Early decrease of type 1 cannabinoid receptor binding and phosphodiesterase 10A activity in vivo in R6/2 Huntington mice

Highlights

• We quantified brain type 1 cannabinoid (CB1) and phosphodiesterase 10A (PDE10A) alterations in Huntington disease in vivo.

• R6/2 mice were investigated in presymptomatic, early symptomatic and late symptomatic phases.

• CB1 and PDE10A binding decreased in basal ganglia areas upon disease progression.

• CB1 and PDE10A binding correlated to anomalies in motor function.

• [¹⁸F]MK-9470 and [¹⁸F]JNJ42259152 may be useful as early biomarkers of Huntington disease.

Abstract

Several lines of evidence imply early alterations in endocannabinoid and phosphodiesterase 10A (PDE10A) signaling in Huntington disease (HD). Using [¹⁸F]MK-9470 and [¹⁸F]JNJ42259152 small-animal positron emission tomography (PET), we investigated for the first time cerebral changes in type 1 cannabinoid (CB1) receptor binding and PDE10A levels in vivo in presymptomatic, early symptomatic, and late symptomatic HD (R6/2) mice, in relation to glucose metabolism ([¹⁸F]FDG PET), brain morphology (magnetic resonance imaging) and motor function. Ten R6/2 and 16 wild-type (WT) mice were investigated at 3 different time points between the age of 4 and 13 weeks. Parametric CB1 receptor and PDE10A images were anatomically standardized to Paxinos space and analyzed voxelwise. Volumetric microMRI imaging was performed to assess HD pathology. In R6/2 mice, CB1 receptor binding was decreased in comparison with WT in a cluster comprising the bilateral caudate-putamen, globus pallidus, and thalamic nucleus at week 5 (−8.1% ± 2.6%, p = 1.7 × 10⁻⁵). Longitudinal follow-up showed further progressive decline compared with controls in a cluster comprising the bilateral hippocampus, caudate-putamen, globus pallidus, superior colliculus, thalamic nucleus, and cerebellum (late vs. presymptomatic age: −13.7% ± 3.1% for R6/2 and +1.5% ± 4.0% for WT, p = 1.9 × 10⁻⁵). In R6/2 mice, PDE10A binding potential also decreased over time to reach significance at early and late symptomatic HD (late vs. presymptomatic age: −79.1% ± 1.9% for R6/2 and +2.1% ± 2.7% for WT, p = 1.5 × 10⁻⁴). The observed changes in CB1 receptor and PDE10A binding were correlated to anomalies exhibited by R6/2 animals in motor function, whereas no correlation was found with magnetic resonance imaging-based striatal volume. Our findings point to early regional dysfunctions in endocannabinoid and PDE10A signaling, involving the caudate-putamen and lateral globus pallidus, which may play a role in the progression of the disease in R6/2 animals. PET quantification of in vivo CB1 and/or PDE10A binding may thus be useful early biomarkers for HD. Our results also provide evidence of subtle motor deficits at earlier stages than previously described.

Introduction

Huntington disease (HD) is an autosomal dominant inherited neurodegenerative disease, characterized by motor dysfunctions, behavioral changes, and cognitive decline. The causative mutation is an expanded CAG repeat in exon 1 of the gene encoding the protein huntingtin (htt) (The Huntington's Disease Collaborative Research Group, 1993). Adult onset is usually observed with trinucleotide repeat blocks between 40 and 50 units, whereas more than 60 repeat results in more severe and much less frequently observed juvenile, and even infantile, forms. The most striking pathophysiological feature of HD affected brains is the progressive atrophy of the caudate nucleus and the putamen, accompanied by a secondary enlargement of the lateral ventricles, and cortical degeneration in some patients (Vonsattel et al., 1985). Despite progress in elucidating the molecular pathology of HD, therapeutic benefit for patients in terms of effective pharmacotherapy with either symptomatic or protective effects, has been scarce.

The type 1 cannabinoid (CB1) receptor and phosphodiesterase 10A (PDE10A) enzyme play a role in changed neurotransmission in HD (Hebb et al., 2004, Katona and Freund, 2008). CB1 receptors presynaptically modulate the release of other neurotransmitters (Goutopoulos and Makriyannis, 2002) and are found at high densities on GABA-ergic striatal projection neurons (Richfield and Herkenham, 1994). PDE10A hydrolyzes the important second messengers cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate and is characterized by a restricted distribution, predominantly in medium spiny neurons of the striatum (Seeger et al., 2003). In HD, loss of CB1 receptor binding and/or signaling from the basal ganglia nuclei is one of the earliest neurochemical alterations observed in humans (Allen et al., 2009, Glass et al., 2000, Richfield and Herkenham, 1994) and experimental models (Blazquez et al., 2011, Centonze et al., 2005, Denovan-Wright and Robertson, 2000, Dowie et al., 2009, Lastres-Becker et al., 2002, Lastres-Becker et al., 2004), preceding the development of cerebral metabolic deficits (Antonini et al., 1996) and striatal volume losses (Glass et al., 2000). Also, decreased levels of PDE10A expression occur before the onset of motor-related HD symptoms in transgenic HD mice (Hebb et al., 2004). In addition, genetic ablation of CB1 receptors showed to accelerate the onset of HD-like symptoms in transgenic R6/2 mice (Blazquez et al., 2011), whereas PDE10A inhibition ameliorated striatal and cortical pathology in experimental HD (Giampa et al., 2010, Giralt et al., 2013, Kleiman et al., 2011). Furthermore, pharmacologic studies demonstrated crosstalk of CB1 and PDE10A with striatal dopaminergic signaling (Chiang et al., 2013, Nishi et al., 2008). In symptomatic HD patients, we found a profound cortical and subcortical loss of CB1 receptor availability in vivo (Van Laere et al., 2010).

So far, only in vitro and ex vivo data exist on the CB1 receptor binding and PDE10A activity in genetic HD models. Ex vivo studies of CB1 receptor changes in transgenic mouse models of HD have focused on the basal ganglia, hippocampus, and motor cortex, but have not assessed other brain regions (Dowie et al., 2009, Dowie et al., 2010 ). Also it remains unknown whether CB1 receptor and PDE10A levels are changed in vivo, and to what spatial and temporal extent. Thanks to the development of selective CB1 ([¹⁸F]MK-9470; Burns et al., 2007) and PDE10A radioligands ([¹⁸F]JNJ42259152; Celen et al., 2013), in vivo imaging of these proteins became feasible.

Our primary objective was thus to investigate CB1 receptor and PDE10A changes in vivo throughout the disease process of R6/2 transgenic HD mice. As the secondary objective, motor function, brain glucose metabolism, and morphology, which are shown to be altered (early) in patients and animal models of HD (Antonini et al., 1996, Carter et al., 1999, Cowin et al., 2011, Rattray et al., 2013), were investigated in the same animals and correlated to the regional CB1 receptor binding and PDE10A levels.