Interactions of antagonists with subtypes of inositol 1,4,5‐trisphosphate (IP₃) receptor

Abstract

Background and PurposeInositol 1,4,5-trisphosphate receptors (IP3Rs) are intracellular Ca2+ channels. Interactions of the commonly used antagonists of IP3Rs with IP3R subtypes are poorly understood.Experimental ApproachIP3-evoked Ca2+ release from permeabilized DT40 cells stably expressing single subtypes of mammalian IP3R was measured using a luminal Ca2+ indicator. The effects of commonly used antagonists on IP3-evoked Ca2+ release and 3H-IP3 binding were characterized.Key ResultsFunctional analyses showed that heparin was a competitive antagonist of all IP3R subtypes with different affinities for each (IP3R3 > IP3R1 ≥ IP3R2). This sequence did not match the affinities for heparin binding to the isolated N-terminal from each IP3R subtype. 2-aminoethoxydiphenyl borate (2-APB) and high concentrations of caffeine selectively inhibited IP3R1 without affecting IP3 binding. Neither Xestospongin C nor Xestospongin D effectively inhibited IP3-evoked Ca2+ release via any IP3R subtype.Conclusions and ImplicationsHeparin competes with IP3, but its access to the IP3-binding core is substantially hindered by additional IP3R residues. These interactions may contribute to its modest selectivity for IP3R3. Practicable concentrations of caffeine and 2-APB inhibit only IP3R1. Xestospongins do not appear to be effective antagonists of IP3Rs.

Basal activity of voltage-gated Ca2+ channels controls the IP3-mediated contraction by α1-adrenoceptor stimulation of mouse aorta segments

“…IP3-sensitive phenylephrine-induced phasic contractions do not involve extracellular Ca 2 þ influx α 1 -Adrenoceptor stimulation of mouse aortic segments with phenylephrine in the absence of extracellular Ca 2 þ caused transient isometric contractions, which were inhibited with 2-APB. The IC 50 of 2-APB was below the concentration of 50 mM, which is the highest concentration that avoided inhibition of Ca 2 þ uptake to the SR (SERCA), but caused a sevenfold decrease in IP 3 -sensitivity of the IP 3 receptor (Saleem et al, 2014). Therefore, the phenylephrineinduced phasic contraction in 0Ca is due to Ca 2 þ release through IP 3 receptor activation via IP 3 formation (Manolopoulos et al, 1991;Yamamura et al, 2012).…”

“…2), a putative inhibitor of the IP 3 -receptor and the IP 3 -mediated Ca 2 þ release from the SR(Lamont and Wier, 2004;Saleem et al, 2014). 2-APB inhibited the transient contraction with an IC 50 of 327 4 mM (n ¼4,Fig.…”

The type 2 inositol 1,4,5-trisphosphate receptor, emerging functions for an intriguing Ca2+-release channel

“…Moreover, the quality of the antibodies raised against IP 3 R2 was also generally poor. Finally, a recent analysis of classically used IP 3 R inhibitors demonstrated that IP 3 R2 expressed in DT40 triple knockout (DT40 TKO) chicken B lymphocytes was the least sensitive of all the IP 3 R isoforms to heparin, caffeine and 2-aminoethoxydiphenyl borate [44]. This latter observation may explain why pharmacological approaches to discern IP 3 R2 function have been largely unsuccessful.…”

Oocyte pre-IVM with caffeine improves bovine embryo survival after vitrification

“…It is possible that additional pathways may have been modified by caffeine such as Ca 2þ and oocyte kinases. High concentrations (>10 mM) of caffeine have antagonistic effects on inositol triphosphate receptors (IP 3 R) [25]. Inositol triphosphate receptors are intracellular Ca 2þ channels located in the endoplasmic reticulum that release Ca 2þ into the cytoplasm in response to different stimuli in many cell types, including oocytes [25,53].…”

“…Other known caffeine mechanisms of action include a competitive antagonism of adenosine effects via adenosine receptors (A 1 , A 2A , and A 2B ) [24], inositol triphosphate receptor one antagonism [25] and ryanodine receptor activation [26]. In oocytes, caffeine raises intracellular cAMP levels, and thereby increases cAMP-dependent protein kinase activity, which in turn inhibits meiotic resumption [21,27].…”