Diurnal rhythms across the human dorsal and ventral striatum

The human striatum can be subdivided into the caudate, putamen, and nucleus accumbens (NAc). Each of these structures have some overlapping and some distinct functions related to motor control, cognitive processing, motivation, and reward. Previously, we used a “time of death” approach to identify diurnal rhythms in RNA transcripts in human cortical regions. Here, we identify molecular rhythms across the three striatal subregions collected from postmortem human brain tissue in subjects without psychiatric or neurological disorders. Core circadian clock genes are rhythmic across all three regions and show strong phase concordance across regions. However, the putamen contains a much larger number of significantly rhythmic transcripts than the other two regions. Moreover, there are many differences in pathways that are rhythmic across regions. Strikingly, the top rhythmic transcripts in NAc (but not the other regions) are predominantly snoRNAs and lncRNAs, suggesting that a completely different mechanism might be used for the regulation of diurnal rhythms in translation and/or RNA processing in the NAc versus the other regions. Further, although the NAc and putamen are generally in phase with regards to timing of expression rhythms, the NAc and caudate, and caudate and putamen, have several clusters of discordant rhythmic transcripts, suggesting a temporal wave of specific cellular processes with the caudate preceding the other regions. Taken together, these studies reveal distinct transcriptome rhythms across the human striatum and are an important step in helping to understand the normal function of diurnal rhythms in these regions and how disruption could lead to pathology. Overall design: NAc, caudate, and putamen tissue samples were obtained through the University of Pittsburgh Brain Tissue Donation Program and the NIH NeuroBioBank. Brain tissue was obtained, following consent from the next of kin, during autopsies conducted at the Allegheny County (Pittsburgh, PA) or Davidson County (Nashville, TN) Medical Examiner's Office. All procedures were approved by the University of Pittsburgh Institutional Review Board for Biomedical Research and Committee for Oversight of Research and Clinical Training Involving Decedents. The absence of lifetime psychiatric disorders was determined by an independent committee of experienced clinicians using information from clinical records, toxicology results, and standardized psychological autopsy. Subjects were included based on three criteria: 1) known time of death (TOD) within a 4-hour window and meeting the criteria of rapid death; 2) age less than 65 years; 3) postmortem interval (PMI) less than 30 hours. A total of 60 subjects met these criteria. One subject (ID: 13250) was excluded from all analyses due to overall low expression levels and low correlation with the other subjects (n=59 for all analyses). The right hemisphere of each brain was blocked coronally, immediately frozen, and stored at -80° C. Tissue blocks containing the body of the caudate, putamen, and NAc were selected for analysis. The medial-lateral border between the caudate and putamen was scored to specifically exclude the internal capsule. The ventral border of the caudate and putamen, and accordingly the dorsal border of the NAc, was defined by the anterior thalamic radiation of the internal capsule or the anterior commissure. Cryostat sections were cut at 40 µm thickness, and each striatal subregion from an individual section was placed into its respective collection tube until a total of volume of 35 mm3 was collected from each region. Total RNA was extracted from the tissue samples using a combination of Trizol (Invitrogen, Carlsbad, CA) and RNeasy Lipid Tissue Mini Kit (Qiagen, Hilden, Germany). RNA quantity and quality were assessed using fluorometry (Qubit RNA Broad Range Assay Kit and Fluorometer; Invitrogen, Carlsbad, CA) and chromatography (Bioanalyzer and RNA 6000 Nano Kit; Agilent, Santa Clara, CA), respectively. Please note that 'Corrected.TOD' is time of death corrected by sunrise and sunset time. 'HU' is sample ID which is one-to-one matched to "pair" numbers. 'PMI' is postmortem interval.