No abstract

SignificanceNeurons and nonneuronal cells in the developing brain dynamically regulate gene expression as neural connectivity is established. However, the specific gene programs activated in distinct cell populations during the assembly and refinement of many intact neuronal circuits have not been thoroughly characterized. In this study, we take advantage of recent advances in transcriptomic profiling techniques to characterize gene expression in the postnatal developing lateral geniculate nucleus (LGN) at single-cell resolution. Our data reveal that genes involved in brain development are dynamically regulated in all major cell types of the LGN, suggesting that the establishment of neural connectivity depends upon functional collaboration between multiple neuronal and nonneuronal cell types in this brain region.

Sensory experience influences the establishment of neural connectivity through molecular mechanisms that remain unclear. Here, we employ single-nucleus RNA sequencing to investigate the contribution of sensory-driven gene expression to synaptic refinement in the dorsal lateral geniculate nucleus of the thalamus, a region of the brain that processes visual information. We find that visual experience induces the expression of the cytokine receptor Fn14 in excitatory thalamocortical neurons. By combining electrophysiological and structural techniques, we show that Fn14 is dispensable for early phases of refinement mediated by spontaneous activity but that Fn14 is essential for refinement during a later, experience-dependent period of development. Refinement deficits in mice lacking Fn14 are associated with functionally weaker and structurally smaller retinogeniculate inputs, indicating that Fn14 mediates both functional and anatomical rearrangements in response to sensory experience. These findings identify Fn14 as a molecular link between sensory-driven gene expression and vision-sensitive refinement in the brain.

Features of the fission yeast Schizosaccharomyces pombe cAMP/PKA pathway make S. pombe particularly amenable for heterologous expression of cAMP pathway proteins such as GαS subunits and their cognate adenylyl cyclases, PKA catalytic and regulatory subunits, and cyclic nucleotide phosphodiesterases. We have constructed two PKA-repressed reporters for use in high-throughput screens to detect compounds that elevate or reduce PKA activity, thus facilitating the discovery of both inhibitors and activators of these target proteins. Here, we describe steps to construct screening strains and to optimize and conduct these screens.