Quantification of receptor activation by oxytocin and vasopressin in endocytosis-coupled bioluminescence reduction assay using nanoKAZ

Kii, Isao; Hirahara-Owada, Shino; Yamaguchi, Masataka; Niwa, Takashi; Koike, Y.; Sonamoto, Rie; Ito, Hiromitsu; Takahashi, Kayo; Yokoyama, Chihiro; Hayashi, Takuya; Hosoya, Takamitsu; Watanabe, Yasuyoshi

doi:10.1016/j.ab.2018.04.001

Cited by 6 publications

(3 citation statements)

References 43 publications

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“…nLuc-GLP-1R internalization is implicated by a decrease in luminescence due to the reduced availability of luciferase substrate inside the cell versus at the cell surface. 41 Luciferase substrate was introduced 30 min after addition of excess (1 µM) GLP-1 or peptides 1-7 . As expected, after GLP-1 treatment a substantial decline in luminescence was observed, which is consistent with extensive nLuc-GLP-1R internalization induced by the native hormone (Figures 2C, S3A).…”

Section: Resultsmentioning

confidence: 99%

Prolonged signaling of backbone-modified glucagon-like peptide-1 analogues with diverse receptor trafficking

Cary,

Hager,

Morris

et al. 2024

Preprint

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Signal duration and subcellular location are emerging as important facets of G protein-coupled receptor (GPCR) function. The glucagon-like peptide-1 receptor (GLP-1R), a clinically relevant class B1 GPCR, stimulates production of the second messenger cAMP upon activation by the native hormone, GLP-1. cAMP production continues after the hormone-receptor complex has been internalized via endocytosis. Here, we report GLP-1 analogues that induce prolonged signaling relative to GLP-1. A single beta-amino acid substitution at position 18, with the residue derived from (S,S)-trans-2-aminocyclopentanecarboxylic acid (ACPC), enhances signaling duration with retention of receptor endocytosis. Pairing ACPC at position 18 with a second substitution, amino-aminoisobutyric acid (Aib) at position 16, abrogates endocytosis, but prolonged signaling is maintained. Prolonged signaling is sensitive to the structure of the beta residue at position 18. Cryo-electron microscopy (cryo-EM) structures of two GLP-1 analogues bound to the GLP-1R:Gs complex suggest substantial alterations to bound peptide structure and dynamics compared to the GLP-1:GLP-1R complex. These structural findings strengthen an emerging view that agonist dynamics in the receptor-bound state influence signaling profile. Our results advance understanding of the structural underpinnings of receptor activation and introduce new tools for exploring the impact of spatiotemporal signaling profiles following GLP-1R activation.

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Section: Resultsmentioning

confidence: 99%

Prolonged signaling of backbone-modified glucagon-like peptide-1 analogues with diverse receptor trafficking

Cary,

Hager,

Morris

et al. 2024

Preprint

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show abstract

“…While pre-treatment with 1 μM atosiban decreased the level of spontaneous contractility, secondary treatment with 1 nM analogs partially recovered contractility in the following order: HPOT > OT > FBOT. Thus, the ability of the analogs to counter the effects of atosiban did not match the potency or affinity orders as atosiban is known to be a non-selective antagonist acting via OT receptors and arginine vasopressin V1A or V1B receptors [ 36 ], one of the main differences between OT and its analogs may be selectivity for the V1A receptors [ 7 ]. HPOT was demonstrated to be more selective in the activation of OT receptors, especially at lower concentrations, while FBOT activated V1A receptors at higher concentrations [ 7 ].…”

Section: Discussionmentioning

confidence: 99%

Two oxytocin analogs, N-(p-fluorobenzyl) glycine and N-(3-hydroxypropyl) glycine, induce uterine contractions ex vivo in ways that differ from that of oxytocin

et al. 2023

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Contraction of the uterus is critical for parturient processes. Insufficient uterine tone, resulting in atony, can potentiate postpartum hemorrhage; thus, it is a major risk factor and is the main cause of maternity-related deaths worldwide. Oxytocin (OT) is recommended for use in combination with other uterotonics for cases of refractory uterine atony. However, as the effect of OT dose on uterine contraction and control of blood loss during cesarean delivery for labor arrest are highly associated with side effects, small amounts of uterotonics may be used to elicit rapid and superior uterine contraction. We have previously synthesized OT analogs 2 and 5, prolines at the 7th positions of which were replaced with N-(p-fluorobenzyl) glycine [thus, compound 2 is now called fluorobenzyl (FBOT)] or N-(3-hydroxypropyl) glycine [compound 5 is now called hydroxypropyl (HPOT)], which exhibited highly potent binding affinities for human OT receptors in vitro. In this study, we measured the ex vivo effects of FBOT and HPOT on contractions of uteri isolated from human cesarean delivery samples and virgin female mice. We evaluated the potency and efficacy of the analogs on uterine contraction, additivity with OT, and the ability to overcome the effects of atosiban, an OT antagonist. In human samples, the potency rank judged by the calculated EC50 (pM) was as follows: HPOT (189) > FBOT (556) > OT (5,340) > carbetocin (12,090). The calculated Emax was 86% for FBOT and 75% for HPOT (100%). Recovery from atosiban inhibition after HPOT treatment was as potent as that after OT treatment. HPOT showed additivity with OT. FBOT (56 pM) was found to be the strongest agonist in virgin mouse uterus. HPOT and FBOT demonstrated high potency and partial agonist efficacy in the human uterus. These results suggested that HPOT and FBOT are highly uterotonic for the human uterus and performed better than OT, indicating that they may prevent postpartum hemorrhage.

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“…Oxytocin-and vasopressin-activated receptors are coupled with G q/11 proteins and take part in regulating intracellular Ca 2+ contents and in activating protein kinase C [Kii et al, 2018]. In accordance with the modulation rules [Silkis, 2002], Avpr1b receptor agonists, like oxytocin, aid induction of LTP of the effi ciency of the synaptic inputs to PN in fi eld Ca 2+ , such that this NMDA-dependent LTP is linked Involvement of the Hypothalamic Nuclei [2014].…”

mentioning

confidence: 99%

Involvement of the Hypothalamic Nuclei in Forming Object–Place Associations in Neurons of Hippocampal Field CA2 (a hypothetical mechanism)

Силькис

2021

Neurosci Behav Physi

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A possible mechanism for the formation of representations of object-place associations in hippocampal neurons based on plastic rearrangements of the effi ciency of synaptic transmission, which depends on neuromodulators synthesized in neurons in various hypothalamic nuclei projecting to hippocampal fi eld CA2 is proposed. The afferent input from the supramammillary nucleus to fi eld CA2 promotes formation of representations of associations in neurons of fi elds CA2 and CA1, as it facilitates induction of long-term potentiation of transmission effi ciency in the CA2-CA1 pathway and summation of arousal arriving from fi elds CA3 and CA2 to CA1. Representations of object-place associations in fi eld CA2 include information on odors arriving in fi eld CA2 from the olfactory bulb via the paraventricular and supraoptic nuclei of the hypothalamus. Neurons in these nuclei synthesize vasopressin and oxytocin, which facilitate induction of long-term potentiation of synaptic inputs to pyramidal neurons in fi eld CA2, which promotes formation of representations of object-place associations on these neurons and their target cells in fi eld CA1, connected with the prefrontal cortex via the thalamic reuniens nucleus. As signals progress from the dentate gyrus through fi elds CA3 and CA2 to fi eld CA1, increasingly elaborate representations of object-place associations form in neurons in these areas. The unique nature of the connections of fi eld CA2 allow it to make a signifi cant contribution to encoding and memorizing not only contextual, but also social information. The consequences of the mechanism proposed here are consistent with results reported from experimental studies. An understanding of the mechanisms of hippocampus-dependent memory may be useful for targeted searches for drugs weakening those symptoms of Parkinson's disease and Alzheimer's disease induced by atrophy of pyramidal neurons in fi eld CA2.

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Quantification of receptor activation by oxytocin and vasopressin in endocytosis-coupled bioluminescence reduction assay using nanoKAZ

Cited by 6 publications

References 43 publications

Prolonged signaling of backbone-modified glucagon-like peptide-1 analogues with diverse receptor trafficking

Prolonged signaling of backbone-modified glucagon-like peptide-1 analogues with diverse receptor trafficking

Two oxytocin analogs, N-(p-fluorobenzyl) glycine and N-(3-hydroxypropyl) glycine, induce uterine contractions ex vivo in ways that differ from that of oxytocin

Involvement of the Hypothalamic Nuclei in Forming Object–Place Associations in Neurons of Hippocampal Field CA2 (a hypothetical mechanism)

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