radioimmunoassayable ACTH. Plasma ACTH is markedly elevated at 2 h, returns almost to normal at 20 h, and is again markedly elevated 92 h after adrenalectomy. We have examined brain serotonin (5HT) turnover during this period using two nonsteady state methods: accumulation of 5HT after monoamine oxidase inhibition with pargyline and decline of 5-hydroxyindoleacetic acid (5HIAA) after pargyline. Both endpoints demonstrated decreases in hypothalamic and brain stem 5HT turnover 2 and 92 h after adrenalectomy, but normal 5HT turnover 20 h after adrenalectomy. Thus, we demonstrated inverse relationships between 5HT turnover in both the hypothalamus and brain stem and the plasma ACTH concentration throughout the period of triphasic change after adrenalectomy. The adrenalectomy-induced increases in plasma ACTH and decreases in brain 5HT turnover both 2 and 92 h after adrenalectomy are inhibited by treatment with small doses of corticosterone. The data strongly suggest that the activity of some brain 5HT neurons changes after adrenalectomy in a triphasic pattern and, further, that these changes are related in part to glucocorticoid withdrawal. 5HT receptor antagonists blunted corticosterone inhibition of the adrenalectomy-induced decreases in brain 5HT turnover, providing further evidence for an interaction between glucocorticoids and brain 5HT neurons. The data are consistent with a role of brain 5HT neurons in the adrenalectomy-induced triphasic changes in ACTH secretion.

Nucleus-dependent regulation of tyrosine aminotransferase degradation in hepatoma tissue culture cells.

Upon removal of the nucleus from rat hepatoma tissue culture cells, levels of the enzyme tyrosine aminotransferase no longer change in response to withdrawal of glucocorticoids. The rate of tyrosine aminotransferase degradation is drastically reduced in rat hepatoma tissue culture cytoplasts leading to stabilization of pre-existing levels of tyrosine aminotransferase. Moreover, the rate of synthesis of the enzyme in cytoplasts is very low near that observed in uninduced whole cells. These effects of enucleation occur very rapidly and appear to be specific for tyrosine aminotransferase and a small number of other unstable hepatoma proteins. A nuclear effect is thus directly involved in the control of tyrosine aminotransferase degradation and synthesis.