Probes for SST

Converging evidence suggests that deficits in somatostatin (SST)-expressing neuron signaling contributes to major depressive disorder. Preclinical studies show that enhancing this signaling, specifically at α5 subunit-containing γ-ami­nobutyric acid subtype A receptors (α5-GABAARs), provides a potential means to overcome low SST neuron function. The cortical microcircuit comprises multiple subtypes of inhibitory γ-aminobutyric acid (GABA) neurons and excitatory pyramidal cells (PYCs). In this study, multilabel fluorescence in situ hybridization was used to characterize α5-GABAAR gene expression in PYCs and three GABAergic neuron subgroups – vasoactive intestinal peptide (VIP)-, SST-, and parvalbumin (PV)-expressing cells – in the human and mouse frontal cortex. Across species, we found the majority of gene expression in PYCs (human: 39.7%; mouse: 54.14%), less abundant expression in PV neurons (human: 20%; mouse: 16.33%), and no expression in VIP neurons (0%). Only human SST cells expressed GABRA5, albeit at low levels (human: 8.3%; mouse: 0%). Together, this localization suggests potential roles for α5-GABAARs within the cortical microcircuit: (1) regulators of PYCs, (2) regulators of PV cell activity across species, and (3) sparse regulators of SST cell inhibition in humans. These results will advance our ability to predict the effects of pharmacological agents targeting α5-GABAARs, which have shown therapeutic potential in preclinical animal models.

The spinal dorsal horn (SDH) is comprised of distinct neuronal populations that process different somatosensory modalities. Somatostatin (SST)-expressing interneurons in the SDH have been implicated specifically in mediating mechanical pain. Identifying the transcriptomic profile of SST neurons could elucidate the unique genetic features of this population and enable selective analgesic targeting. To that end, we combined the Isolation of Nuclei Tagged in Specific Cell Types (INTACT) method and Fluorescence Activated Nuclei Sorting (FANS) to capture tagged SST nuclei in the SDH of adult male mice. Using RNA-sequencing (RNA-seq), we uncovered more than 13,000 genes. Differential gene expression analysis revealed more than 900 genes with at least 2-fold enrichment. In addition to many known dorsal horn genes, we identified and validated several novel transcripts from pharmacologically tractable functional classes: Carbonic Anhydrase 12 (Car12), Phosphodiesterase 11 A (Pde11a), and Protease-Activated Receptor 3 (F2rl2). In situ hybridization of these novel genes showed differential expression patterns in the SDH, demonstrating the presence of transcriptionally distinct subpopulations within the SST population. Overall, our findings provide new insights into the gene repertoire of SST dorsal horn neurons and reveal several novel targets for pharmacological modulation of this pain-mediating population and treatment of pathological pain.

KCNQ2/3 channels, ubiquitously expressed neuronal potassium channels, have emerged as indispensable regulators of brain network activity. Despite their critical role in brain homeostasis, the mechanisms by which KCNQ2/3 dysfunction lead to hypersychrony are not fully known. Here, we show that deletion of KCNQ2/3 channels changed PV+ interneurons', but not SST+ interneurons', firing properties. We also find that deletion of either KCNQ2/3 or KCNQ2 channels from PV+ interneurons led to elevated homeostatic potentiation of fast excitatory transmission in pyramidal neurons. Pvalb-Kcnq2 null-mice showed increased seizure susceptibility, suggesting that decreases in interneuron KCNQ2/3 activity remodels excitatory networks, providing a new function for these channels.

Cell-specific molecular investigations of the human brain are essential for understanding the neurobiology of diseases, but are hindered by postmortem conditions and technical challenges. To address these issues we developed a multi-label fluorescence in situ hybridization protocol and a novel optical filter device to identify cell types and control for tissue autofluorescence. We show that these methods can be used with laser-capture microdissection for human brain tissue cell-specific molecular analysis.

The motor cortico-basal ganglion loop is critical for motor planning, execution, and learning. Balanced excitation and inhibition in this loop is crucial for proper motor output. Excitatory neurons have been thought to be the only source of motor cortical input to the striatum. Here, we identify long-range projecting GABAergic neurons in the primary (M1) and secondary (M2) motor cortex that target the dorsal striatum. This population of projecting GABAergic neurons comprises both somatostatin-positive (SOM+) and parvalbumin-positive (PV+) neurons that target direct and indirect pathway striatal output neurons as well as cholinergic interneurons differentially. Notably, optogenetic stimulation of M1 PV+ and M2 SOM+ projecting neurons reduced locomotion, whereas stimulation of M1 SOM+ projecting neurons enhanced locomotion. Thus, corticostriatal GABAergic projections modulate striatal output and motor activity.

The human brain has enormously complex cellular diversity and connectivities fundamental to our neural functions, yet difficulties in interrogating individual neurons has impeded understanding of the underlying transcriptional landscape. We developed a scalable approach to sequence and quantify RNA molecules in isolated neuronal nuclei from a postmortem brain, generating 3227 sets of single-neuron data from six distinct regions of the cerebral cortex. Using an iterative clustering and classification approach, we identified 16 neuronal subtypes that were further annotated on the basis of known markers and cortical cytoarchitecture. These data demonstrate a robust and scalable method for identifying and categorizing single nuclear transcriptomes, revealing shared genes sufficient to distinguish previously unknown and orthologous neuronal subtypes as well as regional identity and transcriptomic heterogeneity within the human brain.