The affinity and selectivity of α‐adrenoceptor antagonists, antidepressants and antipsychotics for the human α2A, α2B, and α2C‐adrenoceptors and comparison with human α1 and β‐adrenoceptors

Abstract

α2‐Adrenoceptors, subdivided into α2A, α2B, and α2C subtypes and expressed in heart, blood vessels, kidney, platelets and brain, are important for blood pressure, sedation, analgesia, and platelet aggregation. Brain α2C‐adrenoceptor blockade has also been suggested to be beneficial for antipsychotic action. However, comparing α2‐adrenoceptor subtype affinity is difficult due to significant species and methodology differences in published studies. Here, 3H‐rauwolscine whole cell binding was used to determine the affinity and selectivity of 99 α‐antagonists (including antidepressants and antipsychotics) in CHO cells expressing human α2A, α2B, or α2C‐adrenoceptors, using an identical method to β and α1‐adrenoceptor measurements, thus allowing direct human receptor comparisons. Yohimbine, RX821002, RS79948, and atipamezole are high affinity non‐selective α2‐antagonists. BRL44408 was the most α2A‐selective antagonist, although its α1A‐affinity (81 nM) is only 9‐fold greater than its α2C‐affinity. MK‐912 is the highest‐affinity, most α2C‐selective antagonist (0.15 nM α2C‐affinity) although its α2C‐selectivity is only 13‐fold greater than at α2A. There are no truely α2B‐selective antagonists. A few α‐ligands with significant β‐affinity were detected, for example, naftopidil where its clinical α1A‐affinity is only 3‐fold greater than off‐target β2‐affinity. Antidepressants (except mirtazapine) and first‐generation antipsychotics have higher α1A than α2‐adrenoceptor affinity but poor β‐affinity. Second‐generation antipsychotics varied widely in their α2‐adrenoceptor affinity. Risperidone (9 nM) and paliperidone (14 nM) have the highest α2C‐adrenoceptor affinity however this is only 5‐fold selective over α2A, and both have a higher affinity for α1A (2 nM and 4 nM, respectively). So, despite a century of yohimbine use, and decades of α2‐subtype studies, there remains plenty of scope to develop α2‐subtype selective antagonists.

The signaling and selectivity of α‐adrenoceptor agonists for the human α2A, α2B and α2C‐adrenoceptors and comparison with human α1 and β‐adrenoceptors

“…Both inhibitory and stimulatory parts of the response were inhibited by yohimbine to yield K D values of −7.62 ± 0.14 and −7.66 ± 0.03 respectively ( n = 8; Figure 4A ), very similar to that obtained from whole cell binding (log K D −7.66). 40 As expected, Gs‐stimulatory responses were seen in the absence of forskolin (Figure 4B ). Similar responses were also obtained from cAMP accumulation in the presence (log IC 50 −8.19 ± 0.11, log EC 50 −6.56 ± 0.08, n = 7) and absence (log EC 50 −6.09 ± 0.11, 163.0 ± 15.2% 10 μM forskolin, n = 7) of forskolin and the ERK1/2‐phosphorylation response closely resembled the IC 50 obtained from Gi‐inhibition (log −7.78, Table 2 ; Figure 4D ).…”

“…26 This affinity is similar to the affinity obtained for yohimbine from whole cell binding in these cells (log K D −8.48). 40 A similar high affinity for yohimbine was seen with the stimulatory brimonidine response in the presence of PTX (yohimbine log K D −8.48 ± 0.13, n = 15; Figure 1B ), and in the absence of forskolin (whether that be without PTX, Figure 1C , −8.61 ± 0.06, n = 14 or in the presence of PTX (Figure 1D , −8.54 ± 0.04, n = 12). Finally no response was seen to brimonidine in cells without the transfected receptor (see later).…”

“…CHO‐K1 (RIDD: CVCL_0214) stably transfected with a CRE‐SPAP reporter gene and the human α2A‐adrenoceptor (CHO‐α2A), human α2B‐adrenoceptor (CHO‐α2B) or human α2C‐adrenoceptor (CHO‐α2C) were used 40 as were lines expressing the same CRE‐SPAP reporter and human β1‐adrenoceptor (CHO‐β1) or human β2‐adrenoceptor (CHO‐β2, 38 ). The parental cell line, which expresses the CRE‐SPAP reporter but no transfected receptor, and from which these lines were generated, was also used.…”

“…From the IC 50 value, the known concentration of 3 H‐rauwolscine and the known K D 3 H‐rauwolscine (determined from saturation binding), 40 a K D value (concentration at which half the receptors are bound by the competing agonist ligand) was calculated using the Cheng‐Prusoff equation: In some cases the maximum concentration of competing ligand was not able to inhibit all of the specific 3 H‐rauwolscine binding. Where no inhibition of radioligand binding was seen, even with maximum concentration of competing ligand possible, “no binding” is given in the tables.…”

Alpha-2 Adrenoreceptor Antagonist Yohimbine Potentiates Consolidation of Conditioned Fear

“…Several studies suggest that, in addition to NE and DA, modulations of the serotonergic system affect fear conditioning and extinction ( Bauer, 2015 ). In light of the limited specificity of yohimbine, future studies should try to replicate our findings with a higher affinity and more selective α2-adrenoreceptor antagonist, such as atipamezole or MK-912 ( Pettibone et al, 1989 ; Pertovaara et al, 2005 ; Proudman et al, 2022 ).…”