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IMPORTANCE: Urate elevation, despite associations with crystallopathic, cardiovascular, and metabolic disorders, has been pursued as a potential disease-modifying strategy for Parkinson disease (PD) based on convergent biological, epidemiological, and clinical data. OBJECTIVE: To determine whether sustained urate-elevating treatment with the urate precursor inosine slows early PD progression. DESIGN, PARTICIPANTS, AND SETTING: Randomized, double-blind, placebo-controlled, phase 3 trial of oral inosine treatment in early PD. A total of 587 individuals consented, and 298 with PD not yet requiring dopaminergic medication, striatal dopamine transporter deficiency, and serum urate below the population median concentration (

Background DBS in the ventral intermediate nucleus (VIM) of the thalamus has been a revolutionary treatment for patients with essential tremor (ET) by reducing tremor. Unfortunately, some patients develop habituation to DBS and thus experience reduced efficacy and loss of tremor control. There are no standardized methods of addressing habituation to DBS. We propose alternating stimulation patterns as a way to reduce habituation. Methods This was a randomized, placebo-controlled trial for patients with VIM DBS for ET. Patients were randomized to either experimental treatment arm of alternating stimulation patterns on a weekly basis or standard care arm of continuous stimulation settings for 12 weeks. Primary outcome was change in the performance subscale of The Essential Tremor Rating Assessment Scale (TETRAS), which was performed at initial visit and 12-week follow-up. Secondary outcome included change in the activities of daily living subscale of TETRAS. Results Twenty-two patients were enrolled in the trial, and 16 were analyzed at follow-up. Experimental treatment subjects displayed sustained tremor control compared to standard care, as measured by the change in TETRAS performance subscale (-0.6 vs. 6.7 point change, respectively) with a 7.3 difference between the arms (P = 0.006). Conclusion Alternating stimulation patterns on a weekly basis for ET patients with VIM DBS reduced habituation in this pilot study. This study suggests that exposure to different stimulation groups may maintain better tremor control compared to constant stimulation parameters.

5-HT(6) (serotonin) receptors are promising targets for a variety of neuropsychiatric disorders and have been linked to several cellular signaling cascades. Endogenous 5-HT(6) receptors are restricted to the primary neuronal cilium, a small sensory organelle stemming from the cell body that receives numerous extrasynaptic signals. Inhibition of 5-HT(6) receptors decreases cilia length in primary neuronal cultures, but the signaling mechanisms involved are still unclear. Intense overexpression of exogenous 5-HT(6) receptors increases the probability for receptors to localize outside the primary cilium and have been associated with changes in cilia morphology and dendritic outgrowth. In the present study, we explore the role of 5-HT(6)R rescue on neuronal morphology in primary neuronal cultures from 5-HT(6)R-KO mice, at the same time maintaining a more physiologic level of expression, wherein the receptor localizes to cilia in 80%–90% of neurons (similar to endogenous 5-HT(6)R localization). We found that rescue of 5-HT(6)R expression is sufficient to increase cilia length and dendritic outgrowth, but primarily in neurons in which the receptor is located exclusively in the primary cilia. Additionally, we found that expression of 5-HT(6)R mutants deficient in agonist-stimulated cAMP or without the predicted Fyn kinase binding domain maintained constitutive activity for stimulating cAMP and still increased the length of cilia, and that the proposed Fyn kinase domain was required for stimulating dendritic outgrowth. These findings highlight the complexity of 5-HT(6)R function and localization, particularly with the use of exogenous overexpression, and provide greater understanding and potential mechanisms for 5-HT(6)R drug therapies.

Parkinson disease (PD) is a progressive neurodegenerative disorder characterized by motor and non-motor symptoms and signs. Many reports suggest that diminished heart rate variability occurs early, even prior to the cardinal signs of PD. In a longitudinal study of PD, we evaluated whether heart rate variability (HRV) obtained using a 10 second ECG tracing, and the electrocardiographic QT-interval would be associated with PD severity and progression. Subjects were derived from a longitudinal study of 1741 individuals with early, stable PD. The severity of PD was measured using the global statistical test (GST). In a subset, the heart rate corrected QT-interval (QTcB) was calculated for each electrocardiogram (ECG). The HRV was measured for each ECG and then transformed to fit a normal distribution. The baseline analysis included 653 subjects, with 256 completing the 5-year follow up study. There was an association (P

The 5-HT6 receptor has been implicated in a variety of cognitive processes including habitual behaviors, learning, and memory. It is found almost exclusively in the brain, is expressed abundantly in striatum, and localizes to neuronal primary cilia. Primary cilia are antenna-like, sensory organelles found on most neurons that receive both chemical and mechanical signals from other cells and the surrounding environment; however, the effect of 5-HT6 receptor function on cellular morphology has not been examined. We confirmed that 5-HT6 receptors were localized to primary cilia in wild-type (WT) but not 5-HT6 knockout (5-HT6KO) in both native mouse brain tissue and primary cultured striatal neurons then used primary neurons cultured from WT or 5-HT6KO mice to study the function of these receptors. Selective 5-HT6 antagonists reduced cilia length in neurons cultured from wild-type mice in a concentration and time-dependent manner without altering dendrites, but had no effect on cilia length in 5-HT6KO cultured neurons. Varying the expression levels of heterologously expressed 5-HT6 receptors affected the fidelity of ciliary localization in both WT and 5-HT6KO neurons; overexpression lead to increasing amounts of 5-HT6 localization outside of the cilia but did not alter cilia morphology. Introducing discrete mutations into the third cytoplasmic loop of the 5-HT6 receptor greatly reduced, but did not entirely eliminate, trafficking of the 5-HT6 receptor to primary cilia. These data suggest that blocking 5-HT6 receptor activity reduces the length of primary cilia and that mechanisms that regulate trafficking of 5-HT6 receptors to cilia are more complex than previously thought.

Genetic studies have shown an association between smoking and variation at the CHRNA5/A3/B4 gene locus encoding the α5, α3, and β4 nicotinic receptor subunits. The α5 receptor has been specifically implicated because smoking-associated haplotypes contain a coding variant in the CHRNA5 gene. The Chrna5/a3/b4 locus is conserved in rodents and the restricted expression of these subunits suggests neural pathways through which the reinforcing and aversive properties of nicotine may be mediated. Here, we show that, in the interpeduncular nucleus (IP), the site of the highest Chrna5 mRNA expression in rodents, electrophysiological responses to nicotinic acetylcholine receptor stimulation are markedly reduced in α5-null mice. IP neurons differ markedly from their upstream ventral medial habenula cholinergic partners, which appear unaltered by loss of α5. To probe the functional role of α5-containing IP neurons, we used BAC recombineering to generate transgenic mice expressing Cre-recombinase from the Chrna5 locus. Reporter expression driven by Chrna5Cre demonstrates that transcription of Chrna5 is regulated independently from the Chrna3/b4 genes transcribed on the opposite strand. Chrna5-expressing IP neurons are GABAergic and project to distant targets in the mesopontine raphe and tegmentum rather than forming local circuits. Optogenetic stimulation of Chrna5-expressing IP neurons failed to elicit physical manifestations of withdrawal. However, after recent prior stimulation or exposure to nicotine, IP stimulation becomes aversive. These results using mice of both sexes support the idea that the risk allele of CHRNA5 may increase the drive to smoke via loss of IP-mediated nicotine aversion.SIGNIFICANCE STATEMENT Understanding the receptors and neural pathways underlying the reinforcing and aversive effects of nicotine may suggest new treatments for tobacco addiction. Part of the individual variability in smoking is associated with specific forms of the α5 nicotinic receptor subunit gene. Here, we show that deletion of the α5 subunit in mice markedly reduces the cellular response to nicotine and acetylcholine in the interpeduncular nucleus (IP). Stimulation of α5-expressing IP neurons using optogenetics is aversive, but this effect requires priming by recent prior stimulation or exposure to nicotine. These results support the idea that the smoking-associated variant of the α5 gene may increase the drive to smoke via loss of IP-mediated nicotine aversion.

A 63-year-old man with 6 months of mild left hand rest tremor and bradykinesia and subtle left wrist cogwheel rigidity was diagnosed with idiopathic Parkinson disease (PD). The most profound neuronal degeneration in PD occurs in nigrosome 1, a lens-shaped substructure of the substantia nigra containing approximately 22,000 cell bodies in each hemi-midbrain, measuring 6 × 6 × 1 mm.1,2 A 7T MRI at the caudal level of red nucleus (figure, A) shows nigrosome 1 signal present in the left nigra and absent in the right, consistent with the clinically affected side. 7T MRI of a healthy 63-year-old with normal bilateral nigrosome 1 signal is shown for comparison (figure, B).

Medium spiny neurons (MSNs) constitute 95% of neurons in the dorsal striatum subdivided into direct (striatonigral) and indirect (striatopallidal) pathways. Whereas D1 and D2 receptors and several neuropeptides, including dynorphin and enkephalin, are differentially expressed in these neurons, 5-hydroxytryptamine 6 receptors (5-HT6) are expressed in both pathways. Previous results demonstrate that concurrent 5-HT6 receptor overexpression in MSNs of both pathways in the dorsomedial striatum (DMS) interferes with instrumental learning and that 5-HT6 overexpression in the dorsolateral striatum (DLS) relieves rats from inflexible habitual behaviors. We hypothesized that 5-HT6 receptor-mediated co-activation of both pathways interferes with the differential activation/inhibition of direct/indirect pathways by dopamine. To test this idea, we cloned novel viral vectors to selectively overexpress 5-HT6 receptors in direct or indirect pathway MSNs to deconstruct their role in modulating instrumental learning and habitual responding. We found that increasing 5-HT6 receptor expression in either direct or indirect pathway MSNs of the posterior DMS selectively enhanced or impaired initial acquisition of a discrete instrumental learning task respectively, though all rats were ultimately able to learn the task. In a separate set of experiments, 5-HT6 receptor overexpression in indirect pathway MSNs of the DLS facilitated behavioral flexibility in rats overtrained on a repetitive pressing task using a variable interval schedule of reinforcement, during an omission contingency training session and subsequent probe testing. Together these findings further the notion that 5-HT6 signaling causes balanced activation of opposing MSN pathways by serotonin in sub-regions of the dorsal striatum allowing for more reflective modalities of behavior.

The 5-HT6 receptor has been implicated in a variety of cognitive processes including habitual behaviors, learning, and memory. It is found almost exclusively in the brain, is expressed abundantly in striatum, and localizes to neuronal primary cilia. Primary cilia are antenna-like, sensory organelles found on most neurons that receive both chemical and mechanical signals from other cells and the surrounding environment; however, the effect of 5-HT6 receptor function on cellular morphology has not been examined. We confirmed that 5-HT6 receptors were localized to primary cilia in wild-type (WT) but not 5-HT6 knockout (5-HT6KO) in both native mouse brain tissue and primary cultured striatal neurons then used primary neurons cultured from WT or 5-HT6KO mice to study the function of these receptors. Selective 5-HT6 antagonists reduced cilia length in neurons cultured from wild-type mice in a concentration and time-dependent manner without altering dendrites, but had no effect on cilia length in 5-HT6KO cultured neurons. Varying the expression levels of heterologously expressed 5-HT6 receptors affected the fidelity of ciliary localization in both WT and 5-HT6KO neurons; overexpression lead to increasing amounts of 5-HT6 localization outside of the cilia but did not alter cilia morphology. Introducing discrete mutations into the third cytoplasmic loop of the 5-HT6 receptor greatly reduced, but did not entirely eliminate, trafficking of the 5-HT6 receptor to primary cilia. These data suggest that blocking 5-HT6 receptor activity reduces the length of primary cilia and that mechanisms that regulate trafficking of 5-HT6 receptors to cilia are more complex than previously thought.

Currently there is no general approach for achieving specific optogenetic control of genetically defined cell types in rats, which provide a powerful experimental system for numerous established neurophysiological and behavioral paradigms. To overcome this challenge we have generated genetically restricted recombinase-driver rat lines suitable for driving gene expression in specific cell types, expressing Cre recombinase under the control of large genomic regulatory regions (200-300 kb). Multiple tyrosine hydroxylase (Th)::Cre and choline acetyltransferase (Chat)::Cre lines were produced that exhibited specific opsin expression in targeted cell types. We additionally developed methods for utilizing optogenetic tools in freely moving rats and leveraged these technologies to clarify the causal relationship between dopamine (DA) neuron firing and positive reinforcement, observing that optical stimulation of DA neurons in the ventral tegmental area (VTA) of Th::Cre rats is sufficient to support vigorous intracranial self-stimulation (ICSS). These studies complement existing targeting approaches by extending the generalizability of optogenetics to traditionally non-genetically-tractable but vital animal models.

The nucleus accumbens (NAc) in the ventral striatum integrates many neurochemical inputs including dopamine and serotonin projections from midbrain nuclei to modulate drug reward. Although D1 and D2 dopamine receptors are differentially expressed in the direct and indirect pathway medium spiny neurons (dMSNs and iMSNs, respectively), 5-HT6 receptors are expressed in both pathways, more strongly than anywhere else in the brain, and are an intriguing target for neuropsychiatric disorders. In the present study, we used viral vectors utilizing dynorphin or enkephalin promoters to drive expression of 5-HT6 receptors or green fluorescent protein (GFP) selectively in the dMSNs or iMSNs of the NAc shell. Rats were then trained to self-administer cocaine. Increased 5-HT6 receptor expression in dMSNs did not change any parameter of cocaine self-administration measured. However, increasing 5-HT6 receptors in iMSNs reduced the amount of cocaine self-administered under fixed-ratio schedules, especially at low doses, increased the time to the first response and the length of the inter-infusion interval, but did not alter motivation as measured by progressive ratio 'break point' analysis. Modeling of cocaine pharmacokinetics in NAc showed that increased 5-HT6 receptors in iMSNs reduced the rat's preferred tissue cocaine concentration at each dose. Finally, increased 5-HT6 receptors in iMSNs facilitated conditioned place preference for a low dose of cocaine. We conclude that 5-HT6 receptors in iMSNs of NAcSh increase the sensitivity to the reinforcing properties of cocaine, particularly at low doses, suggesting that these receptors may be a therapeutic target for the treatment of cocaine addiction.

Primary neuronal cultures are a useful tool for measuring pharmacological- and transgene-regulated gene expression; however, accurate measurements can be confounded by heterogeneous cell types and inconsistent transfection efficiency. Here we describe our adaptation of a ribosomal capture strategy that was designed to be used in transgenic mice expressing tagged ribosomal subunits (RiboTag) in specific cell types, thereby allowing measurement of translating RNAs from desired cell types within complex tissues. Using this strategy we were able to isolate and analyze neuron-specific RNA despite the presence of glia by co-transfecting experimental plasmids with plasmids that selectively express RiboTag in neurons. RiboTag immunoprecipitation was capable of recovering high integrity RNA from small numbers of transfected cells that can then be interrogated by a variety of methods (e.g., RT-qPCR, PCR array, RNA-Seq) and compared with basal RNA expression of the entire culture. Additionally, we demonstrate how co-transfection of RiboTag with small hairpin RNA (shRNA) constructs can validate and accurately assess the degree of gene expression knockdown, and how RiboTag can be used to measure receptor-mediated gene regulation with transiently expressed designer receptors exclusively activated by designer drugs (DREADDs). RiboTag co-transfection represents a convenient and powerful tool to isolate RNA from a specific subset of cultured cells with a variety of applications for experiments in vitro.

A new study indicates that deep brain stimulation (DBS) plus best medical therapy markedly improves quality of life of patients with advanced Parkinson disease compared with best medical therapy alone. The frequency of serious adverse events related to DBS was similar to that reported in other DBS trials, underscoring the need for careful patient selection and counseling for this invasive therapy.

Few But Powerful Drug activation of the different types of acetylcholine receptors in cholinergic neurons often generates opposing or conflicting effects. Using optogenetic techniques in transgenic mice, Witten et al. (p. 1677 ) investigated the function of a rather enigmatic subpopulation of cholinergic neurons, the giant interneurons of the nucleus accumbens. Their excitation paradoxically reduced neighboring medium spiny neuron firing, while their inhibition increased medium spiny neuron firing. Furthermore, the giant interneurons were directly activated by cocaine, and silencing their drug-induced activity during cocaine exposure in freely behaving animals disrupted cocaine reward.

Background Treatment of Parkinson disease commonly includes levodopa and dopamine agonists; however, the interaction of these 2 drugs is poorly understood. Objective To examine the effects of a dopamine agonist on the motor response to levodopa. Design Double-blind, randomized, placebo-controlled, crossover clinical trial. Setting Ambulatory academic referral center. Patients Thirteen patients with idiopathic Parkinson disease taking levodopa and experiencing motor fluctuations and dyskinesia. Interventions Eligible individuals were randomly assigned to receive pramipexole dihydrochloride or placebo for 4 weeks followed by a 2-hour intravenous levodopa infusion on consecutive days at 2 rates and with blinded assessments. They were then crossed over to the alternate oral therapy for 4 weeks followed by levodopa infusion and reassessment. Main Outcome Measures Change in finger-tapping speed, measured using the area under the curve (AUC) for finger taps per minute across time; peak finger-tapping speed; duration of response; time to “ON” (defined as a 10% increase in finger-tapping speed above baseline); walking speed; and dyskinesia AUC. Results Pramipexole with levodopa infusion increased finger-tapping speed beyond the change in baseline by a mean (SE) of 170 (47.2) per minute × minutes ( P = .006) and more than doubled the AUC for finger-tapping speed. Pramipexole increased peak finger-tapping speed by a mean (SE) of 18 (8.5) taps per minute ( P = .02) and improved mean (SE) walking speed (15.9 [0.70] vs 18.9 [0.70] seconds, P = .004). Pramipexole prolonged duration of response after levodopa infusion and shortened time to ON. Pramipexole increased mean (SE) baseline dyskinesia scores (26.0 [5.85] vs 12.1 [5.85] points, P = .05) and peak dyskinesia scores with levodopa infusion. Conclusions Pramipexole augmented the motor response to levodopa beyond a simple additive effect and increased the severity of levodopa-induced dyskinesia. When considering a combination of these therapies, an appropriate balance should be maintained regarding gain of motor function vs worsening of dyskinesia. Trial Registration clinicaltrials.gov Identifier:NCT00666653

Importance Coenzyme Q10 (CoQ10), an antioxidant that supports mitochondrial function, has been shown in preclinical Parkinson disease (PD) models to reduce the loss of dopamine neurons, and was safe and well tolerated in early-phase human studies. A previous phase II study suggested possible clinical benefit. Objective To examine whether CoQ10 could slow disease progression in early PD. Design, Setting, and Participants A phase III randomized, placebo-controlled, double-blind clinical trial at 67 North American sites consisting of participants 30 years of age or older who received a diagnosis of PD within 5 years and who had the following inclusion criteria: the presence of a rest tremor, bradykinesia, and rigidity; a modified Hoehn and Yahr stage of 2.5 or less; and no anticipated need for dopaminergic therapy within 3 months. Exclusion criteria included the use of any PD medication within 60 days, the use of any symptomatic PD medication for more than 90 days, atypical or drug-induced parkinsonism, a Unified Parkinson’s Disease Rating Scale (UPDRS) rest tremor score of 3 or greater for any limb, a Mini-Mental State Examination score of 25 or less, a history of stroke, the use of certain supplements, and substantial recent exposure to CoQ10. Of 696 participants screened, 78 were found to be ineligible, and 18 declined participation. Interventions The remaining 600 participants were randomly assigned to receive placebo, 1200 mg/d of CoQ10, or 2400 mg/d of CoQ10; all participants received 1200 IU/d of vitamin E. Main Outcomes and Measures Participants were observed for 16 months or until a disability requiring dopaminergic treatment. The prospectively defined primary outcome measure was the change in total UPDRS score (Parts I-III) from baseline to final visit. The study was powered to detect a 3-point difference between an active treatment and placebo. Results The baseline characteristics of the participants were well balanced, the mean age was 62.5 years, 66% of participants were male, and the mean baseline total UPDRS score was 22.7. A total of 267 participants required treatment (94 received placebo, 87 received 1200 mg/d of CoQ10, and 86 received 2400 mg/d of CoQ10), and 65 participants (29 who received placebo, 19 who received 1200 mg/d of CoQ10, and 17 who received 2400 mg/d of CoQ10) withdrew prematurely. Treatments were well tolerated with no safety concerns. The study was terminated after a prespecified futility criterion was reached. At study termination, both active treatment groups showed slight adverse trends relative to placebo. Adjusted mean changes (worsening) in total UPDRS scores from baseline to final visit were 6.9 points (placebo), 7.5 points (1200 mg/d of CoQ10; P = .49 relative to placebo), and 8.0 points (2400 mg/d of CoQ10; P = .21 relative to placebo). Conclusions and Relevance Coenzyme Q10 was safe and well tolerated in this population, but showed no evidence of clinical benefit. Trial Registration clinicaltrials.gov Identifier:NCT00740714

SummaryPrevailing theory suggests that long-term memories are encoded via a two-phase process requiring early involvement of the hippocampus followed by the neocortex. Contextual fear memories in rodents rely on the hippocampus immediately following training but are unaffected by hippocampal lesions or pharmacological inhibition weeks later. With fast optogenetic methods, we examine the real-time contribution of hippocampal CA1 excitatory neurons to remote memory and find that contextual fear memory recall, even weeks after training, can be reversibly abolished by temporally precise optogenetic inhibition of CA1. When this inhibition is extended to match the typical time course of pharmacological inhibition, remote hippocampus dependence converts to hippocampus independence, suggesting that long-term memory retrieval normally depends on the hippocampus but can adaptively shift to alternate structures. Further revealing the plasticity of mechanisms required for memory recall, we confirm the remote-timescale importance of the anterior cingulate cortex (ACC) and implicate CA1 in ACC recruitment for remote recall.