Gonadotropin-releasing hormone (GnRH) is a decapeptide widely known for its role in regulating reproduction by serving as a signal from the hypothalamus to pituitary gonadotropes. In addition to hypothalamic GnRH (GnRH-I), a second GnRH form (pGln-His-Trp-Ser-His-GlyTrp-Tyr-Pro-Gly; GnRH-II) with unknown function has been localized to the midbrain of many vertebrates. We show here that a gene encoding GnRH-II is expressed in humans and is located on chromosome 20p13, distinct from the GnRH-I gene that is on 8p21-p11.2. The GnRH-II genomic and mRNA structures parallel those of GnRH-I. However, in contrast to GnRH-I, GnRH-II is expressed at significantly higher levels outside the brain (up to 30؋), particularly in the kidney, bone marrow, and prostate. The widespread expression of GnRH-II suggests it may have multiple functions. Molecular phylogenetic analysis shows that this second gene is likely the result of a duplication before the appearance of vertebrates, and predicts the existence of a third GnRH form in humans and other vertebrates.Gonadotropin-releasing hormone (GnRH) is a decapeptide widely known for its role in regulating reproduction. Release of GnRH from the hypothalamus controls the production of pituitary gonadotropins responsible for gonadal development and growth in all vertebrates. This function for GnRH has been highly conserved during 500 million years of vertebrate evolution despite the fact that its amino acid sequence varies by 50% (1).In addition to the hypothalamic GnRH of variable sequence, many vertebrate species have been shown to express a second, invariant GnRH form (pGln-His-Trp-Ser-His-Gly-Trp-TyrPro-Gly; GnRH-II) (2). By using antibody staining, this form of GnRH has been found in the midbrain in all species where its location has been described (reviewed in ref. 1). Furthermore, nucleic acid probes have been used to identify GnRH-II expression in the midbrain of several fish species and one mammal (1).Recently, a cDNA encoding this second form of GnRH was found in a placental mammal, the tree shrew Tupaia glis (1), thus leading us to search for it in humans. Here we describe the cloning of a cDNA encoding a second form of GnRH in humans and the subsequent isolation and sequencing of the complete human GnRH-II gene, the first description of a nonhypothalamic GnRH gene form in any species. In addition, the structure and chromosomal location of the new GnRH-II gene is compared with that of the previously described form in humans. Finally, we have constructed a molecular phylogeny of GnRH evolution, incorporating new sequence data for GnRH-II cDNAs from three placental mammals: human (this paper), tree shrew (1), and musk shrew (Suncus murinus; R.B.W., T.L.K., S. White, and R.D.F., unpublished data). MATERIALS AND METHODSLibrary Screen. A 270-nt partial cDNA for the putative human GnRH-II was cloned from human thalamus poly(A) RNA (CLONTECH) by using reverse transcription-PCR (RT-PCR) and 3Ј-RACE (rapid amplification of cDNA ends) as described (1). Oligomers flanking putative...

Corticosteroid hormones, including the mineralocorticoids and the glucocorticoids, regulate diverse physiological functions in vertebrates. These hormones act through two classes of corticosteroid receptors (CR) that are ligand-dependent transcription factors: type I or mineralocorticoid receptor (MR) and type II or glucocorticoid receptor (GR). There is substantial overlap in the binding of these two receptor types to hormones and to DNA. In fish, the overlap in processes controlled by CRs may be different from that in other vertebrates, as fish are thought to synthesize only glucocorticoids, whereas they express both GR and MR. Here we describe the characterization of four CRs in a cichlid fish, Haplochromis burtoni: a previously undescribed GR (HbGR1), another GR expressed in two splice isoforms (HbGR2a and HbGR2b), and an MR (HbMR). Sequence comparison and phylogenetic analysis showed that these CRs sort naturally into GR and MR groups, and that the GR duplication we describe will probably be common to all teleosts. Quantitative PCR revealed differential patterns of CR tissue expression in organs dependent on corticosteroid action. Trans-activation assays demonstrated that the CRs were selective for corticosteroid hormones and showed that the HbMR was similar to mammalian MRs in being more sensitive to both cortisol and aldosterone than the GRs. Additionally, the two HbGR2 isoforms were expressed uniquely in different tissues and were functionally distinct in their actions on classical GR-sensitive promoters. The identification of four CR subtypes in teleosts suggests a more complicated corticosteroid signaling in fish than previously recognized.

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Acute suppression of dipeptidyl peptidase IV (DPP-IV) activity improves glucose tolerance in the Zucker fatty rat, a rodent model of impaired glucose tolerance, through stabilization of glucagon-like peptide (GLP)-1. This study describes the effects of a new and potent DPP-IV inhibitor, FE 999011, which is able to suppress plasma DPP-IV activity for 12 h after a single oral administration. In the Zucker fatty rat, FE 999011 dose-dependently attenuated glucose excursion during an oral glucose tolerance test and increased GLP-1(7-36) release in response to intraduodenal glucose. Chronic treatment with FE 999011 (10 mg/kg, twice a day for 7 days) improved glucose tolerance, as suggested by a decrease in the insulin-to-glucose ratio. In the Zucker diabetic fatty (ZDF) rat, a rodent model of type 2 diabetes, chronic treatment with FE 999011 (10 mg/kg per os, once or twice a day) postponed the development of diabetes, with the twice-a-day treatment delaying the onset of hyperglycemia by 21 days. In addition, treatment with FE 999011 stabilized food and water intake to prediabetic levels and reduced hypertriglyceridemia while preventing the rise in circulating free fatty acids. At the end of treatment, basal plasma GLP-1 levels were increased, and pancreatic gene expression for GLP-1 receptor was significantly upregulated. This study demonstrates that DPP-IV inhibitors such as FE 999011 could be of clinical value to delay the progression from impaired glucose tolerance to type 2 diabetes.

Androgens are an important output of the hypothalamic-pituitary-gonadal (HPG) axis that controls reproduction in all vertebrates. In male teleosts two androgens, testosterone and 11-ketotestosterone, control sexual differentiation and development in juveniles and reproductive behavior in adults. Androgenic signals provide feedback at many levels of the HPG axis, including the hypothalamic neurons that synthesize and release gonadotropin-releasing hormone 1 (GnRH1), but the precise cellular site of androgen action in the brain is not known. Here we describe two androgen receptor subtypes, ARα and ARβ, in the cichlid Astatotilapia burtoni and show that these subtypes are differentially located throughout the adult brain in nuclei known to function in the control of reproduction. ARα was expressed in the ventral part of the ventral telencephalon, the preoptic area (POA) of the hypothalamus and the ventral hypothalamus, whereas ARβ was more widely expressed in the dorsal and ventral telencephalon, the POA, and the ventral and dorsal hypothalamus. We provide the first evidence in any vertebrate that the GnRH1-releasing neurons, which serve as the central control point of the HPG axis, express both subtypes of AR. Using quantitative real-time PCR, we show that A. burtoni AR subtypes have different expression levels in adult tissue, with ARα showing significantly higher expression than ARβ in the pituitary, and ARβ expressed at a higher level than ARα in the anterior and middle brain. These data provide important insight into the role of androgens in regulating the vertebrate reproductive axis. Keywordsin situ hybridization; gonadotropin-releasing hormone; androgen receptor subtypes; teleost Androgens are one endocrine output of the hypothalamic-pituitary-gonadal (HPG) axis that controls reproduction in all vertebrates. In male fish, the major androgens testosterone and 11-ketotestosterone (11-KT) are important for regulating sexual differentiation and functional development at all levels of the HPG axis (Borg, 1994). For example, exogenous androgens accelerate gonadal differentiation, spermatogenesis, and maturation of the hypothalamic and pituitary cells that synthesize and release the signaling peptides to mediate reproductive function (Amano et al., 1994;Goos et al., 1986;Miura et al., 1991a;Montero et al., 1995;Piferrer et al., 1993;Schreibman et al., 1986). If administered early enough in development in some teleost species, sex steroids can induce complete gonadal sex inversion in either direction (Hunter and Donaldson, 1983). In addition androgens also influence the adult HPG axis, mediating important social and reproductive behaviors, including courtship, territoriality, and aggression (Borg, 1994;Breton and Sambroni, 1996;Trudeau et al., 1991).Many physiological actions of androgens are mediated by the androgen receptor (AR) family. As with other steroid hormone receptors, ARs are ligand-dependent transcription factors, containing a highly conserved DNA-binding domain (DBD) and a moderately well cons...