No abstract

A remarkable molecular and functional heterogeneity of the primary sensory neurons and dorsal horn interneurons transmits pain- and or itch-relevant information, but the molecular signature of the projection neurons that convey the messages to the brain is unclear. Here, using retro-TRAP (translating ribosome affinity purification) and RNA sequencing, we reveal extensive molecular diversity of spino- and trigeminoparabrachial projection neurons. Among the many genes identified, we highlight distinct subsets of Cck+-, Nptx2+-, Nmb+-, and Crh+-expressing projection neurons. By combining in situ hybridization of retrogradely labeled neurons with Fos-based assays, we also demonstrate significant functional heterogeneity, including both convergence and segregation of pain- and itch-provoking inputs into molecularly diverse subsets of NK1R- and non–NK1R-expressing projection neurons.

The barrel cortex is within the primary somatosensory cortex of the rodent, and processes signals from the vibrissae. Much focus has been devoted to the function of neurons, more recently, the role of glial cells in the processing of sensory input has gained increasing interest. Microglia are the principal immune cells of the nervous system that survey and regulate the cellular constituents of the dynamic nervous system. We investigated the normal and disrupted development of microglia in barrel cortex by chronically depriving sensory signals via whisker trimming for the animals’ first postnatal month. Using immunohistochemistry to label microglia, we performed morphological reconstructions as well as densitometry analyses as a function of developmental age and sensory experience. Findings suggest that both developmental age and sensory experience has profound impact on microglia morphology. Following chronic sensory deprivation, microglia undergo a morphological transition from a monitoring or resting state to an altered morphological state, by exhibiting expanded cell body size and retracted processes. Sensory restoration via whisker regrowth returns these morphological alterations back to age‐matched control values. Our results indicate that microglia may be recruited to participate in the modulation of neuronal structural remodeling during developmental critical periods and in response to alteration in sensory input.

The implementation of negative consequences for drug seeking can result in its cessation just as they might in human addicts. Similarly, exposure to drug cues can lead to resumption of drug seeking. This model may be useful for studying the mechanisms underlying abstinence and relapse and for developing strategies to prevent relapse.

A remarkable molecular and functional heterogeneity of the primary sensory neurons and dorsal horn interneurons transmits pain-and or itch-relevant information, but the molecular signature of the projection neurons that convey the messages to the brain is unclear. Here, using retro-TRAP (translating ribosome affinity purification) and RNA-seq we reveal extensive molecular diversity of spino-and trigeminoparabrachial projection neurons, which to date are almost exclusively defined by their expression of the neurokinin 1 receptor (NK1R). Among the many genes identified, we highlight distinct subsets of Cck+, Nptx2+, Nmb+, and Crh+ expressing projection neurons. By combining in situ hybridization of retrogradely labeled neurons with Fos-based assays we also demonstrate significant functional heterogeneity, including both convergence and segregation of pain-and itch-provoking inputs onto molecularly diverse subsets of NK1R-and non-NK1R-expressing projection neurons. The current study provides the first comprehensive investigation into the molecular profiles and functional properties of projection neuron subtypes.2007; Jansen and Giesler, 2015;Moser and Giesler, 2014;Simone et al., 2004), with separate populations, in primates, responding to the pruritogens, histamine and cowage (Davidson et al., 2012(Davidson et al., , 2007. From a molecular perspective, however, the projection neurons are often regarded as relatively homogeneous. With few exceptions, the neurokinin 1 receptor (NK1R) defines projection neurons in lamina I and the lateral spinal nucleus (LSN) (Carstens et al., 2010; Mantyh et al., 1997). On the other hand, there is evidence for subsets of NK1R neurons, based on projection target (Huang et al., 2019) or molecular make-up (Blomqvist and Mackerlova, 1995; Cameron et al., 2015; Gamboa-Esteves et al., 2004; Huang et al., 2019). Most recently, using unbiased single cell transcriptomics, Häring et al. (2018) identified an excitatory neuron cluster (Glut15) that includes NK1R-expressing spinoparabrachial neurons. This cluster included Lypd1, a forebrain protein implicated in anxiety disorders (Tekinay et al., 2009). However, as Lypd1 labels ~95% of spinoparabrachial neurons (Häring et al., 2018), it likely does not define a functionally distinct subset.The approach that we took involved a sequential series of filtering steps. First, we performed projection neuron-centric RNA-sequencing. Using retro-TRAP (Translating Ribosome Affinity Purification), we purified lateral parabrachial (LPb)-projecting neurons from the spinal cord and TNC, and generated RNA-seq datasets of candidate projection neuron marker genes. As many of these genes are also expressed by spinal cord and TNC interneurons, in the next filtering step we verified the projection neuron hits by combining retrograde tracing and multiplexed in situ hybridization. The latter studies were qualitative in nature, but they identified genes that establish molecular heterogeneity of projection neurons. Lastly, we performed functional studies using TRAP2 mice...