State-dependent alterations in sleep/wake architecture elicited by the M4 PAM VU0467154 – Relation to antipsychotic-like drug effects
Introduction
Abnormal sleep architecture, including insomnia, reduced quality or duration of slow wave sleep (SWS) and increased Rapid Eye Movement (REM) sleep, are commonly reported in both medicated and un-medicated patients with schizophrenia and linked with the prevalence and severity of the other symptom clusters (for reviews see Cohrs, 2008, Sprecher et al., 2015). Sleep disturbances and decreased REM latency often precede and are present during acute psychosis (Pritchett et al., 2012, Sprecher et al., 2015), while decreases in SWS duration or quality are associated with the negative and cognitive symptoms of the disorder (Goder et al., 2006, Goder et al., 2004, Keshavan et al., 1995b, Yang and Winkelman, 2006). Recent studies suggest that normalizing sleep disturbances associated with schizophrenia may also improve other symptom clusters (Manoach and Stickgold, 2009). For example, declarative memory, one of the most disrupted cognitive domain in schizophrenia (Green et al., 2000) is improved following SWS-rich sleep compared to REM-rich sleep in healthy subjects (Plihal and Born, 1997). Moreover, declarative memory is improved by increasing slow wave activity (SWA), a neurophysiological correlate of sleep quality (Cohrs, 2008, Steiger and Kimura, 2010) during SWS via transcranial direct current stimulation in patients with schizophrenia (Goder et al., 2013). Cognitive performance is a critical predictor of overall functional outcome in schizophrenia patient populations (Bobes et al., 2007, Green, 1996, Green et al., 2004). Thus, strategies to improve cognitive performance, including modifying abnormal sleep architecture, may represent a novel treatment approach for schizophrenia.
In addition to polysomnography, quantitative electroencephalography (qEEG) approaches have correlated aberrant brain oscillation patterns with specific symptoms in unmedicated schizophrenia patients and adverse side effect profiles of antipsychotic drugs (APDs) in medicated patients (Boutros et al., 2008, Boutros et al., 2014, Goder et al., 2006, Keshavan et al., 1995a, Wichniak et al., 2006). For example, elevated power in the high frequency gamma band (e.g., >30 Hz in humans) is associated with positive symptoms, whereas lower gamma power at rest and reduced elevations in gamma power during cognitive testing is linked to cognitive impairments (Baldeweg et al., 1998, Chen et al., 2014, Lee et al., 2003, Uhlhaas and Singer, 2014). To date, clinically available APDs provide modest to no therapeutic benefit for many of the symptoms observed in schizophrenia patients, including negative symptoms and cognitive deficits (Krystal et al., 2008, Reichenberg and Harvey, 2007). At therapeutically relevant doses, many APDs, including the atypical APD clozapine, increase total sleep time and SWA. However, these potentially beneficial effects during sleep persist during wake as a general EEG slowing (e.g. shift in power from high to low frequencies). This EEG slowing is correlated with problematic sedating effects in healthy subjects and schizophrenia patients (Freudenreich et al., 1997, Roubicek and Major, 1977, Yoshimura et al., 2007) that may contribute to cognitive impairments. These studies highlight the importance of understanding state-dependent alterations in sleep architecture and arousal relative to APD-like and cognition enhancing activity in the development of novel APDs.
One promising approach for the development of novel APDs involves modulation of the muscarinic cholinergic system (Jones et al., 2012), which is regulated by five different subtypes of muscarinic acetylcholine receptors (mAChRs), termed M1-M5 (Bonner et al., 1987, Bonner et al., 1988) and plays a critical role in regulating arousal, mood, cognition, and sleep architecture (Graef et al., 2011, Platt and Riedel, 2011). Nonselective mAChR agonists and acetylcholinesterase inhibitors increase arousal, cognition, and REM bouts and/or duration, but reduce NREM sleep duration (Nissen et al., 2006, Riemann et al., 1994). In clinical studies, the M1/M4-preferring mAChR agonist xanomeline reduced the behavioral disturbances and psychotic symptoms observed in Alzheimer's disease and schizophrenia patients, respectively (Bodick et al., 1997a, Bodick et al., 1997b, Shekhar et al., 2008). However, xanomeline and other mAChR agonists failed in clinical development due to dose-limiting adverse side effects attributed to nonspecific activation of peripheral mAChRs (McArthur et al., 2010). Over the last decade, we and others have developed a novel strategy for activation of individual mAChR subtypes, particularly the M4 subtype, using highly selective positive allosteric modulators (PAMs) (Brady et al., 2008, Bubser et al., 2014, Byun et al., 2014). These M4 PAMs do not activate the M4 mAChR directly, but potentiate the response of the receptor to ACh, thereby enhancing activity-dependent signaling through allosteric binding sites that are more topographically distinct and less highly-conserved than the orthosteric ACh binding site (Conn et al., 2009). Selective M4 PAMs produce the APD-like profile previously reported with xanomeline (Brady et al., 2008, Bubser et al., 2014, Byun et al., 2014, Mirza et al., 2003). In addition, the M4 PAM VU0467154 produces cognitive-enhancing effects when administered alone on measures of learning and memory and reverses MK-801-induced hyperlocomotion and cognitive disruptions (Bubser et al., 2014). MK-801, an antagonist of the N-methyl-d-aspartate subtype of the glutamate receptor (NMDAR), is often used to model neurophysiological, neurochemical, and behavioral disruptions associated with NMDAR hypofunction, which are thought to underlie many of the symptoms in schizophrenia (Anticevic et al., 2015, Blot et al., 2013, Coyle et al., 2012). The above studies suggest that M4 PAMs may provide efficacy for multiple symptom clusters associated with schizophrenia. However, the effects of selective M4 PAMs on sleep architecture and arousal relative to antipsychotic-like and cognitive enhancing activity remain unknown.
In the present studies, we examined the effects of the M4 PAM VU0467154, in comparison with clozapine and xanomeline, on sleep architecture and arousal using EEG in freely moving rats. Specifically, we confined our analyses to examine aspects of sleep/wake architecture known to be disrupted in patients with schizophrenia and/or directly affected by APDs. Compounds were evaluated during the light period to examine wake-promoting or arousal enhancing effects and to examine changes in duration of REM/non-REM sleep and SWS quality. Compounds were also evaluated during the dark period to examine sleep-promoting or sedative effects. In addition we extended previous studies to examine the ability of these compounds to attenuate MK-801-induced elevations in high frequency gamma power (50–100 Hz), a putative biomarker of NMDAR hypofunction and potential antipsychotic-like activity. Previous studies have demonstrated that NMDAR antagonists induce cognitive impairments and increase high frequency gamma power in healthy humans and exacerbate cognitive impairments in patients with schizophrenia (Coyle et al., 2012, Hong et al., 2010, Kocsis et al., 2013, Tsai and Coyle, 2002). VU0467154 produced selective increases in the duration, without disrupting quality, of NREM sleep and, in contrast to clozapine, increased arousal during wake. VU0467154 also reversed MK-801-induced elevations in gamma power consistent with APD-like activity (Hiyoshi et al., 2014). The state-dependent actions of the M4 PAM VU0467154 represent a potentially unique therapeutic profile for treating many of the symptoms in schizophrenia patients, including the abnormalities in sleep architecture and arousal, without the sedation observed with other APDs.
Section snippets
Subjects
Male Sprague–Dawley rats (Harlan, Indianapolis, IN) were individually housed and maintained on a 12-h light:12-h dark cycle with ad libitum access to food and water served as subjects. All experiments were approved by the Vanderbilt University Animal Care and Use Committee, and experimental procedures conformed to guidelines established by the National Research Council Guide for the Care and Use of Laboratory Animals.
Surgery
Rats (250–275 g) were surgically implanted under isoflurane anesthesia with a
VU0467154 increases arousal during wake but does not promote wakefulness
Consistent with the nocturnal nature of rodents, ∼70% of time during the light phase was spent sleeping and ∼70% of time during the dark phase was spent awake (Fig. 1). Consistent with the literature, compound administration 2 h into the light cycle resulted in a transient increase in time awake that dissipated within ∼30 min following vehicle administration. There was a significant effect of VU0467154 on percent time awake across the 24-h period (Fig. 1A; dose [F3,42 = 3.14, p < 0.05], time [F
Discussion
Discovery and characterization of the M4 PAM VU0467154 represents a critical breakthrough in the development of subtype-selective mAChR ligands that can enhance and maintain activity-dependent signaling at the M4 mAChR for the potential treatment of the complex symptoms of schizophrenia, including disruptions in sleep architecture, arousal and cognition. Here, we examined the potential impact of selective allosteric potentiation of M4 on state-dependent alterations on sleep architecture,
Financial disclosures
The authors declare the following competing financial interest(s): Over the past two years, C.W.L. consulted for Abbott. M.B., T.M.B., M.J.N., C.M.N., J.S.D, M.R.W., C.W.L., P.J.C., and C.K.J. received research/salary support from AstraZeneca and/or Bristol Myers Squibb. T.M.B., C.M.N., J.S.D., M.R.W., C.W.L., C.K.J., and P.J.C. are inventors on multiple composition of matter patents protecting allosteric modulators of GPCRs. M.E.D., J.D., N.J.B., and M.W.W. are employees of AstraZeneca. M.I.
Acknowledgments
This work was supported by grants from the National Institute of Mental Health (MH086601, C.K.J.; MH073676, MH087965, MH093366, P.J.C), by AstraZeneca and from a PhRMA Foundation postdoctoral fellowship in Pharmacology and Toxicology (RWG). We thank Dina McGinnis and Weimin Peng for surgical expertise, Dr. Ariel Deutch and Dr. David Devilbiss for helpful comments on data analysis and interpretation, and Frank Byers for conducting the in-life phase of the pharmacokinetic studies. All
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