For the TSH receptor signaling at the Golgi trans
For the TSH receptor, signaling at the Golgi/trans-Golgi network appears required for both rapid effects of TSH − such as cgrp antagonist synthesis depolymerization, which is implicated in thyroglobulin reuptake and, thus, thyroid hormone release − and late ones, such as those on gene transcription. Continued signaling by TSH receptors after internalization might contribute to hyperthyroidism in Grave's disease, where autoantibodies chronically activate the TSH receptor. Moreover, it might play a role in the pathogenesis of toxic thyroid adenomas and congenital/familial non-autoimmune hyperthyroidism, which are caused by activating TSH receptor mutations that are often associated with intracellular receptor accumulation , . For the PTH receptor, which plays a critical role in regulating Ca2+ homeostasis and bone turnover and is a major pharmacological target for the therapy of osteoporosis, it has been shown that PTH1-34 but not the PTH related peptide PTHrP1-36 − which activates the PTH receptor in a paracrine fashion − is capable of inducing persistent cAMP signaling . Moreover, a PTH analog (M-PTH1–34) that produces a more sustained cAMP response than PTH1-34 has been shown to induce larger increases in trabecular bone volume and cortical bone turnover, although the responsible mechanisms have not been fully elucidated . Similarly, vasopressin and oxytocin can both induce cAMP/PKA signaling upon binding to the V2 receptor, but only vasopressin leads to a strong antinatriuretic and antidiuretic effect , , . Feinstein et al. showed that this difference in signaling strength possibly results from different spatial signaling patterns induced by these two ligands . These examples also suggest the possibility of designing GPCR agonists capable of preferentially inducing cell-surface vs. intracellular signaling. This might allow developing a new generation of GPCR agonists with tailored biological effects, and thus, potentially improved efficacy and tolerability. More recently, our group took advantage of mice expressing a FRET sensor for cAMP to investigate cAMP signaling in intact ovarian follicles . We found that activation of LH receptors with LH induces two waves of cAMP production that propagate within the follicles. Importantly, blocking receptor internalization prevented the second phase and partially inhibited the LH-induce resumption of meiosis in the oocyte . These data indicate that LH receptor internalization plays an important role in mediating the biological effects of LH. Future studies appear required to further investigate the role of LH receptor signaling at intracellular sites in both female and male reproduction and its alterations in gonadal disorders. With the growing number of studies investigating GPCR signaling at intracellular sites, the physiological implications of this phenomenon are increasing. These include a role in insulin secretion for the GLP1 receptor , , in renal water and sodium reuptake for the vasopressin V2 receptor  and in the excitability of cardiac neurons for the PACAP1 receptor . Whereas receptor internalization has been mostly associated with prolonged cAMP signaling from intracellular sites, and thus mostly with slow biological effects, in the case of dopamine D1 receptors, it has been shown that these receptors are internalized very rapidly after agonist stimulation (within 1 min) and that the resulting cAMP signaling from endosomal membranes increases neuronal excitability in striatal neurons . So far, endosomal GPCR signaling has been mostly investigated in cellular models or using ex vivo preparations. Whereas these studies indicate that receptor internalization is required to mediate the biological effects of several hormones and neurotransmitters, further studies are required to investigate these processes in vivo. Interestingly, two recent studies have provided first in vivo evidence for a relevant physiological role of endosomal GPCR signaling. A first study investigated the role of internalization of the neurokinin 1 (NK1) receptor, which mediates the effects of substance P, on pain sensing . As a result of pain stimuli, substance P is released from the terminals of primary sensory neurons in the dorsal horn of the spinal cord, where it induces activation and internalization of NK1 receptors expressed in second-order neurons , . The results of the study indicate that inhibiting NK1 internalization and the resulting endosomal signaling attenuate nociception in vivo. This study also reports an innovative pharmacological strategy to selectively inhibit receptor endosomal signaling. For this purpose, the authors developed a cholestenol-conjugated antagonist, which accumulates in endosomes and is capable of inhibiting endosomal NK1 receptor signaling − which is required for nociception − without affecting NK1 receptors at the cell surface. Similar results were obtained by the same group for the calcitonin receptor-like receptor, which binds the calcitonin-gene related peptide (CGRP), and is also implicated in pain transmission .