TSH Pulses Finely Tune Thyroid Hormone Release and TSH Receptor Transduction.

Detection of circulating TSH is a first-line test of thyroid dysfunction, a major health problem (affecting about 5% of the population) that, if untreated, can lead to a significant deterioration of quality of life and adverse effects on multiple organ systems. Human TSH levels display both pulsatile and (nonpulsatile) basal TSH secretion patterns; however, the importance of these in regulating thyroid function and their decoding by the thyroid is unknown. Here, we developed a novel ultra-sensitive ELISA that allows precise detection of TSH secretion patterns with minute resolution in mouse models of health and disease. We characterized the patterns of ultradian TSH pulses in healthy, freely behaving mice over the day-night cycle. Challenge of the thyroid axis with primary hypothyroidism because of iodine deficiency, a major cause of thyroid dysfunction worldwide, results in alterations of TSH pulsatility. Induction in mouse models of sequential TSH pulses that mimic ultradian TSH profiles in periods of minutes were more efficient than sustained rises in basal TSH levels at increasing both thyroid follicle cAMP levels, as monitored with a genetically encoded cAMP sensor, and circulating thyroid hormone. Hence, this mouse TSH assay provides a powerful tool to decipher how ultradian TSH pulses encode thyroid outcomes and to uncover hidden parameters in the TSH-thyroid hormone set-point in health and disease.

Sex differences in pituitary corticotroph excitability.

Stress-related illness represents a major burden on health and society. Sex differences in stress-related disorders are well documented, with women having twice the lifetime rate of depression compared to men and most anxiety disorders. Anterior pituitary corticotrophs are central components of the hypothalamic-pituitary-adrenal (HPA) axis, receiving input from hypothalamic neuropeptides corticotrophin-releasing hormone (CRH) and arginine vasopressin (AVP), while regulating glucocorticoid output from the adrenal cortex. The dynamic control of electrical excitability by CRH/AVP and glucocorticoids is critical for corticotroph function; however, whether corticotrophs contribute to sexually differential responses of the HPA axis, which might underlie differences in stress-related disorders, is very poorly understood. Using perforated patch clamp electrophysiology in corticotrophs from mice expressing green fluorescent protein under the control of the Pomc promoter, we characterized basal and secretagogue-evoked excitability. Both male and female corticotrophs show predominantly single-spike action potentials under basal conditions; however, males predominantly display spikes with small-amplitude (<20 mV) afterhyperpolarizations (B-type), whereas females displayed a mixture of B-type spikes and spikes with a large-amplitude (>25 mV) afterhyperpolarization (A-type). In response to CRH, or CRH/AVP, male cells almost exclusively transition to a predominantly pseudo-plateau bursting, whereas only female B-type cells display bursting in response to CRH±AVP. Treatment of male or female corticotrophs with 1 nM estradiol (E2) for 24-72 h has no effect on the proportion of cells with A- or B-type spikes in either sex. However, E2 results in the cessation of CRH-induced bursting in both male and female corticotrophs, which can be partially reversed by adding a BK current using a dynamic clamp. RNA-seq analysis of purified corticotrophs reveals extensive differential gene expression at the transcriptional level, including more than 71 mRNAs encoding ion channel subunits. Interestingly, there is a two-fold lower level (p < 0.01) of BK channel pore-forming subunit (Kcnma1) expression in females compared to males, which may partially explain the decrease in CRH-induced bursting. This study identified sex differences at the level of the anterior pituitary corticotroph ion channel landscape and control of both spontaneous and CRH-evoked excitability. Determining the mechanisms of sex differences of corticotroph and HPA activity at the cellular level could be an important step for better understanding, diagnosing, and treating stress-related disorders.

An expression and function analysis of the CXCR4/SDF-1 signalling axis during pituitary gland development.

The chemokine SDF-1 (CXCL12) and its receptor CXCR4 control several processes during embryonic development such as the regulation of stem cell proliferation, differentiation, and migration. However, the role of this pathway in the formation of the pituitary gland is not understood. We sought to characterise the expression patterns of CXCR4, SDF-1 and CXCR7 at different stages of pituitary gland development. Our expression profiling revealed that SDF-1 is expressed in progenitor-rich regions of the pituitary anterior lobe, that CXCR4 and CXCR7 have opposite expression domains and that CXCR4 expression is conserved between mice and human embryos. We then assessed the importance of this signalling pathway in the development and function of the murine pituitary gland through conditional deletion of CXCR4 in embryonic pituitary progenitors. Successful and specific ablation of CXCR4 expression in embryonic pituitary progenitors did not lead to observable embryonic nor postnatal defects but allowed the identification of stromal CXCR4+ cells not derived from HESX1+ progenitors. Further analysis of constitutive SDF-1, CXCR7 and CXCR4 mutants of the pathway indicates that CXCR4 expression in HESX1+ cells and their descendants is not essential for normal pituitary development in mice.

Corticotroph isolation from Pomc-eGFP mice reveals sustained transcriptional dysregulation characterising a mouse model of glucocorticoid-induced suppression of the hypothalamus-pituitary-adrenal axis.

Glucocorticoids (GC) are prescribed for periods > 3 months to 1%-3% of the UK population; 10%-50% of these patients develop hypothalamus-pituitary-adrenal (HPA) axis suppression, which may last over 6 months and is associated with morbidity and mortality. Recovery of the pituitary and hypothalamus is necessary for recovery of adrenal function. We developed a mouse model of dexamethasone (DEX)-induced HPA axis dysfunction aiming to further explore recovery in the pituitary. Adult male wild-type C57BL6/J or Pomc-eGFP transgenic mice were randomly assigned to receive DEX (approximately 0.4 mg kg-1 bodyweight day-1 ) or vehicle via drinking water for 4 weeks following which treatment was withdrawn and tissues were harvested after another 0, 1, and 4 weeks. Corticotrophs were isolated from Pomc-eGFP pituitaries using fluorescence-activated cell sorting, and RNA extracted for RNA-sequencing. DEX treatment suppressed corticosterone production, which remained partially suppressed at least 1 week following DEX withdrawal. In the adrenal, Hsd3b2, Cyp11a1, and Mc2r mRNA levels were significantly reduced at time 0, with Mc2r and Cyp11a1 remaining reduced 1 week following DEX withdrawal. The corticotroph transcriptome was modified by DEX treatment, with some differences between groups persisting 4 weeks following withdrawal. No genes supressed by DEX exhibited ongoing attenuation 1 and 4 weeks following withdrawal, whereas only two genes were upregulated and remained so following withdrawal. A pattern of rebound at 1 and 4 weeks was observed in 14 genes that increased following suppression, and in six genes that were reduced by DEX and then increased. Chronic GC treatment may induce persistent changes in the pituitary that may influence future response to GC treatment or stress.

A New Perspective on Regulation of Pituitary Plasticity: The Network of SOX2-Positive Cells May Coordinate Responses to Challenge.

Plasticity of function is required for each of the anterior pituitary endocrine axes to support alterations in the demand for hormone with physiological status and in response to environmental challenge. This plasticity is mediated at the pituitary level by a change in functional cell mass resulting from a combination of alteration in the proportion of responding cells, the amount of hormone secreted from each cell, and the total number of cells within an endocrine cell population. The functional cell mass also depends on its organization into structural and functional networks. The mechanisms underlying alteration in gland output depend on the strength of the stimulus and are axis dependent but in all cases rely on sensing of output of the functional cell mass and its regulation. Here, we present evidence that the size of pituitary cell populations is constrained and suggest this is mediated by a form of quorum sensing. We propose that pituitary cell quorum sensing is mediated by interactions between the networks of endocrine cells and hormone-negative SOX2-positive (SOX2+ve) cells and speculate that the latter act as both a sentinel and actuator of cell number. Evidence for a role of the network of SOX2+ve cells in directly regulating secretion from multiple endocrine cell networks suggests that it also regulates other aspects of the endocrine cell functional mass. A decision-making role of SOX2+ve cells would allow precise coordination of pituitary axes, essential for their appropriate response to physiological status and challenge, as well as prioritization of axis modification.

Cell Networks in Endocrine/Neuroendocrine Gland Function.

Reproduction, growth, stress, and metabolism are determined by endocrine/neuroendocrine systems that regulate circulating hormone concentrations. All these systems generate rhythms and changes in hormone pulsatility observed in a variety of pathophysiological states. Thus, the output of endocrine/neuroendocrine systems must be regulated within a narrow window of effective hormone concentrations but must also maintain a capacity for plasticity to respond to changing physiological demands. Remarkably most endocrinologists still have a "textbook" view of endocrine gland organization which has emanated from 20th century histological studies on thin 2D tissue sections. However, 21st -century technological advances, including in-depth 3D imaging of specific cell types have vastly changed our knowledge. We now know that various levels of multicellular organization can be found across different glands, that organizational motifs can vary between species and can be modified to enhance or decrease hormonal release. This article focuses on how the organization of cells regulates hormone output using three endocrine/neuroendocrine glands that present different levels of organization and complexity: the adrenal medulla, with a single neuroendocrine cell type; the anterior pituitary, with multiple intermingled cell types; and the pancreas with multiple intermingled cell types organized into distinct functional units. We give an overview of recent methodologies that allow the study of the different components within endocrine systems, particularly their temporal and spatial relationships. We believe the emerging findings about network organization, and its impact on hormone secretion, are crucial to understanding how homeostatic regulation of endocrine axes is carried out within endocrine organs themselves. © 2022 American Physiological Society. Compr Physiol 12:3371-3415, 2022.

Chronic stress facilitates bursting electrical activity in pituitary corticotrophs.

Coordination of an appropriate stress response is dependent upon anterior pituitary corticotroph excitability in response to hypothalamic secretagogues and glucocorticoid negative feedback. A key determinant of corticotroph excitability is large conductance calcium- and voltage-activated (BK) potassium channels that are critical for promoting corticotrophin-releasing hormone (CRH)-induced bursting that enhances adrenocorticotrophic hormone secretion. Previous studies revealed hypothalamic-pituitary-adrenal axis hyperexcitability following chronic stress (CS) is partly a function of increased corticotroph output. Thus, we hypothesise that chronic stress promotes corticotroph excitability through a BK-dependent mechanism. Corticotrophs from CS mice displayed significant increase in spontaneous bursting, which was suppressed by the BK blocker paxilline. Mathematical modelling reveals that the time constant of BK channel activation, plus properties and proportion of BK channels functionally coupled to L-type Ca2+ channels determines bursting activity. Surprisingly, CS corticotrophs (but not unstressed) display CRH-induced bursting even when the majority of BK channels are inhibited by paxilline, which modelling suggests is a consequence of the stochastic behaviour of a small number of BK channels coupled to L-type Ca2+ channels. Our data reveal that changes in the stochastic behaviour of a small number of BK channels can finely tune corticotroph excitability through stress-induced changes in BK channel properties. Importantly, regulation of BK channel function is highly context dependent allowing dynamic control of corticotroph excitability over a large range of time domains and physiological challenges in health and disease. This is likely to occur in other BK-expressing endocrine cells, with important implications for the physiological processes they regulate and the potential for therapy. KEY POINTS: Chronic stress (CS) is predicted to modify the electrical excitability of anterior pituitary corticotrophs. Electrophysiological recordings from isolated corticotrophs from CS male mice display spontaneous electrical bursting behaviour compared to the tonic spiking behaviour of unstressed corticotrophs. The increased spontaneous bursting from CS corticotrophs is BK-dependent and mathematical modelling reveals that the time constant of activation, properties and proportion of BK channels functionally coupled to L-type calcium channels determines the promotion of bursting activity. CS (but not unstressed) corticotrophs display corticotrophin-releasing hormone-induced bursting even when the majority of BK channels are pharmacologically inhibited, which can be explained by the stochastic behaviour of a small number of BK channels with distinct properties. Corticotroph excitability can be finely tuned by the stochastic behaviour of a small number of BK channels dependent on their properties and functional co-localisation with L-type calcium channels to control corticotroph excitability over diverse time domains and physiological challenges.

Renewing an old interest: Pituitary folliculostellate cells.

Anterior pituitary folliculostellate (FS) cells, first described almost 50 years ago, have a wide range of functions with respect to supporting and coordinating endocrine cell function, in particular through paracrine and gap junction-mediated signalling. Our previous studies identified the morphological organisation of FS cells, which mediates coordinated calcium activity throughout the homotypic FS network and allows signalling across the whole pituitary gland. It is also clear that FS cells can modify endocrine output and feedback on pituitary axes over a range of timescales. Recently, several studies have defined FS cells as a source of anterior pituitary endocrine cell renewal, which has resulted in a renaming of FS cells as "Sox2+ve stem cells". Here, we highlight the broader potential of the FS cell population in fine-tuning and coordinating pituitary axes function. In addition, we identify a need for: the definition of the possible subtypes of FS cell and their relationship with the stem cell population; the potential role of FS cells in pulsatile hormone secretion and coordination of heterotypic cell networks; and the roles that FS cells may play in both early-life programming of pituitary axes and in memory, or anticipation, of demand. Further studies of FS cells may demonstrate the fundamental importance of this cell type and its potential as a therapeutic target to correct pituitary gland dysfunction, one of which is stem cell therapy. Clearly, a thorough understanding of all of these interactions and relationships of FS and endocrine cells is required whatever therapeutic use is suggested by their various roles.

Hyperprolactinemia in a male pituitary androgen receptor knockout mouse is associated with female-like lactotroph development.

BACKGROUND: Circulating prolactin concentration in rodents and humans is sexually dimorphic. Oestrogens are a well-characterised stimulator of prolactin release. Circulating prolactin fluctuates throughout the menstrual/oestrous cycle of females in response to oestrogen levels, but remains continually low in males. We have previously identified androgens as an inhibitor of prolactin release through characterisation of males of a mouse line with a conditional pituitary androgen receptor knockout (PARKO) which have an increase in circulating prolactin, but unchanged lactotroph number. OBJECTIVES: In the present study, we aimed to specify the cell type that androgens act on to repress prolactin release. MATERIALS AND METHODS: PARKO, lactotroph-specific, Pit1 lineage-specific and neural-specific conditional androgen receptor knockout male mice were investigated using prolactin ELISA, pituitary electron microscopy, immunohistochemistry and qRT-PCR. RESULTS: Lactotroph-specific, Pit1 lineage-specific and neural-specific conditional AR knockouts did not duplicate the high circulating prolactin seen in the PARKO line. Using electron microscopy to examine ultrastructure, we showed that pituitary androgen receptor knockout male mice develop lactotrophs that resemble those seen in female mice. Castrated PARKO males have significantly reduced circulating prolactin compared to intact males. When expression of selected oestrogen-regulated anterior pituitary genes was examined, there were no differences in expression level between controls and knockouts. DISCUSSION: The cell type that androgens act on to repress prolactin release is not the lactotroph, cells in the Pit1-lineage, or the dopaminergic neurons in the hypothalamus. PARKO males develop a female-specific lactotroph ultrastructure that this is likely to contribute to the increase in circulating prolactin. Castrated PARKO males have significantly reduced circulating prolactin compared to intact males, which suggests that removal of both circulating oestrogens and androgens reduces the stimulation of pituitary prolactin release. CONCLUSION: Further investigation is needed into prolactin regulation by changes in androgen-oestrogen balance, which is involved sexual dimorphism of development and diseases including hyperprolactinemia.

Characterisation of a mural cell network in the murine pituitary gland.

The anterior and intermediate lobes of the pituitary are composed of endocrine cells, as well as vasculature and supporting cells, such as folliculostellate cells. Folliculostellate cells form a network with several postulated roles in the pituitary, including production of paracrine signalling molecules and cytokines, coordination of endocrine cell hormone release, phagocytosis, and structural support. Folliculostellate cells in rats are characterised by expression of S100B protein, and in humans by glial fibrillary acid protein. However, there is evidence for another network of supporting cells in the anterior pituitary that has properties of mural cells, such as vascular smooth muscle cells and pericytes. The present study aims to characterise the distribution of cells that express the mural cell marker platelet derived growth factor receptor beta (PDGFRß) in the mouse pituitary and establish whether these cells are folliculostellate. By immunohistochemical localisation, we determine that approximately 80% of PDGFRß+ cells in the mouse pituitary have a non-perivascular location and 20% are pericytes. Investigation of gene expression in a magnetic cell sorted population of PDGFRß+ cells shows that, despite a mostly non-perivascular location, this population is enriched for mural cell markers but not enriched for rat or human folliculostellate cell markers. This is confirmed by immunohistochemistry. The present study concludes that a mural cell network is present throughout the anterior pituitary of the mouse and that this population does not express well-characterised human or rat folliculostellate cell markers.

IGSF1 Deficiency Results in Human and Murine Somatotrope Neurosecretory Hyperfunction.

CONTEXT: The X-linked immunoglobulin superfamily, member 1 (IGSF1), gene is highly expressed in the hypothalamus and in pituitary cells of the POU1F1 lineage. Human loss-of-function mutations in IGSF1 cause central hypothyroidism, hypoprolactinemia, and macroorchidism. Additionally, most affected adults exhibit higher than average IGF-1 levels and anecdotal reports describe acromegaloid features in older subjects. However, somatotrope function has not yet been formally evaluated in this condition. OBJECTIVE: We aimed to evaluate the role of IGSF1 in human and murine somatotrope function. PATIENTS, DESIGN, AND SETTING: We evaluated 21 adult males harboring hemizygous IGSF1 loss-of-function mutations for features of GH excess, in an academic clinical setting. MAIN OUTCOME MEASURES: We compared biochemical and tissue markers of GH excess in patients and controls, including 24-hour GH profile studies in 7 patients. Parallel studies were undertaken in male Igsf1-deficient mice and wild-type littermates. RESULTS: IGSF1-deficient adult male patients demonstrated acromegaloid facial features with increased head circumference as well as increased finger soft-tissue thickness. Median serum IGF-1 concentrations were elevated, and 24-hour GH profile studies confirmed 2- to 3-fold increased median basal, pulsatile, and total GH secretion. Male Igsf1-deficient mice also demonstrated features of GH excess with increased lean mass, organ size, and skeletal dimensions and elevated mean circulating IGF-1 and pituitary GH levels. CONCLUSIONS: We demonstrate somatotrope neurosecretory hyperfunction in IGSF1-deficient humans and mice. These observations define a hitherto uncharacterized role for IGSF1 in somatotropes and indicate that patients with IGSF1 mutations should be evaluated for long-term consequences of increased GH exposure.

Imaging and Manipulating Pituitary Function in the Awake Mouse.

Extensive efforts have been made to explore how the activities of multiple brain cells combine to alter physiology through imaging and cell-specific manipulation in different animal models. However, the temporal regulation of peripheral organs by the neuroendocrine factors released by the brain is poorly understood. We have established a suite of adaptable methodologies to interrogate in vivo the relationship of hypothalamic regulation with the secretory output of the pituitary gland, which has complex functional networks of multiple cell types intermingled with the vasculature. These allow imaging and optogenetic manipulation of cell activities in the pituitary gland in awake mouse models, in which both neuronal regulatory activity and hormonal output are preserved. These methodologies are now readily applicable for longitudinal studies of short-lived events (e.g., calcium signals controlling hormone exocytosis) and slowly evolving processes such as tissue remodeling in health and disease over a period of days to weeks.

The Processes of Anterior Pituitary Hormone Pulse Generation.

More than 60 years ago, Geoffrey Harris described his "neurohumoral theory," in which the regulation of pituitary hormone secretion was a "simple" hierarchal relationship, with the hypothalamus as the controller. In models based on this theory, the electrical activity of hypothalamic neurons determines the release of hypophysiotropic hormones into the portal circulation, and the pituitary simply responds with secretion of a pulse of hormone into the bloodstream. The development of methodologies allowing the monitoring of the activities of members of the hypothalamic-vascular-pituitary unit is increasingly allowing dissection of the mechanisms generating hypothalamic and pituitary pulses. These have revealed that whereas hypothalamic input is required, its role as a driver of pulsatile pituitary hormone secretion varies between pituitary axes. The organization of pituitary cells has a key role in the modification of their response to hypophysiotropic factors that can lead to a memory of previous demand and enhanced function. Feedback can lead to oscillatory hormone output that is independent of pulses of hypophysiotropic factors and instead, results from the temporal relationship between pituitary output and target organ response. Thus, the mechanisms underlying the generation of pulses cannot be generalized, and the circularity of feedforward and feedback interactions must be considered to understand both normal physiological function and pathology. We describe some examples of the clinical implications of recognizing the importance of the pituitary and target organs in pulse generation and suggest avenues for future research in both the short and long term.

Autocrine actions of prolactin contribute to the regulation of lactotroph function in vivo.

Prolactin (PRL), whose principal role is regulation of lactation, is mainly synthesized and secreted by lactotroph anterior pituitary cells. Its signaling is exerted via a transmembrane PRL receptor (PRLR) expressed in a wide variety of tissues, including the anterior pituitary. Dopamine, which is secreted by tuberoinfundibular hypothalamic neurons, is the major inhibitory regulator of prolactin secretion. Although PRL is well established to stimulate hypothalamic dopamine secretion, thereby exerting a negative feedback regulation on its own release, autocrine or paracrine actions of PRL on lactotroph cells have also been suggested. Within the pituitary, PRL may inhibit both lactotroph proliferation and secretion, but in vivo evaluation of these putative functions is limited. To determine whether the autocrine actions of prolactin have a significant role in the physiologic function of lactotrophs in vivo, we examined the consequences of conditional deletion of Prlr in lactotroph cells using a novel mouse line with loxP sites flanking the Prlr gene ( Prlrlox/lox) and Cre-recombinase (Cre) expressed under the control of the pituitary-specific Prl promoter. Prlrlox/lox/Prl-Cre mice have normal PRL levels and did not develop any pituitary lactotroph adenoma, even at 20 mo of age. Nevertheless, Prlrlox/lox/Prl-Cre mice displayed an increased dopaminergic inhibitory tone compared with control Prlrlox/lox mice. These results elegantly confirm an autocrine/paracrine feedback of PRL on lactotroph cells in vivo, which can be fully compensated by an intact hypothalamic feedback system.-Bernard, V., Lamothe, S., Beau, I., Guillou, A., Martin, A., Le Tissier, P., Grattan, D., Young, J., Binart, N. Autocrine actions of prolactin contribute to the regulation of lactotroph function in vivo.

NOTCH activity differentially affects alternative cell fate acquisition and maintenance.

The pituitary is an essential endocrine gland regulating multiple processes. Regeneration of endocrine cells is of therapeutic interest and recent studies are promising, but mechanisms of endocrine cell fate acquisition need to be better characterised. The NOTCH pathway is important during pituitary development. Here, we further characterise its role in the murine pituitary, revealing differential sensitivity within and between lineages. In progenitors, NOTCH activation blocks cell fate acquisition, with time-dependant modulation. In differentiating cells, response to activation is blunted in the POU1F1 lineage, with apparently normal cell fate specification, while POMC cells remain sensitive. Absence of apparent defects in Pou1f1-Cre; Rbpjfl/fl mice further suggests no direct role for NOTCH signalling in POU1F1 cell fate acquisition. In contrast, in the POMC lineage, NICD expression induces a regression towards a progenitor-like state, suggesting that the NOTCH pathway specifically blocks POMC cell differentiation. These results have implications for pituitary development, plasticity and regeneration. Activation of NOTCH signalling in different cell lineages of the embryonic murine pituitary uncovers an unexpected differential sensitivity, and this consequently reveals new aspects of endocrine lineages development and plasticity.

Heterogeneity of Calcium Responses to Secretagogues in Corticotrophs From Male Rats.

Heterogeneity in homotypic cellular responses is an important feature of many biological systems, and it has been shown to be prominent in most anterior pituitary hormonal cell types. In this study, we analyze heterogeneity in the responses to hypothalamic secretagogues in the corticotroph cell population of adult male rats. Using the genetically encoded calcium indicator GCaMP6s, we determined the intracellular calcium responses of these cells to corticotropin-releasing hormone and arginine-vasopressin. Our experiments revealed marked population heterogeneity in the response to these peptides, in terms of amplitude and dynamics of the responses, as well as the sensitivity to different concentrations and duration of stimuli. However, repeated stimuli to the same cell produced remarkably consistent responses, indicating that these are deterministic on a cell-by-cell level. We also describe similar heterogeneity in the sensitivity of cells to inhibition by corticosterone. In summary, our results highlight a large degree of heterogeneity in the cellular mechanisms that govern corticotroph responses to their physiological stimuli; this could provide a mechanism to extend the dynamic range of the responses at the population level to allow adaptation to different physiological challenges.

Complementary actions of dopamine D2 receptor agonist and anti-vegf therapy on tumoral vessel normalization in a transgenic mouse model.

Angiogenesis contributes in multiple ways to disease progression in tumors and reduces treatment efficiency. Molecular therapies targeting Vegf signaling combined with chemotherapy or other drugs exhibit promising results to improve efficacy of treatment. Dopamine has been recently proposed to be a novel safe anti-angiogenic drug that stabilizes abnormal blood vessels and increases therapeutic efficacy. Here, we aimed to identify a treatment to normalize tumoral vessels and restore normal blood perfusion in tumor tissue with a Vegf receptor inhibitor and/or a ligand of dopamine G protein-coupled receptor D2 (D2R). Dopamine, via its action on D2R, is an endogenous effector of the pituitary gland, and we took advantage of this system to address this question. We have used a previously described Hmga2/T mouse model developing haemorrhagic prolactin-secreting adenomas. In mutant mice, blood vessels are profoundly altered in tumors, and an aberrant arterial vascularization develops leading to the loss of dopamine supply. D2R agonist treatment blocks tumor growth, induces regression of the aberrant blood supply and normalizes blood vessels. A chronic treatment is able to restore the altered balance between pro- and anti-angiogenic factors. Remarkably, an acute treatment induces an upregulation of the stabilizing factor Angiopoietin 1. An anti-Vegf therapy is also effective to restrain tumor growth and improves vascular remodeling. Importantly, only the combination treatment suppresses intratumoral hemorrhage and restores blood vessel perfusion, suggesting that it might represent an attractive therapy targeting tumor vasculature. Similar strategies targeting other ligands of GPCRs involved in angiogenesis may identify novel therapeutic opportunities for cancer.