Dense functional and molecular readout of a circuit hub in sensory cortex

INTRODUCTION: The diversity of cell types is a defining feature of the neuronal circuitry that makes up the areas and layers of the mammalian cortex. At a molecular level, the extent of this diversity is now better appreciated through recent efforts to census all potential cortical cell types through single-cell transcriptional profiling. Cortical populations can be hierarchically subdivided into multiple putative transcriptomic cell classes, subclasses, and types. This new catalog of neuronal subclasses and subtypes opens up new questions and avenues of investigation for how these cell types are collectively organized into circuits that function to process information and adapt to changes in experience. RATIONALE: We investigated the function of newly identified cell types in layers 2 or 3 (L2/3) of the primary somatosensory cortex, a region that integrates bottom-up sensory information with top-down internal representations. Current in vivo methods primarily allow cell types to be investigated one at a time and have limited ability to label cell types defined by combinations of expressed genes. To densely survey these cell types and investigate how they interact during task behavior, we developed a platform, Comprehensive Readout of Activity and Cell Type Markers (CRACK), that combines population calcium imaging with subsequent multiplexed fluorescent in situ hybridization. Multiplexed labeling of mRNA transcripts is critical to deciphering the identity of cell types defined by combinatorial patterns of gene expression. RESULTS: We profiled the functional responses of three excitatory cell types and eight inhibitory subclasses in L2/3 as mice performed a whisker-based tactile working memory task. Task-related properties of both excitatory and inhibitory neurons continue to differentiate as they are segregated into increasingly discrete molecular types. Our analysis revealed that the excitatory cell type, L2/3 intratelencephalic Baz1a (Baz1a), functions as a highly active detector of tactile features. Simultaneous imaging across identified cell types enabled measurements of functional connectivity between subpopulations. Functional connectivity analysis indicated that Baz1a neurons orchestrate local network activity patterns. We found that Baz1a neurons show strong functional connections with dendrite-targeting, somatostatin-expressing (Sst) inhibitory neurons. Trans-monosynaptic viral tracing confirmed that Baz1a neurons preferentially synapse onto Sst neurons. Baz1a neurons also show enrichment of select plasticity-related, immediate early genes, including Fos. To determine whether the expression pattern of immediate early genes is a stable property of Baz1a neurons and how this relates to neuronal plasticity, we tracked Fos expression and neuronal activity in mice subjected to whisker deprivation. We found that Baz1a neurons homeostatically adapt to sensory deprivation while stably maintaining Fos expression. CONCLUSION: These results demonstrate that Baz1a neurons are a component of a molecularly defined circuit motif that is capable of recruiting local circuits for sensory processing when salient features are encountered during behavior. This cell type also functions to preserve sensory representations during ongoing and altered sensory experience. This builds on our knowledge for how local circuits in somatosensory cortex are implemented to negotiate bottom-up and top-down information. The ability to map functional and transcriptional relationships across neuronal populations provides insight into how the organizing principles of the cortex give rise to the computations it performs.