Virus-induced secretion of proinflammatory chemokines (e.g., regulated on activation, normal T cells expressed and secreted [RANTES], interleukin [IL]-8) by airway epithelial cells helps to initiate antiviral responses and airway inflammation by enhancing inflammatory cell recruitment. To define mechanisms for virus-induced chemokine secretion, monolayers of nontransformed bronchial epithelial cells were transfected or incubated with polydeoxyinosinic-deoxycytidylic acid (synthetic double-stranded [ds] RNA), rhinovirus dsRNA, or single-stranded RNA (ssRNA), and the secretion of selected chemokines was determined. Transfection or incubation with dsRNA, but not ssRNA, significantly enhanced secretion of RANTES and IL-8, but not eotaxin or macrophage inflammatory protein-1alpha. Mechanistically, dsRNA induced and activated dsRNA-dependent protein kinase (PKR), and activated nuclear factor-kappaB and p38 mitogen-activated protein kinase. Furthermore, the PKR inhibitor 2-aminopurine significantly blocked dsRNA-induced RANTES and IL-8 secretion, whereas the p38 mitogen-activated protein kinase inhibitor SB203580 suppressed dsRNA-induced IL-8, but not RANTES. These findings indicate that dsRNA selectively induce the secretion of chemokines such as IL-8 and RANTES, and implicate dsRNA-sensitive signaling proteins in this process. Moreover, these data suggest that this may be an important mechanism for the selective secretion of chemokines by viruses (e.g., rhinovirus, respiratory syncytial virus, influenza) that synthesize dsRNA during replication.

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Contributions RK, ASF, JAO, PHR and NHK conceptualized the study. NHK and ASF oversaw the study. RK, MKR, and EMF collected behavioral data. RK and DAF developed the rapid staining laser-capture microdissection (LCM) microscopy method and collected the RNA data. DAF performed RNA extractions. JAK and his group performed RNA-Seq. TS aligned the RNA-Seq data. RK and TS analyzed the RNA-Seq data. RK, MKR, and PHR collected tissue and PHR assessed cortisol. JLF, NHK, and RK conceptualized the stereology study. RK and CEG collected and analyzed the stereology data. JLF performed retrograde tracer surgeries and collected injected tissue. RK performed triple labeling of tracing experiments and microscopic analysis. RK and NHK wrote the paper..

Alterations in central extended amygdala (EAc) function have been linked to anxiety, depression, and anxious temperament (AT), the early-life risk to develop these disorders. The EAc is composed of the central nucleus of the amygdala (Ce), the bed nucleus of the stria terminalis (BST), and the sublenticular extended amygdala (SLEA). Using a non-human primate model of AT and multimodal neuroimaging, the Ce and the BST were identified as key AT-related regions. Both areas are primarily comprised of GABAergic neurons and the lateral Ce (CeL) and lateral BST (BSTL) have among the highest expression of neuropeptides in the brain. Somatostatin (SST) is of particular interest because mouse studies demonstrate that SST neurons, along with corticotropin releasing factor (CRF) neurons, contribute to a threat-relevant EAc microcircuit. Although the distribution of CeL and BSTL SST neurons has been explored in rodents, this system is not well described in non-human primates. In situ hybridization demonstrated an anteriorposterior gradient of SST mRNA in the CeL but not the BSTL of non-human primates. Triple labeling immunofluorescence staining revealed that SST protein expressing cell bodies are a small proportion of the total CeL and BSTL neurons and have considerable co-labeling with CRF. The SLEA exhibited strong SST mRNA and protein expression, suggesting a role for SST in mediating

ContributionsRK, ASF, JAO, PHR and NHK conceptualized the study. NHK and ASF oversaw the study. RK, MKR, and EMF collected behavioral data. RK and DAF developed the rapid staining laser-capture microdissection (LCM) microscopy method and collected the RNA data. DAF performed RNA extractions. JAK and his group performed RNA-Seq. TS aligned the RNA-Seq data. RK and TS analyzed the RNA-Seq data. RK, MKR, and PHR collected tissue and PHR assessed cortisol. JLF, NHK, and RK conceptualized the stereology study. RK and CEG collected and analyzed the stereology data. JLF performed retrograde tracer surgeries and collected injected tissue. RK performed triple labeling of tracing experiments and microscopic analysis. RK and NHK wrote the paper. SummaryChildren exhibiting extreme anxious temperament (AT) are at an increased risk to develop anxiety and depression. Work in young rhesus monkeys mechanistically links the central nucleus of the amygdala (Ce) to AT. Here, we used laser capture microscopy and RNA sequencing in 47 young rhesus monkeys to investigate AT's molecular underpinnings by focusing on lateral Ce (CeL) neurons. We found 528 AT-related transcripts, including protein kinase C type-delta (PKCd), a CeL microcircuit cell marker implicated in rodent threat processing. We characterized PKCd neurons in the rhesus CeL, compared their distribution to the mouse, and demonstrated that a subset of these neurons project to the laterodorsal bed nucleus of the stria terminalis (BSTLd).These findings present evidence in the primate of a CeL to BSTLd circuit that maybe relevant to understanding human anxiety and points to specific molecules within this circuit that could serve as potential treatment targets for anxiety disorders. Keywordsfear, anxiety, stress, central nucleus of the amygdala, bed nucleus of the stria terminalis, microcircuitry, protein kinase C type delta, PKCδ, somatostatin, retrograde tracing