[Mechanism of vascular desensitization to agonists].
1
Citation
0
Reference
10
Related Paper
Citation Trend
Abstract:
The patterns and mechanism of vascular desensitization to agonists were studied in isolated arteries of Wistar and SHR rats. The results showed that: (1) Arteries were much easier to desensitize peptidergic agonists (A II, AVP) than catecholaminer gic agonists (NE, PE); (2) the desensitization induced by peptidergic agonists and catecholaminer gic agonists were homologous within 30 minutes; (3) the endothelium-EDRF-cGMP pathway was not involved in agonist-induced desensitization; (4) inhibition of surface receptor internalization by phenylarsine oxide (PAO, 10(-5)mol/L) could transiently reverse the desensitization and the blocking of intracellular receptor re-insertion into the membrane by chloroauine (10(-5)mol/L) could accelerate the desensitization in varying degrees with different arteries, which demonstrated the involvement of receptor cycling in homologous desensitization, but was not the unique mechanism; (5) there was no change in the function of voltage-dependent Ca2+ channel during homologous desensitization; (6) the concentration-contraction curve of GTP gamma S shifted down-ward during homologous desensitization, suggesting the functional change of G protein (s); (7) the desensitization to agonists didn't decrease in arteries of SHR compared with the arteries of Wistar rats.Keywords:
Homologous desensitization
Internalization
Cite
In this study we examined agonist-induced internalization of the cloned human nociceptin receptor (hNOP) expressed in CHOK1 cells. Internalization was proven by receptor binding assay on viable cells and confocal microscopy. The agonists nociceptin/orphanin FQ (NC), NC-NH2, NC(1-13)-NH2, [(pF)Phe4]NC-NH2 and RO 64-6198 promote a rapid, concentration-dependent internalization of the hNOP receptor. Under the same conditions, [Phe1,ψ (CH2NH)Gly2]NC(1-13)-NH2 and [Phe1, (CH2NH)Gly2,Arg14,Lys15]NC(1-13)-NH2 failed to induce significant, concentration-dependent NOP receptor endocytosis; even when present at high concentrations (up to 1 mM) they promoted only an approximately 25-30% internalization of hNOP receptors. We also investigated hNOP receptor desensitization upon agonist challenge: ligand efficacy to inhibit forskolin-stimulated cAMP production. After 1 h exposure to NC, NC-NH2, NC(1-13)- NH2, [(pF)Phe4]NC-NH2 and RO 64-6198 (5 μM) ≉20 to 30% of receptor desensitization was observed. Moreover, we found that the blockade of hNOP receptor recycling by monensin would cause a more prolonged and relevant desensitization of this receptor. The noninternalizing agonists [Phe1,ψ (CH2NH)Gly2]NC(1-13)-NH2 and [Phe1, (CH2NH)Gly2,Arg14,Lys15]NC(1-13)-NH2 (100 μM) resulted in a strong (67 and 74 %, respectively) receptor desensitization which was not influenced by monensin. Finally, CHO-hNOP cells exposed to the receptor-internalizing agonists for 24 h resulted in a significantly higher cAMP accumulation (defined supersensitization) compared with the non-internalizing agonists. In addition, blocking of receptor recycling by monensin led to a decrease of the cAMP accumulation only in cells exposed to internalizing agonists. These data show that prolonged receptor signaling mediated by receptor endocytosis and recycling/reactivation might reduce the development of tolerance but can enhance compensatory mechanisms that lead to supersensitivity of specific signaling pathways. Keywords: Internalization, recycling, CHO-K1 cells, nociceptin, nociceptin receptor, desensitization, cAMP
Internalization
Nociceptin receptor
NOP
Homologous desensitization
Cite
Citations (23)
A perifusion method consisting of dispersed chicken anterior pituitary cells suspended in columns of Bio-Gel was developed to monitor the dynamics of LH release. The perifused cells responded to chicken I GnRH (Gln8-GnRH) in a dose-dependent manner. The ED50 was 3 × 10−10m, and maximal LH release occurred in response to 4 × 10−9m Gln8-GnRH. Continuous administration of 10−7m Gln8-GnRH and agonist stimulated an initial 8- to 10-fold increase in LH release within minutes. LH release then declined rapidly, reaching basal levels within 100 min. A biphasic response was noted. Calcium ionophore A23187 was effective in releasing additional LH from cells desensitized to 10−7m Gln8-GnRH and agonist, indicating that total cellular LH was not depleted. In contrast, delivery of 2-min pulses of 10−7m and 10−9m Gln8-GnRH at a frequency of one pulse every 30 or 60 min for 3–5 h maintained pituitary responsiveness. Exposure to 10−7m Gln8-GnRH for 20 min was sufficient to desensitize pituitary cells to subsequent Gln8-GnRH stimulation. However, 20-min exposure to 10−7m GnRH antagonist neither evoked LH release nor had a desensitizing effect on subsequent stimulation by 10−7m Gln8-GnRH, indicating that receptor activation, not merely receptor binding, is necessary for Gln8-GnRH-mediated homologous desensitization. Pituitary cells desensitized by 20-min exposure to 10−8m Gln8- GnRH maintained responsiveness to a higher dose (10−6m) of Gln8-GnRH, suggesting that down-regulation of pituitary GnRH receptors might play a part in desensitization. Calcium ionophore A23187 partially desensitized pituitary cells to subsequent stimulation with Gln8-GnRH, probably due to depletion of releasable LH or desensitization of calcium-coupled secretory mechanisms. In calcium-free medium, 10−7m Gln8-GnRH did not evoke LH release, but nevertheless partially desensitized cells to subsequent 10−7m Gln8-GnRH stimulation. Thus desensitization is partially calcium-dependent. These findings demonstrate that the phenomenon of GnRH-mediated desensitization of gonadotrophs is a characteristic of chicken pituitary cells as in the mammal. However, chicken pituitary cells differ from mammalian cells in that desensitization is more rapid and partially dependent on extracellular calcium. (Endocrinology119: 1510–1518, 1986)
Homologous desensitization
Cite
Citations (40)
Following agonist binding, neurokinin-1 receptors undergo rapid desensitization followed by internalization and recycling. Desensitization requires receptor phosphorylation but does not require internalization, whereas resensitization is thought to require internalization and recycling. Our previous data, however, have suggested that, following activation and desensitization, the return of responsiveness to the neurokinin-1 agonist substance P (termed "resensitization") occurs hours before internalized receptors are recycled back to the plasma membrane. To further investigate this novel mechanism of neurokinin-1 receptor resensitization, we have studied the time courses of neurokinin-1 receptor responsiveness, recycling, and dephosphorylation by measuring cellular Ca(2+) responses, ligand-receptor binding, and receptor phosphorylation, respectively. Concentration-response curves and competition binding curves were obtained at various times following desensitization. The effects of the nonhydrolyzable GTP analog Gpp(NH)p on substance P binding were also studied to assess receptor-G protein coupling. After receptor activation and desensitization, Ca(2+) signaling in response to substance P occurred within 90 min, whereas the return of receptor binding required 240 min. Receptor dephosphorylation was greater than 90% complete 20 min after agonist washout. In addition, the return of substance P responsiveness coincided with a return in sensitivity of substance P binding to Gpp(NH)p, indicating a return in receptor-G protein coupling. These data show that the resensitization of responsiveness to substance P precedes receptor recycling. This may result from a conversion of nonfunctional neurokinin-1 receptors to functional receptors at the plasma membrane.
Tachykinin receptor 1
Cite
Citations (12)
Many G protein-coupled receptors (GPCRs) are known to internalize following agonist exposure, however the relative importance of this mechanism for the desensitization and resensitization of different GPCRs is unclear. In the present study, we have pretreated NG108-15 cells with hypertonic sucrose or concanavalin A (con A), to investigate the effects of these inhibitors of internalization on the agonist-induced desensitization and subsequent resensitization of three Gs-coupled receptor responses. Incubation of cells with sucrose or con A did not affect subsequent acute agonist stimulation of the A2A adenosine receptor or the agonist-induced desensitization of this receptor response. However, the resensitization of the A2A adenosine receptor response following agonist removal was abolished in the presence of sucrose or con A. Sucrose or con A treatment affected neither the desensitization nor resensitization of IP-prostanoid receptor responsiveness. On the other hand con A but not sucrose reduced the agonist-induced desensitization of secretin receptor responsiveness. However, secretin receptor responsiveness did not resensitize within the time period studied whether or not inhibitors of internalization were present. These results indicate that receptor internalization appears to subserve different functions for different GPCRs.
Internalization
Homologous desensitization
Cite
Citations (48)
We injected hCG into pseudopregnant rabbits on day 7 of pseudopregnancy and analyzed changes in the components of luteal adenylyl cyclase system in order to determine which components are responsible for altered hormonal responsiveness upon desensitization. hCG-induced desensitization was homologous (loss of responsiveness to LH) early (first 6 h), then became heterologous (partial loss of responsiveness to catecholamines) later (12-48 h). The total number of LH receptors was reduced approximately 30% 3 h after treatment at a time when LH stimulation of adenylyl cyclase activity was not altered. Total LH receptors remained at this level until 24 h, when total receptors were reduced by 88%. While total LH receptor number remained constant, LH-stimulated adenylyl cyclase activity was declining to 57% of the control value at 12 h. Available unoccupied LH receptors were reduced by 96% at 12 h. The affinity of the occupied receptors was reduced 4-fold before down-regulation. The changes in β-adrenergic receptor number paralleled the changes in catecholamine responsiveness. hCG treatment also altered luteal G-protein function, as assessed by reconstitution of adenylyl cyclase activity in S49 cyc- lymphoma membranes and ADP ribosylation by cholera and pertussis toxins. Isoproterenol (ISO)-reconstituting activities of luteal G8 (the stimulatory G-protein of adenylyl cyclase) were depressed by 65% 12-48 h after hCG treatment, the same time as reduced catecholamine responsiveness. In contrast, NaFreconstituting activites were at control levels at 12 h and reduced by 55% at 24 and 48 h. Pertussis toxin's ability to ADP ribosylate ai 40 was increased 3 and 6 h after treatment, while cholera toxin's ability to ADP ribosylate as 45 was reduced throughout the study period. These studies demonstrate that hCG-induced heterologous desensitization results in a complex series of changes in β-adrenergic and LH receptors as well as G-protein function, which account for the altered hormonal responsiveness. (Endocrinology124: 1932-1941,1989)
Cholera toxin
Homologous desensitization
ADCY10
Gonadotropin
Cite
Citations (11)
Prolonged or repeated activation of many G protein-coupled receptors induces rapid desensitization followed by a period during which receptors are resensitized. In this study, concanavalin A (Con A) and monensin were used to investigate the mechanisms of desensitization and resensitization of the neurokinin-1 receptor. Con A inhibits internalization, whereas monensin prevents receptor recycling. The effects of Con A and monensin on desensitization, resensitization, receptor phosphorylation, endocytosis, and recycling of the neurokinin-1 receptor were assessed. Desensitization was defined as the decrease in the ability of substance P (SP) to elicit an intracellular Ca2+ response after a prolonged SP exposure. Resensitization was characterized as the return of SP responsiveness. Under control conditions, desensitization occurred after a 5-min exposure to agonist. Resensitization was evident 30 min after agonist washout. Neither monensin nor Con A prevented desensitization. Monensin completely inhibited resensitization, whereas Con A decreased but did not completely block resensitization. Receptor phosphorylation was increased after agonist activation and returned to basal levels after a recovery period. Neither Con A nor monensin altered the amount of agonist-specific receptor phosphorylation. Receptor binding analysis showed that plasma membrane receptors were internalized after a 5-min agonist exposure. Receptor recycling was not observed after a 1-h recovery period; however, resensitization was apparent. Taken together, these results suggest that rapid neurokinin-1 receptor desensitization can occur without receptor internalization and that resensitization occurs before receptor recycling.
Internalization
Homologous desensitization
Cite
Citations (13)
Homologous desensitization
Internalization
Phenylephrine
Glycogenolysis
Cite
Citations (5)
The glucagon-like peptide-1 receptor (GLP-1R) is an important target in the treatment of type 2 diabetes mellitus. The aim of this study was to compare the rate of agonist stimulated desensitization and internalization of GLP-1R. To this end, an N-terminally myc-tagged GLP-1R was stably expressed in HEK-293 cells. Homologous desensitization was assessed by measuring the cAMP response to agonist stimulation following pre-incubation with agonist for up to 120 min. Receptor internalization was monitored using an indirect ELISA-based method and confocal microscopy. Pre-incubation with GLP-1 resulted in a time-dependent loss of response to a second stimulation. Washing cells following pre-incubation failed to bring cAMP levels back to basal. Taking this into account, two desensitization rates were calculated: “apparent” (t1/2 = 19.27 min) and “net” (t1/2 = 2.99 min). Incubation of cells with GLP-1 also resulted in a time-dependent loss of receptor cell surface expression (t1/2 = 2.05 min). Rapid agonist-stimulated internalization of GLP-1R was confirmed using confocal microscopy. Stimulation of GLP-1R with GLP-1 results in rapid desensitization and internalization of the receptor. Interestingly, the rate of “net” desensitization closely matches the rate of internalization. Our results suggest that agonist-bound GLP-1R continues to generate cAMP after it has been internalized.
Internalization
Homologous desensitization
Cite
Citations (43)
Cite
Citations (8)
Internalization
Homologous desensitization
Nociceptin receptor
Cite
Citations (22)