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GPR139, a Gq-coupled receptor that is activated by the essential amino acids L-tryptophan and L-phenylalanine, is predominantly expressed in the brain and pituitary. The physiological function of GPR139 remains elusive despite the availability of pharmacological tool agonist compounds and knock-out mice. Whole tissue RNA sequencing data from human, mouse and rat tissues revealed that GPR139 and the dopamine D2 receptor (DRD2) exhibited some similarities in their distribution patterns in the brain and pituitary gland. To determine if there was true co-expression of these two receptors, we applied double in situ hybridization in mouse tissues using the RNAscope® technique. GPR139 and DRD2 mRNA co-expressed in a majority of same cells within part of the dopaminergic mesolimbic pathways (ventral tegmental area and olfactory tubercle), the nigrostriatal pathway (compact part of substantia nigra and caudate putamen), and also the tuberoinfundibular pathway (arcuate hypothalamic nucleus and anterior lobe of pituitary). Both receptors mRNA also co-express in the same cells of the brain regions involved in responses to negative stimulus and stress, such as lateral habenula, lateral septum, interpeduncular nucleus, and medial raphe nuclei. GPR139 mRNA expression was detected in the dentate gyrus and the pyramidal cell layer of the hippocampus as well as the paraventricular hypothalamic nucleus. The functional interaction between GPR139 and DRD2 was studied in vitro using a calcium mobilization assay in cells co-transfected with both receptors from several species (human, rat, and mouse). The dopamine DRD2 agonist did not stimulate calcium response in cells expressing DRD2 alone consistent with the Gi signaling transduction pathway of this receptor. In cells co-transfected with DRD2 and GPR139 the DRD2 agonist was able to stimulate calcium response and its effect was blocked by either a DRD2 or a GPR139 antagonist supporting an in vitro interaction between GPR139 and DRD2. Taken together, these data showed that GPR139 and DRD2 are in position to functionally interact in native tissue.

IntroductionAntipsychotic-induced hyperprolactinemia is associated with relevant side effects: short-term as hypogonadism, gynecomastia, amenorrhoea, sexual dysfunction and galactorrhoea; long-term as cardiovascular disease, bone demineralization and breast and prostate tumors.AimsTo evaluate the effect of switching to long-acting injectable aripiprazole on long-lasting antypsychotic-induced hyperprolactinemia.MethodsThis was a prospective observational 1-year study carried out in 125 outpatients with schizophrenia who were clinically stabilized but a switching to another antipsychotic was indicated. We measured the basal prolactine at the start of the study and 1 year after switching to long acting injecatable (LAI) aripiprazole.ResultsIn basal analytic, 48% had hyperprolactinemia (21.8–306.2 ng/mL) and 66.5% of them described side effects: 78% sexual dysfunction (72% men), 11% galactorrhoea (100% women), 5.5% amenorrhoea and 5.5% bone pain (100% women). In 48% of patients with hyperprolactinemia, the previous antipsychotics comprised: LAI-paliperidone (65,7%), oral-risperidone (7%), oral-olanzapine (6.1%), oral-paliperidone (5.2%), LAI-risperidone (4%) and others (12%). One year after switching to LAI-aripiprazole, prolactine levels were lower in all patients and in 85% prolactine levels were normalized. Overall, 72% described a clinical improvement, especially in terms of sexual dysfunction.ConclusionsSeveral studies have described an improvement of drug-induced hyperprolactinemia after switching to or adding oral aripiprazole. In our study, we observed that levels of prolactine were normalized in 85% of patients with a clinical improvement in almost all of cases. These findings suggest that switching to LAI aripiprazole may be an effective alternative for managing antipsychotic-induced hyperprolactinemia due to its partial agonism in D2 brain receptors, especially in tuberoinfundibular pathway.Disclosure of interestThe authors have not supplied their declaration of competing interest.

The central and peripheral nervous systems are important factors influencing the functioning of liver cytochrome P450 (CYP). It has been shown that changes in the brain monoaminergic systems affect liver cytochrome P450 (CYP) expression (CYP1A, CYP2B, CYP2C11 and CYP3A). The brain dopaminergic system has been established as an important center regulating the liver CYP. This regulation proceeds via the tuberoinfundibular pathway and the dopaminergic D2 receptors of the pituitary, as well as the mesolimbic pathway engaging the D2 receptors of the nucleus accumbens (conveying a message to the paraventricular nucleus of the hypothalamus). These two dopaminergic pathways stimulate the secretion of pituitary hormones, which directly (GH) or indirectly (ACTH, TSH) activate hepatic nuclear/ cytosolic receptors controlling CYP genes. Recent preliminary studies with selective noradrenaline or serotonin neurotoxins suggest also involvement of the brain noradrenergic and serotonergic systems in the regulation of liver CYP. Moreover, the influence of the peripheral nervous system involving several neurotransmitters (acetylcholine, noradrenaline, adrenaline, dopamine, serotonin) on liver function may also be important for the physiological regulation of hepatic CYP activity. The hypothalamus controls liver function not only by releasing hormones from the pituitary gland but also by stimulating the autonomic sympathetic and parasympathetic projections to the liver. In addition to direct neural connections, the autonomic nervous system can indirectly affect liver function via the hypothalamus-adrenal axis and the hypothalamus- pancreas axis. Therefore, the estimation of neuroactive drug action on hepatic CYP requires an in vivo model which allows the central neuroendocrine and peripheral autonomic regulation of genes coding for CYP isoforms.

The aim of the present study was to investigate the influence of activation of brain dopaminergic system by different dopaminomimetics on the level and activity of liver cytochrome P450 (CYP) isoforms. Studies into the identification of hormones and cytokines which are known to mediate liver CYP expression were also simultaneously carried out. Stimulation of dopaminergic receptors in the pituitary, a target for the tuberoinfundibular pathway, by dopamine (a D(1)/D(2) receptor agonist) administered intraperitoneally caused a significant increase in the activities and protein levels of CYP2B, CYP2C11 and CYP3A, a substantial increase in the blood plasma level of growth hormone (GH) and a significant decrease in triiodothyronine (T(3)) level. Local stimulation of dopaminergic receptors in the nucleus accumbens, a target for the mesolimbic pathway, by apomorphine (a D(1)/D(2) receptor agonist), amphetamine (an indirect D(1)/D(2) dopaminemimetic) and quinpirole (a D(2) receptor agonist) produced a substantial rise in CYP3A activity and protein level, caused a large increase in corticosterone concentration and a moderate drop in T(3) level. SKF82958 (a D(1) receptor agonist) did not significantly affect the CYP isoforms or hormones studied. In both cases (activation of the tuberoinfundibular or mesolimbic pathway), the activity and the protein level of CYP1A considerably decreased. Plasma levels of thyroxine, testosterone, interleukin-2 and interleukin-6 were not changed after activation of the two pathways. The obtained results establish the brain dopaminergic system as a physiological centre regulating cytochrome P450 (engaging D(2) receptors and pituitary hormones) and demonstrate new pharmacological aspects of neuroactive drugs that affect this system.

Tuberoinfundibular peptide (TIP39) was initially identified as a neurotransmitter and agonist of the PTH2 receptor, which is expressed in the cardiovascular system. This study documents for the first time the cardiac expression and function of TIP39. Expression was analyzed via RT-PCR. Function was characterized on Langendorff-perfused rat hearts as left ventricular developed pressure (LVDP) and on isolated cells via a cell edge detection system. cGMP levels were detected with RIA. Tuberoinfundibular peptide (TIP39) mRNA was found to be constitutively expressed in coronary endothelium cells, isolated cardiomyocytes, ventricles, atria, and aorta. At first we investigated the vasodilatory properties of TIP39 (100 nM) in the presence of L-nitro-arginine (L-NA, 100 microM). Surprisingly, TIP39 had no vasodilatory effect but decreased LVDP by 35 +/- 7%. In the absence of L-NA, addition of TIP39 decreased LVDP by 8 +/- 2%. The PTH2 receptor antagonist Trp23-Tyr36-PTHrP (1-36, 100 nM) abolished this TIP39 effect in the presence of L-NA. The experiments with isolated cardiomyocytes provided similar results. TIP39 (10 nM) lowered the contraction amplitude by 6 +/- 3%. In the presence of L-NA (100 micromol/liter), TIP39 lowered the amplitude by 34 +/- 6%. cGMP concentration in cardiomyocytes was stimulated by TIP39 (10 nM) in the same range as by the nitric oxide (NO) donor SNAP (100 microM). In the presence of L-NA, this increase was abolished. These results suggest that an inhibition of endogenous NO production unmasks a profound negative inotropic effect of TIP39 that is mediated by an activation of the PTH2 receptor. The results obtained with isolated cardiomyocytes suggest that myocyte-derived NO, rather than vascular NO, is responsible for this effect. cGMP seems to be the downstream signal of produced NO.

The widespread use of the selective serotonin reuptake inhibitors (SSRIs) has been accompanied by numerous reports describing a potential association with hyperprolactinemia. Antipsychotics are commonly known to elevate serum prolactin (PRL) through blockade of dopamine receptors in the pituitary. However, there is little awareness of the mechanisms by which SSRIs stimulate PRL release. Hyperprolactinemia may result in overt symptoms such as galactorrhea, which may be accompanied by impaired fertility. Long-term clinical sequelae include decreased bone density and the possibility of an increased risk of breast cancer. Through literature review, we explore the possible pathways involved in serotonin-induced PRL release. While the classic mechanism of antipsychotic-induced hyperprolactinemia directly involves dopamine cells in the tuberoinfundibular pathway, SSRIs may act on this system indirectly through GABAergic neurons. Alternate pathways involve serotonin stimulation of vasoactive intestinal peptide (VIP) and oxytocin (OT) release. We conclude with a comprehensive review of clinical sequelae associated with hyperprolactinemia, and the potential role of SSRIs in this phenomenon.

Editorial Sexual dysfunction is a common side effect of psychotropic drugs and has received much attention because of its negative effects on quality of life and adherence to antidepressant agents. In patients with depression, sexual dysfunction can be attributed to the underlying disorder or to antidepressant drugs. Sexual dysfunction also can occur with first generation antipsychotic drugs and second-generation antipsychotic drugs (e.g., thioridazine and risperidone, respectively). In patients with schizophrenia, sexual dysfunction can be attributed to the underlying illness as well as to antipsychotic induced hyperprolactinemia due to dopamine blockade in tuberoinfundibular pathway [1].

Dopamine agonists with activity at both dopamine D 2 and D 3 receptor subtypes stimulate tuberoinfundibular dopamine neurons and inhibit prolactin secretion from the anterior pituitary. The purpose of the present study was to identify the dopamine receptor subtypes mediating these effects using recently developed selective agonists for dopamine D 2 (PNU-95,666) and D 3 (PD128907) receptors. The activity of tuberoinfundibular dopamine neurons was estimated by measuring either the synthesis (accumulation of 3,4-dihydroxyphenylalanine [DOPA] following inhibition of decarboxylase activity) or metabolism (3,4-dihydroxyphenylacetic acid [DOPAC] concentrations) of dopamine in the median eminence, the region of the hypothalamus containing axon terminals of these neurons. In one experiment, the activity of mesolimbic dopamine neurons was also determined by measuring DOPA accumulation in terminals of these neurons in the nucleus accumbens. Activation of dopamine D 2 receptors with PNU-95,666 caused dose- and time-related increases in DOPAC concentrations in median eminence which were temporally correlated with decreases in plasma prolactin concentrations. Activation of dopamine D 3 receptors with PD128907 decreased DOPA concentrations in the nucleus accumbens, but had no effect on concentrations of DOPAC or DOPA in the median eminence or prolactin in plasma. These results reveal that tuberoinfundibular dopamine neurons are regulated by dopamine D 2 rather than D 3 receptors, and suggest that the ability of mixed dopamine D 2 /D 3 receptor agonists to increase the activity of these neurons is mediated by an action at dopamine D 2 receptors. Furthermore, these results confirm that tuberoinfundibular dopamine neurons are not regulated by inhibitory dopamine D 2 or D 3 autoreceptors.

To the Editor: We describe a case report of galactorrhea with prolactin level within normal limits in a healthy 16-year-old African American girl after administration of quetiapine. Prolactin is an endogenous hormone from the anterior pituitary gland that promotes lactation. Dopamine predominantly inhibits prolactin. Antipsychotic medications block the dopamine-2 (D2) receptor via the tuberoinfundibular dopamine pathway, which can result in hyperprolactinemia.1 Serotonin is an indirect modulator of prolactin secretion.2 Hyperprolactinemia can cause galactorrhea, gynecomastia, sexual dysfunction, menstrual irregularities, infertility, hirsutism, and acne in women due to hypoestrogenism; hypogonadism and impaired growth and sexual maturation in adolescents; and potential osteoporosis.3,4 Women have greater prolactin elevations from antipsychotics than do men.5,6 Children and adolescents may also have greater prolactin elevations in response to antipsychotics.7 Conventional antipsychotics, risperidone, zotepine, and amisulpride are known to elevate prolactin more than other atypical antipsychotic medications.4,8–10 The median time to onset of galactorrhea is 20 days (range, 7–75 days) after commencement of an antipsychotic medication.8 Compared to typical antipsychotics with tighter D2 receptor binding, most atypical antipsychotics are less likely to cause hyperprolactinemia and thus galactorrhea. Quetiapine is known to have overall weaker dopamine binding activity; positron emission tomography scans showed transiently high striatal D2 receptor occupancy. Quetiapine’s transient association with the D2 receptor is postulated to allow normal dopaminergic neurotransmission in the tuberoinfundibular pathway and thus avoid hyperprolactinemia. This may explain only a temporary elevation of prolactin levels. D2 receptor occupancy decreases to 0%–27% in 12 hours.11 Quetiapine is less likely to elevate serum prolactin levels than risperidone.10,12 Fluoxetine also has been described to cause hyperprolactinemia and galactorrhea, possibly by stimulating prolactin release from pituitary lactotrophs.13 Hyperprolactinemia and galactorrhea from selective serotonin reuptake inhibitor use tended to correlate with above average dosages.14 Our patient was taking fluoxetine at 15 mg daily. Oral fluoxetine at 60 mg daily for 6 days increased prolactin levels in various studies.14,15 Case report. Mary, a 16-year-old African American adolescent girl, presented to the adolescent inpatient unit in 2011 for a chief complaint of “hearing voices and seeing spirits that tell her to kill herself.” As a child, she had delays in meeting her milestones in walking, talking, and toileting. She denied any substance abuse history. Her urine drug screen was negative. Her medical history was unremarkable except for previous diagnosis of schizoaffective disorder. Mary started hearing voices at 12 years of age. Previously, the voices commanded her to hurt others. She was in the 11th grade and receiving special education for behavior issues when she lost her grandfather and started hearing voices again. This time, the voices consisted of the voice of her deceased grandfather, the voice of her aunt, and an unidentified male voice. The voices were instructing her to kill herself now. She had multiple attempts of suicide by cutting herself. Sometimes, she “blacked out” in severe distress from the voices. She reported memory lapses and recalls poor choices and actions made during such episodes. Her medications at admission included oral fluoxetine 10 mg daily and oral ziprasidone 60 mg twice daily. After reviewing her history, symptoms, and medication trials, we decided to treat her with oral fluoxetine 15 mg daily and oral quetiapine extended release (XR) 100 mg daily. Ziprasidone was discontinued because it was not effective. She reported bilateral mastalgia, breast enlargement, and “milk discharge” from her breasts 3 days after starting quetiapine. Galactorrhea was confirmed by physical examination. She denied having headaches and vision changes. Her serum prolactin level was 19.9 ng/mL (reference range is 3.4 to 24.1 ng/mL) 6 days after starting fluoxetine and quetiapine XR. Prolactin levels > 100 ng/mL are characteristic of tumors secreting prolactin.4 We checked her serum prolactin level on 2 different occasions; findings of both serum prolactin assays were not elevated beyond a normal reference range (reference range for a 13- to 15-year-old girl is < 60 ng/mL16). Both prolactin assays were obtained in a fasting state. Quetiapine XR was discontinued at this time. Her serum prolactin level was 19.2 ng/mL 7 days after stopping quetiapine XR. It can take 3 days to 3 weeks for serum prolactin levels to normalize after stopping antipsychotic medications.4 Mary stated that her menstrual cycle was regular over the last 3 months. She did endorse vaginal discharge. The pelvic examination showed pelvic tenderness. Urinalysis confirmed elevated white blood cells and nitrites. We tested her for gonorrhea and chlamydia, and the results were negative. We treated her with doxycycline and azithromycin. Her urine culture was positive for diphtheroid bacilli. Her urine pregnancy test was negative, and her serum beta human chorionic gonadotropin test was also negative. Her thyroid-stimulating hormone and free T4 levels were within normal limits. Euprolactinemic galactorrhea has been associated with thyroid abnormalities.17 Her blood urea and creatinine levels were within normal limits. Prolactin can be high in patients with chronic renal failure.18 Gonadotropin-associated protein and acetylcholine have been identified as prolactin-inhibiting factors in animals.19 A computed tomography scan of the head, performed after hospital discharge, showed no mass lesion in the brain. Mary exhibited hostile behavior and mood alterations with other females that were problematic. These behaviors can be a clinical manifestation of hyperprolactinemia,20 but our patient had prolactin levels within normal limits. Quetiapine was discontinued, and after 2 weeks her galactorrhea resolved. Prolactin levels can be high in patients with chronic renal failure.17 Her serum prolactin level was checked again 2 weeks after discontinuation of quetiapine, and it was still within the normal range. Drug-induced hyperprolactinemia is postulated to occur through various mechanisms. One mechanism decreases dopamine, another increases serotonin. Prolactin can be increased by various medications and mechanisms. Lithium acts on serotoninergic pathways to elevate prolactin. Estrogen potentiation of vasoactive intestinal peptide mediates hypothalamic synthesis of prolactin. Estrogen also has actions on the pituitary lactotroph cells to stimulate prolactin release. Opiates can inhibit dopamine synthesis and raise prolactin. H2 antagonists can inhibit dopamine release. Alprazolam raises prolactin levels through an unclear mechanism. Most psychotropic medications increase prolactin by inhibiting dopaminergic pathways.2,4 Thyrotropin-releasing hormone promotes prolactin release.21 There is a trend to apply atypical antipsychotics to treat a broad spectrum of psychiatric diseases. Quetiapine has a lower affinity for dopamine D2 receptors and appears to be selective for mesolimbic and mesocortical dopamine receptors, with relative sparing of the tuberoinfundibular system.22 Quetiapine has also been used to correct olanzapine-induced galactorrhea.23 However, there is a case report documenting dose-dependent quetiapine-induced galactorrhea.24 Female adolescents are more vulnerable to the side effect of galactorrhea caused by antipsychotic medications. In this case, typical and atypical antipsychotic medications most likely contributed to the galactorrhea, although fluoxetine may have contributed as well. Our patient had a serum prolactin level within normal limits despite the galactorrhea. Our patient was reluctant to reveal that she had galactorrhea; clinicians should screen female adolescents for galactorrhea when prescribing psychotropic medications and administer atypical antipsychotics cautiously.

We previously observed that morphine markedly amplifies LH secretion following intracerebroventricular (i.c.v.) norepinephrine (NE) infusions. Based on additional evidence, we hypothesized that perhaps these morphine effects were due to suppression of tuberoinfundibular dopamine (TIDA) secretion thus allowing NE to evoke a greater release of LHRH from axon terminals in the median eminence than would otherwise occur. In the present studies, we examined whether apomorphine (a DA receptor agonist) would suppress and haloperidol (a DA receptor antagonist) would mimic these enhancing effects of morphine on NE-induced LH secretion. Estrogen-treated ovariectomized rats were used in these studies. NE, when infused i.c.v. (45 micrograms) evoked a modest increase in plasma LH (1.1 +/- 0.2 to 2.2 +/- 0.2 ng/ml) within 15 min. When morphine sulfate (10 mg/kg s.c.) was given 15 min prior to NE, LH peak values of 11 +/- 2 ng/ml were obtained by 60 min. Treatment of rats with apomorphine (1.5 mg/kg s.c.) at -15 min, morphine at 0 min and i.c.v. NE at 15 min resulted in a significant blunting of morphine's effect on NE-induced LH release. Moreover, in all morphine-treated rats, plasma prolactin (PRL) increased significantly within 10 min, peaked at 30 min and declined towards basal values by 90 min. Apomorphine completely blocked this morphine effect of PRL release. Haloperidol (HAL; 2.5 mg/kg s.c.) treatment had no effect on basal LH release but resulted in a significant increase in PRL which remained elevated up to 180 min.(ABSTRACT TRUNCATED AT 250 WORDS)

A major goal in assessing biological determinants of behavior lies in studying the effect(s) of rearing on the development of the central nervous system. Specifically, a series of neuroanatomic analyses have been undertaken to identify potential neuropathological changes seen in monkeys exposed to early social deprivation, which leads to profound psychopathology and inappropriate responses to stress. The animals used in this study were either raised with their mother and peers (socially reared) or raised without maternal/peer contact (socially deprived). Within this context, the distribution of tuberoinfundibular dopaminergic neurons in the hypothalamic paraventricular nucleus and arcuate nucleus of rhesus monkeys was determined by immunohistochemistry using an antibody against the enzyme tyrosine hydroxylase, a marker for dopamine-containing systems. Additionally, the distribution of corticotropin-releasing factor-containing neurons in the paraventricular nucleus was assessed immunohistochemically. The majority (97.5%) of dopaminergic neurons in the paraventricular nucleus were parvicellular, with a small (2.5%), but consistently observed population of magnocellular neurons immunoreactive for tyrosine hydroxylase. Within the arcuate nucleus, tyrosine hydroxylase-immunoreactive neurons were similar in morphology to the parvicellular neurons of the paraventricular nucleus. Qualitative assessment of corticotropin-releasing factor-immunoreactive neurons in the paraventricular nucleus revealed a parvicellular population of neurons located in medial aspects of the nucleus, similar to what has been observed in the rat. Quantitative analysis revealed no differences in the number of tyrosine hydroxylase- and corticotropin-releasing factor-immunoreactive neurons between rearing conditions, suggesting that these neurons were not affected, in terms of overall cell counts, by the early environmental insult of social deprivation.

This paper reviews evidence that changes in the functioning of the brain dopaminergic system affect liver cytochrome P450 (CYP) expression (CYP1A, CYP2B, CYP2C11 and CYP3A in the case of the tuberoinfundibular pathway or CYP1A and CYP3A in the case of the mesolimbic pathway), as well as blood plasma concentration of the respective pituitary hormones in the rat. Thus, the brain dopaminergic system has been established as an important center regulating the liver CYP. This regulation proceeds through the dopaminergic D(2) receptors of the pituitary (activated by the tuberoinfundibular pathway) and the D(2) receptors of the nucleus accumbens (activated by the mesolimbic pathway and conveying a message from the nucleus accumbens to the paraventricular nucleus of the hypothalamus). These receptors directly (GH) or indirectly (CRH --> ACTH --> corticosterone; TRH --> TSH --> T(3)) stimulate the secretion of hormones, which activate nuclear/cytosolic receptors controlling CYP genes. Thus, the prediction of neuroactive drug action on hepatic CYP and drug-drug interactions on the basis of in vitro studies only is not sufficient, because such an experimental model does not allow the central neuroendocrine regulation of the enzyme.

The effect of bombesin on the activity of dopamine (DA) neurons comprising the nigrostriatal, mesolimbic, tuberoinfundibular and tuberohypophysial systems in the male rat was determined by measuring: (1) the accumulation of 3,4-dihydroxyphenylalanine (DOPA) after administration of a decarboxylase inhibitor, and (2) the concentration of the DA metabolite 3,4-dihydroxyphenylacetic acid (DOPAC) in brain (striatum, nucleus accumbens, median eminence) and pituitary regions (intermediate and neural lobes) containing terminals of these neurons. Intracerebroventricular (i.c.v.) injection of bombesin caused a dose- and time-related increase in the activity of DA neurons projecting to the median eminence and intermediate lobe of the pituitary, and a corresponding decrease in the concentrations of prolactin and alpha-melanocyte-stimulating hormone (alpha MSH) in the plasma. In contrast, doses of bombesin up to 10 ng i.c.v. failed to alter the activity of DA neurons terminating in the striatum, nucleus accumbens or neural lobe of the pituitary gland. Equimolar doses of bombesin and gastrin-releasing peptide (GRP), a bombesin-like peptide, increased the concentrations of DOPAC in the median eminence and intermediate lobe of the pituitary, suggesting that GRP-preferring receptors may be responsible for the stimulatory effects of bombesin on DA neuronal activity in these regions. The results of these studies suggest that bombesin increases the activity of tuberoinfundibular and tuberohypophysial DA neurons projecting to the median eminence and intermediate lobe of the pituitary, respectively, and thereby inhibits the secretion of prolactin and alpha MSH.

Stimulation of the ventral noradrenergic ascending bundle (VNAB) at low frequencies (0.5/5 Hz) excited the majority (37/46, 80%) of single paraventricular nucleus (PVN) tuberoinfundibular neurones, with high frequency (50 Hz) trains of stimuli reversing the direction of the response to inhibition for 7/16 (44%) of these excited cells. Iontophoretic application of noradrenaline, or the alpha 1-adrenoceptor agonist 1-phenylephrine, increased the spontaneous electrical activity of most of the cells tested (94% and 72%), whilst application of the alpha 1-antagonist, ergotamine reduced the spontaneous activity of 44% of the cells tested and prevented the excitation following VNAB stimulation for 84% of the cells examined. Application of the beta-adrenoceptor antagonist, propranolol, increased the spontaneous activity of 77% of cells and prevented the inhibitory PVN neuronal responses following high frequency VNAB stimulation of 94% of the cells, often reversing the response to excitation similar to that observed following low frequency VNAB stimulation. The alpha 2-adrenoceptor antagonist, tolazoline, was found to evoke mixed responses from the cells examined but a trend towards a suppression of spontaneous activity and potentiation of VNAB stimulation-evoked responses was observed. The alpha 2-adrenoceptor agonist, clonidine, elicited an initial excitation from the majority of cells tested, with most of the cells then exhibiting an inhibition, either with or without continued application. Excitatory responses following stimulation of the sciatic nerve were recorded from the majority of cells (82.5%) and ergotamine was able to suppress this response for all four cells so tested.(ABSTRACT TRUNCATED AT 250 WORDS)

The effect of cerebroventricular anterior pituitary (AP) grafts on brain tyrosine hydroxylase (TH) expression in the median eminence (ME), substantia nigra (SN), and corpus striatum (CS) has been investigated in young and aged female rats. TH expression was studied using the following indices: in situ TH activity, TH mass, and quantity of TH mRNA. The rate of synthesis of dihydroxyphenylalanine (DOPA) was evaluated by measuring its accumulation in the ME, SN, and CS. TH mRNA and TH mass were quantified by an S1 nuclease protection assay and an immunoblot assay, respectively. Viability of the grafts was demonstrated by histological examination and by their ability to secrete PRL into the cerebrospinal fluid (CSF). Liver grafts served as controls. In castrated young and castrated aged animals with AP grafts, the PRL concentrations in the CSF were 204 +/- 49 (mean +/- SE) and 345 +/- 83 ng/ml, respectively, compared to control values of 14 +/- 9 and 23 +/- 9. In intact aged animals, the concentration of PRL was 729 +/- 180 ng/ml in CSF of rats with AP grafts and 223 +/- 62 ng/ml in the controls. DOPA synthesis in the ME of castrated young rats and castrated aged rats with AP grafts was significantly (P less than 0.01) greater than that in controls with liver grafts. AP grafts did not stimulate DOPA synthesis in the ME of intact aged animals. The synthesis of DOPA in the SN and CS was not affected by AP grafts, regardless of the status of the animal. The amount of TH mRNA and the quantity of TH were not influenced by the AP grafts in any of the animal models. However, the in situ molar activity of TH in the ME was significantly greater in castrated young rats and castrated aged rats bearing AP grafts than in animals with liver grafts. The in situ molar activity of TH in the ME of castrated young rats was greater than that of castrated or intact aged animals. It is concluded that AP grafts secrete a substance that stimulates DOPA synthesis in the ME of young as well as aged castrated animals, but not in the nigrostriatal system. This stimulation is due to increased catalytic activity of a fixed number of TH molecules rather than an increase in the mass of TH.

The effect of the kappa opioid receptor antagonist nor-binaltorphimine (NOR-BNI) was examined on the activity of dopamine (DA) neurons comprising the nigrostriatal, mesolimbic, and tuberohypophysial systems in the male rat. DA neuronal activity was estimated by measuring: (1) the concentration of the DA metabolite 3,4-dihydroxyphenylacetic acid and, (2) the accumulation of 3,4-dihydroxyphenylalanine after administration of a decarboxylase inhibitor in brain (striatum, nucleus accumbens) and pituitary regions (intermediate lobe, neural lobe) containing terminals of these neurons. The intracerebroventicular administration of NOR-BNI produced a dose- and time-related increase in the activity of tuberohypophysial DA neurons, but failed to alter the activity of nigrostriatal or mesolimbic DA neurons. The ability of NOR-BNI to enhance the activity of tuberohypophysial DA neurons was blocked by the kappa opioid agonist U-50,488. These results indicate that NOR-BNI, acting on kappa opioid receptors, activates tuberohypophysial DA neurons projecting to the neural and intermediate lobes of the pituitary.

Genes encoding different cytochrome P450 (CYP) isoforms are regulated by endogenous hormones (e.g. pituitary hormones, thyroid hormones, glucocorticoids) which are all under control of the central nervous system. The aim of the present study was to investigate the influence of lesions of brain dopaminergic pathways on the level and the activity of CYP isoforms (1A, 2A, 2B, 2C6, 2C11, 2D, 3A) in rat liver. At 48 h after lesion of the tuberoinfundibular pathway, only the activity and the protein level of CYP2B were significantly decreased. Seven days after lesion of the above-mentioned pathway, significant inhibition of CYP2B, CYP2C11 and CYP3A activities and a decrease in CYP protein levels were observed. At the same time, the activity and the protein level of CYP1A considerably increased. Fourteen days after damage of the mesolimbic pathway, the activity and the protein level of CYP3A were significantly reduced, while those of CYP1A were substantially elevated. In contrast, lesion of the nigrostriatal pathway did not affect any CYP isoforms studied. The obtained results provide the first direct evidence for the influence of brain dopaminergic system on the level and the activity of CYP in the liver, which is pathway- and isoform-dependent. Hence stimulation or inhibition of the brain dopaminergic system (e.g. by dopamine receptor-blocking neuroleptics) may cause changes in CYP activity of physiological, pharmacological and toxicological significance, since CYP isoforms that are regulated by the dopaminergic system catalyze the metabolism of endogenous substances (e.g. steroids), clinically important drugs (e.g. psychotropics, calcium channel antagonists, antibiotics) and toxins.

Received April 11, 2006; revised April 23, 2006; accepted May 5, 2006. From the Dept. of Psychiatry and Behavioral Sciences, Eastern Virginia Medical School, Norfolk, VA. Send correspondence and reprint requests to Dr. Steinhagen. e-mail: steinhck@evms.edu 2007 The Academy of Psychosomatic Medicine Antipsychotic medications are thought to exert their benefit on psychosis by the mechanism of dopamine D2 receptor antagonism in the mesolimbic pathway. Antipsychotic medications also indiscriminately cause dopamine-receptor blockade in the tuberoinfundibular pathway, which can lead to elevated prolactin levels. Hyperprolactinemia can lead to clinical presentations that include galactorrhea. Pituitary adenomas, which secrete prolactin, can cause hyperprolactinemia, and, by virtue of the pituitary’s location, enlargement of this gland can also cause symptoms such as headaches and visual disturbances. Treatment includes dopamine agonists. Aripiprazole is an antipsychotic medication that is a dopamine D2-receptor partial agonist. It has been shown to effectively treat psychosis and decrease serum prolactin levels. This case report illustrates the use of aripiprazole to effectively treat a patient with psychotic depression, hyperprolactinemia, and a pituitary microadenoma.

The biology of social phobia has been little studied, but a possible role for dopamine has been implicated in this disorder. The aim of this study was to examine central dopaminergic function in patients with generalised social phobia using the prolactin response to quinagolide, a dopamine D2 receptor agonist, and to compare responses with those of normal controls. The study included 14 patients with moderate or severe generalised social phobia and 14 healthy age- and gender-matched comparison subjects. Quinagolide (0.5 mg) was administered orally and prolactin responses were measured over 4 h. There was no significant difference between prolactin responses in patients and healthy controls, nor was there a correlation between prolactin response and age, sex, or severity of illness. This would suggest that tuberoinfundibular dopamine D2 receptor sensitivity is normal in this disorder.

Neuropeptide Y (NPY) has been implicated in the sexual maturation process in chicks and other species. This study attempted to identify specific neural sites where NPY could influence gonadotrophin-releasing hormone (GnRH) neurons during maturation as evidenced by immunocytological changes. To do this sulfamethazine (SMZ), an antibiotic known to advance sexual maturation in male chicks, was used to elicit early testes development. One week posthatch, 20 chicks were fed either a control ration or one with 0.2% SMZ for 2 weeks or 5 weeks. At 3 and 6 weeks of age, birds were perfused through the heart, brains removed, and prepared for immunocytochemical assay for either NPY or GnRH. Results showed that chicks given SMZ had significantly greater testes weight compared to controls at both ages (por = 0.05) while body weight did not differ significantly between these groups at either age. The results of cell quantification showed no difference in the number of GnRH neurons between the SMZ chicks and controls in either the bed nucleus of the pallial commissure or the anterior lateral thalamic nucleus (LA) at 3 or 6 weeks of age. There was a significantly greater number of GnRH neurons found within the LA of 6-week compared to 3-week-old control chicks. A ratio of the mean width of the external zone (EZ) of median eminence (ME) to the total width of ME showed no significant difference between the experimental and control animals at either 3 or 6 weeks of age. The ratio EZ/ME did, however, increase as a function of age. The results of the assay for immunoreactivity to NPY showed no significant difference in the number of NPY neurons found within the nucleus marginalis tractus opticus between treatment groups at either age. A highly significant increase in NPY neurons was found within the infundibular nucleus (IN) and the internal zone of the ME of SMZ-treated chicks compared to controls at 3 weeks of age. Data suggest that NPY-like neurons within the IN complex may serve an important role regarding the early stages of sexual maturation in chicks.

Anatomical relationships between tachykinin-containing terminals and neurons of the medial preoptic area that innervate the arcuate nucleus were studied using silver staining of the retrograde tracer wheat germ agglutinin-apoperoxidase-gold (WGA-ApoHRP-gold) complex injected in the arcuate nucleus and pre-embedding immunocytochemistry for neurokinin A (NKA). At the histological level, retrogradely labeled cells not stained for NKA were seen to be surrounded by numerous NKA-immunopositive punctate profiles, in particular in the dorsal part of the medial preoptic area. At the ultrastructural level, retrogradely labeled cell bodies and dendritic profiles displayed highly electron-dense silver particle accumulations over the cytoplasm. The were seen in synaptic contact with one or several NKA-immunoreactive axon terminals containing small clear vesicles and dense-cored vesicles. Such synapses were either symmetrical or asymmetrical. The occurrence of synaptic contacts between tachykinin terminals and cells innervating the arcuate nucleus in the medial preoptic region provides a morphological support for a tachykinergic regulation of preoptic afferences to the arcuate nucleus. These results suggest that tachykinins are implicated in the indirect control of neuronal activity in the arcuate nucleus notably via the preoptic area. Consequently, tachykinins are potentially able to regulate indirectly numerous neuroendocrine events involving the tuberoinfundibular system.

In male rats anesthetized with urethane (600 mg/kg) and alpha-chloralose (60 mg/kg), extracellular recordings were made from tuberoinfundibular (TI) neurons in the paraventricular nucleus (PVN) identified by antidromic stimulation of the median eminence. Electrical stimulation of the gastric branches of the vagus nerve excited 26 (72%) of 36 TI neurons tested. I.v. (0.5-1 microgram/rat) or i.p. (5 micrograms/rat) administration of cholecystokinin (CCK-8) excited 10 (63%) of 16 TI neurons tested. The excitatory responses induced by CCK-8 were abolished by bilateral cervical vagotomy. These results suggest that TI neurons in the PVN receive excitatory inputs from gastric afferents via vagus nerves.

In this study we examined electrophysiologically the involvement of the intrinsic GABAergic system of the accessory olfactory bulb (AOB) in controlling the activity of tuberoinfundibular (TI) arcuate neurons in anaesthetized female mice. Local infusions of the gamma-aminobutyric acid-A (GABAA) receptor antagonist, bicuculline into the AOB enhanced the spontaneous firing activity of TI arcuate neurons with excitatory inputs from the AOB. This finding reveals a neural mechanism responsible for the pregnancy blocking effect of this drug in freely behaving female mice and, taken together with the cytoarchitecture of the AOB, suggests that the reciprocal dendrodendritic interaction between mitral cells and GABAergic granule cells in the AOB is critical to control of AOB output to TI arcuate neurons as part of the final common pathway of the accessory olfactory system.

Tyrosine hydroxylase (TH) activity was measured in discrete brain regions of rats during short-term forced running stress (FRS). TH activity was also determined in a depression-like state and in a recovered state after a long-term FRS. Under the short-term FRS, the TH activity showed a significant increase in the locus ceruleus, certain limbic regions and tuberoinfundibular system. In the depression-like state, however, there was a significant decrease in the locus ceruleus and certain limbic regions, but a significant increase was seen in the median eminence. The TH activity in recovered rats showed no difference from the level in the controls. These findings demonstrate an adaptive increase in the TH activity in relation to stress, and may also indicate a failure of adaptation in the depression-like state.

In female guinea pigs, a combination of retrograde tracing and immunofluorescence for progesterone receptors (PR) was applied to determine if PR-immunoreactive (PR-IR) neurons in the arcuate nucleus (AR) send their axons directly to the median eminence (ME). Axonal projections to the ME were studied by different techniques using fluorescent dyes. From 31 adult animals, ovariectomized and primed by estradiol, small deposits of Lucifer Yellow (LY) were made on the cut surface of the ME, either by direct application of LY crystals or by iontophoresis. These techniques were carried out on excised mediobasal hypothalamus maintained in vitro and allowed visualization of AR perikarya projecting to the ME after dye diffusion in the severed axons. In another group of ten immature animals primed by estradiol, Granular Blue (GB) was injected in the jugular vein. Blood-borne GB was taken up in the ME by intact nerve endings and retrogradely transported to the perikarya of origin. PR-IR neurons and perikarya filled with LY or retrogradely labeled by GB were intermingled with each other throughout the rostrocaudal extent of the AR. Double-labeled cells, displaying PR immunoreactivity and dye labeling, were observed consistently, but their number was small. This result demonstrates that some AR neurons sending axonal projections to the ME are target cells for progesterone. As the majority of PR-IR neurons in the AR do not project to the ME, it is suggested that most PR-IR neurons present in this nucleus form local circuit projections or project to distant areas of the central nervous system.

Insulin (63 microM) stimulated endogenous dopamine (DA) release from tuberoinfundibular neurons. This effect was independent on the presence of extracellular glucose and did not involve the outward transport of DA, mediated by its membrane carrier. By contrast this effect was completely prevented by the removal of extracellular Ca++ ions in presence of the Ca(++)-chelator ethyleneglycol-2-(2-aminoethyl)-tetracetic acid (EGTA). Furthermore 1-(5-isoquinolinyl-sulfonyl)-2-methyl-piperazine (H7), a compound which behaves as a putative inhibitor of protein kinase C (PK-C) (10 microM), completely counteracted the stimulation of endogenous DA release induced by insulin. Amiloride (300 microM) and its 5-amino nitrogen atom-substituted derivative, 5-(N-methyl-N-(guanidinocarbonylmethyl) amiloride (MGCMA) (10 microM), a highly selective inhibitor of the Na(+)-H+ membrane antiporter, were both able to prevent the stimulatory action exerted by insulin on endogenous DA release. Collectively, these results suggest that the transductional events by which insulin stimulated endogenous DA release from TIDA neurons may involve the activation of PK-C, the enhancement of Ca++ influx and the stimulation of the Na(+)-H+ exchange system.

The role of ascending noradrenergic projections of medullary origin in regulating the activity of tuberoinfundibular neurons of the hypothalamic paraventricular nucleus (PVN) was examined in pentobarbital-anesthetized male Sprague-Dawley rats. Discrete electrical stimulation of either the A1 or the A2 noradrenaline cell group areas of the caudal medulla enhanced the probability of firing in a substantial proportion of antidromically identified tuberoinfundibular PVN cells tested. Notably, no inhibitory effects were observed. Destruction of the PVN noradrenergic terminal plexus by local application of the neurotoxin 6-hydroxydopamine 1 day prior to electrophysiological experiments abolished the effects of both A1 and A2 stimulation. These findings indicate that noradrenergic afferents can exert a facilitatory influence on the activity of a population of tuberoinfundibular PVN neurons, thus supporting earlier suggestions that central noradrenergic structures can enhance the release of certain anterior pituitary hormones.

The corticotropin releasing factor (CRF)-immunoreactive paraventriculo-infundibular neuronal system of long-term adrenalectomized and adrenalectomized-short term dexamethasone treated rats was analyzed at the ultrastructural level using the preembedding peroxidase anti-peroxidase complex (PAP)-immunohistological method. In both groups of animals, parvocellular neurons located in the medial and dorsal subnuclei of the paraventricular nucleus (PVN) showed CRF-like immunoreactivity. The perikarya contained hypertrophied rough endoplasmic reticulum (rER) with dilated cisternae, active Golgi-complexes and numerous neurosecretory granules. The majority of the neurosecretory granules measured 80-120 nm. Dendrites of CRF-immunoreactive neurons contained labeled vesicles, secretory granules, bundles of microtubules, a well-developed smooth endoplasmic reticulum (sER) complex and free ribosomes. Unlabeled terminal boutons of axons were observed to synapse on dendrites and somata of CRF-neurons. In addition, CRF perikarya were found in direct somato-somatic apposition with both CRF-immunopositive and immunonegative parvocellular cells. Retraction of glial processes and the existence of puncta adherentia between the cell membranes characterized these appositions. Varicose CRF axons within the median eminence contained hypertrophied sER, labeled vesicles and neurosecretory granules. The preterminal portions of the CRF-axons were dilated and possessed many labeled 80-120 nm diameter granules. CRF-terminals were greatly enlarged and established direct neurohemal contacts with the external limiting basal lamina of portal vessels without the interposition of tanycytic ependymal foot-processes. These tanycytes were not CRF immunopositive. CRF positive terminals contained clusters of microvesicles, labeled small vesicles and multivesicular bodies, but fewer granular elements than were observed within the the preterminals. Many of the labeled organelles were attached to tubules of sER. Occasionally, CRF-axons were observed within the pericapillary space adjacent to portal vessels. The ultrastructural features of CRF-neurons, obtained from adrenalectomized and adrenalectomized plus short-term dexamethasone treated rats did not differ significantly from each other. The hormone content of the entire CRF-neuron was greater in the steroid treated group. Adrenocorticotrophic hormone (ACTH) synthesizing cells in the pars distalis of adrenalectomized-dexamethasone treated rats also showed increased numbers of immunopositive secretory granules (150-320 nm in diameter).(ABSTRACT TRUNCATED AT 400 WORDS)

The regulation of prolactin secretion by tuberoinfundibular dopamine (DA) neurons appears to be altered in the aged rat: the concentration of prolactin in the serum increases and the activity of the tuberoinfundibular DA neurons decreases. In the young female rat a brief period of stress reduces the tuberoinfundibular DA neurosecretory activity and increases the secretion of prolactin. The purpose of the present study was to determine if the responsiveness of tuberoinfundibular DA neurons to restraint stress is altered in the aged female rat. The activity of these neurons was estimated from the rate of DA synthesis in their terminals in the median eminence, as measured by the rate of accumulation of dihydroxyphenylalanine (DOPA) after the administration of a decarboxylase inhibitor. Thirty minutes of restraint stress increased serum prolactin concentrations in both young (3 months) and aged (26 months) constant estrous rats, but reduced the rate of DOPA accumulation in the median eminence of only the young rats. Restraint also decreased the rate of DOPA accumulation in the median eminence of intermediate-aged rats (14 months) independently of whether the rats were exhibiting normal ovarian cycles (measured on the day of estrus) or were in a constant estrus. This suggests that the loss of ovarian cyclicity per se is not associated with the age-related change in the response of tuberoinfundibular DA neurons to restraint stress.(ABSTRACT TRUNCATED AT 250 WORDS)

The effects of acute stress on serum prolactin concentrations and tuberoinfundibular dopaminergic (TIDA) neuronal activity were studied in female rats. TIDA neuronal activity was estimated by measuring the rate of dihydroxyphenylalanine (DOPA) accumulation after the administration of a decarboxylase inhibitor (NSD 1015) and the rate of decline of dopamine (DA) after the administration of a tyrosine hydroxylase inhibitor (alpha-methyltyrosine) in the median eminence. Serum prolactin concentrations were increased following 30 min of supine immobilization (restraint stress), but returned to control levels by 2, 8, and 16 h after the onset of this stress. The rate of DOPA accumulation was decreased during the 30 min of restraint; it was still further reduced 2 h later but had returned to control levels 8 and 16 h later. No change in the rate of DOPA accumulation was observed in the striatum or neurointermediate lobe of the pituitary at any time after the start of restraint. Restraint stress also decreased the rate of DA turnover in the median eminence, but was without effect on the rates of DA turnover in the striatum or neurointermediate lobe. These results suggest that restraint stress activates an inhibitory neuronal pathway which decreases the activity of TIDA neurons and may be responsible, at least in part, for the increase in serum prolactin concentrations. The responsiveness of TIDA neurons to the stress-induced decrease in activity was not influenced by the time of day or the stage of the estrous cycle. Not all stressful manipulations decreased TIDA neuronal activity.(ABSTRACT TRUNCATED AT 250 WORDS)

In human fetus, newborn, infant and adult hypothalami, antibodies to ovine corticoliberin-41 stain a paraventriculo-infundibular neuroglandular pathway. The perikarya are located in the paraventricular nucleus, they mainly project to the ventral and lateral areas of the median eminence. Eminential corticoliberin-positive fibres appear during the 16th week of fetal life, and increase in number during the following weeks. Perikarya were first revealed in the 19th week. In some areas of the median eminence, corticoliberin-, vasopressin- or [Met]enkephalin-immunoreactive terminals are similarly distributed. Sequential stainings or staining comparison of contiguous semi-thin sections failed to prove the coexpression of corticoliberin and [Met]enkephalin immunoreactivities in fibres, but indicated that corticoliberin and vasopressin immunoreactivities may be coexpressed in a few fibres. Those methods enabled us to observe, in the paraventricular nucleus, perikarya revealed by corticoliberin and vasopressin antisera. Our results suggest a possible release of corticoliberin in portal vessels of the median eminence beginning in the 16th week of fetal life i.e. 8 weeks later than appearance of the corticotrophs in the pituitary. Establishment of a corticoliberin hypothalamic control of pituitary corticotrophs at mid gestation agrees with previous physiological and teratological studies. Abundance, as well as immunostaining intensity of the corticoliberin processes, in the infant and adult median eminence attest to the physiological importance of this system. Close vicinity of corticoliberin, vasopressin and [Met]enkephalin fibres, in some eminential areas and coexpression of corticoliberin and vasopressin immunoreactivities in some neurons, are morphological correlates of functional relations which were reported.

The possible involvement of calmodulin in the process of endogenous dopamine (DA) release from arcuate-periventricular nuclei-median eminence fragments, containing tuberoinfundibular dopaminergic (TIDA) neurons, has been investigated in an in vitro incubation system. For this purpose the basal and K+-stimulated DA release was examined in the presence and in the absence of the different putative calmodulin antagonists, pimozide, trifluoperazine, penfluridol and N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide (W-7). Trifluoperazine and pimozide in concentrations up to 100 microM were both uneffective in blocking K+-evoked DA release. Penfluridol in doses of 5 and 10 microM, did not prevent 35 mM K+-induced endogenous DA release. It was able to reduce K+-stimulated DA release only at the very large concentration of 100 microM. W-7 added in vitro to the hypothalamic fragments, prevented endogenous DA release evoked by 35 mM K+ in a dose-dependent manner. W-5, a chlorine deficient analogue of W-7, that interacts only weakly with calmodulin, failed to modify K+-stimulated endogenous DA release in doses up to 200 microM. All the putative calmodulin antagonists used in the present study did not induce any change of basal DA release. In conclusion the fact that most of the agents, except W-7, known to antagonize calmodulin-dependent processes in many biological systems failed to interfere with the release of endogenous DA from TIDA neurons seems to suggest that calmodulin does not play a crucial role in the process of DA release and that the inhibitory effect of W-7 on endogenous DA release may be better attributed to other mechanisms different from its anticalmodulin action.

The effect of long-term decreases in circulating concentrations of prolactin was determined on the responsiveness of tuberoinfundibular dopamine (DA) neurons to this hormone. The activity of these neurons in ovariectomized rats was estimated by measuring the rate of DA synthesis (DOPA accumulation after the administration of a decarboxylase inhibitor) in the median eminence at various times after serum concentrations of prolactin had been reduced by hypophysectomy or the chronic administration of a DA agonist (bromocriptine, 3 mg/kg/day). The concentration of DA in the median eminence, but not in striatum, declined progressively up to 12 days after hypophysectomy, but did not change at any time during bromocriptine treatment. On the other hand, norepinephrine concentrations in the median eminence were increased 12 days after both treatments. Within 24 h after hypophysectomy or the first injection of bromocriptine the rate of DA synthesis in the median eminence was decreased; this decrease was maintained for at least 12 days suggesting that tuberoinfundibular DA neuronal activity is normally maintained by endogenous prolactin. Intracerebroventricular (ICV) injections of prolactin (10 µg, 12 h prior to sacrifice) increased the rate of DA synthesis in the median eminence of control, 24-hour hypophysectomized and 24-hour bromocriptine-treated rats. After longer periods (6–12 days) of bromocriptine treatment or after hypophysectomy the responsiveness of tuberoinfundibular DA neurons to prolactin was reduced. Dose-response studies revealed that the sensitivity and magnitude of response to ICV prolactin was markedly reduced in 12-day hypophysectomized rats. The decreased responsiveness of tuberoinfundibular DA neurons to the actions of prolactin following long-term bromocriptine treatment and hypophysectomy is most likely due to the prolonged decrease in circulating prolactin, since the concurrent daily administration of bromocriptine and ICV prolactin for 12 days maintained the ability of tuberoinfundibular DA neurons to respond to prolactin. The decreased basal activity and responsiveness of tuberoinfundibular DA neurons to prolactin induced by hypoprolactinemia is reversible; 12 days after terminating chronic bromocriptine treatment basal activity and the responsiveness of tuberoinfundibular DA neurons to prolactin return to control values. These results suggest that the response of tuberoinfundibular DA neurons to prolactin is determined, in part, by the preceding circulating levels of this hormone.

The secretion of prolactin from the adenohypophysis is tonically inhibited by dopamine that is released into the hypophysial portal blood from terminals of tuberoinfundibular neurons located in the external layer of the median eminence. These tuberoinfundibular neurons are unique among other dopaminergic neurons in the brain (including the well-characterized nigrostriatal neurons) in that they are not directly regulated by dopaminergic receptor-mediated mechanisms, but instead are selectively responsive to changes in prolactin concentrations in blood and cerebrospinal fluid. In the rat, the activity of the tuberoinfundibular dopaminergic neurons is higher in the female than in the male, exhibits a characteristic cyclical pattern during the first half of pregnancy and is constantly high as a result of stimulation by placental lactogen during the last 9 days of pregnancy, and is reduced in lactating animals and acutely inhibited during suckling.

It is well established that opiate agonists alter tuberoinfundibular dopaminergic activity and consequently prolactin release. The purpose of this study was to characterize the effects of morphine on prolactin secretion and tuberoinfundibular dopaminergic neuronal activity with respect to time after administration. Additionally, the effect of an initial morphine injection on the response produced by a second injection of morphine was determined. The rate of depletion of median eminence dopamine content following synthesis inhibition byα-methyl-p-tyrosine was used as an index of dopaminergic neuronal activity. Male rats given a single injection of morphine sulfate (15 mg/kg, s.c.) showed a significant increase in circulating prolactin levels and had a lower rate of median eminence dopamine turnover 1 h after injection. Four hours after injection, circulating prolactin levels were similar to those in vehicle treated rats, while dopamine turnover was significantly higher than controls. When two injections of morphine sulfate (15 mg/kg, s.c.) were given 4 h apart, the stimulation of prolactin release produced by the second injection was significantly attenuated. Although this second injection caused a significant decrease in dopamine turnover, the turnover rate following this injection was significantly greater than that following the initial injection. The combination of fluoxetine and 5-hydroxytryptophan (FLX/5-HTP) caused an initial increase in prolactin secretion with plasma values returning to basal levels by 4 h. When rats were pretreated with FLX/5-HTP instead of morphine, the prolactin response to an injection of morphine 4 h later was not attenuated. Similarly a FLX/5-HTP pretreatment had no influence on a second injection of FLX/5-HTP administered 4 h later. The attenuation of a secretory response to a second morphine injection is specific to prolactin, since the growth hormone responses to the initial and the second injections of morphine were the same. These results indicate that the response of tuberoinfundibular dopaminergic neurons to morphine is biphasic. Following morphine administration, dopamine turnover is initially decreased but by 4 h turnover is increased above pretreatment rates. These alterations in dopamine turnover affect prolactin secretion such that release is initially stimulated, but 4 h later the response to a second morphine stimulation is attenuated.

Previously, we demonstrated that morphine (MOR) and opioid peptides inhibit the turnover of dopamine (DA) in the tuberoinfundibular pathway (TIP) in a dose-dependent, naloxone (NAL)-reversible manner. Subsequent studies showed that tolerance to both the effects of MOR on PRL release and DA turnover in the median eminence (ME) develops during the same period of chronic MOR treatment, which supports the idea that opioids increase the rate of PRL release by inhibiting the activity of the DA neurons of the TIP, which inhibit the release of PRL. We now report that cross-tolerance develops between the effects of ether stress and MOR on plasma PRL levels, which strongly supports the idea that endogenous opioids are involved in the stress-induced activation of PRL release. However, our data do not support the idea that opioid pathways are involved in the estrogen-induced activation of PRL release. The evidence against opioid mediation of estrogen's action on the release of PRL follows. 1.) The effects of 17-beta-estradiol (EB) and of a maximally effective dose of MOR on plasma PRL concentration are additive, which suggests that these agents act by different mechanisms. 2.) EB increases the rate of DA turnover in the ME of ovariectomized (OVX) rats treated for 3 days with 20 micrograms/kg of EB per day, while MOR inhibits DA turnover in this group and all other groups studied. 3.) Although NAL decreases EB-elevated plasma PRL levels at one of two time points that were studied, it also inhibits the effect of alpha-methyl-rho-tyrosine (AMT) on the concentration of PRL in the plasma, and AMT acts by a non-opioid mediated mechanism to activate the release of PRL.

A new method for estimation of in vitro neurosecretory activity of tuberoinfundibular dopaminergic (TIDA) neurons was developed by measuring the rate of synthesis of dihydroxyphenylalanine (DOPA) in the median eminence of hypothalamic slices. Sagittal hypothalamic slices of ovariectomized rats were incubated in a medium containing 3-hydroxybenzylhydrazine (NSD 1015), an inhibitor of DOPA decarboxylase. DOPA accumulated in the median eminence following incubation with NSD 1015 was determined by high-performance liquid chromatography with electro-chemical detection. The amount of DOPA accumulated in vitro in the median eminence was maximal in a medium containing 10 mM NSD 1015 and linear up to 120 min at 37 degrees C. Increasing the concentration of tyrosine in medium stimulated the synthesis of DOPA in the median eminence. The synthesis of DOPA was blocked by 1 mM alpha-methyltyrosine, an inhibitor of tyrosine hydroxylase. The rate of in vitro synthesis of DOPA in the median eminence was 33% of that of in vivo synthesis. Incubation in a medium containing 50 mM K+ to depolarize neurons caused a 2.4-fold increase in DOPA synthesis in the median eminence. The high K+-induced increase in DOPA synthesis was blocked by omission of Ca2+ and addition of 1 mM EGTA into the medium, suggesting Ca2+ dependency of depolarization-activated DOPA synthesis. These results indicate that this in vitro assay is a useful means to study the regulatory mechanisms of TIDA neurons.

In fetal and postnatal rats, serum melanophore-stimulating hormone (MSH) levels, pituitary 3H-spiperone binding, pituitary catecholamine content, and other parameters were measured to characterize the development of pars intermedia pituitary control by the tubero-hypophyseal dopamine system. Bioassay of serum for MSH revealed low but significant levels 6 days before birth (gestational day 17), rising to a peak 4 days after birth (postnatal day 5). Levels declined after PN 6, reached a low 2 weeks after birth, and then rose gradually throughout the next several weeks until adult values were attained. From excised pituitaries, significant catecholamine fluorescence in the neurointermediate lobe was first demonstrated on the 2nd postnatal day, with the highest density of fibers being attained at 2 weeks after birth. Thereafter, the density of terminals decreased in the pars intermedia until the adult pattern of innervation was observed at day 60. Total dopamine content of the neurointermediate lobe roughly paralleled the morphological observations through the first 2 weeks after birth. Later, content was seen to stabilize, while concentration decreased in line with the reduction in terminal density. Dopamine levels exceeded norepinephrine concentration by a factor of 10. 3H-spiperone binding sites were present in the pituitary before birth. Their density increased rapidly in the neurointermediate lobe during the first 2 postnatal weeks, then diminished until adult densities were reached at day 30. Binding in the anterior pituitary slowly rose throughout the first 30 days after birth, although values were only about one-quarter of that found in the neurointermediate lobe. In vitro incubation of neurointermediate lobes with dopamine showed a dose-dependent inhibition of spontaneous MSH secretion as early as 2 days after birth. Blockade of dopaminergic binding sites by i.p. administration of cis-flupenthixol (1 mg/kg) to rat pups had no effect until days 8, 15, and 22 where there was a significant rise in serum MSH levels as compared with controls. These data indicate that the intermediate lobe is responsive to dopamine at around birth, while the inhibitory dopaminergic control develops only during the 1st postnatal week, in conjunction with marked changes in circulating MSH levels.

In the tubero-hypophyseal system of the rat the approximate concentrations of immunoreactive substance P (I-SP), expressed as fmol/mg tissue, were 100 in the medial basal hypothalamus (MBH), 20 in the anterior lobe (AL), 20 in the neural lobe (NL) and 4 in the intermediate lobe of the hypophysis. These values were not altered by treatment with capsaicin on day 2 after birth. In rats in which the secretion of neurohormones from the NL was increased by dehydration followed by sodium loading there was a fall by 70% in the I-SP concentration of the NL; no change occurred in the AL or the MBH.

Electrical stimulation in the subfornical organ (SFO) alters the excitability of antidromically identified paraventricular nucleus neurons. Extracellular recordings demonstrate that the dominant effect of single stimuli delivered to the SFO on neurohypophyseal oxytocin and vasopressin containing neurons is an increase in excitability. In 35% of cells tested, this excitation showed a long latency (44.3 +/- 3.4 ms) prolonged duration (208.7 +/- 23.5 ms), while in 16% of the neurons the excitation observed may be described as short latency (24.7 +/- 1.8 ms) short duration (11.6 +/- 1.4 ms). Of the remaining cells antidromically identified as projecting to the posterior pituitary, 12% showed initial decreases in excitability following SFO stimulation while the remaining 37% were unaffected. Evidence is presented demonstrating that stimulation in the region of the SFO results in short latency (27.9 +/- 2.4 ms) short duration (7.8 +/- 0.7 ms) increases in excitability in 22% of antidromically identified PVN tuberoinfundibular neurons tested. These data provide electrophysiological evidence in support of the proposed role of the subfornical organ in the control of posterior and anterior pituitary function.

Single unit activity was recorded from tuberoinfundibular neurones in urethane-anaesthetized pro-oestrous rats. Responses following single shock (0.2 Hz) stimulation of the preoptic area/anterior hypothalamus (PO/AH) or nucleus accumbens (ACB) were recorded and computed for 137 single units. More cells were shown, by antidromic activation, to project to ACB (4 of 77) than to PO/AH (1 of 60). Fewer units were responsive to stimulation of ACB (p less than 0.02), those that were responsive showing a longer latency to the onset of response compared to that following PO/AH stimulation. Significantly (p less than 0.02) more slowly firing cells were found in the group of animals tested with stimulation of PO/AH. Delivery of high frequency stimulation to PO/AH resulted in a 21/2 fold increase in plasma luteinizing hormone (LH) levels (p less than 0.05), whereas ACB-stimulated and control animals showed no increases in plasma LH concentrations. During delivery of the high frequency stimulation only one cell in the ACB group of animals was orthodromically affected (out of five tested). However, in the PO/AH group, five of the seven cells tested showed either excitation or inhibition during stimulation. The possibility that the functional connections demonstrated between ACB and the endocrine hypothalamus are involved in the expression of 'higher' brain functions, recognised during changing endocrine states, is discussed.

The basal activity o f tuberoinfundibular dopaminergic (TIDA) neurons is higher and the response of these neurons to the stimulatory actions of prolactin is greater in the female than in the male rat. In the female rat, the restraint-stress-induced increase in serum prolactin concentrations is accompanied by a concurrent decrease in the activity of TIDA neurons. The purpose of the present study was to compare these effects of restraint in male and female rats. TIDA neuronal activity was estimated by measuring the rate of dopamine (DA) synthesis (DOPA accumulation after the administration of a decarboxylase inhibitor, NSD 1015) and the rate of DA turnover (decline of DA after administration of a tyrosine hydroxylase inhibitor; alpha-methyltyrosine) in the median eminence. Thirty minutes of restraint increased serum prolactin concentrations in both male and female rats, but a greater response was observed in the females. Restraint also decreased the rates of synthesis and turnover of DA in the median eminence of the female but not the male rat. The difference in the response of TIDA neurons in male and female rats to restraint is not the consequence of neuronal differentiation resulting from neonatal androgen exposure, because restraint aso decreased the activity of TIDA neurons in androgen-sterilized female rats. The inability of restraint stress to reduce TIDA neuronal activity in the male rat appears to be the consequence of testosterone, since TIDA neurons were responsive to restraint following castration of the males.(ABSTRACT TRUNCATED AT 250 WORDS)

The autoregulatory feedback control of prolactin, which is mediated by tuberoinfundibular dopamine (DA) neurons, is altered in the aged rat; this is evidenced by increased circulating concentrations of prolactin and decreased activity of these neurons. In the present study the action of prolactin on tuberoinfundibular DA neurons in young and aged female rats was estimated by measuring the rate of DA synthesis (dopa accumulation following the administration of a decarboxylase inhibitor) in the median eminence. The rate of dopa accumulation in the median eminence of the aged (26 months) rat was reduced to 50-60% of that in the young (3 months) rat. The acute systemic administration of haloperidol, a DA antagonist which increases serum concentrations of prolactin or intracerebroventricular infusions of prolactin increased the rate of dopa accumulation in the median eminence of both young and aged rats by the same relative amount. The administration of haloperidol and prolactin increased the rate of DA synthesis to a greater extent in young than in aged rats. The administration of bromocriptine, a DA agonist which reduces serum concentrations of prolactin, decreased the rate of dopa accumulation in the median eminence of both young and aged rats. In young animals the intracerebroventricular administration of prolactin reversed the bromocriptine-induced decrease in DA synthesis in the median eminence after 4 h and caused a further increase after 12 h. Qualitatively similar effects were seen in the aged rats; however, prolactin-treated young rats had much higher levels of DA synthesis than aged rats.(ABSTRACT TRUNCATED AT 250 WORDS)

Immunoreactive substance P (SPi) was measured in the various hypothalamic structures and amygdala one week after placing a cut around the medial basal hypothalamus (MBH). SPi content decreased in the stalk-median eminence (SME), anterior and posterior parts of the arcuate nucleus (inside the isolated region) as well as in the paraventricular nucleus (outside the cut), while there was no change in the posteromedial amygdaloid nucleus. We suggest that there is a tuberoinfundibular neuronal system containing SPi with cell bodies in the MBH and terminals in the SME. In addition, nerve fibers containing SPi from outside the MBH may reach both the SME and the arcuate nucleus and might influence neuroendocrine regulation.

Previous results demonstrated that GABA exerts a dual control on PRL secretion, one excitatory mediated in part by the impairment of the tubero-infundibular dopaminergic (TIDA) system function, the other inhibitory occurring at the level of the anterior pituitary (AP), where 3H-GABA and 3H-Muscimol (3H-M) recognition sites have been described. This report provides evidence for a physiological role of the tubero-infundibular GABAergic system (TI-GABA) on PRL secretion in the rat. In lactating rats separated for 4 h from their pups reinstitution of suckling for different periods resulted in an increase either in glutamic acid decarboxylase (GAD) activity in the mediobasal hypothalamus (MBH) or in AP-GABA content. Dynamic changes of the GABAergic function in the MBH-AP system seemed to have a certain degree of specificity because suckling did not affect GAD activity in the caudate nucleus. In lactating rats 2, 4, 8 and 24 h after removal of the offsprings AP-GABA concentrations and plasma PRL titers significantly decreased with respect to values present in rats never separated from their pups. Since it has been demonstrated that the PRL lowering effect of GABA is a receptor-mediated event, we have investigated the plasticity of AP-GABA receptors during suckling. The inhibitory action of GABA seems to be mediated mainly by the activation of the high affinity binding sites. This proposition is supported by the fact that in lactating rats, where only the high affinity receptor population is present, M was still able to decrease significantly plasma PRL concentrations. These results indicate that the MBH-AP GABAergic system could play a role on PRL secretion, being involved in mechanisms which dampens PRL output during continuous suckling stimulation.

Immunohistochemistry was used to detect tyrosine hydroxylase (TH)-containing neurons in the hypothalamic arcuate nucleus in male rats. Two weeks following castration, the number of TH-positive cells was significantly greater than in intact controls. The castration-responsive TH-positive cells were uniformly distributed throughout the mediolateral extent of the arcuate nucleus. Treatment with testosterone significantly suppressed the castration response, whereas neither estradiol nor 5α-dihydrotestosterone were effective. The number of TH-positive arcuate neurons in the female was similar to that in the male. Ovariectomy did not affect the number of TH-positive neurons. These findings indicate that TH expression is tonically inhibited in the tuberoinfundibular dopaminergic system of the male rat by testosterone.

Using an antiserum which reacts with the carboxyl terminus of GRF(1–44)NH 2 , the distribution of immunoreactive growth hormone releasing factor (GRF) in the rhesus monkey hypothalamus was delineated by peroxidase immunocytochemistry. Immunoreactive material was present in dense terminal fields in the median eminence closely associated with portal capillaries but in a location distinct from that noted for immunoreactive thyrotropin-releasing hormone (TRH) or somatostatin. GRF-immunoreactive cell bodies were identified in the arcuate nucleus and ventromedial nucleus. These studies provide evidence for the presence of GRF(1–44)NH 2 in the primate brain and demonstrate that in the hypothalamus it is localized exclusively in cells and fibers corresponding to the tuberoinfundibular system.

To address the issue of whether, after morphine treatment, the reduced release of dopamine (DA) into portal blood is entirely responsible for the increased prolactin (PRL) release, the following study was conducted. The concentration of DA in plasma from a single portal vessel of untreated, ovariectomized rats was 1.85 +/- 0.33 ng/ml (mean +/- SE). Treatment of ovariectomized rats with alpha-methyl-p-tyrosine (alpha MT) caused a 91% reduction in the concentration of DA in portal plasma. Infusion of DA (0.4 microgram/min/kg BW) into a jugular vein of rats pretreated with alpha MT restored the DA concentration in portal plasma to that seen in untreated rats. Injection of morphine sulfate elicited a marked increase in the concentration of PRL in plasma. Infusion of DA at rates of 0.4 and 0.8 microgram/min/kg BW suppressed by 52 and 75%, respectively, the secretion of PRL after morphine treatment. Infusion of DA had no effect on the release of PRL induced by intracerebroventricularly administered beta-endorphin. Although treatment of rats with alpha MT caused release of PRL that was similar to that seen in rats treated with morphine, infusions of DA at 0.4 and 0.8 microgram/min/kg BW into alpha MT-treated, ovariectomized rats suppressed the secretion of PRL by 89 and 96%, respectively. Thus, after morphine treatment, the decreased release of DA into portal blood is not in itself sufficient to account for the increase seen in the secretion of PRL. It is suggested that morphine and opiate-like peptides induce the release of a hypothalamic substance(s) that stimulates PRL release.