Project description:L-3,4-dihydroxyphenylalanine (levodopa) treatment is the major pharmacotherapy for Parkinson's disease. However, almost all patients receiving levodopa eventually develop debilitating involuntary movements (dyskinesia). While it is known that striatal spiny projection neurons (SPNs) are involved in the genesis of this movement disorder, the molecular basis of dyskinesia is not understood. In this study, we identify distinct cell-type-specific gene expression changes that occur in sub-classes of SPNs upon induction of a parkinsonian lesion followed by chronic levodopa treatment. We identify several hundred genes whose expression is correlated with levodopa dose, many of which are under the control of AP-1 and ERK signaling. In spite of homeostatic adaptations involving several signaling modulators, AP-1-dependent gene expression remains highly dysregulated in direct pathway SPNs (dSPNs) upon chronic levodopa treatment. We also discuss which molecular pathways are most likely to dampen abnormal dopaminoceptive signaling in spiny projection neurons, hence providing potential targets for antidyskinetic treatments in Parkinson's disease. To profile the cell-type-specific responses of striatal spiny projection neurons (SPNs) to striatal dopamine depletion, we conducted TRAP analysis of the two major classes of these neurons: dSPNs that express the dopamine receptor 1a (Drd1a), and iSPNs that express the dopamine receptor 2 (Drd2). To disrupt dopamine innervation to both of these SPN populations that reside in the striatum, we injected the neurotoxin 6-hydroxydopamine (6-OHDA), unilaterally, in the medial forebrain bundle (MFB) in hemizygous Drd1-TRAP and Drd2-TRAP adult (9-14 weeks) male mice (kept on a C57BL/6J genetic background). This lesion procedure causes nigral dopamine cell death within a few days, along with a widespread and near-complete loss of dopaminergic innervation to the entire dorsal striatum on one side of the brain (a hemiparkinsonian model). We first examined the effects of dopamine depletion alone, compared to a mock lesion (ascorbate / saline injected). We then examined the effects of chronic levodopa treatment upon the molecular profiles of dopamine- depleted dSPNs and iSPNs, with two dose regimens. The ‘high-dose’ L-DOPA regimen (3 mg/kg on days 1-3, followed by 6 mg/kg on days 4-9) was expected to induce severe dyskinesia in all MFB-lesioned mice. The low-dose L-DOPA regimen (1 mg/kg on days 1-3, followed by 2 mg/kg on days 4-9) was expected to reverse limb use asymmetry without causing conspicuous dyskinesias. To equalize the effects of stress and handling across all groups, including control groups, all mice were equally handled and thus received saline injections when not receiving levodopa injections. Each treatment group contained 7-10 replicates. TRAP-purified mRNAs from either Drd1a- or Drd2-expressing SPNs were reverse-transcribed, amplified, and used to interrogate Affymetrix 430_2.0 GeneChip microarrays.

Project description:The brain executes control of nearly all bodily functions via spinal projecting neurons (SPNs) that carry command signals from numerous supraspinal regions to the spinal cord. Despite their physiological and clinical relevance, a comprehensive molecular characterization of SPNs is still lacking. Here, we use retrograde labeling, whole-brain imaging, and high-throughput transcriptional profiling to generate a unified brain-wide anatomic and transcriptomic atlas of adult mouse SPNs at single-cell resolution. We transcriptionally profiled a total of 65,002 SPNs, identified 76 region-specific SPN types, and mapped these types into a companion atlas of the whole mouse brain generated by the Allen Institute for Brain Science within the BRAIN Initiative Cell Census Network (BICCN). This SPN taxonomy reveals a three-component organization of SPNs: (1) molecularly homogeneous excitatory SPNs from the cortex, red nucleus, and cerebellum with somatotopic spinal terminations suitable for point-to-point communication; (2) highly heterogeneous excitatory and inhibitory populations in the reticular formation with broad spinal termination patterns, thus suitable for relaying commands related to the activities of the entire spinal cord; and (3) modulatory neurons expressing slow-acting neurotransmitters and/or neuropeptides in the hypothalamus, midbrain, and reticular formation for gain control of brain-spinal signals. Within each of these components, this atlas revealed additional insights. From components (2) and (3), we discovered a LIM homeobox transcription factor code that parcellates the most transcriptionally complex population, reticulospinal neurons, into five molecularly distinct and spatially segregated populations. For neurons in component (1), we found transcriptional signatures of a subset of SPNs with large soma size and correlated these with fast-firing electrophysiological properties; thus, at least two different cable lines (namely, Pvalb/Kcng4/Spp1 positive and negative) with different electrophysiological properties might underlie the transmission of brain signals to the spinal cord. Together, by integrating the anatomy, molecular identity, and physiological properties, this study establishes a comprehensive taxonomy of brain-wide SPNs and provides insight into the functional organization of SPNs in mediating brain control of bodily functions.

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A Spatiomolecular Map of the Striatum

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