CoChat
Sign Up
Cited by 17Neuroendocrine regulation and behaviorNeurotransmitter Receptor Influence on BehaviorNeuropeptides and Animal Physiology

Evidence for the existence of facilitatory interactions between the dopamine D2 receptor and the oxytocin receptor in the amygdala of the rat relevance for anxiolytic actions

Open access
Hernández-Mondragón, Juan Carlos, Hernández-Hernández, Dexter A., Crespo-Ramírez, Minerva, Prospero-García, Oscar, Rocha-Arrieta, Luisa, Fuxe, Kjell, Borroto-Escuela, Dasiel O., Perez de la Mora, Miguel·2023·Frontiers in Pharmacology
DOI 10.3389/fphar.2023.1251922PMC PMC10603234
Open this paper in CoChat — free accountOpen PDF

Abstract

Introduction: The amygdala is a limbic region of high value for understanding anxiety and its treatment. Dopamine D2 receptors (D2Rs) and oxytocin receptors (OXTRs) have both been shown to participate in modulating anxiety involving effects in the amygdala. The goal is to understand if D2R–OXTR heterocomplexes exist in the central amygdala and if, through enhancing allosteric receptor–receptor interactions, may enhance anxiolytic actions. Methods: The methods used involve the shock-probe burying test, the in situ proximity ligation assay (PLA), image acquisition and analysis, and the BRET2 assay. Bilateral cannulas were introduced into the amygdala, and the effects of the coadministration of oxytocin and the D2R-like agonist quinpirole into the amygdala were studied. Results: The combination treatment enhanced the anxiolytic effects compared to the single treatment. The D2R/D3R antagonist raclopride blocked the effects of the combination treatment of oxytocin and the D2R agonist, although oxytocin is regarded as a distinct modulator of fear-mediating anxiolytic effects. In situ PLA results indicate the existence of D2R–OXTR heteroreceptor complexes and/or the co-location of OXTR and D2R within the same cell membrane nanodomains in the central amygdala. With BRET2, evidence is given for the existence of D2R–OXTR heteromers in HEK293 cells upon co-transfection. Discussion: The enhanced behavioral effects observed upon co-treatment with OXTR and D2R agonists may reflect the existence of improved positive receptor–receptor interactions in the putative D2R–OXTR heterocomplexes in certain neuronal populations of the basolateral and central amygdala. The D2R–OXTR heterocomplex, especially upon agonist co-activation in the central amygdala, may open a new pharmacological venue for the treatment of anxiety.

AI snapshot

Written from the paper’s full text.

TL;DR

In male Wistar rats, intra-central amygdala co-administration of sub-effective oxytocin and the D2R-like agonist quinpirole produced anxiolytic-like effects in the shock-probe burying test, and this was blocked by the D2R/D3R antagonist raclopride. In situ PLA and BRET2 assays provided evidence for D2R–OXTR heterocomplexes/heteromers in the amygdala and in co-transfected HEK293T cells.

Key findings

  • Bilateral quinpirole infusion into the central amygdala dose-dependently reduced burying time (one-way ANOVA F3,42 = 3.9; p < 0.01), reaching significance at 0.3 µg/side vs control (p < 0.01), with no significant effect on burying latency and no effect on open-field locomotion.
  • Co-administration of sub-optimal oxytocin (3 ng/side) and quinpirole (0.15 µg/side) markedly decreased burying behavior vs control (Kruskal–Wallis F3,47 = 10.92; p < 0.05; Dunn’s test p < 0.05), whereas each alone had no significant effect; latency, shocks, and locomotion were not significantly changed.
  • Prior intra-amygdaloid raclopride (1 µg/side) blocked the anxiolytic-like effect of the oxytocin + quinpirole combination (Figure 4; p < 0.05), with no significant changes in latency, shocks, or locomotion.
  • In situ PLA showed D2R–OXTR puncta in CeA and BLA, with higher density in CeA (4.12 ± 0.47 puncta/nucleus/sample field) than BLA (2.19 ± 0.53); negative controls averaged 0.62 ± 0.04 and about 10% false-positive clusters. BRET2 in HEK293T cells showed saturable D2R–OXTR signals under vehicle (BRET2max 0.065 ± 0.002), increased by oxytocin (100 nM) + quinpirole (200 nM) to BRET2max 0.119 ± 0.007.

Methods

  • Male Wistar rats (250–270 g) received bilateral cannulas targeting the central nucleus of the amygdala; drugs were microinjected, and behavior was assessed with the shock-probe burying test followed by an open-field locomotion test.
  • In situ proximity ligation assay (PLA) used free-floating fixed rat brain sections at bregma −2.2 mm with anti-OXTR and anti-D2R antibodies, confocal imaging, and Duolink ImageTool quantification.
  • BRET2 assays used HEK293T cells co-transfected with D2R-Rluc and OXTR-GFP2 for saturation and ligand-induced BRET2 experiments; statistics used one-way ANOVA/Dunnett or Kruskal–Wallis/Dunn for non-normal data, with significance at p ≤ 0.05.

Limitations

  • The paper states that in situ PLA resolution is about 25–30 nm, so it cannot exclude detection of D2R–OXTR heteroreceptor complexes versus co-location of D2R and OXTR within the same cell membrane nanodomains.
  • The paper notes that increased BRET signal after oxytocin and quinpirole co-stimulation could also reflect co-location/co-presence in small endocytic vesicles, though this is considered less likely; negative controls showed about 10% false-positive PLA clusters.
  • Evident from the text: experiments used only male Wistar rats, behavioral conclusions rely mainly on the shock-probe burying test, group sizes were small (e.g., n = 7–15 in several experiments), and some statistical reporting appears inconsistent (e.g., raclopride ANOVA reported as F2,21 = 7.7 with p > 0.05 in text while Figure 4 indicates p < 0.05).

Model-written summary. Check it against the paper before relying on it.

Integrity signals

Registry

Crossref · 10.3389/fphar.2023.1251922

Cites

58 of 58 references verified0 citation errors · 0 not checkable

Statistics

Nothing to recompute.

Claims the paper makes

0 of 45 cited sources checked · 0 supported · 0 flagged

  • Findingown data

    The D2R/D3R antagonist raclopride blocked the effects of the combination treatment of oxytocin and quinpirole on burying behavior in the shock-probe burying test.

    “The D2R/D3R antagonist raclopride blocked the effects of the combination treatment of oxytocin and the D2R agonist”
    • Also stated in: § 3.1 Animal experiments · plain
      “a dramatic blockade of the co-administration of oxytocin and quinpirole by raclopride on the burying behavior in the shock-probe burying test (Figure 4).”
    Quote in the paperSubstantial claim
  • Findingown data

    In co-transfected cells, BRET2 signaling exhibited a hyperbolic response curve with increasing OXTR-GFP2 concentrations, reaching an asymptote at highest concentrations.

    “The BRET2 signaling in co-transfected cells exhibited a hyperbolic response curve with increasing concentrations of the OXTR–GFP2 fusion construct, reaching an asymptote at the highest concentrations tested.”
    Quote in the paperSubstantial claim
  • Findingown data

    Quinpirole (0.075-0.3 μg/side) into the CeA did not significantly affect the latency to bury the probe.

    “No significant effects were, however, observed in the latency to bury the probe (one-way ANOVA F3,42 = 1.2; p > 0.05).”
    Quote in the paperNot a substantial claim
  • Findingown data

    Co-administration of oxytocin and quinpirole into the CeA did not significantly affect the latency to the first burying episode.

    “No significant changes were instead observed by either oxytocin or quinpirole alone treatment or its combination on both the latency to the first burying episode (Kruskal–Wallis test F3,47 = 2.58; p > 0.05) and the number of shocks received by the rats during the test (Kruskal–Wallis test F3,47 = 2.58; p > 0.05).”
    Quote in the paperNot a substantial claim
  • Findingown data

    Locomotion activity in the open-field test was not significantly affected by any treatment (oxytocin, quinpirole, or their combination).

    “Likewise, locomotion activity in the open-field test was not significantly affected by any treatment (Kruskal–Wallis test F3,47 = 2.58; p > 0.05).”
    Quote in the paperSubstantial claim
  • May combine two claims. 20
  • Classified as probably not a substantial claim. 15

  • May combine two claims.

    Findingown data

    Co-administration of oxytocin (3 ng/side) and quinpirole (0.15 μg/side) into the central nucleus of the amygdala decreased burying behavior in the shock-probe burying test relative to control, whereas either treatment alone did not significantly change burying behavior.

    “The combination treatment enhanced the anxiolytic effects compared to the single treatment.”
    • Also stated in: § 3.1 Animal experiments · boosted
      “its combination triggered a remarkable decrease in burying activity, relative to the control group (Figure 3).”
    • Also stated in: § 3.1 Animal experiments · plain
      “although no significant changes in burying behavior were induced by the administration of low doses of both quinpirole and oxytocin”
    Quote in the paperSubstantial claim
    • Rests on an assumption: Quinpirole and oxytocin act selectively on D2R and OXTR, respectively, at the doses used.

  • May combine two claims.

    Findingown data

    Quinpirole (0.075-0.3 μg/side) into the CeA reduced the time spent burying the electrified probe, with a dose-dependent trend that reached statistical significance only at the highest dose (0.3 μg/side) compared to control.

    “A main effect of quinpirole was observed (one-way ANOVA F3,42 = 3.9; p < 0.01) on the time rats spent burying the electrified probe of the test. The reductive effect seems to be dose-dependent (Figure 2), although it only reached statistical significance (p < 0.01) at the higher dose used concerning the control group (Dunnett’s test).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    BRET2 assays on HEK293T cells co-transfected with D2R-Rluc and OXTR-GFP2 demonstrated the presence of D2R-OXTR heteromers, with significant, strong, saturable BRET2 signals under vehicle-treated conditions (BRET2max 0.065 ± 0.002).

    “As shown in Figure 6A, we observed significant, strong, and saturable BRET2 signals for the D2R–OXTR complex under vehicle-treated conditions (BRET2max of 0.065 ± 0.002).”
    • Also stated in: § 4 Discussion · boosted
      “In our current study, we successfully demonstrated the presence of D2R–OXTR heteromers in HEK293 cells using the BRET2 method.”
    Quote in the paperSubstantial claim
    • Rests on an assumption: The BRET2 signal reflects direct physical interaction (heterodimerization) between D2R and OXTR.

  • May combine two claims.

    Findingown data

    In situ PLA indicates the existence of D2R-OXTR heteroreceptor complexes and/or co-localization of OXTR and D2R within the same cell membrane nanodomains in the central amygdala and basolateral amygdala.

    “In situ PLA results indicate the existence of D2R–OXTR heteroreceptor complexes and/or the co-location of OXTR and D2R within the same cell membrane nanodomains in the central amygdala.”
    • Also stated in: § 3.2 Neurochemical experiments · hedged
      “While our in situ PLA data indicate the existence of D2R–OXTR heteroreceptor complexes and/or the co-localization of OXTR and D2R receptors within the same cell membrane nanodomains in both the CeA and BLA regions of the amygdala”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    D2R-OXTR heteroreceptor complexes exist in the rat amygdala, with moderate densities in the central amygdala and low densities in the basolateral amygdala, as indicated by in situ PLA.

    “Detection of D2R–OXTR heteroreceptor complexes in the different subfields of rat amygdala by in situ proximity ligation assay (in situ PLA).”
    • Also stated in: § 4 Discussion · hedged
      “D2R–OXTR heterocomplexes were indicated to exist using in situ PLA, in moderate densities in the central amygdala, and in low densities in the basolateral amygdala.”
    Quote in the paperSubstantial claim
    • Rests on an assumption: The in situ PLA signal specifically detects D2R–OXTR heteroreceptor complexes.

  • May combine two claims.

    Findingown data

    No changes in latency to burying were observed in any experiments, and only a modest but not reproducible increase in number of shocks was observed with a single intermediate dose of quinpirole.

    “No changes in latency to burying were observed in any of our experiments, and only a modest but otherwise not reproducible increase was observed in the number of shocks elicited by a single intermediate dose of quinpirole”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation

    Local infusion of D2R antagonists triggers anxiogenic nonconditioned responses and a reduction in contextual conditioned fear.

    “Thus, it has been demonstrated that local infusion of D2R antagonists triggers both anxiogenic nonconditioned responses (Perez de la Mora et al., 2012) and a reduction in contextual conditioned fear (de Souza Caetano et al., 2013).”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites Perez de la Mora et al., 2012 source

      Not checked in the free run.

    • cites de Souza Caetano et al., 2013 source

      Not checked in the free run.

  • Classified as probably not a substantial claim.

    Study design and conductcitation

    The primary antibodies used for in situ PLA were previously validated for immunohistochemistry in rat brain tissue and HEK293 cell lines.

    “These primary antibodies were previously validated for immunohistochemistry in rat brain tissue and HEK293 cell lines ((Borroto-Escuela et al., 2017b; Borroto-Escuela et al., 2018; Feltmann et al., 2018)).”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites Borroto-Escuela et al., 2017b

      Not checked in the free run.

    • cites Borroto-Escuela et al., 2018 source

      Not checked in the free run.

    • cites Feltmann et al., 2018 source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    Modulating D2Rs in the central amygdala plays a vital role in fear response consolidation, with stimulation aiding discriminative learning and blockade inducing generalized threat responses.

    “Modulating D2Rs in the central amygdala plays a vital role in fear response consolidation, with stimulation aiding discriminative learning and blockade of inducing generalized threat responses (De Bundel et al., 2016).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites De Bundel et al., 2016 source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductreasoning

    Ketamine was unlikely to produce differential effects because it was administered to both experimental and control rats.

    “It is worth noting that ketamine, used as an anesthetic agent in this study, was unlikely to produce differential effects since it was administered to both the experimental and control rats.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    PET imaging shows that elevated D2R levels contribute to amygdala dysfunction under stress.

    “PET imaging shows that elevated D2R levels contribute to amygdala dysfunction under stress (Yan et al., 2021).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Yan et al., 2021 source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Interpretationcitation

    An increase in the BRET signal after OXT and quinpirole co-stimulation could also be explained by co-location/co-presence of OXTR and D2R in small endocytic vesicles, but this is considered less likely.

    “An increase in the BRET signal after OXT and quinpirole co-stimulation could also be explained by the co-location/co-presence of OXTR and D2R in small endocytic vesicles (Gomes et al., 2016). However, while this possibility cannot be excluded, it is considered less likely in view of the experimental data reported previously (Romero-Fernandez et al., 2013).”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites Gomes et al., 2016 source

      Not checked in the free run.

    • cites Romero-Fernandez et al., 2013 source

      Not checked in the free run.

  • May combine two claims.

    Interpretationown data

    D2R-OXTR dimers are formed constitutively and activation or inhibition of each protomer by agonist or antagonist treatments does not affect their oligomerization state.

    “Surprisingly, none of the administered ligands consistently induced changes in the BRET2 ratio, suggesting that the dimers are formed constitutively and that the activation or inhibition of each protomer by agonist or antagonist treatments does not affect their oligomerization state (Figure 6B).”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductown data

    In situ PLA quantification was performed by an experimenter blind to the experiment conditions.

    “PLA was quantified as PLA (red puncta/blobs) per nuclei per field by an experimenter blind to the experiment conditions.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Study design and conductnone given

    The rats were randomly assigned to different groups and tested in a randomized manner.

    “The rats were randomly assigned to different groups and tested in a randomized manner.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S4 (based).

    Premisereasoning

    The allosteric receptor-receptor interactions within OXTR heteroreceptor complexes allow for bidirectional modulation between the OXTR protomer and other participating receptor protomers such as D2R.

    “The exploration of these heteroreceptor complexes will significantly contribute to oxytocin research, as the allosteric receptor–receptor interactions within OXTR heteroreceptor complexes allow for bidirectional modulation between the OXTR protomer and other participating receptor protomers such as D2R.”
    Quote in the paperNot a substantial claim

  • Classified as probably not a substantial claim.

    Premisecitation

    GPCR heteroreceptor complexes exert allosteric receptor-receptor interactions that modulate the recognition, signaling, and trafficking of the participating receptor protomers.

    “These complexes exert allosteric receptor–receptor interactions that modulate the recognition, signaling, and trafficking of the participating receptor protomers, thereby influencing other associated proteins (Fuxe et al., 2010; Borroto-Escuela et al., 2014; Fuxe et al., 2014; Borroto-Escuela et al., 2015b; Fuxe and Borroto-Escuela, 2016; Borroto-Escuela et al., 2017a).”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites Fuxe et al., 2010 source

      Not checked in the free run.

    • cites Borroto-Escuela et al., 2014 source

      Not checked in the free run.

    • cites Fuxe et al., 2014 source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Study design and conductcitation

    In situ PLA has a resolution of 25-30 nm, so the authors cannot exclude that they may detect D2R-OXTR heteroreceptor complexes and/or co-location within the same cell membrane nanodomains.

    “As in situ PLA typically provides a resolution in the range of 25–30 nm when secondary IgG antibodies are used (Soderberg et al., 2006; Soderberg et al., 2007), we cannot exclude the possibility that in our current experiments, we may detect D2R–OXTR heteroreceptor complexes and/or the co-location of D2R and OXTR within the same cell membrane nanodomains (hereafter referred to as heterocomplexes).”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites Soderberg et al., 2006 source

      Not checked in the free run.

    • cites Soderberg et al., 2007 source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Oxytocin is a critical modulator of fear and anxiety and in the vast majority of cases elicits anxiolytic effects.

    “Oxytocin, on its side, has been widely demonstrated to be a critical modulator of fear and anxiety (Jurek and Neumann, 2018), which in the vast majority of cases, elicits anxiolytic effects (see Jang et al. (2021); Chun et al. (2022)).”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites Jurek and Neumann, 2018 source

      Not checked in the free run.

    • cites Jang et al. (2021) source

      Not checked in the free run.

    • cites Chun et al. (2022) source

      Not checked in the free run.

  • May combine two claims.

    Interpretationcitation

    The paper proposes that D2R-OXTR heterocomplexes may exist in the basolateral amygdala and in the central nucleus of the amygdala, including the contralateral (CeL) and centro medial (CeM) amygdala.

    “We propose that in the basolateral amygdala (Perez de la Mora et al., 2012) as well as in the central nucleus of the amygdala, including the contralateral (CeL) and centro medial (CeM) amygdala, D2R–OXTR heterocomplexes may exist.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Perez de la Mora et al., 2012 source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

    Interpretationnone given

    These potential D2R-OXTR heterocomplexes may predominantly modulate the signaling of GABA interneurons and projection neurons.

    “These potential D2R–OXTR heterocomplexes may predominantly modulate the signaling of GABA interneurons and projection neurons.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Intra-BLA D2R stimulation by quinpirole elicited anxiogenic effects in the elevated plus maze test.

    “in the work of Bananej et al. (2012), the intra-BLA D2R stimulation by quinpirole (Eilam and Szechtman, 1989) elicited anxiogenic effects in the elevated plus maze test”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites Bananej et al., 2012 source

      Not checked in the free run.

    • cites Eilam and Szechtman, 1989

      Not checked in the free run.

  • May combine two claims.

    Interpretationown data

    The reduction in burying behavior is the most sensitive measure of anxiety observed in our work and is consistent with the triggering of anxiolytic effects.

    “The reduction in burying behavior (the most sensitive measure of anxiety observed in our work) is consistent with the triggering of anxiolytic effects.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data

    The interaction between D2R and OXTR in the amygdala is facilitatory, as ineffective doses of quinpirole and oxytocin potentiate each other to elicit anxiolytic effects.

    “such interaction seems to be of a facilitatory nature since, as indicated before, ineffective doses of both quinpirole and oxytocin potentiate each other to elicit anxiolytic effects.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S2 (checkable), S3 (load-bearing), S4 (based).

    Interpretationreasoning

    Such interactions may occur through allosteric changes at the interface of the potential D2R-OXTR heterocomplexes.

    “Such interactions may occur through allosteric changes occurring at the interface of the potential D2R–OXTR heterocomplexes.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation

    Co-injection of oxytocin with raclopride and prior raclopride administration prevented oxytocin-induced anxiolytic effects.

    “both the co-injection of oxytocin with raclopride (de la Mora et al., 2016) and the previous raclopride administration (Laszlo et al., 2020) prevented the oxytocin-induced anxiolytic effects”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites de la Mora et al., 2016 source

      Not checked in the free run.

    • cites Laszlo et al., 2020 source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    D2R blockade within CeA in the shock-probe burying test and elevated plus maze test produced anxiogenic and lack of effects, respectively, in our work.

    “although both anxiogenic and lack of effects have been reported in our work (Perez de la Mora et al., 2012) following the D2R blockade within the CeA in the shock-probe burying test and the elevated plus maze test, respectively”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Perez de la Mora et al., 2012 source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    D2R-OXTR heterocomplexes were previously shown to occur in the nucleus accumbens shell and dorsal striatum of the rat in terms of D2R signaling and recognition.

    “Previously, heterocomplexes formed by D2R and OXTR protomers were shown to take place in the nucleus accumbens shell and the dorsal striatum of the rat in terms of D2R signaling and recognition (Romero-Fernandez et al., 2013; Borroto-Escuela et al., 2022).”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites Romero-Fernandez et al., 2013 source

      Not checked in the free run.

    • cites Borroto-Escuela et al., 2022 source

      Not checked in the free run.

  • May combine two claims.

    Interpretationreasoning

    Enhanced activity in inhibitory amygdala GABA projection neurons to the brainstem could be an important mechanism involved in inhibiting local GABA interneurons, setting free increased activation in monoamine ascending pathways, reducing stress and anxiety.

    “it is proposed that an enhanced activity in the inhibitory amygdala GABA projection neurons to the brainstem could be an important mechanism involved in inhibiting local GABA interneurons in this region, setting free increased activation in, e.g., monoamine ascending pathways from the lower brainstem and the hypothalamus.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Interpretationcitation

    The potential receptor-receptor interactions may involve an enhancing effect of D2R-OXTR heterocomplexes on CREB, MAPK, and PLC signaling pathways.

    “it may also be possible that in agreement with de la Mora et al. (2016), the potential receptor–receptor interactions may involve an enhancing effect of D2R–OXTR heterocomplexes on CREB, MAPK, and PLC signaling pathways”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites de la Mora et al., 2016 source

      Not checked in the free run.

  • May combine two claims.

    Interpretationown data

    The paper's findings strongly indicate the existence of D2R-OXTR heteroreceptor complexes and/or populations of OXTRs and D2Rs co-located within the same cell membrane nanodomains in the amygdala.

    “Our findings strongly indicate the existence of D2R–OXTR heteroreceptor complexes and/or populations of OXTRs and D2Rs co-located within the same cell membrane nanodomains.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data

    The findings provide robust indications for the existence of D2R-OXTR heterocomplexes and highlight their potential functional significance in cellular signaling.

    “These findings provide robust indications for the existence of D2R–OXTR heterocomplexes and highlight their potential functional significance in cellular signaling.”
    Quote in the paperSubstantial claim
32 of 32

  • The paper relies on this without support.

    The D2R/D3R antagonist raclopride's blockade of the combination treatment is attributable to D2R antagonism rather than D3R antagonism.

    “The D2R/D3R antagonist raclopride blocked the effects of the combination treatment of oxytocin and the D2R agonist”
    “raclopride (1 µg/side), a D2R/D3R dopamine receptor antagonist”

  • The paper relies on this without support.

    Quinpirole and oxytocin act selectively on D2R and OXTR, respectively, at the doses used.

    “quinpirole (0.075–0.3 µg/side) or saline”
    “OXT (3 ng/side)”

  • The paper relies on this without support.

    The in situ PLA signal specifically detects D2R–OXTR heteroreceptor complexes.

    “Detection of D2R–OXTR heteroreceptor complexes in the different subfields of rat amygdala by in situ proximity ligation assay (in situ PLA).”
    “D2R–OXTR heterocomplexes were indicated to exist using in situ PLA, in moderate densities in the central amygdala, and in low densities in the basolateral amygdala.”

  • The paper relies on this without support.

    The BRET2 signal reflects direct physical interaction (heterodimerization) between D2R and OXTR.

    “BRET2 analysis was conducted to examine the heterodimerization of D2R and OXTR receptors.”
    “In our current study, we successfully demonstrated the presence of D2R–OXTR heteromers in HEK293 cells using the BRET2 method.”

  • The paper relies on this without support.

    Raclopride is a selective D2R/D3R antagonist.

    “raclopride, a selective D2R/D3R antagonist.”

  • The paper relies on this without support.

    Quinpirole is a D2R-like agonist.

    “quinpirole, a D2R-like agonist”
6 of 6

Claims other papers make about this one

Each is checked against this paper’s own text: does it say what the citing paper says it does?

  • In the striatum, OXTR activation on astrocytes inhibits glutamate release, and D2R-OXTR heterocomplexes facilitate dopaminergic signaling.

    “In the striatum, OXTR activation on astrocytes inhibits glutamate release (Amato et al., 2022), and D2R-OXTR heterocomplexes facilitate dopaminergic signaling (Hernandez-Mondragon et al., 2023; Pérez de la Mora et al., 2016).”

    From Music's context-dependent influence on oxytocin, social bonding, and emotion regulation: a systematic review (2026) · cites this paper as Hernandez-Mondragon et al., 2023

    The Discussion (page unavailable) reports D2R–OXTR heterocomplexes in the rat dorsal striatum in connection with D2R signaling, but does not say they facilitate signaling there. Its evidence for enhanced interactions concerns the amygdala and co-transfected cells. The astrocyte claim is attached to a different citation and is not assessed here. (LLM, conf=0.84)

    Experimental — accuracy not yet measured

Self-check

Percentages, sums and reported differences re-computed from the paper’s own numbers.

Each contradiction between two passages listed here was confirmed by an independent check.

  • BRET2 experiment counts conflict

    Major

    The BRET2 methods report three independent experiments, but the Figure 6B caption says its ratios came from at least six experiments.

  • Latency result is nonsignificant but reports p < 0.05

    Minor

    The text says there was no statistically significant change in burying latency, while its accompanying test reports p < 0.05.

  • Shock-count result is nonsignificant but reports p < 0.05

    Minor

    The text says the number of shocks did not change significantly, while the reported ANOVA gives p < 0.05.

  • PLA brain-section bregma levels conflict

    Minor

    The PLA methods place the rat brain sections at −2.2 mm, while the caption for the PLA images specifies 1.00 mm.

  • Raclopride effect is called significant despite p > 0.05

    Minor

    The reported ANOVA p-value is greater than 0.05, but the text calls the effect statistically significant and the figure caption reports p < 0.05 for the raclopride effect.

  • Locomotion result reports no effect alongside p < 0.05

    Minor

    The text reports no locomotion effect, but gives p < 0.05 for its ANOVA.

  • BRET2 drug-incubation times differ

    Minor

    The methods specify 15 minutes of drug incubation before BRET2 measurement, whereas the Figure 6 caption specifies 20 minutes of stimulation.

7 of 7

Cites

Flagged first, then by citations

Cited by

Sorted by citations

Related

Authors
Hernández-Mondragón, J. C., Hernández-Hernández, D. A., Crespo-Ramírez, M., Prospero-García, O., Rocha-Arrieta, L., Fuxe, K., Borroto-Escuela, D. O., & Perez de la Mora, M.
Affiliations
  • Universidad Nacional Autónoma de México (MX)
  • Centro de Investigación y de Estudios Avanzados del Instituto Politécnico Nacional (MX)
ORCID
Hernández-Mondragón, Juan Carlos
Venue
Frontiers in Pharmacology
Publisher
Frontiers Media SA
Published
2023-10-12
Volume
14
Pages
1251922-1251922
ISSN
1663-9812 (ISSN-L)
DOI
10.3389/fphar.2023.1251922
PMC
PMC10603234
OpenAlex
W4387568593
Semantic Scholar
188fc4b1009b6932c6a4d0f91b770180c49fd5be
Cited by
17
References
58
Citations by year
2026: 7 · 2025: 6 · 2024: 4
FWCI
5.03
Type
journal article
Language
en
Open access
Licence
Locations
  • Frontiers in Pharmacology (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
Funders
  • Olle Engkvists Stiftelse
  • Hjärnfonden
Topics
  • Social Sciences › Psychology › Social Psychology › Neuroendocrine regulation and behavior
  • Life Sciences › Neuroscience › Cellular and Molecular Neuroscience › Neurotransmitter Receptor Influence on Behavior
  • Life Sciences › Neuroscience › Cellular and Molecular Neuroscience › Neuropeptides and Animal Physiology
Keywords
dopamine D2 receptor, oxytocin receptor, receptor heteromers, amygdala, anxiety, anxiolytic effects, quinpirole, oxytocin, raclopride, allosteric interactions, basolateral amygdala, central amygdala, membrane nanodomains, proximity ligation assay

About these checks

Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.

Claims read by deepseek-v4.1-flash under definition version 3 (extract-v3) on 2026-10-10.

Classified by jev-1.13-20260917 (gold-guideline-v0.2) on 2026-10-10.

Statistics and arithmetic are recomputed, not judged.

Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.

What counts as a claim ›How we measure confidence ›How we check this ›

Are you an author, or do you see a problem with this page? Request a review