Differential effect of aging on serum levels of prolactin and alpha-melanotropin in rats.

Prolactin (PRL) and alpha-melanocyte-stimulating hormone (alpha-MSH) are the only two pituitary hormones whose basal secretion is under tonic dopaminergic inhibition exerted by the hypothalamus. In the female rat, continuous exposure to estrogens is believed to depress hypothalamic dopaminergic activity and lead to the appearance of PRL-secreting pituitary adenomas during aging. Since there is no information about the impact of aging on circulating alpha-MSH levels, it was of interest to assess and compare the serum levels of PRL and alpha-MSH in male and female rats of different ages. Young (3-4 months) and old (24-25 months) male and female Sprague-Dawley rats as well as senescent (33-35 months) females were killed by decapitation between 10 AM and 1 PM, and pituitaries were immediately removed and dissected. Hormones were measured in unextracted trunk serum by radioimmunoassay. Serum PRL levels were (mean +/- SE), 18.4 +/- 2.0, 26.8 +/- 3.8, 19.8 +/- 2.5, 43.0 +/- 7.5, and 193.5 +/- 47.6 ng/ml for young and old males, and young, old, and senescent females, respectively. Serum alpha-MSH levels were 243.2 +/- 15.2, 252.9 +/- 24.8, 320.0 +/- 31.3, 234.7 +/- 19.1, and 374.0 +/- 29.7 pg/ml for young and old males, and young, old and senescent females, respectively. Anterior pituitary and neurointermediate lobe weights increased significantly with age in both sexes, although the change was particularly conspicuous in the females. We conclude that aging does not have a major impact on circulating alpha-MSH levels in rats and that melanotrophs probably have a greater ability than prolactotrophs to withstand age-associated alterations in central regulatory mechanisms.     

Sexual differences in the activity of periventricular-hypophysial dopaminergic neurons in rats.

The activities of periventricular-hypophysial dopaminergic (DA) neurons were compared in male and female rats by measuring dopamine synthesis (accumulation of 3,4-dihydroxyphenylalanine [DOPA] after inhibition of L-aromatic amino acid decarboxylase) and metabolism (concentrations of 3,4-dihydroxyphenylacetic acid [DOPAC]) in terminals of these neurons in the intermediate lobe of the pituitary. For comparison, the synthesis and metabolism of dopamine in the neural lobe of the pituitary and median eminence were also determined. The concentrations of DOPAC and accumulation of DOPA were higher in females than in males in both the intermediate lobe and median eminence, revealing a sexual difference in the basal activity of periventricular-hypophysial and tuberoinfundibular DA neurons. In contrast, there were no differences between male and female rats in activity of DA neurons terminating in the neural lobe. One week following gonadectomy, DOPA accumulation in the median eminence was decreased in females and increased in males, but remained unchanged in the intermediate lobe. These results indicate that sexual differences in the activity of periventricular-hypophysial DA neurons terminating in the intermediate lobe are not dependent upon the presence of circulating gonadal steroids, and in this respect, these neurons differ from tuberoinfundibular DA neurons.     

Association of dopaminergic fibers with corticotropin releasing hormone (CRH)-synthesizing neurons in the paraventricular nucleus of the rat hypothalamus

Summary Catecholamines are known to exert a central influence on the hypothalamo-hypophyseal-adrenal neuroendocrine system. The selective dopaminergic innervation of the hypothalamic paraventricular nucleus (PVN) and putative relationships between dopaminergic fibers and corticotropin releasing hormone (CRH)-synthesizing neurons were studied in the male rat by means of immunocytochemistry following the elimination of noradrenergic and adrenergic inputs to the hypothalamus. A 3.0-mm-wide coronal cut was placed unilaterally in the brain at the rostral level of the mesencephalon. All neuronal structures from the cortex to the ventral surface of the brainstem, including the ascending catecholaminergic fiber bundles were transected. This surgical intervention resulted in the accumulation of dopamine--hydroxylase (DBH)-immunoreactivity in axons proximal to the cut, and an almost complete disappearance of DBH activity in those located distal to the lesion. Two weeks following the operation, DBH immunoreactivity was significantly diminished in the PVN located on the side of lesion, while tyrosine hydroxylase (TH)-immunoreactivity was present in a substantial number of fibers in the same nucleus. Both DBH- and TH-immunoreactive axons were preserved in the contralateral PVN. Simultaneous immunocytochemical localization of either DBH- or TH-IR fibers and corticotropin releasing hormone-synthesizing neurons in the hypothalami from brainstem-lesioned, colchicine treated animals revealed that the distribution of catecholaminergic fibers and CRH neurons is homologous within the PVN of the intact side. Only a few scattered DBH-immunoreactive axons were detected among CRH-producing neurons in the PVN on the side of the lesion. In contrast, many tyrosine hydroxylase containing neurons and neuronal processes were observed on the lesioned side and the TH-IR fibers established juxtapositions with CRH-synthesizing neurons.These morphological data demonstrate that following the surgical ablation of noradrenergic and adrenergic afferents to the PVN, a substantial number of tyrosine hydroxylase-IR fibers remained in the nucleus and they were associated with corticotropin releasing hormone synthesizing neurons. Therefore, it is hypothesized that the paraventricular nucleus receives a selective dopaminergic innervation and these dopaminergic axons might influence the function of the pituitary and adrenal glands via the hypothalamic CRH system.Supported by grants from the National Science Foundation (NSF INT 8703030), the Hungarian Academy of Sciences (OTKA 104), the National Institutes of Health (NS 19266) and the National Foundation of Technical Development (OKKFT Tt 286/1986)     

Effects of atropine sulfate on the hypothalamic LH-RH cells of the guinea-pig]

A single 30 mg/kg., s.c. injection of atropine sulfate into male guinea pigs produces a storage of LH-RH immunoreactive material in hypothalamic neurons. In diestrus female there is no increase of immunoreactive LH-RH substance in neuron and in eminential terminals; the rate of LH-RH synthesis may be still slight. The LH-RH immunoreactive neuronal increase, in male may be related to an atropine blockade of LH-RH portal cession, perhaps via infundibular dopaminergic neurons.     

Dissecting the genetic effect of the CRH system on anxiety and stress-related behaviour

Corticotropin-releasing hormone (CRH) plays a central role in the adaptation of the body to stress. CRH integrates the endocrine, autonomic and behavioural responses to stress acting as a secretagogue within the line of the hypothalamic pituitary adrenocortical (HPA) system and as a neurotransmitter modulating synaptic transmission in the central nervous system. Accumulating evidence suggests that the neuroendocrine and behavioural symptoms observed in patients suffering from major depression are at least in part linked to a hyperactivity of the CRH system. Genetic modifications of the CRH system by conventional and conditional gene targeting strategies in the mouse allowed us to study the endogenous mechanisms underlying HPA system regulation and CRH-related neuronal circuitries involved in pathways mediating anxiety and stress-related behaviour.     

Is progesterone a pre-hormone in the CNS?

In this paper, experimental evidences have been presented indicating that progesterone per se appears to be a powerful modulatory steroid of presynaptic striatal dopaminergic terminals of the central nervous system of the rat. This effect of the progesterone signal is concentration as well as infusion mode dependent. Low pulsatile doses of the steroid positively modulate the mechanism by which dopamine terminals respond to amphetamine stimulation and increase tissue dopamine concentration. Whereas, continuous and/or high doses of this steroid negatively modulate the response of the dopamine terminals to amphetamine stimulation and decreases tissue dopamine concentration. This effects occurs through a membrane mediated mechanism either upon the dopamine neuron directly and/or upon an interneuron. Pregnanolone a 5- beta-3 beta-metabolite of progesterone known to activate the hypothalamic LHRH neural apparatus at the level of the hypothalamus of ovariectomized estrogen primed rats in both in vitro as well as in vivo preparations was completely ineffective at the level of the corpus striatum of similar animal preparations. Therefore, it is reasonable to assume that site specific mechanisms exist within the central nervous system which may control differentially the final action of progesterone. In the hypothalamus, pregnanolone appears to be the final signal for its action on the LHRH neural apparatus, whereas in the corpus striatum, the steroid per se, and dependent on the modality and/or the strength of the signal can either directly or indirectly up-regulate (stimulatory component) or down-regulate (inhibitory component) the activity of striatal dopaminergic terminals.     

Adrenal inhibition of corticotropin-releasing hormone-induced thyrotropin release: a comparative study in pre- and posthatch chicks

Recent evidence indicates that corticotropin-releasing hormone (CRH) acts as a potent stimulator of thyrotropin (TSH) release in the chicken. In this study adrenal and thyroidal feedback mechanisms were studied. Administration of corticosterone 30 min prior to an ovine CRH (oCRH) challenge diminished the in vivo sensitivity of thyrotrophs to oCRH in 19-day-old chicken embryos (E19) (20 micrograms corticosterone; 2 micrograms oCRH) but not in 8-day-old chickens (C8) (40 micrograms corticosterone; 4 micrograms oCRH). At both ages studied, corticosterone (0.01 and 1 microM) did not alter the in vitro TSH response to oCRH (100 nM) indicating that an indirect mechanism is involved at the embryonic stage which is no longer present in posthatch chickens. In vitro, 3,5,3'-triiodothyronine (T3) pretreatment (0.01 and 1 microM) resulted at both ages studied in a dose-dependent drop in the in vitro oCRH-induced TSH release. As recorded previously, corticosterone treatment provoked a rise in plasma T3 in embryonic but not in posthatch chickens. The presence of an indirect adrenal feedback mechanism in chicken embryos may therefore be linked to the increase in plasma T3 which will alter the sensitivity of thyrotrophs to hypothalamic releasing factors. In conclusion, corticosterone does not directly modulate the responsiveness of thyrotrophs to CRH, but its feedback mechanism may be dependent on the evoked increase in plasma T3 which is only present in embryonic chickens. Corticosterone may in this regard play an essential role during embryonic development by coordinating thyroidal feedback mechanisms at the level of the chicken pituitary.     

Neurotensin-induced activation of hypothalamic dopaminergic neurons is accompanied by a decrease in pituitary secretion of prolactin and alpha-melanocyte-stimulating hormone.

The effects of neurotensin on the activity of hypothalamic tuberoinfundibular and periventricular-hypophysial dopaminergic (DA) neurons, and on the secretion of pituitary hormones that are tonically regulated by these neurons (i.e. prolactin and alpha-melanocyte-stimulating hormone [alpha MSH], respectively) were examined in estrogen-primed ovariectomized rats. The activity of tuberoinfundibular and periventricular-hypophysial DA neurons was estimated by measuring concentrations of the dopamine metabolite 3,4-dihydroxyphenylacetic acid (DOPAC) in the terminals of these neurons in the median eminence and intermediate lobe of the posterior pituitary, respectively. Intracerebroventricular administration of neurotensin caused a dose- and time-related increase in DOPAC concentrations in both the median eminence and intermediate lobe, and a concurrent decrease in plasma levels of prolactin and alpha MSH. These results suggest that neurotensin-induced inhibition of secretion of prolactin and alpha MSH from the pituitary may be due to the stimulatory action of this neuropeptide on the release of dopamine from tuberoinfundibular and periventricular-hypophysial neurons.     

Administration of gonadal steroids to neonatal rats affects beta-endorphin levels in the adult

Administration of gonadal steroids to neonatal rats has a profound effect on the function of the neuroendocrine system in the adult animal. Considering that gonadal steroids modulate hypothalamic and pituitary levels of beta-endorphin (BE) in adult male and female rats, the effects of neonatal gonadal steroid treatment on BE levels in the adult animal were investigated. Neonatal male rats were administered testosterone and neonatal female rats were treated with estrogen. Matched control littermates received vehicle. All animals were sacrificed at 90 days of age. Neonatal gonadal steroid treatment did not affect the level of immunoreactive beta-endorphin (IR-BE) in the anterior pituitary (AP) of male rats but did result in a significant increase in IR-BE in the AP of female rats. Neonatal administration of gonadal steroids produced a significant decrease in IR-BE in the neurointermediate lobe of the pituitary (NIL) of both male and female rats, with the magnitude of the decrease being greater in the NIL of the female rats. IR-BE levels in the hypothalamus of male or female rats were not altered by the treatments. Column chromatography indicated that the increase in IR-BE in the AP represented a proportional increase in BE and beta-lipotropin, while the reduction in IR-BE in the NIL of the treated rats represented a reduction in BE. These findings suggest that gonadal steroids may influence the development of the neurotransmitter systems which regulate BE levels in the adult pituitary, the development of the biosynthetic mechanisms of the adult pituitary, or both.     

Localization and pharmacology of some dopamine receptor complexes in the striatum and the pituitary gland: synaptic and son-synaptic communication

The striatum receives massive dopaminergic projections from neurons in the ventral tegmental area, the substantia nigra and the retro-rubral cell group. Dopaminergic neurons in the arcuate nucleus and periventricular hypothalamic nuclei project to the median eminence and the neuro-intermediate lobe of the pituitary gland. The anterior lobe of the pituitary gland is not innervated by dopaminergic neurons, but receives dopamine via a vascular route from the median eminence. Two categories of dopamine receptors (D-1 and D-2) can be identified on the basis of the ability of various drugs to discriminate between these two entities. Dopamine stimulates both D-1 and D-2 receptors. The affinity of dopamine for the D-2 receptor is approximately 1000 times higher than for the D-1 receptor. Dopamine is involved in synaptic as well as non-synaptic communication. Examples of non-synaptic communication via D-2 receptors are the dopamine induced inhibition of prolactin release from the anterior pituitary gland and most likely the D-2 receptor mediated inhibition of the release of acetylcholine in the striatum. Examples of synaptic communication have been found in the striatum where (with ultrastructural techniques) synaptic contacts between dopaminergic nerve terminals and elements from cells containing GABA, substance P or enkephalin have been demonstrated. It is tempting to speculate that synaptic and non-synaptic communication occurs via D-1 and D-2 receptors respectively.     

Modulation of hypothalamic galanin gene expression by estrogen in peripubertal rats.

Hypothalamic galanin gene expression was investigated during reproductive maturation in peripubertal rats. Rat galanin-like immunoreactivity (rGAL-LI) increased in the median eminence and anterior pituitary during the extended first estrous and diestrous phases relative to anestrous phase female or male rats. In the neurointermediate lobe, rGAL-LI was elevated in first diestrous phase compared to anestrous phase females or males. In a second study, hypothalamic tissue was divided into quadrants for analysis of rat galanin (rGAL) mRNA by Northern blot hybridization. Two days after the injection of pregnant mare's serum gonadotropin (PMSG), rGAL mRNA increased approximately twofold in the paraventricular area and preoptic area quadrants. No effects of PMSG on galanin gene expression were found in medial basal or supraoptic hypothalamic quadrants. Because PMSG acts through the stimulation of ovarian estrogen secretion, these studies conclude that galanin gene expression in dorsal hypothalamic nuclei is under the stimulatory influence of estrogen and suggest that galanin may be a mediator of central ovarian steroid feedback.     

In vivo correlation between c-Fos expression and corticotroph stimulation by adrenocorticotrophic hormone secretagogues in rat anterior pituitary gland

In the anterior pituitary gland, c-Fos expression is evoked by various stimuli. However, whether c-Fos expression is directly related to the stimulation of anterior pituitary cells by hypothalamic secretagogues is unclear. To confirm whether the reception of hormone-releasing stimuli evokes c-Fos expression in anterior pituitary cells, we have examined c-Fos expression of anterior pituitary glands in rats administered with synthetic corticotrophin-releasing hormone (CRH) intravenously or subjected to restraint stress. Single intravenous administration of CRH increases the number of c-Fos-expressing cells, and this number does not change even if the dose is increased. Double-immunostaining has revealed that most of the c-Fos-expressing cells contain adrenocorticotrophic hormone (ACTH); corticotrophs that do not express c-Fos in response to CRH have also been found. However, restraint stress evokes c-Fos expression in most of the corticotrophs and in a partial population of lactotrophs. These results suggest that c-Fos expression increases in corticotrophs stimulated by ACTH secretagogues, including CRH. Furthermore, we have found restricted numbers of corticotrophs expressing c-Fos in response to CRH. Although the mechanism underlying the different responses to CRH is not apparent, c-Fos is probably a useful immunohistochemical marker for corticotrophs stimulated by ACTH secretagogues. This work was supported by the Jichi Medical University young investigator award.     

Mediation of the behavioral, endocrine and thermoregulatory actions of ghrelin

The action of ghrelin on telemetrically recorded motor activity and the transmission of the effects of this neuropeptide on spontaneous and exploratory motor activity and some related endocrine and homeostatic parameters were investigated. Different doses (0.5-5 microg) of ghrelin administered intracerebroventricularly caused significant increases in both square crossing and rearing activity in the "open-field" apparatus, while only the dose of 5 microg evoked a significant increase in the spontaneous locomotor activity recorded by telemetry. Ghrelin also induced significant increases in corticosterone release and core temperature. To determine the transmission of these neuroendocrine actions, the rats were pretreated with different antagonists, such as a corticotropin-releasing hormone (CRH) antagonist (alpha-helical CRH(9-41)), the nitric oxide synthase inhibitor Nomega-nitro-L-arginine-methyl ester (L-NAME), haloperidol, cyproheptadine or the cyclooxygenase inhibitor noraminophenazone (NAP). The open-field and biotelemetric observations revealed that the motor responses were diminished by pretreatment with the CRH antagonist and haloperidol. In the case of HPA (hypothalamic pituitary adrenal) activation, only cyproheptadine pretreatment proved effective; haloperidol and L-NAME did not modify the corticosterone response. NAP had only a transient, while cyproheptadine elicited a more permanent impact on the hyperthermic response evoked by ghrelin; the other antagonists proved to be ineffective. The present data suggest that both CRH release and dopaminergic transmission may be involved in the ghrelin-evoked behavioral responses. On the other hand, ghrelin appears to have an impact on the HPA response via a serotonergic pathway and on the hyperthermic response via a cyclooxygenase and a serotonergic pathway.     

Coexisting peptides in hypothalamic neuroendocrine systems: Some functional implications

1. Coexisting with oxytocin or vasopressin in the cell bodies and nerve terminals of the hypothalamic-neurohypophysial system are smaller amounts of other peptides. For a number of these "copeptides" there is strong evidence of corelease with the major magnocellular hormones. Guided by the location of their specific receptors we have studied the effects of three copeptides, dynorphin, cholecystokinin (CCK), and corticotropin releasing hormone (CRH), on the secretion of oxytocin and vasopressin from isolated rat neural lobe or neurointermediate lobe preparations in vitro. 2. Dynorphin is coreleased with vasopressin from neural lobe nerve terminals and acts on neural lobe kappa-opiate receptors to inhibit the electrically stimulated secretion of oxytocin. Naloxone augments oxytocin release from the neural lobe in a manner directly proportional to the amount of vasopressin (and presumably dynorphin) released. 3. Cholecystokinin, coreleased with oxytocin by neural lobe terminals, has been shown to have high-affinity receptors located in the NL and to stimulate secretion of both oxytocin and vasopressin. CCK's secretagogue effect was independent of electrical stimulation and extracellular Ca2+ and was blocked by an inhibitor of protein kinase C. 4. CRH, coreleased with OT from the neural lobe, has receptors in the intermediate lobe of the pituitary, but not in the neural lobe itself. CRH stimulates the secretion of oxytocin and vasopressin from combined neurointermediate lobes but not from isolated neural lobes. Intermediate lobe peptides, alpha and gamma melanocyte stimulating hormone, induced secretion of oxytocin and vasopressin from isolated neural lobes. Their effect was, like that of CCK, independent of electrical stimulation and extracellular Ca2+ and blocked by an inhibitor of protein kinase C. 5. Among the CRH-producing parvocellular neurons of the paraventricular nucleus, in the normal rat, approximately half also produce and store vasopressin. After removal of glucocorticoid influence by adrenalectomy, virtually all of the CRH neurons contain vasopressin. 6. The two subtypes of CRH neurosecretory cells found in the normal rat possess different topographical distributions in the paraventricular nucleus, suggesting the possibility of differential innervation. Stress selectively activates the vasopressin containing subpopulation of CRH neurons, indicating that there are separate channels of regulatory input controlling the two components of the parvocellular CRH neurosecretory system.     

Ovine corticotropin-releasing hormone stimulation test in patients with chronic renal failure: pharmacokinetic properties, and plasma adrenocorticotropic hormone and serum cortisol responses

The data on the status of the hypothalamic-pituitary-adrenal (HPA) axis in haemodialysis (HD) patients are conflicting. Moreover, a state reminiscent of Cushing's syndrome has been reported in this group of patients. Corticotropin-releasing hormone (CRH), that is produced by the hypothalamus and modulates the secretion of adrenocorticotropic hormone (ACTH), has been shown to be useful as a provocative test of the HPA axis. We investigated the effect of exogenous ovine CRH (oCRH) on plasma levels of ACTH and cortisol in 13 chronic HD patients. The plasma concentrations of immunoreactive CRH following oCRH administration were similar in patients and controls. In all patients, oCRH given intravenously as bolus injection caused a further increase in the already elevated levels of cortisol. The mean basal plasma levels of ACTH were within the normal range. There was, however, a blunted ACTH response to oCRH. We conclude that the HPA axis in chronic HD patients retains the ability to respond to exogenous oCRH. The patterns of the ACTH and cortisol response to this peptide resemble those observed in chronic stress (depression, anorexia nervosa). Besides, the kinetics of disappearance of oCRH indicate that the kidney may not be the major organ that metabolizes oCRH.     

Effects of p-chloroamphetamine, a serotonin-depleting drug, on the median eminence and pituitary pars intermedia

p-Chloroamphetamine (PCA), an agent known to cause depletion of levels of brain serotonin in rodents, was administered to rats in three sequential injections (10mg/kg) to study effects on the hypothalamic median eminence and pituitary gland. One week following the initial sequence of injections of PCA, light and electron micrographs revealed degenerate fibers in the outer zone of the median eminence. Lower drug doses or single 10-mg/kg doses did not lead to morphologic changes. Neuronal processes located in the pituitary intermediate lobe appeared normal although there was a significant increase in the numbers of secretory granules contained within intermediate lobe cells drug-treated rats, as compared to controls. Fluorometric analysis of levels of catecholamine and indoleamine showed a decrease in serotonin in median eminence and pons-medulla, but no change in that of the pituitary. Levels of dopamine and norepinephrine remained unchanged after PCA treatment. The data suggest that fibers affected in the median eminence contain serotonin. Processes in the intermediate lobe may be resistant to the serotonin-lowering effects of PCA observed in brain tissue. In addition, PCA may directly affect granule release from pituitary cells, or may alternatively act on hypothalamic regions which affect the release of intermediate lobe cell hormones.     

Effects of neonatal administration of monosodium glutamate and castration on neurokinin A levels in the hypothalamus and anterior pituitary of rats.

The effects of neonatal administration of monosodium glutamate (MSG) and castration on hypothalamic and anterior pituitary levels of neurokinin A (NKA) were studied in male and female rats killed at 46 days of age. In male rats treated neonatally with MSG, body, anterior pituitary, testis, ventral prostate, and seminal vesicle weights and serum testosterone levels were significantly lower than in saline-injected controls. Hypothalamic NKA was significantly lower in MSG-treated male rats as compared with the controls, and no apparent changes were recorded in anterior pituitary NKA. Orchidectomy was followed by a significant decrease in hypothalamic NKA in saline controls, but not in MSG-treated rats. In female rats treated with MSG, there was a significant decrease in body, anterior pituitary, and ovarian weights, as compared with saline-injected controls, but no significant differences were observed in uterine weights and serum estradiol levels. Hypothalamic NKA was lower, although not significantly, in MSG-treated rats as compared with the respective controls, and no differences were recorded in anterior pituitary NKA levels. Ovariectomy was followed by a significant decrease in hypothalamic NKA in both MSG-treated and control rats, but NKA in the anterior pituitary was significantly increased after ovariectomy only in saline-treated controls, whereas MSG-treated females failed to show this response. It is concluded that neonatal MSG treatment resulted in a decrease of hypothalamic NKA, which was particularly pronounced in male rats without any significant change in anterior pituitary NKA levels. The response of hypothalamic NKA to castration and the response of anterior pituitary NKA to ovariectomy were also altered in MSG-treated rats; this may reflect a functional block of some neuroendocrine functions of the hypothalamus that resulted from the neuronal lesions induced by MSG.     

CRH and lysine-vasopressin stimulation tests in the diagnosis of hypoadrenalism secondary to hypothalamic or pituitary disorders

Ten patients with secondary hypoadrenalism have been tested with corticotropin releasing hormone (CRH) and lysine-vasopressin (LVP). One patient had isolated ACTH deficiency; 9 had deficiency of other pituitary hormones attributable to a primary pituitary disease in 3 and to an hypothalamic disorder in 6. After CRH administration, a definite increase in plasma ACTH was observed in all 6 patients with hypothalamic disorder. No response was elicited in the 3 patients with pituitary disease and in the patient with isolated ACTH deficiency. In the responsive patients. ACTH showed a delayed and prolonged pattern of response. Lysine-vasopressin administration produced an increase in plasma ACTH in 4 of the 6 hypothalamic patients and no response in those with pituitary disease and in the patient with isolated ACTH deficiency. These findings suggest that CRH represents a reliable test in differentiating hypothalamic from pituitary adrenal failure; LVP appeared a less sensitive diagnostic test.