G-protein–coupled receptors (GPCRs) are generally thought to signal to second messengers like cyclic AMP (cAMP) from the cell surface and to become internalized upon repeated or prolonged stimulation. Once internalized, they are supposed to stop signaling to second messengers but may trigger nonclassical signals such as mitogen-activated protein kinase (MAPK) activation. Here, we show that a GPCR continues to stimulate cAMP production in a sustained manner after internalization. We generated transgenic mice with ubiquitous expression of a fluorescent sensor for cAMP and studied cAMP responses to thyroid-stimulating hormone (TSH) in native, 3-D thyroid follicles isolated from these mice. TSH stimulation caused internalization of the TSH receptors into a pre-Golgi compartment in close association with G-protein α_s-subunits and adenylyl cyclase III. Receptors internalized together with TSH and produced downstream cellular responses that were distinct from those triggered by cell surface receptors. These data suggest that classical paradigms of GPCR signaling may need revision, as they indicate that cAMP signaling by GPCRs may occur both at the cell surface and from intracellular sites, but with different consequences for the cell. Cells respond to many environmental cues through the activity of cell surface receptor proteins, which sense these cues and convey that information to signaling molecules inside the cell. G-protein–coupled receptors (GPCRs) form the largest eukaryotic family of plasma membrane receptors. They convert the information provided by extracellular stimuli into intracellular second messengers, like cyclic AMP (cAMP). After prolonged stimulation, they are internalized inside cells, an event that to date has been thought to terminate the production of second messengers. Though many of the key steps of GPCR signaling are known in detail, precisely how signaling and termination actually occur in time and space (i.e., in subcellular compartments or microdomains) is still largely unexplored. To observe GPCR signaling in living cells, we generated mice expressing a fluorescent sensor that allows monitoring the intracellular levels of cAMP with a microscope. We utilized this system to study, directly in native thyroid follicles, the signal sent by the receptor for thyroid-stimulating hormone (TSH). Our findings indicate that TSH receptors are internalized rapidly after activation but continue to stimulate cAMP production inside cells and that this sustained, cAMP production is apparently required for localized activation of downstream components. These data challenge the current model of the GPCR-cAMP pathway by suggesting the existence of previously unrecognized intracellular site(s) for cAMP generation and of differential signaling outcomes as a result of intracellular GPCR signaling. Such intracellular site(s) may provide specialized signaling platforms, thus contributing to the spatiotemporal regulation of cAMP production and to signaling specificity within the GPCR family.

Central hypothyroidism (CH) is a particular hypothyroid condition due to an insufficient stimulation by TSH of an otherwise normal thyroid gland. This condition raises several challenges for clinicians; therefore, a review of the most relevant findings on CH epidemiology, pathogenesis, and clinical management has been performed. The relevant papers were selected by a PubMed search using appropriate key words. CH can be the consequence of various disorders affecting either the pituitary gland or the hypothalamus, but most frequently affecting both of them. CH is about 1000-fold rarer than primary hypothyroidism. Except for the neonatal CH due to biallelic TSHβ mutations, the thyroid hormone defect is rarely as profound as can be observed in some primary forms. In contrast with primary hypothyroidism, CH is most frequently characterized by low/normal TSH levels, and adequate thyroid hormone replacement is associated with the suppression of residual TSH secretion. Thus, CH often represents a clinical challenge because physicians cannot rely on the systematic use of the "reflex TSH strategy." The clinical management of CH is further complicated by the frequent combination with other pituitary deficiencies and their substitution.

Premature ovarian failure (POF) is a primary ovarian defect characterized by absent menarche (primary amenorrhea) or premature depletion of ovarian follicles before the age of 40 years (secondary amenorrhea). It is a heterogeneous disorder affecting approximately 1% of women <40 years, 1:10,000 women by age 20 and 1:1,000 women by age 30. The most severe forms present with absent pubertal development and primary amenorrhea (50% of these cases due to ovarian dysgenesis), whereas forms with post-pubertal onset are characterized by disappearance of menstrual cycles (secondary amenorrhea) associated with premature follicular depletion. As in the case of physiological menopause, POF presents by typical manifestations of climacterium: infertility associated with palpitations, heat intolerance, flushes, anxiety, depression, fatigue. POF is biochemically characterized by low levels of gonadal hormones (estrogens and inhibins) and high levels of gonadotropins (LH and FSH) (hypergonadotropic amenorrhea). Beyond infertility, hormone defects may cause severe neurological, metabolic or cardiovascular consequences and lead to the early onset of osteoporosis. Heterogeneity of POF is also reflected by the variety of possible causes, including autoimmunity, toxics, drugs, as well as genetic defects. POF has a strong genetic component. X chromosome abnormalities (e.g. Turner syndrome) represent the major cause of primary amenorrhea associated with ovarian dysgenesis. Despite the description of several candidate genes, the cause of POF remains undetermined in the vast majority of the cases. Management includes substitution of the hormone defect by estrogen/progestin preparations. The only solution presently available for the fertility defect in women with absent follicular reserve is ovum donation.

A large number of studies have contributed to understanding the general mechanisms driving ovarian folliculogenesis in humans and show a complex endocrine dialog between the central nervous system, the pituitary and the ovary, integrated by various intraovarian paracrine messages. The role of intraovarian paracrine regulation has acquired more relevance in the recent years owing to the discovery of previously unknown factors, such as the oocyte-derived bone morphogenetic protein (BMP)15. A thorough literature search was carried out in order to summarize what has been reported so far on the role of BMP15, and the BMP15 paralog, growth and differentiation factor 9 (GDF9), in ovarian function and female fertility. Research articles published in English until March 2014 were included. The biological actions of BMP15 include: (i) the promotion of follicle growth and maturation starting from the primary gonadotrophin-independent phases of folliculogenesis; (ii) the regulation of follicular granulosa cell (GC) sensitivity to FSH action and the determination of ovulation quota; (iii) the prevention of GC apoptosis and (iv) the promotion of oocyte developmental competence. The existence of biologically active heterodimers with GDF9, and/or the synergistic co-operation of BMP15 and GDF9 homodimers are indeed relevant in this context. Experimental disruption of the bmp15 gene in mice resulted in a mild fertility defect limited to females, whereas natural missense mutations in ewes cause variable phenotypes (ranging from hyperprolificacy to complete sterility) depending on a fine gene dosage mechanism also involving GDF9. Strong evidence supports the concept that such a mechanism plays an important role in the regulation of ovulation rate across mammalian and non-mammalian species. Following the discovery of sheep fecundity genes, several research groups have focused on alterations in human BMP15 associated with primary ovarian insufficiency (POI) or polycystic ovary syndrome. Several variants of BMP15 are significantly associated with POI supporting their pathogenic role, but the underlying biological mechanism is still under investigation and of great interest in medicine. BMP15 maps to the Xp locus involved in the determination of the ovarian defect in Turner syndrome and significantly contributes to the determination of ovarian reserve. Pioneering studies in women undergoing controlled ovarian stimulation indicate that BMP15 may represent a marker of ovarian response or oocyte quality. BMP15, an oocyte-derived growth and differentiation factor, is a critical regulator of folliculogenesis and GC activities. Variations in BMP15 gene dosage have a relevant influence on ovarian function and can account for several defects of female fertility. The modulation of BMP15 action may have interesting pharmacological perspectives and the analysis of BMP15 may become a useful marker in IVF procedures. Recent outcomes indicate that the close interactions of BMP15/GDF9 have a critical biological impact that should be taken into account in future studies.