Developing Brain as an Endocrine Organ: A Paradoxical Reality

The maintaining of homeostasis in the organism in response to a variable environment is provided by the highly hierarchic neuroendocrine-immune system. The crucial component of this system is the hypothalamus providing the endocrine regulation of key peripheral organs, and the adenohypophysis. In this case, neuron-derived signaling molecules (SM) are delivered to the blood vessels in hypothalamic “neurohaemal organs” lacking the blood–brain barrier (BBB), the posterior lobe of the pituitary and the median eminence. The release of SM to the blood vessels in most other brain regions is prohibited by BBB. According to the conventional concept, the development of the neuroendocrine system in ontogenesis begins with the “maturation” of peripheral endocrine glands which first are self-governed and then operate under the adenohypophysial control. Meantime, the brain maturation is under the control of SM secreted by endocrine glands of the developing organism and coming from the placenta and maternal organism. The hypothalamus is involved in the neuroendocrine regulation only after its full maturation that is followed by the conversion of the opened-looped neuroendocrine system to the closed-looped system as in adulthood. Neurons of the developing brain begin to secrete SM shortly after their origin and long before the establishment of specific interneuronal relations providing initially autocrine and paracrine morphogenetic influence on differentiating target neurons. Taking into account that the brain lacks BBB over this ontogenetic period, we hypothesized that it operates as the multipotent endocrine gland secreting SM to the general circulation and thereby providing the endocrine regulation of peripheral organs and the brain. The term “multipotent” means that the spectrum of the brain-derived circulating SM and their occupancy at the periphery in the developing organism should greatly exceed those in adulthood. In order to test this hypothesis, gonadotropin-releasing hormone (GnRH), dopamine (DA), and serotonin (5-hydroxytryptamine, 5-HT) were chosen as the markers of the presumptive endocrine function of the brain in ontogenesis. According to our data, the concentrations of GnRH, DA, and 5-HT in the rat general circulation during the perinatal period, i.e. before the establishment of BBB, was as high as those in the portal circulation in adulthood. The concentrations of circulating GnRH and DA dropped to almost undetectable level after the development of BBB suggesting their brain origin. This suggestion has been proven by showing an essential decrease of GnRH, DA, and 5-HT concentrations in general circulation of perinatal rats after microsurgical elimination of synthesizing neurons or the inhibition of specific syntheses in the brain before the establishment of BBB. GnRH, DA, and 5-HT apparently as dozens of other brain-derived SM appear to be capable of providing the endocrine influence on their peripheral targets like the adenohypophysis, gonads, kidney, heart, blood vessels, and the brain (endocrine autoregulation). Although the ontogenetic period of the brain operation as the multipotent endocrine gland is relatively short, the brain-derived SM are thought to be capable of providing long-lasting morphogenetic effects on peripheral targets and the brain. Thus, the developing brain operates as the multipotent endocrine gland from the onset of neurogenesis to the establishment of BBB providing the endocrine regulation of the developing organism.     

The brain as an endocrine gland in adult and in developing ofranism

It is suggested that the abult mammals' brain is, on the one hand, a conglomerate of neuronal assemblies within which information is being transmitted with the aid of chemical signals, and on the other hand, a considerable part of the neurons are secreting cells and their accumulations by their functional characteristics are identical to endocrine glands. Proceeding from the brain dualism, a question arises: which of the two functions is the pimary one? It seems that, prior to forming specific interneuronal links and blood-brain barrier, the neurons function as secreting cells and the brain--as an endocrine gland. The physiologically active substances (FASs) entering the general circulation system from the brain seem to take part in regulation of development of the visceral target organs and the brain itself. Forming of the interneuronal links and blood-brain barrier leads to a qualitative jump in the brain development followed by a considerable limitation of its endocrine functions. According to this concept, the brain, prior to the moment of forming of the interneuronal links and blood-brain barrier, does not influence the development of the whole organism whereas development of the brain itself is regulated by the hormones of the foetus and placenta's endocrine glands.     

Drug metabolizing enzymes in cerebrovascular endothelial cells afford a metabolic protection to the brain.

The brain is partially protected from chemical insults by a physical barrier mainly formed by the cerebral microvasculature, which prevents penetration of hydrophilic molecules in the cerebral extracellular space. This results from the presence of tight junctions joining endothelial cells, and from a low transcytotic activity in endothelial cells, inducing selective permeability properties of cerebral microvessels that characterize the blood-brain barrier. The endothelial cells provide also, as a result of their drug-metabolizing enzymes activities, a metabolic barrier against potentially penetrating lipophilic substances. It has been established that in cerebrovascular endothelial cells, several families of enzymes metabolize potentially toxic lipophilic substrates from both endogenous and exogenous origin to polar metabolites, which may not be able to penetrate further across the blood-brain barrier. Enzymes of drug metabolism present at brain interfaces devoid of blood-brain barrier, like circumventricular organs, pineal gland, and hypophysis, that are potential sites of entry for xenobiotics, display higher activities than in cerebrovascular endothelial cells, and conjugation activities are very high in the choroid plexus. Finally, xenobiotic metabolism normally results in detoxication, but also in some cases in the formation of pharmacologically active or neurotoxic products, possibly altering some blood-brain barrier properties.     

Development of central and peripheral serotonin-producing systems in rats in ontogenesis

The work deals with study of development of central and peripheral serotonin-producing systems in rat ontogenesis before and after formation of the blood-brain barrier. By the method of highly efficient liquid chromatography it has been shown that the serotonin level in peripheral blood before formation of the blood-brain barrier (in fetuses and neonatal rats) is sufficiently high for realization of physiological effect on target cells and organs. At the period of formation of the blood-brain barrier the serotonin level in brain sharply rises, whereas the serotonin concentration and amount in blood plasma and duodenum increase insignificantly. Completion of formation of the blood-brain barrier is accompanied by a significant increase of the serotonin content in duodenum, probably for maintenance of the high serotonin level in blood. To evaluate secretory activity, the mean rate of daily serotonin increment in the studied tissues was calculated. In brain, this parameter was maximal at the period of formation of the blood-brain barrier-from the 4th to the 16th postnatal days. This allows thinking hat brain before formation of the blood-brain barrier is the most important source of serotonin in peripheral blood.     

Brain is a source of blood serotonin in rats during perinatal development

The aim of this study was to test our hypothesis that the brain functions as an endocrine organ before the blood-brain barrier is formed. A model of drug-inhibited serotonin synthesis in the brain using a single stereotactic administration of p-chlorophenylalanine, an inhibitor of serotonin synthesis, was developed. The inhibitor dose inducing the maximum effect in the brain and no effect on serotonin synthesis in the periphery was experimentally selected. The concentration of serotonin and its metabolites (5-hydroxytryptophan and 5-hydroxy-indoleacetic acid) was studied by high performance liquid chromatography in the brain, duodenum, and blood (separately in plasma and platelets). The optimal p-chlorophenylalanine dose (200 mg/kg) was shown to induce a sharp decrease in the brain level of serotonin (70%), a moderate decrease in plasma (16%) and platelets (26%), and an insignificant decrease in the duodenum (12%). At the same time, this dose did not decrease the 5-hydroxytryptophan level in the intestine. This suggests that the decrease in the blood level of serotonin was due to the inhibition of its synthesis in the brain, whereas the decrease in the duodenum level of serotonin was due to the compensatory release to blood while its synthetic rate remained unaltered. Thus, the developing brain before the blood-brain barrier formation was shown to secrete serotonin into blood.     

Ancestral TSH mechanism signals summer in a photoperiodic mammal

In mammals, day-length-sensitive (photoperiodic) seasonal breeding cycles depend on the pineal hormone melatonin, which modulates secretion of reproductive hormones by the anterior pituitary gland [1]. It is thought that melatonin acts in the hypothalamus to control reproduction through the release of neurosecretory signals into the pituitary portal blood supply, where they act on pituitary endocrine cells [2]. Contrastingly, we show here that during the reproductive response of Soay sheep exposed to summer day lengths, the reverse applies: Melatonin acts directly on anterior-pituitary cells, and these then relay the photoperiodic message back into the hypothalamus to control neuroendocrine output. The switch to long days causes melatonin-responsive cells in the pars tuberalis (PT) of the anterior pituitary to increase production of thyrotrophin (TSH). This acts locally on TSH-receptor-expressing cells in the adjacent mediobasal hypothalamus, leading to increased expression of type II thyroid hormone deiodinase (DIO2). DIO2 initiates the summer response by increasing hypothalamic tri-iodothyronine (T3) levels. These data and recent findings in quail [3] indicate that the TSH-expressing cells of the PT play an ancestral role in seasonal reproductive control in vertebrates. In mammals this provides the missing link between the pineal melatonin signal and thyroid-dependent seasonal biology.     

Glutaminyl cyclase knock-out mice exhibit slight hypothyroidism but no hypogonadism: implications for enzyme function and drug development

Glutaminyl cyclases (QCs) catalyze the formation of pyroglutamate (pGlu) residues at the N terminus of peptides and proteins. Hypothalamic pGlu hormones, such as thyrotropin-releasing hormone and gonadotropin-releasing hormone are essential for regulation of metabolism and fertility in the hypothalamic pituitary thyroid and gonadal axes, respectively. Here, we analyzed the consequences of constitutive genetic QC ablation on endocrine functions and on the behavior of adult mice. Adult homozygous QC knock-out mice are fertile and behave indistinguishably from wild type mice in tests of motor function, cognition, general activity, and ingestion behavior. The QC knock-out results in a dramatic drop of enzyme activity in the brain, especially in hypothalamus and in plasma. Other peripheral organs like liver and spleen still contain QC activity, which is most likely caused by its homolog isoQC. The serum gonadotropin-releasing hormone, TSH, and testosterone concentrations were not changed by QC depletion. The serum thyroxine was decreased by 24% in homozygous QC knock-out animals, suggesting a mild hypothyroidism. QC knock-out mice were indistinguishable from wild type with regard to blood glucose and glucose tolerance, thus differing from reports of thyrotropin-releasing hormone knock-out mice significantly. The results suggest a significant formation of the hypothalamic pGlu hormones by alternative mechanisms, like spontaneous cyclization or conversion by isoQC. The different effects of QC depletion on the hypothalamic pituitary thyroid and gonadal axes might indicate slightly different modes of substrate conversion of both enzymes. The absence of significant abnormalities in QC knock-out mice suggests the presence of a therapeutic window for suppression of QC activity in current drug development.     

Comparative evaluation of the biogenic amine level of the brain and the ethanol kinetics in the blood of inbred C57B1/6 and CBA mice

The kinetics of ethanol in blood and the levels of serotonin, noradrenaline, and dopamine in the cortex of cerebral hemispheres, hypothalamus, thalamus, and brain stem were studied in male C57C1/6 and CBA mice characterized by predisposition and non-predisposition to the development of experimental alcoholism. C57B1/6 mice characterized by predisposition to the development of experimental alcoholism demonstrated a high level of serotonin and noradrenaline in the hypothalamus and brain stem and high rate of ethanol elimination from the blood. CBA mice non-predisposed to the development of experimental alcoholism were characterized by a low level of serotonin and noradrenaline in the hypothalamus and brain stem and by low rate of ethanol elimination from blood.     

Neuroendocrine aspects of aging: experimental data

Aging is characterized by changes in neuroendocrine/endocrine functions which are manifest in female reproductive physiology and less perceptible in other functions such as thyroid, adrenal or growth/metabolic functions. The contribution of each level of the axis - hypothalamus, adenohypophysis or peripheral tissues - is not clearly established. Functional impairments with age are recognized in the peripheral glands (gonad, thyroid, adrenal) as well as in the pituitary, but increasing evidence is accumulating for a marked contribution of the hypothalamus in the age-associated endocrine changes observed in animals and humans. In old rats, multineuronal dysfunctions are demonstrated in the hypothalamus, with a documented decline in the activity of the neurons producing dopamine and thyrotropin-releasing hormone, and to a lesser extent luteinizing hormone- and growth hormone-releasing hormones, and alterations in regulatory mechanisms of these neurons are disclosed. Moreover, impairments are observed in the processing - binding, accumulation and intracellular distribution - of hypothalamic hormones in the adenohypophysis of old rats. Taken together, these observations are supportive of the view that the neuroendocrine/endocrine changes appearing with age result from a complex balance of functional alterations occurring at each level - central and peripheral - of the axis.     

Hypocretin/orexin suppresses corticotroph responsiveness in vitro

The hypocretin/orexins (Hcrts/ORXs) are peptides produced in neurons in the lateral hypothalamic area that project to neuroendocrine centers in the hypothalamus. Hcrt/ORX receptors are present in the hypothalamus and anterior pituitary gland. We examined the possibility that the Hcrts/ORXs, which we have demonstrated previously to act in the brain to stimulate sympathetic function, could alter stress hormone secretion by a direct pituitary action. In vitro studies revealed a dose-related inhibitory effect of the Hcrts/ORXs on corticotropin-releasing hormone-stimulated ACTH secretion that appeared to be mediated via the orexin-1 receptor and to be expressed at doses (threshold dose 1 nM orexin A) similar to the affinity constant for the receptor. The effect was not due to abrogation of the cAMP response of the corticotroph to corticotropin-releasing hormone and was not pertussis toxin sensitive, suggesting a non-G(i)-mediated mechanism. Instead, a G(q)-mediated signaling mechanism was indicated by the ability of protein kinase C blockade with calphostin C to reverse the inhibitory action of orexin A. Orexin A and orexin B did not significantly alter basal ACTH secretion in vitro and did not alter basal or releasing factor-stimulated secretion of luteinizing hormone, prolactin, thyroid-stimulating hormone or growth hormone from cells harvested from male or random-cycle female donors. Our data suggest a direct, pituitary action of the Hcrts/ORXs to modulate the endocrine response to stress and identify the potential cellular mechanism of a unique biological action of the peptides in the anterior pituitary gland.     

Morphofunctional Characteristics of Gonadotropin-Releasing Hormone Immunoreactive Structures in the Brain of the Sturgeon Acipenser güldenstädti before Spawning

Light-microscopic immunohistochemical study of the brain with use of the unlabeled antibody to gonadotropin-releasing hormone (GnRH) was carried out on sexually mature individuals of the sturgeon of stage IV of gonadal maturity (before spawning). The brain was examined as a whole; the GnRH-immunoreactive (GnRH-IR) structures were revealed in the olfactory bulb, forebrain, hypothalamus, and neuropituitary. In the fish studied at prespawning period, the highest density of GnRH-IR structure was noticed in the ventral region of forebrain, preoptic region, and anterior neuropituitary. The GnRH-IR cells of the forebrain ventral region, preoptic, and tuberal nuclei send their axons to the region of anterior neuropituitary, in which they contact vessels of the primary portal pituitary system. Thereby, the gonadotropin secretion regulation is performed by corresponding cells of adenopituitary. Rare GnRH-IR fibers in the posterior pituitary lobe contact the general circulation vessels. The dendrites of the GnRH-IR cells that we have revealed in the preoptic region and in the region of tuberal nucleus are located very close to the preoptic bay cavity and to the cerebral III ventricle, respectively. This indicates a possibility of secretion of the neurohormone into the cerebrospinal fluid.     

Transport processes through the blood-brain barrier

The existence of the blood-brain barrier is due to tight junctions between endothelial cells preventing the passage of liquid and solute material at the capillary level. Substances can thus pass across the blood-brain barrier if they are lipophilic or if they have transport systems in the membranes of endothelial cells. The luminal membrane brings metabolites needed for the brain function, the abluminal one plays an important part in removing substances from brain, this can happen against a concentration gradient and thus needs energy. Ions are transported differently by the 2 membranes. Sodium and chloride have carriers and potassium is transported very actively by the sodium-potassium ATPase of the abluminal membrane. Blood-brain glucose influx is very important and happens by carrier transport at the 2 membranes. Efflux seems to use the same transport system as the influx. Transport of ketone bodies seems to happen only from blood to brain, the carriers being reversibly used for brain-blood transport of pyruvic and lactic acid. Amino-acid transport is very different on the luminal and abluminal membranes. On the luminal membrane there are 2 transport systems, one for basic amino acids, the other one, the L system, for neutral amino-acids. All neutral amino-acids are transported through the abluminal membrane by the L, A and ASC systems. There exists a system of transport for basic amino-acids, and a very active one for acid amino-acids. Some systems for the transport of hormones, vitamins and for some peptides exist also at the blood-brain barrier which thus plays a very important role in the regulation of brain metabolism.     

Prolactin-releasing peptides

Physiologic control of prolactin (PRL) secretion is largely dependent upon levels of dopamine accessing the adenohypophysis via the hypophysial portal vessels. However, it is clear that other factors of hypothalamic origin can modulate hormone secretion in the absence or presence of dopamine. Several neuropeptides have been identified as PRL releasing factors (PRFs) but none of these peptides appears to be a major determinant of PRL secretion in vivo. There remain uncharacterized activities in hypothalamic extracts that can alter secretion and production of the hormone. In addition, there exist a wide variety of substances (neurotransmitters, neuromodulators, neuropeptides) that can act within the hypothalamus to modify the neuroendocrine regulation of PRL secretion. These factors may not be considered true PRFs because their actions are not exerted directly at the level of the lactotroph; however, they can act in brain to stimulate PRL release in vivo and therefore might be considered PRL releasing peptides (PRPs).     

The gonadotropin-releasing hormone receptor: signals involved in gonadotropin secretion and biosynthesis

The neurohormone gonadotropin-releasing hormone (GnRH) is a decapeptide which is synthesized in the hypothalamus and released into the hypophysial portal system in a pulsatile manner. GnRH exerts its effect on the anterior pituitary gonadotrophs where it regulates the secretion and synthesis of gonadotropins (luteinizing hormone and follicle-stimulating hormone) through receptor-mediated actions. The GnRH receptor has been characterized and shown to be coupled to the formation of 'second messengers' which participate in signal transduction mechanisms. GnRH stimulation of luteinizing hormone release is a Ca2(+)-dependent process. G protein, phosphoinositide hydrolysis, protein kinase C as well as arachidonic acid and some of its metabolites were identified as possible mediators in the process.     

Relation of endogenous opioid peptides and morphine to neuroendocrine functions.

Morphine and the endogenous opioid peptides (EOP) exert similar effects on the neuroendocrine system. When adminstered acutely, they stimulate growth hormone (GH), prolactin (PRL), and adrenocorticotropin (ACTH) release, and inhibit release of luteinizing hormone (LH), follicle stimulating hormone (FSH),and thyrotropin (TSH). Recent studies indicate that the EOP probably have a physiological role in regulating pituitary hormone secretion. Thus injection of naloxone (opiate antagonist) alone in rats resulted in a rapid fall in serum concentrations of GH and PRL, and a rise in serum LH and FSH, suggesting that the EOP help maintain basal secretion of these hormones. Prior administration of naloxone or naltrexon inhibited stress-induced PRL release, and elevated serum LH in castrated male rats to greater than normal castrate levels. Studies on the mechanisms of action of the EOP and morphine on hormone secretion indicate that they have no direct effect on the pituitary, but act via the hypothalamus. There is no evidence that the EOP or morphine alter the action of the hypothalamic hypophysiotropic hormones on pituitary hormone secretion; they probably act via hypothalamic neurotransmitters to influence release of the hypothalamic hormones into the pituitary portal vessels. Preliminary observations indicate that they may increase serotonin and decrease dopamine metabolism in the hypothalamus, which could account for practically all of their effects on pituitary hormone secretion.     

Adrenomedullin in the cerebral circulation.

The central nervous system requires an effective autoregulation of cerebral circulation in order to meet the critical and unusual demands of the brain. In addition, cerebral microvessels has a unique feature, the formation of the blood-brain barrier, which contributes to the stability of the brain parenchymal microenvironment. Many factors are known to be involved in the regulation of cerebral circulation and blood-brain barrier functions. In the last few years a new potential candidate, adrenomedullin, a hypotensive peptide was added to this list. Adrenomedullin has a potent vasodilator effect on the cerebral vasculature, and it may be implicated in the pathologic mechanism of cerebrovascular diseases. In this review, we describe current knowledge about the origin and possible role of adrenomedullin in the regulation of cerebral circulation and blood-brain barrier functions.     

Prepubertal changes in the hypothalamic-pituitary axis of Holstein bulls

We investigated the nature and sites of changes in the hypothalamic-pituitary axis associated with the onset of high-frequency, high-amplitude discharges of luteinizing hormone (LH) in young bulls during the transition from the infantile to the prepubertal phase of development. Blood serum and neuroendocrine tissues from bulls killed at 1, 6, 10, 14, or 18 wk of age were evaluated. Concentrations of LH in serum from bulls 1 or 6 wk old averaged less than 0.25 ng/ml and only one episodic discharge of LH was detected for 10 bulls. At 10, 14, or 18 wk, 14 of 15 bulls had episodic discharges of LH. Concentrations of testosterone in serum were progressively higher at 10, 14, and 18 wk, but the concentration of estradiol was maximal at 6 wk. The concentrations of gonadotropin-releasing hormone (GnRH) in the anterior hypothalamus, posterior hypothalamus, or median eminence were not influenced by age. However, concentration of GnRH receptors in the anterior pituitary gland increased 314% between 6 and 10 wk and the concentration of LH increased 67%. Between 6 and 10 wk, concentrations of estradiol receptors in the anterior and posterior hypothalamus declined by 68% and 46%, but the concentration of estradiol receptors in the anterior pituitary gland increased by 103%. For most characteristics, there was no major change between 10 and 18 wk. We postulate that between 6 and 10 wk of age, there is 1) removal of an estradiol-mediated block of GnRH secretion and 2) an estradiol-mediated, and possibly GnRH-mediated, increase in pituitary GnRH receptors. Together, these changes result in greatly increased stimulation of the anterior pituitary gland by GnRH between 6 and 10 wk of age and stimulation of the discharges of LH characteristic of bulls in the early prepubertal phase of development.     

Endogenous dopamine and serotonin release from the explanted rat tuberohypophyseal system: effects of electrical stimulation and neurotensin

Excised blocks of brain tissue encompassing the hypothalamic periventricular nucleus, arcuate nucleus, infundibular stalk and attached pituitary neurointermediate lobe (NIL) were obtained from rats and perifused in vitro. The intact tuberohypophyseal dopaminergic pathway projecting from cell bodies in the hypothalamus to terminations in the pars intermedia and posterior lobes was thus isolated. An electrode placed in the explant delivered electrical pulses at various frequencies to hypothalamic targets. Products released in the immediate vicinity of the NIL were continuously collected and assayed for dopamine and serotonin content. Electrical stimulation of the arcuate nucleus from 1.0 - 10.0 Hz produced a frequency-dependent increase in dopamine, but not serotonin, release from the NIL. Stimulation of the infundibular stalk, however, elevated the release of both neurotransmitters. Addition of neurotensin (0.001 - 1.0 microM) to the bathing media produced a concentration-dependent increase in dopamine, but not serotonin, release. These experiments demonstrate the utility of the tuberohypophyseal explant as a model for use in the study of dopaminergic neuronal function in this neuroendocrine axis.     

Studies on a possible pituitary effect of monoamines on luteinizing hormone release in ovariectomized rats

Incubation of anterior pituitaries from ovariectomized rats with LHRH and various concentrations of dopamine, norepinephrine or serotonin indicated that none of these neurotransmitters could decrease pituitary LH secretion in response to the releasing hormone. This indicated that the inhibitions of pulsatile LH release previously observed in our laboratory in ovariectomized rats in response to intraventricularly administered catecholamines or stimulation of brain serotoninergic neurons are due to central rather than pituitary effects of these transmitters.     

Effect of L-tryptophan and restraint stress on hypothalmic and brain serotonin turnover, and pituitary TSH and prolactin release in rats.

The dose response and time course effects of L-tryptophan and restraint stress on the metabolism of serotonin and release of thyroid stimulating hormone (TSH) and prolactin (PRL) were tested in male rats. Both treatments increased serotonin turnover in the hypothalamus (H) and remaining brain tissue minus the cerebellum (brain) as determined by enhanced accumulation of serotonin following monoamine oxidase (MAO) inhibition. L-tryptophan but not restraint stress elevated levels of tryptophan in the cerebellum. Both L-tryptophan and restraint stress inhibited TSH release and stimulated PRL release. These findings indicate that enhanced rates of serotonin turnover produced by L-tryptophan and physical restraint are associated with inhibition of TSH and stimulation of PRL release from the anterior pituitary.