Ionic strength of assay buffers influences antagonist binding affinity estimates at muscarinic M1–M5 cholinoceptors

Contractions of the Mouse Prostate Elicited by Acetylcholine Are Mediated by M3 Muscarinic Receptors

“…pA 2 values of the antagonists at the muscarinic receptors in the mouse prostate were compared against the mean published antagonist pK D or pK i values at each of the five muscarinic receptor subtypes (Table 1). Ionic strength of assay buffer is known to influence the binding affinity of muscarinic receptor antagonists (Loury et al, 1999). As such, pK D or pK i values were selected from Loury et al (1999) and the International Union of Basic and Clinical Pharmacology G protein-coupled receptor database only from studies using buffers of comparable ionic strength to the Krebs-Henseleit solution used in this study.…”

“…Ionic strength of assay buffer is known to influence the binding affinity of muscarinic receptor antagonists (Loury et al, 1999). As such, pK D or pK i values were selected from Loury et al (1999) and the International Union of Basic and Clinical Pharmacology G protein-coupled receptor database only from studies using buffers of comparable ionic strength to the Krebs-Henseleit solution used in this study. Where different antagonist affinities were given for the same subtype, the mean value was used for the correlation analysis.…”

Competition radioligand binding assays for the investigation of bispyridinium compound affinities to the human muscarinic acetylcholine receptor subtype 5 (hM5)

“…[19][20][21][22][23][24][25][26][27] As already mentioned in the introduction, buffers, incubation temperature and time were different. [41] In conclusion, it is evident that experimental factors (particularly the buffer composition) may influence ligand binding affinity. [23,24] This is in contrast to the results achieved in this work.…”

The muscarinic M5 receptor: a silent or emerging subtype?

“…In lieu of an endogenous correlate of the muscarinic M 5 receptor, one must turn to recombinant systems to characterize the pharmacology. Antagonist affinity data from our group ( Watson et al ., 1999 ; Loury et al ., 1999 ; Table 1) confirms previous reports on the receptor affinity profile. Historically, the M 5 receptor exhibits a low affinity for AF‐DX 116, AQ‐RA 741 and an intermediate affinity for methoctramine and pirenzepine; no compound has been reported that exhibits a preferential high affinity for the receptor.…”

“…In several respects, the profile of antagonist affinities at the M 5 subtype resembles that determined at the muscarinic M 3 receptor (Table 1), a finding highlighted by several workers in the area ( Buckley et al ., 1989 ; Jones et al ., 1991 ; Dorje et al ., 1991 ). Importantly, therefore, several ligands, including AQ‐RA 741, himbacine, and darifenacin are preferential for the M 3 over the M 5 receptor ( Buckley et al ., 1989 ; Jones et al ., 1991 ; Wallis & Napier 1999 , Loury et al ., 1999 ; Watson et al ., 1999 ). Recently, our group has also shown that older compounds, including oxybutynin, racemic secoverine and (S) secoverine, possess similar selectivity ( Choppin et al ., 1999b ).…”