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1 Departments of Internal Medicine, Erasmus Medical Center, Rotterdam, The Netherlands, 2 Molecular and Clinical Endocrinology and Oncology, ‘Federico II’ University, Via Sergio Pansini, 5, 80131 Naples, Italy and 3 Endocrinological and Metabolic Sciences, Centre of Excellence for Biomedical Research, University of Genova, Genova, Italy
(Correspondence should be addressed to R Pivonello; Email: rpivone{at}tin.it)
This paper was presented at a symposium held at the Erasmus Medical Center, Rotterdam, The Netherlands, 2005. The symposium was jointly organized by LJ Hofland, Erasmus Medical Center, and A Colao, Federico II University of Naples, Italy. Ipsen partially supported the publication of these proceedings.
Abstract
The dopaminergic system has a pivotal role in the central nervous system but also plays important roles in the periphery, mainly in the endocrine system. Dopamine exerts its functions via five different receptors, named D1–D5, belonging to the category of G protein coupled membrane receptors. Dopamine receptors are heterogeneously expressed in different cells, tissues and organs, where they stimulate or inhibit different functions, including neurotransmission and hormone synthesis and secretion. In particular, the dopamineric system has a pivotal role in the physiological regulation of the hypothalamus–pituitary–adrenal axis. Recent data have demonstrated the expression and function of dopamine receptors not only in endocrine organs but also in endocrine tumors, mainly those belonging to the hypothalamus–pituitary–adrenal axis, and also in the so-called ‘neuroendocrine’ tumors. These data confirm the important role of the dopaminergic system in this endocrine axis, as well as in the neuroendocrine system. This review summarizes the main structural and functional characteristics of dopamine receptors, emphasizing the most recent novelties, and focused on the physiological and pathological regulation of the hypothalamus–pituitary–adrenal axis by the dopaminergic system. In addition, the recent findings on the relationship between dopamine receptors and neuroendocrine tumors are summarized.
Introduction
Dopamine is the predominant catecholamine neuro-transmitter in the human central nervous system, where it controls a variety of functions including cognition, emotion, locomotor activity, hunger and satiety, and endocrine system regulation. Dopamine also plays multiple roles in the periphery as a modulator of cardiovascular and renal function, gastrointestinal motility, and the endocrine system (1). Dopamine exerts its functions via its binding to dopamine receptors (1). In recent years, the availability of new dopaminergic compounds and new investigations into the dopaminergic system have produced novel information on the physiological role of dopamine and dopamine receptors, mainly in the endocrine system, and their implication in the management of endocrine tumors, especially regarding the hypothalamus–pituitary–adrenal axis, and the neuroendocrine system.
Structural characteristics of dopamine receptors
Dopamine receptors belong to the family of seven transmembrane domain G protein coupled receptors and include five different receptor subtypes, named D1–D5. The members of the dopamine receptor family are encoded by genes localized at different chromosomal loci, displaying a considerable homology in their protein structure and function. The analysis of dopamine receptor structure and function suggests the existence of two different groups of receptors: D1-like, including D1 and D5 receptors, generally associated to a stimulatory function, and D2-like, including D2–D4 receptors, generally associated to an inhibitory function (1). The D1 and D5 receptors are encoded by genes lacking introns and share an 80% homology in their transmembrane domains. The D2 receptor shares a 75% homology with the D3 and a 53% homology with the D4 transmembrane domains, and all three receptor subtypes are encoded by genes which are interrupted by introns (1). The NH2-terminal stretch has a similar number of amino acid residues in all the receptor subtypes and carries a variable number of consensus N-glycosylation sites, which can also be present in the extracellular loops: the D1 and D5 receptors possess two such sites, the D2 has four, the D3 has three, and the D4 possesses only one potential N-glycosylation site. The carboxy terminus, which is about seven times longer for the D1-like than for the D2-like receptors, is rich in serine and threonine residues and contains a cysteine residue, probably involved in the anchorage of the cytoplasmic tail to the membrane. Potential phosphorylation sites are localized not only on the COOH-terminal, but also in intracellular loops, mainly the third loop. Dopamine receptors possess two cysteine residues in the second and third extracellular loops, which probably form a disulfide bridge used for the stabilization of the receptor structure. The D1-like receptors have a short third intracellular loop, typical for receptors interacting with G-stimulatory (Gs) proteins, to stimulate cyclic AMP production, whereas the D2-like receptors have a long third intracellular loop, typical for receptors interacting with G-inhibitory (Gi) proteins, to inhibit cyclic AMP production. The third intracellular loop is the region responsible for the G protein coupling and signal transmission. The hydrophobic transmembrane domains are responsible, through some specific amino acid residues, for the binding of dopamine as well as its agonists and antagonists (1).
The D2 receptor exists in two main variants, the long isoform named D2L and the short isoform named D2S, generated by an alternative splicing of an 87 bp exon. These two D2 receptor isoforms differ for the presence or absence of a stretch of 29 amino acid residues in the third cytoplasmic loop in their protein structure (2). The two D2 receptor isoforms have similar pharmacological but different functional characteristics, since the third cytoplasmic loop plays a crucial role in the signal transmission, particularly in determining the G protein coupling specificity, and consequently in activating different signaling pathways (3). It is noteworthy that the two D2 receptor isoforms have also a distinct regulation of the receptor internalization (4). Splicing variants of the D3 receptor encoding nonfunctional proteins have also been identified. The analysis of the D4 receptor reveals the existence of polymorphic variations within the coding sequence, being a 48 bp sequence existent as a direct repeat sequence (D4.1), fourfold (D4.4), sevenfold (D4.7), or 11-fold (D4.11) repeat sequence. Therefore, the D4 receptor isoforms differ for the length of the third cytoplasmic loop and have one, four, seven, or 11 times the same insert of a stretch of 16 amino acid residues in their protein structure. The D5 receptor has two related pseudogenes, which share a 95% homology with the gene and encode for truncated nonfunctional forms of the receptor (1). The molecular characteristics of human dopamine receptor family are summarized in Table 1
. A schematic representation of the human dopamine receptor is shown in Fig. 1, whereas a schematic representation of the two different D2 receptor isoforms is shown in Fig. 2.
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The pharmacological profile of dopamine receptors displays a difference between D1- and D2-like receptors, mainly a variable binding affinity of certain dopamine agonists and antagonists (1). Dopamine binds all five receptors although among the D1-like receptors it binds the D1 with lower affinity than the D5 receptor, and among the D2-like receptors it binds the D2 with lower affinity than D3 and D4 receptors. Beyond dopamine, different agonists or antagonists preferentially or exclusively bind D1- or D2-like receptors. For instance, among the dopamine agonists, bromocriptine preferentially binds D2-like receptors but it is able to bind D1-like receptors as well, whereas among the dopamine antagonists, sulpiride binds the D2-like receptors exclusively. It is noteworthy that D1 and D5 receptors cannot be clearly differentiated pharmacologically, since no dopamine agonist or antagonist exclusively or preferentially binds D1 or D5 receptors, whereas they generally display similar binding affinities for both receptor subtypes. Conversely, the availability of dopamine agonists and/or antagonists which exclusively or preferentially bind D2, D3, or D4 receptors make it possible to clearly differentiate these pharmacologically. No compound is able to clearly discriminate between D2L and D2S although a marginal difference in the affinities of the two D2 receptor isoforms has been described for the dopamine antagonist sulpiride (1). Among the dopamine agonists, cabergoline is able, like bromocriptine, to bind both D1 and D2 receptors, with higher affinities for both receptors and higher selectivity for D2 receptors (5). Moreover, cabergoline has recently been demonstrated to possess a higher affinity than bromocriptine not only for both D2 receptor isoforms, but also for D3 and D4 receptors (6, 7). No study has ever compared the binding affinity of bromocriptine and cabergoline for the D5 receptor. The different pharmacological characteristics of bromocriptine and cabergoline definitely demonstrate that cabergoline is more potent when compared with bromocriptine in the binding and activation of the D2-like receptors. The pharmacological profile of the D2-like dopamine receptors in relation to dopamine and the dopamine agonists bromocriptine and cabergoline is shown in Table 2.
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Dopamine receptors mediate the effects of dopamine and dopaminergic compounds via a number of different mechanisms of signal transduction (1). Among these, the most important is the modulation of adenylyl cyclase activity resulting in either stimulation or inhibition of cyclic AMP accumulation. This mechanism is mediated via the activation of different G proteins, mainly Gs
for the stimulation and Gi for the inhibition of adenylyl cyclase. D1-like receptors generally stimulate whereas D2-like receptors inhibit adenylyl cyclase activity and cyclic AMP accumulation (1). Most commonly, the result of the stimulation or inhibition of cyclic AMP accumulation is the modulation (activation or inactivation) of protein kinase A, responsible, through phosphorylation or dephosphorylation, for the regulation of the synthesis of different cytoplasmic and nuclear proteins, function of membrane channels, and sensitization or desensitization of different G protein coupled receptors (2). Dopamine receptors are also able to activate different mechanisms of signal transduction, including the modulation of the activity of phospholipase C or the release of arachidonic acid, as well as the activity of the calcium and potassium channels. Moreover, dopamine receptors also seem to modulate the activity of Na/H exchangers and the Na–K ATPase (1). A novel mechanism of signal transmission has been recently demonstrated for the D2S isoform of the D2 receptors. The activation of this receptor is able to induce the stimulation of the phospholipase D, probably through the coupling with the Rho family of G proteins, and the involvement of a specific protein kinase C isoform (8). Phospholipase D is an enzyme able to catalyze the hydrolysis of phosphatidylcholine to phosphatidic acid and choline. The phosphatidic acid, together with the diacylglycerol produced from its own dephosphorylation is an active signaling molecule implicated in a variety of cell-signaling pathways, mainly involved in the regulation of cell metabolism, as well as cell growth and differentiation. The activation of phospholipase D by D2S seems to be associated with an anti-proliferative effect (9). This mechanism of signal transmission has recently been found to be associated with different dopamine receptors. It is noteworthy that dopamine receptors are able to control cell growth and differentiation with different mechanisms (1). Recently, both D2S and D2L receptor isoforms have been demonstrated to activate, through the involvement of Gbg subunits and protein kinase C, the mitogen-activated protein kinase (MAPK) and extracellular signal-regulated kinase (ERK) pathway, generally involved in the regulation of cell growth, differentiation and apoptosis (10). In particular, in pituitary tumor cell lines, dopamine agonists have been found to exert a clear anti-proliferative effect, inducing cell death and apoptosis through the D2 receptor-mediated activation of MAPK and/or ERK pathways (11, 12). The main intracellular pathways connected with the activation of dopamine receptors are summarized in Fig. 3.
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Dopamine receptors, being G protein coupled receptors, are characterized by the phenomenon of oligomerization. They may form homo-oligomers, grouping with other monomers of the same receptor, or hetero-oligomers, grouping with monomers of different dopamine receptor subtypes or different receptors (15). These hetero-oligomers have functional characteristics, which are distinct from those of the single monomers or homo-oligomers, with consequent implications for receptor pharmacology, signaling and trafficking (15). The complete meaning of the homo-and hetero-oligomerization of dopamine receptors require further studies to be completely elucidated.
Distribution and role of dopamine receptors
Dopamine receptors are mainly and widely distributed in the central nervous system (1). In particular, the dopaminergic neurons in the substantia nigra, tegmental area and hypothalamus give origin to three main pathways: the nigrostriatal, the mesolimbocortical, and the tuberoinfundibular pathways. Dopamine receptors are, therefore, mainly localized in the striatum, the limbic system, the brain cortex and the infundibulum. However, the presence of dopamine receptors has been demonstrated in most areas of the central nervous system, where they mediate the effect of dopamine on cognition, emotion, regulation of hunger and satiety, locomotor activity and on the endocrine system (1). Dopamine receptors are also expressed in the pituitary gland, where they mediate the effect of dopamine on the regulation of hormone synthesis and secretion (1). Finally, dopamine receptors are widely distributed in the periphery, mainly at the level of the cardiovascular system, kidney and adrenal gland, beyond the peripheral nervous system. In the cardiovascular system, dopamine is known to induce vasodilation and increase cardiac contractility. In the kidney, dopamine induces an increase in the renal filtration rate and a decrease in salt reabsorption, as well as a stimulation of renin secretion (activating the renin–angiotensin–aldosterone system) and inhibition of vasopressin action. The presence of dopamine receptors in the adrenal gland suggests a role for these receptors in the regulation of adrenal hormone synthesis or secretion (1).
Hypothalamus–pituitary–adrenal axis and dopamine receptors
Dopamine receptors have an important role in the regulation of the hypothalamus–pituitary–adrenal axis. Indeed, dopaminergic pathways are known to control the hypothalamic function. Moreover, increasing amounts of evidence demonstrate that the dopaminergic system also has a pivotal role in the control of the entire pituitary gland, as well as in the adrenal gland.
Dopamine receptors have been clearly demonstrated in the anterior lobe of the pituitary gland. In rats, the D2 receptor is expressed mainly in lactotropes (16), but it is also found in non-lactotrope cells, mainly somatotropes (17) and thyrotropes (18), as well as in gonadotrope cells (19). Moreover, D2 receptor expression in corticotrope cells has been suggested by the demonstration of its expression in AtT20 cells, a mouse pituitary corticotrope cell line (20, 21). Similarly, in humans the D2 receptor is also expressed mainly in lactotrope cells (22), although it was found in more than 75% of cells of the anterior pituitary gland, indicating a broader spectrum of expression, not only confined in lactotrope, but also extended in non-lactotrope cell populations (23). In addition, dopamine receptors, and particularly D2 receptors, have been clearly demonstrated in the intermediate zone of the pituitary gland. Indeed, several studies on experimental animals have shown the presence of D2 receptors in melanotrope cells (24, 25). These data suggest the possible D2 receptors expression in melanotrope cells in humans. With respect to D2 receptor isoforms, several studies on experimental animals have demonstrated that both D2L and D2S receptor isoforms are expressed in lactotrope and melanotrope cells, although D2L is predominantly expressed when compared with the D2S isoform (26). Finally, the D2 receptor is not the only dopamine receptor expressed in the pituitary gland. Indeed, the D4 receptor, in particular its D4.4 variant, is also expressed, although its role in the physiology of the pituitary gland is not known (27). The role of dopamine receptors, in particular the D2 receptor, in the pituitary gland is to mediate the regulatory effects of hypothalamic dopamine on the different pituitary cell populations. In rats and humans, the major role of the pituitary D2 receptor is inhibitory control of prolactin synthesis and secretion, as well as the growth of lactotrope cells (22). Moreover, it has been demonstrated in rats, and hypothesized for humans, that the D2 receptor is also involved in the inhibitory control of the synthesis and/or secretion of melanocyte-stimulating hormone and growth of the melanotrope cells (28, 29). The finding that the expression of the Pit 1 transcription factor, which is involved in pituitary hormone gene expression, is inhibited by activation of D2 receptors in transfected cell lines supports the existence of a dopaminergic control on pituitary hormone gene expression (30). In addition, a series of in vitro and/or in vivo studies on experimental animals and/or in humans suggests a possible modulatory role of dopamine on growth hormone, thyroid-stimulating hormone, and follicle-stimulating hormone/luteinizing hormone release from the somatotrope, thyrotrope and gonadotrope cells respectively (31). Finally, a recent study has clearly demonstrated that dopamine receptors, mainly D2 receptors, are expressed in the majority of corticotrope pituitary tumors, where dopaminergic agents are able to inhibit adrenocorticotropic hormone (ACTH) secretion (32). This evidence suggests a physiological role of dopamine in the regulation of ACTH release.
Dopamine receptors are also expressed in the adrenal cortex. In experimental animals, as well as in humans, receptor ligand binding studies have demonstrated the presence of specific and saturable binding sites for different radiolabeled dopaminergic compounds such as spiperone, which bind both D1- and D2-like receptors, and sulpiride, which selectively binds D2-like receptors (33). These studies demonstrated that both D1-like and D2-like receptors are expressed in the adrenal cortex. In addition, the expression of D1-like and D2-like receptors in the adrenal medulla has been clearly demonstrated (34). A recent study has evaluated the expression of the D2-like receptor subtypes in human normal adrenal glands, demonstrating that D2 and D4 receptors are expressed in all three zones of the adrenal cortex and in the adrenal medulla. In the cortex they are localized mainly in the zona glomerulosa, responsible for aldosterone synthesis, and zona reticularis, responsible for the synthesis of sex hormones, and to a lesser extent in the zona fasciculata of the adrenal cortex, where cortisol is synthesized (35). The expression of dopamine receptors in the adrenal cortex has been initially hypothesized after showing a role of dopamine in the control of aldosterone secretion by in vivo studies in both experimental animals and humans (36). The administration of the D2 antagonist metoclopramide to both rats and humans increased the plasma aldosterone levels without influencing any stimulator of aldosterone -release. This effect was blocked by the i.v. injection of dopamine (37–39). However, the administration of dopamine or the dopamine agonist bromocriptine did not modify plasma aldosterone levels (40). These observations suggest that aldosterone production was under maximum tonic dopaminergic inhibition. Subsequent studies demonstrated that the sodium balance status is crucial for the effects of dopamine or dopamine agonists on aldosterone secretion. Indeed, dopamine and D2 agonists were shown to inhibit angiotensin-stimulated and upright posture-induced increased aldosterone secretion in sodium-depleted but not in sodium-repleted normal subjects (41, 42). Moreover, in vitro studies with isolated adrenal glomerulosa cells demonstrated that the activation of D2 receptors may result in a remarkable inhibition of angiotensin II-induced aldosterone secretion, whereas it does not influence basal and ACTH-induced aldosterone secretion (43). These studies first suggested the expression of D2 or D2-like receptors in the cells of the zona glomerulosa of the adrenal cortex and a selective functional interaction between dopamine and angiotensin II in the regulation of aldosterone secretion. While no definitive data are available on the role of dopamine and dopamine receptors in the adrenal cortex, their role in controlling catecholamine secretion in the adrenal medulla is well documented (44). A recent study has clearly demonstrated that different dopamine receptors, including D1-like and D2-like, are expressed in normal human adrenal glands, and that D2 receptors are expressed in all three zones of the adrenal glands, although predominantly in the zona glomerulosa and zona reticularis, and in the adrenal medulla (45). In this study, dopaminergic drugs, and mainly cabergoline, showed a modulatory effect on aldosterone, but also on cortisol and androstenedione secretion, displaying a stimulatory effect at lower concentration and an inhibitory effect at higher concentrations of the compounds (45). These evidences demonstrated that dopamine may have an important physiological role in the regulation of adrenal function.
Finally, it is noteworthy that dopamine receptors have been clearly demonstrated to be expressed in corticotrope pituitary tumors (32) and in tumors deriving from the adrenal medulla, the pheochromocytomas (34, 35, 45), and have also been recently found in tumors deriving from the adrenal cortex, mainly aldosterone-secreting and cortisol-secreting tumors (45). In corticotrope pituitary tumors, D2 receptors have an important role in mediating the inhibitory effect of dopaminergic drugs on ACTH and, consequently, cortisol secretion, so that treatment with cabergoline is able to induce remission in one-third of cases with Cushing’s disease (32). Conversely, in the adrenal tumors, the role of dopamine receptors is still unknown, but the documented expression of the D2 receptors permit a hypothesis for a possible role of dopaminergic drugs in the treatment of these tumors as well, especially in those associated to Conn’s or Cushing’s syndrome (45). However, no study has ever investigated the effectiveness of dopaminergic drugs in the treatment of these adrenal diseases.
Neuroendocrine tumors and dopamine receptors
Dopamine receptors have been also demonstrated in the so-called neuroendocrine tumors. Indeed, beyond pituitary tumors, mainly including GH-secreting and PRL-secreting, ACTH-secreting and clinically nonfunctioning tumors as well as pheochromocytomas, dopamine receptors – mostly D2 receptors – have recently been demonstrated to be expressed in a major class of well-differentiated endocrine carcinomas, known as ‘carcinoids’, especially those associated to ectopic ACTH syndrome, or ectopic Cushing’s syndrome (46). It is possible that these receptors mediate the inhibitory effect of dopaminergic drugs in these tumors, since cabergoline has been found to induce normalization of ACTH and consequently cortisol secretion, in one out of three cases of ectopic Cushing’s syndrome due to ACTH-secreting lung carcinoids (46). However, the role of dopaminergic drugs in the treatment of corticotrope ectopic tumors needs to be confirmed by further investigations on a larger number of cases of ectopic Cushing’s syndrome.
Carcinoid tumors are not the only neuroendocrine tumors expressing dopamine receptors. The previous finding of dopamine receptors in gastro-entero-pancreatic cell lines (47) and the recent finding of D2 receptors in gastro-entero-pancreatic tumors (48) suggests a role for these receptors, and possibly a role of dopaminergic drugs in this category of neuroendocrine tumors. Finally, dopamine receptors, and mainly D2 receptors, have been also found to be expressed in a peculiar type of neuroendocrine tumors, the melanomas. However, the role of dopamine receptors in these tumors still needs to be investigated.
Conclusions
Dopamine receptors are a group of five different G protein coupled membrane receptors able to stimulate or inhibit different cell functions through the activation of inhibition of different intracellular pathways. They are heterogeneously expressed mainly in the central and peripheral nervous systems, where they mediate the effect of the neurotransmitter dopamine in several nervous functions, and in the endocrine system where they mediate the effect of dopamine on the control of hormone synthesis and secretion. In particular, the dopaminergic system has been demonstrated to be important in the regulation of the hypothalamus–pituitary–adrenal axis, which is strictly connected with the nervous system through the hypothalamus and the adrenal medulla. The dopamine receptor expression and function in the corticotrope pituitary tumors and adrenal tumors confirms the role of dopaminergic system in the hypothalamus–pituitary–adrenal axis, and suggests novel treatment strategies for Cushing’s syndrome. Finally, the dopamine receptor expression in neuroendocrine tumors confirms the role of the dopaminergic system in the entire neuroendocrine system.
References
1/2-adrenoceptor. Journal of Pharmacology and Experimental Therapeutics 2002 303 805–814.
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