The interactions between TRPV1 and µ-opioid receptors (MOR) have recently attracted much attention because these two receptors play important roles in pain pathways and can apparently modulate each other's functioning. However, the knowledge about signaling interactions and crosstalk between these two receptors is still limited. In this study, we investigated the mutual interactions between MOR and TRPV1 shortly after their activation in HEK293 cells expressing these two receptors. After activation of one receptor we observed significant changes in the other receptor's lateral mobility and vice versa. However, the changes in receptor movement within the plasma membrane were not connected with activation of the other receptor. We also observed that plasma membrane β-arrestin 2 levels were altered after treatment with agonists of both these receptors. Knockdown of β-arrestin 2 blocked all changes in the lateral mobility of both receptors. Furthermore, we found that β-arrestin 2 can play an important role in modulating the effectiveness of ERK1/2 phosphorylation after activation of MOR in the presence of TRPV1. These data suggest that β-arrestin 2 and ERK1/2 are important mediators between these two receptors and their signaling pathways. Collectively, MOR and TRPV1 can mutually affect each other's behavior and β-arrestin 2 apparently plays a key role in the bidirectional crosstalk between these two receptors in the plasma membrane.

• Klíčová slova
• ERK1/2, TRPV1, biased signaling, receptor lateral mobility, β-arrestin 2, μ-opioid receptor,
• MeSH
• arrestiny metabolismus   MeSH
• beta arrestin 2 metabolismus   fyziologie   MeSH
• beta arrestiny metabolismus   MeSH
• buněčná membrána metabolismus   fyziologie   MeSH
• fosforylace MeSH
• HEK293 buňky MeSH
• kationtové kanály TRPV metabolismus   fyziologie   MeSH
• lidé MeSH
• MAP kinasový signální systém fyziologie   MeSH
• morfin metabolismus   MeSH
• opioidní analgetika metabolismus   MeSH
• receptory opiátové mu metabolismus   fyziologie   MeSH
• receptory opiátové metabolismus   MeSH
• signální transdukce MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• arrestiny MeSH
• beta arrestin 2 MeSH
• beta arrestiny MeSH
• kationtové kanály TRPV MeSH
• morfin MeSH
• opioidní analgetika MeSH
• receptory opiátové mu MeSH
• receptory opiátové MeSH
• TRPV1 protein, human MeSH Prohlížeč

There are some indications that biased μ-opioid ligands may diversely affect μ-opioid receptor (MOR) properties. Here, we used confocal fluorescence recovery after photobleaching (FRAP) to study the regulation by different MOR agonists of receptor movement within the plasma membrane of HEK293 cells stably expressing a functional yellow fluorescent protein (YFP)-tagged μ-opioid receptor (MOR-YFP). We found that the lateral mobility of MOR-YFP was increased by (D-Ala2,N-MePhe4,Gly5-ol)-enkephalin (DAMGO) and to a lesser extent also by morphine but decreased by endomorphin-2. Interestingly, cholesterol depletion strongly enhanced the ability of morphine to elevate receptor mobility but significantly reduced or even eliminated the effect of DAMGO and endomorphin-2, respectively. Moreover, the ability of DAMGO and endomorphin-2 to influence MOR-YFP movement was diminished by pertussis toxin treatment. The results obtained by agonist-stimulated [35S]GTPγS binding assays indicated that DAMGO exhibited higher efficacy than morphine and endomorphin-2 did and that the efficacy of DAMGO, contrary to the latter agonists, was enhanced by cholesterol depletion. Overall, our study provides clear evidence that biased MOR agonists diversely affect receptor mobility in plasma membranes as well as MOR/G protein coupling and that the regulatory effect of different ligands depends on the membrane cholesterol content. These findings help to delineate the fundamental properties of MOR regarding their interaction with biased MOR ligands and cognate G proteins.

• Klíčová slova
• Biased agonists *, Cholesterol *, FRAP *, G protein coupling *, μ-Opioid receptor *,
• MeSH
• bakteriální proteiny genetika   metabolismus   MeSH
• buněčná membrána účinky léků   metabolismus   MeSH
• cholesterol nedostatek   MeSH
• enkefalin, Ala(2)-MePhe(4)-Gly(5)- metabolismus   farmakologie   MeSH
• FRAP MeSH
• guanosin 5'-O-(3-thiotrifosfát) metabolismus   MeSH
• HEK293 buňky MeSH
• konfokální mikroskopie MeSH
• lidé MeSH
• ligandy MeSH
• luminescentní proteiny genetika   metabolismus   MeSH
• morfin metabolismus   farmakologie   MeSH
• narkotika - antagonisté farmakologie   MeSH
• oligopeptidy metabolismus   farmakologie   MeSH
• pertusový toxin farmakologie   MeSH
• proteiny vázající GTP - alfa-podjednotky Gi-Go metabolismus   MeSH
• receptory opiátové mu agonisté   genetika   metabolismus   MeSH
• rekombinantní fúzní proteiny metabolismus   MeSH
• transfekce MeSH
• transport proteinů MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• srovnávací studie MeSH
• Názvy látek
• bakteriální proteiny MeSH
• cholesterol MeSH
• endomorphin 2 MeSH Prohlížeč
• enkefalin, Ala(2)-MePhe(4)-Gly(5)- MeSH
• guanosin 5'-O-(3-thiotrifosfát) MeSH
• ligandy MeSH
• luminescentní proteiny MeSH
• morfin MeSH
• narkotika - antagonisté MeSH
• oligopeptidy MeSH
• pertusový toxin MeSH
• proteiny vázající GTP - alfa-podjednotky Gi-Go MeSH
• receptory opiátové mu MeSH
• rekombinantní fúzní proteiny MeSH
• yellow fluorescent protein, Bacteria MeSH Prohlížeč