Partial or Complete Loss of Norepinephrine Differentially Alters Contextual Fear and Catecholamine Release Dynamics in Hippocampal CA1.

Background: Contextual fear learning is heavily dependent on the hippocampus. Despite evidence that catecholamines contribute to contextual encoding and memory retrieval, the precise temporal dynamics of their release in the hippocampus during behavior is unknown. In addition, new animal models are required to probe the effects of altered catecholamine synthesis on release dynamics and contextual learning. Methods: We generated 2 new mouse models of altered locus coeruleus-norepinephrine (NE) synthesis and utilized them together with GRABNE and GRABDA sensors and in vivo fiber photometry to investigate NE and dopamine (DA) release dynamics in the dorsal hippocampal CA1 during contextual fear conditioning. Results: Aversive foot shock increased both NE and DA release in the dorsal CA1, while freezing behavior associated with recall of fear memory was accompanied by decreased release. Moreover, we found that freezing at the recent time point was sensitive to both partial and complete loss of locus coeruleus-NE synthesis throughout prenatal and postnatal development, similar to previous observations of mice with global loss of NE synthesis beginning postnatally. In contrast, freezing at the remote time point was compromised only by complete loss of locus coeruleus-NE synthesis beginning prenatally. Conclusions: Overall, these findings provide novel insights into the role of NE in contextual fear and the precise temporal dynamics of both NE and DA during freezing behavior and highlight complex relationships between genotype, sex, and NE signaling.

Comprehensive molecular analysis of 61 Egyptian families with hereditary nonsyndromic hearing loss.

Nonsyndromic hearing loss is an extremely heterogeneous disorder. Thus, clinical diagnostics is challenging, in particular due to differences in the etiology of hearing loss between populations. With this study, we wanted to elucidate the genetic basis of hearing loss in 61 consanguineous Egyptian families. In 25 families, linkage analysis was used as a prescreening to identify regions for targeted sequencing of candidate genes. Initially, the coding regions of 12 and later of 94 genes associated with hearing loss were enriched and subjected to massively parallel sequencing (MPS) with diagnostic yields of 36% and 75%, respectively. Causative variants were identified in 48 families (79%). They were found in 23 different genes with the majority being located in MYO15A (15.3%), SLC26A4 (9.7%), GJB2 (8.3%), and MYO7A (6.4%). As many as 32 variants were novel ones at the time of detection. Five variants were shared by two, three, or even four families. Our study provides a first survey of the mutational spectrum of deaf patients in Egypt revealing less GJB2 variants than in many European populations. It underlines the value of targeted enrichment of well-selected deafness genes in combination with MPS in the diagnostics of this frequent and genetically heterogeneous disorder.

Community Based Efforts to Address Infant Mortality and Disparities in Oklahoma.

IMPORTANCE: Oklahoma's infant mortality remains among the highest in the nation.1 Infant mortality rates are highest within the African American community.2 Physician and community partner efforts to decrease infant mortality are discussed to encourage more involvement in addressing infant mortality. The purpose of this article is to describe both provider and community-based efforts to combat infant mortality, particularly those focused on infant mortality disparities. OBSERVATIONS: The leading causes of infant deaths are prematurity, congenital malformations and/or chromosomal anomalies, and unclassified deaths such as Sudden Infant Death Syndrome or accidents. Prematurity accounts for the highest number of infant deaths. Efforts in Oklahoma focus on prematurity and SIDS prevention. Fetal Infant Mortality Review programs in Oklahoma and Tulsa Counties focus on local issues contributing to infant mortality and promote community engagement. In central Oklahoma, an Infant Mortality Alliance (IMA) was formed including over 180 stakeholders focusing on healthcare access, community and faith engagement, and health disparities. In the year following the IMA's initial work, the non-Hispanic African American infant mortality rate in Oklahoma County decreased by 18.8%.12. CONCLUSIONS AND RELEVANCE: Infant mortality is multifactorial and requires multiple strategies to combat. To address infant mortality and disparities, all aspects of the community must be involved. No individual alone can improve infant mortality. Physicians providing prenatal care make an impact by implementing recommended guidelines for progesterone therapy. Physicians seeing infants can encourage safe sleep practices among their families and local hospitals. While progress has been made addressing Oklahoma's infant mortality, much work remains.

Community Engagement: Why Oklahoma? Why Now?

IMPORTANCE: Oklahoma ranked 43rd in the 2017 America's Health Rankings largely due to health behaviors such as adult obesity, lack of physical activity and smoking. Oklahoma children also suffer from adverse childhood experiences that contribute to poor health outcomes. Community engagement, a process that involves people affiliated by geographic location and shared interest working together to address issues affecting community wellbeing, is a common model that has had some success in Oklahoma communities addressing child health. OBSERVATIONS: Factors that contribute to poor health in Oklahoma include not only health behaviors such as obesity and smoking, but also lack of access to care created by a lack of health insurance and primary care providers, compounded by the largely rural nature of the state. The National Institutes of Health is committed to funding research aimed at improving the health of rural and disadvantaged populations. Historically, these populations are difficult to reach and may not be interested in the national health research initiatives, but rather want to focus on health issues important to their communities. CONCLUSIONS AND RELEVANCE: In this article we discuss some of Oklahoma's most pressing pediatric health needs, community engagement efforts to address these issues and a newly funded NIH grant at OUHSC aimed at supporting and learning from these efforts.

Mutations in STX1B, encoding a presynaptic protein, cause fever-associated epilepsy syndromes.

Febrile seizures affect 2-4% of all children and have a strong genetic component. Recurrent mutations in three main genes (SCN1A, SCN1B and GABRG2) have been identified that cause febrile seizures with or without epilepsy. Here we report the identification of mutations in STX1B, encoding syntaxin-1B, that are associated with both febrile seizures and epilepsy. Whole-exome sequencing in independent large pedigrees identified cosegregating STX1B mutations predicted to cause an early truncation or an in-frame insertion or deletion. Three additional nonsense or missense mutations and a de novo microdeletion encompassing STX1B were then identified in 449 familial or sporadic cases. Video and local field potential analyses of zebrafish larvae with antisense knockdown of stx1b showed seizure-like behavior and epileptiform discharges that were highly sensitive to increased temperature. Wild-type human syntaxin-1B but not a mutated protein rescued the effects of stx1b knockdown in zebrafish. Our results thus implicate STX1B and the presynaptic release machinery in fever-associated epilepsy syndromes.

Deciphering the genetic basis of microcystin tolerance.

BACKGROUND: Cyanobacteria constitute a serious threat to freshwater ecosystems by producing toxic secondary metabolites, e.g. microcystins. These microcystins have been shown to harm livestock, pets and humans and to affect ecosystem service and functioning. Cyanobacterial blooms are increasing worldwide in intensity and frequency due to eutrophication and global warming. However, Daphnia, the main grazer of planktonic algae and cyanobacteria, has been shown to be able to suppress bloom-forming cyanobacteria and to adapt to cyanobacteria that produce microcystins. Since Daphnia's genome was published only recently, it is now possible to elucidate the underlying molecular mechanisms of microcystin tolerance of Daphnia. RESULTS: Daphnia magna was fed with either a cyanobacterial strain that produces microcystins or its genetically engineered microcystin knock-out mutant. Thus, it was possible to distinguish between effects due to the ingestion of cyanobacteria and effects caused specifically by microcystins. By using RNAseq the differentially expressed genes between the different treatments were analyzed and affected KOG-categories were calculated. Here we show that the expression of transporter genes in Daphnia was regulated as a specific response to microcystins. Subsequent qPCR and dietary supplementation with pure microcystin confirmed that the regulation of transporter gene expression was correlated with the tolerance of several Daphnia clones. CONCLUSIONS: Here, we were able to identify new candidate genes that specifically respond to microcystins by separating cyanobacterial effects from microcystin effects. The involvement of these candidate genes in tolerance to microcystins was validated by correlating the difference in transporter gene expression with clonal tolerance. Thus, the prevention of microcystin uptake most probably constitutes a key mechanism in the development of tolerance and adaptation of Daphnia. With the availability of clear candidate genes, future investigations examining the process of local adaptation of Daphnia populations to microcystins are now possible.

Genome-wide quantitative analysis of histone H3 lysine 4 trimethylation in wild house mouse liver: environmental change causes epigenetic plasticity.

In mammals, exposure to toxic or disease-causing environments can change epigenetic marks that are inherited independently of the intrauterine environment. Such inheritance of molecular phenotypes may be adaptive. However, studies demonstrating molecular evidence for epigenetic inheritance have so far relied on extreme treatments, and are confined to inbred animals. We therefore investigated whether epigenomic changes could be detected after a non-drastic change in the environment of an outbred organism. We kept two populations of wild-caught house mice (Mus musculus domesticus) for several generations in semi-natural enclosures on either standard diet and light cycle, or on an energy-enriched diet with longer daylight to simulate summer. As epigenetic marker for active chromatin we quantified genome-wide histone-3 lysine-4 trimethylation (H3K4me3) from liver samples by chromatin immunoprecipitation and high-throughput sequencing as well as by quantitative polymerase chain reaction. The treatment caused a significant increase of H3K4me3 at metabolic genes such as lipid and cholesterol regulators, monooxygenases, and a bile acid transporter. In addition, genes involved in immune processes, cell cycle, and transcription and translation processes were also differently marked. When we transferred young mice of both populations to cages and bred them under standard conditions, most of the H3K4me3 differences were lost. The few loci with stable H3K4me3 changes did not cluster in metabolic functional categories. This is, to our knowledge, the first quantitative study of an epigenetic marker in an outbred mammalian organism. We demonstrate genome-wide epigenetic plasticity in response to a realistic environmental stimulus. In contrast to disease models, the bulk of the epigenomic changes we observed were not heritable.

Frequent mutations in chromatin-remodelling genes in pulmonary carcinoids.

Pulmonary carcinoids are rare neuroendocrine tumours of the lung. The molecular alterations underlying the pathogenesis of these tumours have not been systematically studied so far. Here we perform gene copy number analysis (n=54), genome/exome (n=44) and transcriptome (n=69) sequencing of pulmonary carcinoids and observe frequent mutations in chromatin-remodelling genes. Covalent histone modifiers and subunits of the SWI/SNF complex are mutated in 40 and 22.2% of the cases, respectively, with MEN1, PSIP1 and ARID1A being recurrently affected. In contrast to small-cell lung cancer and large-cell neuroendocrine lung tumours, TP53 and RB1 mutations are rare events, suggesting that pulmonary carcinoids are not early progenitor lesions of the highly aggressive lung neuroendocrine tumours but arise through independent cellular mechanisms. These data also suggest that inactivation of chromatin-remodelling genes is sufficient to drive transformation in pulmonary carcinoids.

Fast isogenic mapping-by-sequencing of ethyl methanesulfonate-induced mutant bulks.

Mapping-by-sequencing (or SHOREmapping) has revitalized the powerful concept of forward genetic screens in plants. However, as in conventional genetic mapping approaches, mapping-by-sequencing requires phenotyping of mapping populations established from crosses between two diverged accessions. In addition to the segregation of the focal phenotype, this introduces natural phenotypic variation, which can interfere with the recognition of quantitative phenotypes. Here, we demonstrate how mapping-by-sequencing and candidate gene identification can be performed within the same genetic background using only mutagen-induced changes as segregating markers. Using a previously unknown suppressor of mutants of like heterochromatin protein1 (lhp1), which in its functional form is involved in chromatin-mediated gene repression, we identified three closely linked ethyl methanesulfonate-induced changes as putative candidates. In order to assess allele frequency differences between such closely linked mutations, we introduced deep candidate resequencing using the new Ion Torrent Personal Genome Machine sequencing platform to our mutant identification pipeline and thereby reduced the number of causal candidate mutations to only one. Genetic analysis of two independent additional alleles confirmed that this mutation was causal for the suppression of lhp1.