Selective Activation of M₄ Muscarinic Acetylcholine Receptors Reverses MK-801-Induced Behavioral Impairments and Enhances Associative Learning in Rodents

Abstract

Positive allosteric modulators (PAMs) of the M4 muscarinic acetylcholine receptor (mAChR) represent a novel approach for the treatment of psychotic symptoms associated with schizophrenia and other neuropsychiatric disorders. We recently reported that the selective M4 PAM VU0152100 produced an antipsychotic drug-like profile in rodents after amphetamine challenge. Previous studies suggest that enhanced cholinergic activity may also improve cognitive function and reverse deficits observed with reduced signaling through the N-methyl-d-aspartate subtype of the glutamate receptor (NMDAR) in the central nervous system. Prior to this study, the M1 mAChR subtype was viewed as the primary candidate for these actions relative to the other mAChR subtypes. Here we describe the discovery of a novel M4 PAM, VU0467154, with enhanced in vitro potency and improved pharmacokinetic properties relative to other M4 PAMs, enabling a more extensive characterization of M4 actions in rodent models. We used VU0467154 to test the hypothesis that selective potentiation of M4 receptor signaling could ameliorate the behavioral, cognitive, and neurochemical impairments induced by the noncompetitive NMDAR antagonist MK-801. VU0467154 produced a robust dose-dependent reversal of MK-801-induced hyperlocomotion and deficits in preclinical models of associative learning and memory functions, including the touchscreen pairwise visual discrimination task in wild-type mice, but failed to reverse these stimulant-induced deficits in M4 KO mice. VU0467154 also enhanced the acquisition of both contextual and cue-mediated fear conditioning when administered alone in wild-type mice. These novel findings suggest that M4 PAMs may provide a strategy for addressing the more complex affective and cognitive disruptions associated with schizophrenia and other neuropsychiatric disorders.

Allosteric activation of M 4 muscarinic receptors improve behavioral and physiological alterations in early symptomatic YAC128 mice

“…However, to confirm that the reversal of rotarod deficits could not be attributed to residual VU0467154, we determined the effect of acute administration of 10 mg/kg (i.p. ), VU0467154, given at least 45 min before testing, a time when VU0467154 levels in the CNS are at their peak (30). As in the original cohort of WT and YAC128 mice, Vh-treated YAC128 mice showed deficits in rotarod performance relative to WT controls.…”

“…To directly determine the effective residual brain levels of VU0467154, 48 h after the last dose, mice were injected daily for 14 d with 10 mg/kg VU0467154. Using mass spectrometry, we determined that, at the 48-h time point, the measured unbound concentrations of VU0467154 were 3.15 ± 1.7 nM, which is below the active concentration of this compound at potentiating M 4 (30).…”

“…Although M 4 is responsible for regulation of corticostriatal transmission in prepubertal WT mice (25), it is possible that M 4 is not the predominant mAChR subtype responsible for the increased effect of CCh in older YAC128 mice. To determine whether the enhanced inhibitory effect of CCh in SPNs from 2-mo-old YAC128 mice was mediated by M 4 , we determined the potentiating effect of VU0467154, a highly selective M 4 PAM discovered in our laboratory (30), on the CCh-mediated inhibition of glutamatergic transmission. Bath application of VU0467154 (3 μM) potentiated the inhibitory effect of a submaximal concentration of CCh (1 μM) on peak eEPSC amplitude in YAC128 SPNs (Fig.…”

“…), once daily] on the appearance of synaptic and motor deficits in YAC128 mice. The dose and dosing interval were chosen based on the compound's predicted pharmacokinetics (PK) at steady state using previously reported acute PK data together with concentration-efficacy relationship data (30). Based on this, we determined that dosing animals once daily at a dose of 10 mg/kg should provide constant central nervous system (CNS) exposure for the duration of the dosing period.…”

“…WT and YAC128 animals received their last administration of VU0467154 or Vh at least 48 h before testing to allow clearance of the M 4 PAM. Previous PK analysis of VU0467154 predicts that the compound should be cleared from the brain and plasma by 48 h and that residual levels will be well below those required to potentiate M 4 responses at this time point (30). To directly determine the effective residual brain levels of VU0467154, 48 h after the last dose, mice were injected daily for 14 d with 10 mg/kg VU0467154.…”