Increased prefrontal D2 protein in Tourette syndrome: a postmortem analysis of frontal cortex and striatum

Abstract

The precise neuropathological mechanism underlying Tourette syndrome (TS) is unknown. In order to evaluate a variety of proposed dopaminergic abnormalities, postmortem tissue samples were obtained from three individuals with TS (two typical males with childhood onset, ages 29 and 77, and a 62-year-old female with adult-onset) and three age- and sex-matched controls. Samples from caudate, putamen, ventral striatum, and prefrontal cortex (Brodmann's area 9, BA9) were analyzed by semiquantitative immunoblotting for relative densities of dopamine receptors (D1, D2), transporter (DAT), monoamine terminals (vesicular monoamine transporter type 2), vesicular docking and release proteins (VAMP-2, synaptotagmin, SNAP-25, syntaxin, synaptophysin), and receptors inhibiting dopamine release (alpha 2-adrenergic receptors, α-2A). Concentrations of monoamine neurotransmitters and their metabolites were assessed by high performance liquid chromatography. Data from each TS sample was calculated as a percent value of its control. Results showed that prefrontal cortex, rather than striatum, had the greatest number of changes in the two typical TS cases, including increases for D2, DAT, VAMP-2, and α-2A. All three TS subjects had increased densities of prefrontal D2 receptor protein, greater than 140% of their matched control. These results suggest the presence of a prefrontal-dopaminergic abnormality in TS and emphasize the need for a more specific focus on the frontal lobe.

Introduction

Tourette syndrome (TS) is a neurological disorder characterized by the presence of chronic involuntary motor and vocal tics that wax and wane. Pathophysiologically, cortico-striatal-thalamo-cortical (CSTC) circuits have been identified as the site of neuroanatomic localization, typically with emphasis on subcortical involvement [1]. Alterations of several synaptic neurotransmitter systems have been proposed, with most focusing on dopaminergic abnormalities [2]. The availability of postmortem striatal and frontal cortical tissue from two typical TS cases and one with an atypical (adult-onset) presentation has provided an opportunity to evaluate the role of specific neurotransmitters in this disorder.

Although CSTC circuits contain a wide variety of neurotransmitters, a significant dopaminergic abnormality has been suggested based on the therapeutic response to neuroleptics and results of several neuroimaging protocols [2]. Abnormal dopamine release, dopamine hyperinnervation, and supersensitive dopamine receptors, have all been considered as underlying hypotheses. Dopamine release, measured by PET comparison of [¹¹C]raclopride binding after saline and amphetamine administration, is significantly increased in putamen from TS patients, but not in caudate [3]. Dopaminergic hyperinnervation suggested by findings of elevated dopamine transporter (DAT) binding in postmortem caudate and putamen [4] and two [¹²³I]β-CIT SPECT studies [5], [6], has not been confirmed by measurements of the vesicular monoamine transporter (VMAT-2), which were normal [7]. Lastly, receptor binding studies in postmortem tissue have shown slight increases in D2 receptor binding (measured by [³H]spiperone) [4], but examination by PET and SPECT have produced conflicting findings [8], [9].

Although there is general consensus of a frontal-subcortical abnormality in TS, initial investigations focused primarily on the striatal component. More recently, however, there has been increasing evidence of a significant cortical dysfunction. A volumetric MRI study has shown larger volumes of the dorsolateral prefrontal region in children with TS, but significantly smaller volumes in adults with the disorder [10]. These differences in prefrontal volume size correlated inversely with the severity of tic symptoms, and suggest that larger prefrontal volumes in children could represent adaptive processes that help to reduce tic severity [11]. Cortical white matter in children with TS was increased in the right frontal lobe [12] and decreased in the deep left frontal region [13]. Mid-sagittal measurements of the corpus callosum, which interconnects homologous cortical areas, have shown variable alterations in the size of this structure [14], [15]. Functional MR imaging, in which images acquired during periods of voluntary tic suppression were compared with those during spontaneous expression of tics, suggests that tic suppression involves activation of the prefrontal cortex [16]. Event-related PET techniques revealed correlations between tics and cortical brain activity in many regions, including the dorsolateral–rostral prefrontal cortex [17]. Lastly, transcranial magnetic stimulation studies have suggested that tics may originate from impaired inhibition directly at the level of the motor cortex [18], [19].

In this study, we analyzed samples of caudate, putamen, ventral striatum, and prefrontal cortex (Brodmann's area 9, BA9) from three individuals with TS and three age- and sex-matched controls. Assays, selected for their ability to define dopaminergic abnormalities, included: (a) determination of the following by semi-quantitative immunoblotting: relative densities of dopamine receptors (D1, D2), transporter (DAT), monoamine terminals (vesicular monoamine transporter type 2, VMAT-2), vesicular docking and release proteins (VAMP-2, synaptotagmin, SNAP-25, syntaxin, synaptophysin), and receptors inhibiting dopamine release (alpha 2-adrenergic receptors); and (b) measurement of the concentrations of monoamine neurotransmitters and their metabolites by high performance liquid chromatography (HPLC). We hypothesized that dopaminergic markers would differ between subjects with TS and their matched controls and that changes would be more evident in basal ganglia than in prefrontal cortex.