The growth and development of an organism requires the coordinated actions of many factors. During development individual cells undergo proliferation, migration and differentiation to form the adult organism. Two structurally related members of the glucagon superfamily, growth hormone releasing hormone (GRF) and pituitary adenylate cyclase-activating polypeptide (PACAP), are thought to modulate vertebrate development. In mammals, GRF modulates the development of pituitary somatotrophs and the release of fetal growth hormone. In contrast, PACAP appears to have a more general role during development. PACAP may be involved in the patterning of the embryonic axis and in the development of the neural tube. The objectives of my study were to isolate GRF, PACAP and their receptors from the zebrafish, characterize their expression in the developing embryo and adult embryo and examine the role of PACAP during brain development. To study the role of GRF and PACAP, I isolated a genomic clone encoding the GRF and PACAP peptides from the zebrafish genomic library and characterized its gene copy number and adult tissue expression pattern. The GRF-PACAP gene isolated from the zebrafish was comprised of five exons with the GRF peptide encoded on the fourth exon and the PACAP peptide encoded on the fifth exon. This gene structure is similar to that found in other non-mammalian vertebrates and supports the hypothesis that the gene duplication leading to the encoding of the GRF and PACAP peptides on separate genes occurred later in evolution. In addition, the zebrafish genome was found to contain only one copy of the GRF-PACAP gene. The GRF-PACAP gene was widely expressed in the adult zebrafish in tissues developmentally derived from all three germ layers, suggesting that the gene may be widely expressed in the embryo as well. To examine the functional significance of the co-expression of GRF and PACAP in zebrafish, I isolated the GRF and PACAP receptors and characterized their expression pattern. I isolated three distinct cDNAs from zebrafish encoding the GRF receptor, the PACAP specific PAC1 receptor and the shared vasoactive intestinal peptide/PACAP receptor VPAC1. In addition, four isoforms of the PAC1 receptor were isolated from zebrafish including a novel isoform found in the gill. All three receptors were widely expressed in adult zebrafish and receptors for both GRF and PACAP were found in most tissues. This indicates that GRF and PACAP may modulate each other’s function. To determine the developmental role of GRF and PACAP, I characterized the expression pattern of the GRF-PACAP gene and the GRF, PAC1 and VPAC1 receptors in the zebrafish embryo. The GRF and PAC1 receptors are the earliest to be expressed in development starting at the cleavage stage. Later, the GRF-PACAP gene and the VPAC1 receptor are first expressed at the late blastula/early gastrula stage in the zebrafish and are expressed throughout the developmental period. Strong expression of the GRF, PACAP and their receptors during mid gastrulation indicates that these peptides may be involved in modulating the formation of the embryonic axis. During the segmentation period the GRF-PACAP gene is widely expressed in the zebrafish embryo and the PAC1 receptor short and hop isoforms are differentially expressed. Therefore, PACAP may regulate cell cycle exit or cell proliferation through activation of different PAC1 receptor isoforms during the segmentation stage. In the subsequent pharyngula period, the GRF-PACAP transcript is localized mainly to the hatching gland. However, expression is seen also in tissues that undergo differentiation during this stage. Therefore, the timing of the expression of the GRF-PACAP gene indicates that it may be involved in early patteming events and promoting cell cycle exit prior to differentiation. To investigate the role of GRF and PACAP in the developing brain, I localized the expression of GRF, PACAP and the PAC1 receptor in neuroblasts derived from an embryonic day 3.5 chick. PACAP was found to stimulate the cAMP pathway in these cells, indicating that PACAP may modulate brain development. This work indicates that GRF and PACAP play an important role in vertebrate development. / Graduate

Growth hormone-releasing hormone (GRF) and pituitary adenylate cyclase-activating polypeptide (PACAP) are members of the PACAP/Glucagon superfamily. The family is proposed to have developed from an ancestral PACAP-like molecule in invertebrates. Through successive exon, gene and genome duplications the family has grown to include seven other members. In mammals GRF and PACAP are located on different genes, but in fish, amphibians and birds they are located on the same gene. The main function of GRF is the release of growth hormone (GH) from the pituitary. Also, during development GRF influences the fetal pituitary and stimulates GH release during late gestation. In contrast, the functions of PACAP are extremely varied. PACAP is the newest member of the superfamily and there is still much work to be done before its actions are well understood. Like GRF, PACAP is a releasing hormone acting on the pituitary and in addition, the adrenal gland, pancreas and heart, as well as other organs. Also, PACAP regulates smooth muscle in the vascular system, gut, respiratory tract and reproductive tract During development PACAP affects proliferation, differentiation and apoptosis. GRF and PACAP are expressed throughout development in fish, beginning during the blastula period in rainbow trout and at the end of gastrulation in zebrafish (earliest stage examined). In rainbow trout the grf/pacap gene is expressed as two transcripts, a short and a long transcript. The short transcript is produced by alternative splicing of the gene and does not include the fourth exon which codes for GRF. The long transcript includes the coding regions for both GRF and PACAP. By this means PACAP can be regulated separately from GRF. With the extensive role PACAP appears to play in development, separate regulation of the hormone may be necessary. Expression of the grf/pacap gene in zebrafish is widespread early in development and gradually becomes localized. Of particular interest is the expression of the grf/pacap transcript in regions associated with the prechordal plate, an important organizing center in development. Although it is not yet confirmed, there is evidence to suggest GRF and PACAP are expressed in the prechordal plate and its derivatives in the gut and hatching gland. In addition, expression of the grf/pacap transcript is observed in the neuroectoderm (eye, brain and spinal cord) and the developing heart. Considering the expression pattern of GRF and PACAP, I propose that one of both of these hormones may be involved in patterning during vertebrate embryogenesis. The evolution of gene families is thought to occur through successive exon, gene and genome duplications. Duplicate exons or genes become differentiated and eventually gain new functions or become functionless. During evolution of the grf/pacap lineage, several duplication events have occurred. Analysis of rainbow trout leads me to think that this fish and other salmonids possess two copies of the grf/pacap gene. This is not unexpected considering the tetraploid nature of salmonids. Present day mammals encode GRF and PACAP on separate genes. At some point during the evolution of this lineage a duplication event has occurred, possibly in early mammals or prior to the divergence of birds. The study of multigene families is a useful way to understand evolutionary processes. To this end I examined three members of multigene families from sockeye salmon. Therefore, in addition to the evolutionary mechanisms and pathways that directed grf/pacap gene evolution, I examined the ferritin-H subunit, the alpha-tubulin subunit and the beta-globin subunit. These cDNA sequences are similar to their counterparts in other teleost. The evolution of the ferritin gene family is particularly interesting because it involves the addition or deletion of DNA sequences that affect regulation and cytosolic location. / Graduate

L’antéhypohyse est une glande endocrine maitresse produisant 6 hormones (GH, TSH, ACTH, PRL, FSH et LH) qui régulent des processus vitaux comme la croissance. Chez l’Homme, des mutations des gènes impliqués dans l’ontogénèse pituitaire sont responsables de déficit(s) isolé (IGHD) ou multiples (CPHD) en hormones antéhypophysaires, pouvant être syndromique(s). L’incidence de ces déficits est d’environ 1/8000 naissances. A ce jour, des mutations rapportées dans une trentaine de gènes permettent d’expliquer cette affection chez seulement 25% des patients. Le premier objectif de ce travail de thèse était d’identifier de nouveaux gènes responsables de déficit antéhypophysaire par une approche « gènes candidats », et par des analyses d’exome couplées à une cartographie par homozygotie. Nous avons pu mettre en évidence des variations dans les gènes NOG, VAX1 et GSX1, impliqués dans l’ontogénèse pituitaire murine, et identifier d’autres gènes potentiellement clés dans le développement de l’hypophyse. L’autre objectif était de caractériser la fonction de LHX4, GHRHR, GLI2 et GSX1 à travers l’étude fonctionnelle des variants identifiés au laboratoire. L’implication de nouveaux variants génétiques de LHX4, GHRHR et GLI2 a permis de préciser le spectre phénotypique associé aux mutations de chacun de ces gènes, et aussi de préciser le rôle de LHX4 dans le réseau moléculaire complexe nécessaire à la formation de l’hypophyse. La compréhension des mécanismes moléculaires à l’origine du développement pituitaire devrait permettre à terme une meilleure prise en charge de cette pathologie très hétérogène, incluant le conseil génétique des familles concernées et le traitement des patients. / The anterior pituitary is a master endocrine gland producing 6 hormones (GH, TSH, ACTH, PRL, FSH and LH) that regulate essential life processes such as growth. In the human, mutations in genes encoding morphogenes and transcription factors implicated in pituitary ontogenesis are responsible for isolated (IGHD) or combined (CPHD) pituitary hormone deficit(s) that can be syndromic. The incidence of those deficits is close to 1/8,000 births. So far, mutations reported in about thirty genes explain only 25% of the cases. The first goal of this thesis was to identify new genes responsible for pituitary hormone deficits through a candidate gene approach, and by whole-exome sequencing coupled with homozygosity mapping, using a large cohort of more than 3000 DNA samples collected by the laboratory since 1997.We found variations in NOG, VAX1 and GSX1 genes, all involved in mouse pituitary ontogenesis, and we identified potentially key genes for pituitary developpment.The other aim of this work was to characterize the function of LHX4, GHRHR, GLI2 and GSX1 through the functional evaluation of the variants identified in the laboratory. The implication of new disease-causing genetic variants in LHX4, GHRHR and GLI2 allowed us to delineate the phenotypic spectrum associated with mutations for each gene, and also to precise the role of LHX4 in the complex molecular network required for proper pituitary development. A better understanding of the molecular mecanisms involved in pituitary formation should lead to new ways for management of this heterogenous condition, including genetic counseling and therapeutic decisions in affected families.

No description available.

Indiana University-Purdue University Indianapolis (IUPUI) / LHX3 is a transcription factor important in pituitary and nervous system development. Patients with mutations in coding regions of the gene have combined pituitary hormone deficiency (CPHD) that causes growth, fertility, and metabolic problems. Promoter and intronic elements of LHX3 important for basal gene expression in vitro have been identified, but the key regulatory elements necessary for in vivo expression were unknown. With these studies, I sought to elucidate how LHX3 gene expression is regulated in vivo. Based on sequence conservation between species in non-coding regions, I identified a 7.9 kilobase (kb) region 3' of the human LHX3 gene as a potential regulatory element. In a beta galactosidase transgenic mouse model, this region directed spatial and temporal expression to the developing pituitary gland and spinal cord in a pattern consistent with endogenous LHX3 expression. Using a systematic series of deletions, I found that the conserved region contains multiple nervous system enhancers and a minimal 180 base pair (bp) enhancer that direct expression to both the pituitary and spinal cord in transgenic mice. Within this minimal enhancer, TAAT/ATTA sequences that are characteristic of homeodomain protein binding sites are required to direct expression. I performed DNA binding experiments and chromatin immunoprecipitation assays to reveal that the ISL1 and PITX1 proteins specifically recognize these elements in vitro and in vivo. Based on in vivo mutational analyses, two tandem ISL1 binding sites are required for enhancer activity in the pituitary and spine and a PITX1 binding site is required for spatial patterning of gene expression in the pituitary. Additional experiments demonstrated that these three elements cannot alone direct gene expression, suggesting a combination of factors is required for enhancer activity. This study reveals that the key regulatory elements guiding developmental regulation of the human LHX3 gene lie in this conserved downstream region. Further, this work implicates ISL1 as a new transcriptional regulator of LHX3 and describes a possible mechanism for the regulation of LHX3 by a known upstream factor, PITX1. Identification of important regulatory regions will also enable genetic screening in candidate CPHD patients and will thereby facilitate patient treatment and genetic counseling.

Der GLI2-Transkriptionsfaktor ist eines der Haupt Effektor-Proteine des Sonic Hedgehog (SHH)-Signalweges und hat vermutlich eine Schlüsselfunktion in der Entwicklung der Hypophyse. Genomische GLI2-Veränderungen welche zu abgeschnittenen Proteinen führten, wurden beschrieben als Ursache für Holoprosenzephalie (HPE) oder HPE-ähnliche Veränderungen, teilweise in Verbindung mit einer Hypophysenunterfunktion. Ziel dieser Arbeit war die Ermittlung der Frequenz von GLI2-Mutationen in Patienten mit multiplem hypophysärem Hormonausfall (multiple pituitary hormone deficiency, MPHD) und eine funktionelle Untersuchung der gefunden Mutationen mittels Transkriptionsaktivitäts-Untersuchungen durch funktionelle Luciferase assays. Hierfür wählten wir Teilnehmer der GeNeSIS (Genetics and Neuroendocrinology of Short Stature International Study)-Studie. Patienten bei denen bereits Mutationen eines der etablierten Gene für MPHD nachgewiesen wurde, wurden ausgeschlossen. Insgesamt haben wir 168 Patienten mit MPHD untersucht. Bei allen Patienten waren mindestens ein GH- und ein TSH-Mangel dokumentiert, Auffälligkeiten in der zentralen Bildgebung mittels cMRT wurden bei 96 Patienten angegeben. In fünf Studienteilnehmern wurden vier verschiedene heterozygote missense Varianten nachgewiesen, hiervon wurden zwei bislang noch nicht in der Literatur beschrieben. Eine Variante, pR516P, führte in den in-vitro Experimenten zu einem kompletten Verlust der Proteinaktivität. Zusätzlich zu einem Wachstumshormonmangel hatte der Träger dieser Mutation einen Mangel an TSH und der Gonadotropine, sowie einen nichtdeszendierten Hypophysenhinterlappen und eine Polydaktylie, aber keine ersichtlichen Mittelliniendefekte. Anhand der funktionellen Untersuchung konnten wir erstmalig nachweisen, dass ein heterozygoter Aminosäuren-Austausch im GLI2-Protein zu einer möglichen Funktionseinschränkung der Transkriptionsaktivität führen kann und somit die Ursache für MPHD mit milden extrahypophysären Auffälligkeiten sein könnte. Der Phänotyp von GLI2-Mutationen ist variabel und die Penetranz ist unvollständig. GLI2-Mutationen sind assoziiert mit einer Hypoplasie des Hypophysenvorderlappens und treten gehäuft mit einem ektopen Hypophysenhinterlappen auf.

Made available in DSpace on 2014-06-11T19:25:35Z (GMT). No. of bitstreams: 0 Previous issue date: 2008-11-17Bitstream added on 2014-06-13T18:53:44Z : No. of bitstreams: 1 cruz_jb_me_botfm.pdf: 449482 bytes, checksum: 5627323423c217568fc73820cbf66a6a (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / A hipófise anterior compõe-se de cinco tipos celulares que são definidos pelos hormônios que secretam. A diferenciação desses tipos celulares resulta de uma cascata temporalmente regulada de fatores transcricionais expressos no tecido hipofisário. Mutações de um desses fatores podem resultar tanto em defeitos estruturais da glândula como em deficiências hormonais, que podem ser isoladas (Déficit de hormônio do crescimento - DGH) ou combinadas (DHHC), dependendo do papel do fator transcricional mutado. A Displasia Septo – óptica (DSO) caracteriza-se pela presença de hipoplasia hipofisária, hipoplasia de nervo óptico e/ou má formações de estruturas da linha média. Foram avaliados 11 pacientes com quadro clinico de SOD, DGH e DHHC e realizado o sequenciamento genético do gene HESX1 desses indivíduos. Nos casos de DHHC foi feita uma análise adicional do gene PROP1. A mutação missense em estado de heterozigose A1772G levando a substituição N125S foi identificada em um paciente portador de DSO, no gene HESX1. Essa troca já foi previamente relatada como um polimorfismo na população Afro-Caribenha. Encontramos três pacientes portadores da variante alélica A9A e N20S no exon 1 do gene PROP1, já descritos previamente na literatura como polimorfismos. / The anterior pituitary is made of five types of cell defined according to the hormones they secrete. Differentiation among such cell types derives from a cascade of temporally regulated transcriptional factors expressed in the pituitary tissue. Mutation in one of these factors may result in both structural gland defects and hormonal deficiencies, which may be either isolated (DGH – Growth Hormone Deficiency) or combined, depending on the role of the mutated transcriptional factor. Septo-optic dysplasia (SOD) is characterized by pituitary hypoplasia, optic nerve hypoplasia and/or malformation of medium line structures. Eleven patients with a clinical state of SOD, DGH and CPHD were assessed, and genetically sequenced for the HESX1 gene. For CPHD cases, an additional analysis for the PROP1 gene was also conducted. One SOD patient was found to have a missense mutation in A1772G heterozygosis state, leading to the N125S replacement, in HESX1 gene. Such replacement has already been reported as a polymorphism in the Afro-Caribbean population. We found three patients with the alelic variation A9S and N20A in exon 1 of PROP1 gene, previously described as polymorphisms.

Gonadotropin-releasing hormone (GnRH) and its superactive analogues are currently being used in the treatment of a number of endocrine disorders, such as endometriosis, precocious puberty, infertility and prostatic cancer. Selection of these analogues for clinical use have been previously based on their activities in animal models. This thesis has therefore investigated the binding characteristics of the human GnRH receptor, in comparison to those of the rat receptor, as well as the activities of a number of GnRH analogues for stimulating luteinising hormone (LH) and follicle stimulating hormone (FSH) secretion from cultured human pituitary cells. The establishment of a human pituitary bioassay system has further made possible the investigation of the direct regulatory roles of GnRH and other neuropeptides in man. To date, such studies in man have been performed in vivo and are thus complicated by the simultaneous interactions of numerous modulators.

The control of prolactin secretion is a complex interaction of peptides and neurotransmitters acting either in an inhibitory or stimulating way to effect final secretion of this hormone from the lactotrope cell in the anterior hypothalamus. These factors may act either directly on the lactotrope cell or indirectly by changing either dopamine restraint of prolactin secretion or by modulating peptide substances or neurotransmitters higher up in the hypothalamus. Gonadal steroids may also modulate the effect of peptides or dopamine at the level of the lactotrope. Prolactin's major role in the female rat is one of milk production post - partum, nurturing the young. It probably also has other physiological functions and may play a part in the menstrual cycle although this is controversial. Certainly, pulsatile secretion of prolactin during the menstrual cycle is well established and in the luteal phase this is concomitant with the secretion of luteinizing hormone. Theories explaining the synchronous surges seen during this phase of the menstrual cycle have been proposed and GnRH has been implicated in the genesis of the concordance of these secretory spikes. Using a potent GnRH antagonist an experiment was undertaken to establish the role of GnRH by blocking this hypothalamic peptide and observing the effect that this had on luteinizing hormone, prolactin and follicle stimulating hormone. In the first part of the thesis the control of prolactin secretion is reviewed. In the following section, an experiment was performed using a potent GnRH antagonist. A dose response curve was established for the antagonist action on LH. Then a twice maximum dose of this peptide was administered to three subjects in the midluteal phase of the menstrual cycle and the response of LH, prolactin and FSH was measured. The results indicate that although the GnRH antagonist significantly blocked LH secretory peaks, this action was not observed for either prolactin or FSH. This result is perhaps at variance with previous data which suggested that GnRH was responsible for concordant secretory spikes of LH and prolactin in the midluteal phase of the menstrual cycle.

Η αυξητική ορμόνη (GH), πολυλειτουργική ορμόνη που παράγεται από τα σωματοτρόπα κύτταρα του πρόσθιου λοβού της υπόφυσης, προάγει την μεταγεννητική ανάπτυξη σκελετικών και μαλακών ιστών. Επίσης, ασκεί ποικίλες άλλες βιολογικές δράσεις, όπως ρύθμιση του μεταβολισμού των υδατανθράκων, των πρωτεϊνών και του λίπους. Κατά συνέπεια, η ανεπάρκεια της εκτός από αναπτυξιακά μπορεί να προκαλέσει και σοβαρά μεταβολικά προβλήματα. Η GH δρα στους περιφερικούς ιστούς άμεσα αλλά και έμμεσα μέσω του ινσουλινόμορφου αυξητικού παράγοντα IGF-I. Μετά από πρόσδεση της GH στον υποδοχέα της (GHR), ο IGF-I παράγεται στο ήπαρ, όπου απελευθερώνεται στην γενική κυκλοφορία, αλλά παράγεται και τοπικά στους περιφερικούς ιστούς, όπου δρα με αυτοκρινή ή παρακρινή τρόπο. Η έκκριση της GH από την υπόφυση έχει παλμική μορφή και ρυθμίζεται κυρίως μέσω τριών υποφυσιοτρόπων παραγόντων: εκλυτική ορμόνη της GH (GHRH), σωματοστατίνη (SRIF) και γκρελίνη. Η απελευθέρωση της GHRH και της SRIH από τον υποθάλαμο επηρεάζεται και από μια ποικιλία άλλων νευροδιαβιβαστών, νευροορμονών και νευροπεπτιδίων. Έχει υπολογιστεί σε διάφορες μελέτες ότι κοντό ανάστημα συσχετιζόμενο με ανεπάρκεια της αυξητικής ορμόνης (GHD) παρατηρείται με συχνότητα 1 στις 4000 έως 1 στις 10000 γεννήσεις. Παρόλο που οι περισσότερες περιπτώσεις είναι σποραδικές και θεωρούνται αποτέλεσμα περιβαλλοντικών εγκεφαλικών προσβολών ή αναπτυξιακών ανωμαλιών, γενετική αιτιολογία προτείνεται περίπου στο 10% των GHD περιπτώσεων, λόγω του ότι έχει προσβληθεί ένας τουλάχιστον πρώτου βαθμού συγγενής. Η διάγνωση της GHD είναι μια πολύπλευρη διαδικασία που απαιτεί εκτενή κλινική εκτίμηση, αξιολόγηση σωματομετρικών παραμέτρων, βιοχημικές δοκιμασίες του GH-IGF άξονα, και ακτινολογική εκτίμηση. Η GHD μπορεί να παρουσιάζεται είτε ως μεμονωμένο πρόβλημα (IGHD) είτε σε συνδυασμό με πολλαπλές ορμονικές ανεπάρκειες (CPHD). Μοντέλα ζώων έχουν χρησιμοποιηθεί για μελέτη της φυσιολογικής λειτουργίας του υποθαλαμικού-GH άξονα και των πιθανών διαταραχών που οδηγούν σε IGHD/CPHD στους ανθρώπους. Σύμφωνα με τα κλινικά χαρακτηριστικά, τον τρόπο κληρονομικότητας και την ανταπόκριση στην εξωγενή θεραπεία, τέσσερις τύποι οικογενούς IGHD έχουν περιγραφεί στον άνθρωπο. Μεταλλαγές έχουν βρεθεί να συμβαίνουν στο GH γονίδιο (GH1) και στο γονίδιο του υποδοχέα της GHRH (GHRH-R). Πολυμορφισμοί στον υποκινητή του GH1 γονιδίου μειώνουν επίσης την έκφραση του. Πρόσφατα, μεταλλαγές στο γονίδιο του υποδοχέα της γκρελίνης (GHS-R) συσχετίστηκαν με IGHD. Μεταλλαγές σε διακριτά γονίδια μεταγραφικών παραγόντων, που είναι βασικά για την ανάπτυξη και διαφοροποίηση των κυττάρων του πρόσθιου λοβού της υπόφυσης, όπως Pit1/POU1F1, PROP1, HESX1, LHX3, LHX4, έχουν αναγνωρισθεί μέχρι σήμερα σε ανθρώπους με CPHD. Καθώς μεγάλο ποσοστό οικογενών περιπτώσεων IGHD/CPHD δεν οφείλεται σε μεταλλαγές σε κάποιο από τα ήδη γνωστά γονίδια, φαίνεται να εμπλέκονται μεταλλαγές σε επιπρόσθετα υποψήφια γονίδια. Περαιτέρω γενετικές μελέτες μπορούν να συμβάλλουν σε καλύτερη κατανόηση της GHD, σε πρώιμη διάγνωση και βελτίωση της θεραπευτικής αγωγής στα άτομα με GHD. / Growth hormone (GH), a multifunctional hormone which is synthesized in the somatotrope cells of the anterior pituitary gland, promotes postnatal development of skeletal and soft tissues. In addition, GH exerts multiple biological actions, such as regulating the metabolism of carbohydrates, proteins and fat. Consequently, GH deficiency (GHD) apart from causing developmental disorders can also have a deleterious effect on the body’s metabolism. GH acts on peripheral tissues both directly and indirectly, through the mediation of insulin-like growth factor-1 (IGF-1). Upon binding of GH to its receptor (GHR), IGF-1 is produced both in the liver, from where it is released into the general circulation, and locally in the peripheral tissues, such as bone, cartilage, and muscle, where it acts in an autocrine or paracrine fashion. GH is secreted from the pituitary gland in a pulsatile fashion. Major regulatory factors include three hypophysiotropic factors: GH releasing hormone (GHRH), somatostatin (SRIF), and ghrelin. Moreover, GH secretion can be affected by a variety of other neurotransmitters, neurohormones and neuropeptides. The diagnosis of GHD demands detailed clinical, auxological, radiological and biochemical evaluation of the GH-IGF axis. GHD may occur as isolated GHD (IGHD) or in combination with other pituitary hormone deficiencies (Combined Pituitary Hormone Deficiency, CPHD). The physiological actions of the hypothalamic-GH axis and the possible disorders leading to IGHD/CPHD in humans have been extensively studied in animal models. Short stature associated with GHD has been estimated to occur in about 1/4000-1/10000 in various studies. Whereas most cases are sporadic and believed to result from environmental cerebral insults or developmental anomalies, approximately 10% of the affected individuals have a first-degree relative with the same disorder, suggesting a hereditary trend and genetic factors affecting the disorder. Four types of familial IGHD have been described in humans according to clinical characteristics, the mode of inheritance and the response to exogenous therapy. Mutations reducing gene expression have been described in the GH1 gene and in the GHRH receptor (GHRH-R) gene. Polymorphisms found in the promoter of the GH1 gene can also reduce its expression. Recently, mutations in the ghrelin receptor (GHS-R) gene were associated with IGHD. Mutations in discrete genes of transcriptional factors necessary for the development and differentiation of anterior pituitary cells, such as Pit1/POU1F1, PROP1, HESX1, LHX3, LHX4 have been recognized in individuals with CPHD. Considering that a large proportion of familial cases of IGHD/CPHD are not caused by mutations in any of the known genes, mutations in additional candidate genes may be involved. Further genetic studies may contribute to a better understanding of GHD, earlier diagnosis and better therapeutic approaches for this disorder.