Contribution of neurons that express fruitless and Clock transcription factors to behavioral rhythms and courtship.

Animals need to integrate information across neuronal networks that direct reproductive behaviors and circadian rhythms. In Drosophila, the master regulatory transcription factors that direct courtship behaviors and circadian rhythms are co-expressed in a small set of neurons. In this study we investigate the role of these neurons in both males and females. We find sex-differences in the number of these fruitless and Clock -expressing neurons ( fru ∩ Clk neurons) that is regulated by male-specific Fru. We assign the fru ∩ Clk neurons to the electron microscopy connectome that provides high resolution structural information. We also discover sex-differences in the number of fru -expressing neurons that are post-synaptic targets of Clk -expressing neurons, with more post-synaptic targets in males. When fru ∩ Clk neurons are activated or silenced, males have a shorter period length. Activation of fru ∩ Clk neurons also changes the rate a courtship behavior is performed. We find that activation and silencing fru ∩ Clk neurons impacts the molecular clock in the sLNv master pacemaker neurons, in a cell-nonautonomous manner. These results reveal how neurons that subserve the two processes, reproduction and circadian rhythms, can impact behavioral outcomes in a sex-specific manner.

Utilizing Peripheral Nerve Blocks for Pain Management in Pediatric Patients during Embolization and Sclerotherapy for Vascular Malformations.

Vascular anomalies are a diverse group of abnormal blood vessel developments that can occur at birth or shortly afterward. Embolization and sclerotherapy have been utilized as a treatment option for these malformations but may cause moderate-to-severe pain. This study aims to evaluate the utilization of peripheral nerve blocks in opioid consumption, pain scores, and length of stay. A retrospective chart review was conducted at the UPMC Children's Hospital of Pittsburgh for all patients who underwent embolization and sclerotherapy between 2011 and 2020. Patient data were collected to compare opioid consumption, pain scores, and length of stay. In total, 854 procedures were performed on 347 patients. The morphine milligram equivalent per kilogram mean difference between groups was 0.9 (0.86, 0.95) with a p-value of <0.001. The pain score mean ratio was -1.17 (-2.2, -0.1) with a p-value of 0.027. The length of stay had an incident rate ratio of 0.94 (0.4, 2) and a p-value of 0.875. By decreasing opioid consumption and postoperative pain scores, peripheral nerve blocks may have utility in patients undergoing embolization and sclerotherapy while not clinically increasing the length of stay for patients. Their use should be individualized and carefully discussed with the interventional radiologist.

Dietary Conjugated Linoleic Acid Reduces Body Weight and Fat in Snord116m+/p- and Snord116m-/p- Mouse Models of Prader-Willi Syndrome.

Prader-Willi Syndrome (PWS) is a human genetic condition that affects up to 1 in 10,000 live births. Affected infants present with hypotonia and developmental delay. Hyperphagia and increasing body weight follow unless drastic calorie restriction is initiated. Recently, our laboratory showed that one of the genes in the deleted locus causative for PWS, Snord116, maintains increased expression of hypothalamic Nhlh2, a basic helix-loop-helix transcription factor. We have previously also shown that obese mice with a deletion of Nhlh2 respond to a conjugated linoleic acid (CLA) diet with weight and fat loss. In this study, we investigated whether mice with a paternal deletion of Snord116 (Snord116m+/p-) would respond similarly. We found that while Snord116m+/p- mice and mice with a deletion of both Snord116 alleles were not significantly obese on a high-fat diet, they did lose body weight and fat on a high-fat/CLA diet, suggesting that the genotype did not interfere with CLA actions. There were no changes in food intake or metabolic rate, and only moderate differences in exercise performance. RNA-seq and microbiome analyses identified hypothalamic mRNAs, and differentially populated gut bacteria, that support future mechanistic analyses. CLA may be useful as a food additive to reduce obesity in humans with PWS.

Inactivating NHLH2 variants cause idiopathic hypogonadotropic hypogonadism and obesity in humans.

Metabolism has a role in determining the time of pubertal development and fertility. Nonetheless, molecular/cellular pathways linking metabolism/body weight to puberty/reproduction are unknown. The KNDy (Kisspeptin/Neurokinin B/Dynorphin) neurons in the arcuate nucleus of the hypothalamus constitute the GnRH (gonadotropin-releasing hormone) pulse generator. We previously created a mouse model with a whole-body targeted deletion of nescient helix-loop-helix 2 (Nhlh2; N2KO), a class II member of the basic helix-loop-helix family of transcription factors. As this mouse model features pubertal failure and late-onset obesity, we wanted to study whether NHLH2 represents a candidate molecule to link metabolism and puberty in the hypothalamus. Exome sequencing of a large Idiopathic Hypogonadotropic Hypogonadism cohort revealed obese patients with rare sequence variants in NHLH2, which were characterized by in-silico protein analysis, chromatin immunoprecipitation, and luciferase reporter assays. In vitro heterologous expression studies demonstrated that the variant p.R79C impairs Nhlh2 binding to the Mc4r promoter. Furthermore, p.R79C and other variants show impaired transactivation of the human KISS1 promoter. These are the first inactivating human variants that support NHLH2's critical role in human puberty and body weight control. Failure to carry out this function results in the absence of pubertal development and late-onset obesity in humans.

Snord116 Post-transcriptionally Increases Nhlh2 mRNA Stability: Implications for Human Prader-Willi Syndrome.

The smallest genomic region causing Prader-Willi Syndrome (PWS) deletes the non-coding RNA SNORD116 cluster; however, the function of SNORD116 remains a mystery. Previous work in the field revealed the tantalizing possibility that expression of NHLH2, a gene previously implicated in both obesity and hypogonadism, was downregulated in PWS patients and differentiated stem cells. In silico RNA: RNA modeling identified several potential interaction domains between SNORD116 and NHLH2 mRNA. One of these interaction domains was highly conserved in most vertebrate NHLH2 mRNAs examined. A construct containing the Nhlh2 mRNA, including its 3'-UTR, linked to a c-myc tag was transfected into a hypothalamic neuron cell line in the presence and absence of exogenously-expressed Snord116. Nhlh2 mRNA expression was upregulated in the presence of Snord116 dependent on the length and type of 3'UTR used on the construct. Furthermore, use of actinomycin D to stop new transcription in N29/2 cells demonstrated that the upregulation occurred through increased stability of the Nhlh2 mRNA in the 45 minutes immediately following transcription. In silico modeling also revealed that a single nucleotide variant (SNV) in the NHLH2 mRNA could reduce the predicted interaction strength of the NHLH2:SNORD116 diad. Indeed, use of an Nhlh2 mRNA construct containing this SNV significantly reduces the ability of Snord116 to increase Nhlh2 mRNA levels. For the first time, these data identify a motif and mechanism for SNORD116-mediated regulation of NHLH2, clarifying the mechanism by which deletion of the SNORD116 snoRNAs locus leads to PWS phenotypes.

Effects of copper hydroxychloride on growth performance and abundance of genes involved in lipid metabolism of growing pigs.

An experiment was conducted to test the hypothesis that copper (Cu) hydroxychloride improves growth performance by upregulating the mRNA transcription of genes involved in lipid metabolism of pigs fed a diet based on corn, soybean meal (SBM), and distillers dried grains with solubles (DDGS). Thirty-two pigs (15.05 ± 0.98 kg) were allotted to 2 dietary treatments with 2 pigs per pen for a total of 8 replicate pens per treatment. Pigs were fed a corn-SBM-DDGS control diet that included Cu to meet the requirement. A second diet was formulated by adding 150 mg Cu/kg from copper hydroxychloride to the control diet. On the last day of the experiment, one pig per pen was sacrificed, and samples from liver, skeletal muscle, and subcutaneous adipose tissue were collected to analyze relative mRNA abundance of genes involved in lipid metabolism. Results indicated that overall ADG and G:F were greater (P < 0.05) for pigs fed the diet containing copper hydroxychloride compared with pigs fed the control diet. Pigs fed the diet supplemented with copper hydroxychloride also had increased (P < 0.05) abundance of cluster of differentiation 36 in the liver and increased (P < 0.05) abundance of fatty acid-binding protein 4 and lipoprotein lipase in subcutaneous adipose tissue. Inclusion of copper hydroxychloride also tended to increase (P < 0.10) the abundance of fatty acid-binding protein 1, peroxisome proliferator-activated receptor α, and carnitine palmitoyltransferase 1B in the liver, skeletal muscle, and subcutaneous adipose tissue, respectively. This indicates that dietary Cu may affect signaling pathways associated with lipid metabolism by improving the uptake, transport, and utilization of fatty acids. In conclusion, supplementation of copper hydroxychloride to the control diet improved growth performance and upregulated the abundance of some genes involved in postabsorptive metabolism of lipids.

Reinforcer pathology: Narrative of hurricane-associated loss increases delay discounting, demand, and consumption of highly palatable snacks in the obese.

Reinforcer pathology is derived from the integration of two measures: (a) self-control (i.e., delay discounting), and (b) reward valuation (i.e., behavioral economic demand). Narrative theory asserts that vividly imagining oneself in a hypothetical, yet realistic, scenario can acutely alter decision making, valuation of reinforcers such as food, and how much food is consumed. The present study measured changes in reinforcer pathology for highly palatable snacks following either a negative or neutral scenario in obese individuals. Participants (N = 48), with a body mass index of 30 or greater, rated their liking of 7 calorie-dense snack items and completed discounting and purchase demand tasks for their top-rated snack. Participants then read a randomly assigned hypothetical scenario (i.e., a devastating hurricane [negative] or minor storm [neutral]), completed the tasks again, and were given ad libitum access to their top 3 ranked snack foods. Results indicated that delay discounting, demand for participants' top-rated snack food, and negative affect all increased in the hurricane group compared with the minor storm group. The hurricane group also consumed more calories, even when hunger was standardized with a preload meal bar. Consistent with reinforcer pathology, these results suggest that vivid consideration of a devastating scenario constricts the temporal window and increases demand for hedonic snack foods among obese individuals. (PsycINFO Database Record (c) 2020 APA, all rights reserved).

Effects of copper hydroxychloride and distillers dried grains with solubles on intestinal microbial concentration and apparent ileal and total tract digestibility of energy and nutrients by growing pigs1.

An experiment was conducted to test the hypothesis that Cu hydroxychloride improves nutrient digestibility and alters the concentration of microbial protein in the small intestine or large intestine by pigs fed a corn-soybean meal diet or a diet based on corn, soybean meal, and distillers dried grains with solubles (DDGS). Twenty-four barrows (33.3 ± 3.4 kg) that had a T-cannula installed in the distal ileum were allotted to a 2 × 2 factorial design with 2 levels of DDGS (0% or 45%) and 2 levels of supplemental Cu from Cu hydroxychloride (0 or 150 mg/kg). A 2-period switch back design with the 4 diets and 6 replicate pigs per diet in each period was used resulting in 12 replicate pigs per diet for the 2 periods. The initial 9 d of each period was considered an adaptation period to the experimental diets. For each period, feces were collected on days 10, 11, and 12, and ileal digesta were collected for 8 h on days 13 and 14. Results indicated that inclusion of 45% DDGS to diets reduced (P < 0.05) the apparent ileal digestibility (AID) of AA and the AID and the apparent total tract digestibility (ATTD) of dry matter, gross energy, and crude protein. In contrast, inclusion of DDGS to diets increased (P < 0.05) the AID and the ATTD of acid hydrolyzed ether extract and the concentration of microbial protein in the hindgut (P < 0.05). However, the total concentration of volatile fatty acids (VFA) in ileal digesta and in feces from pigs fed the DDGS diets were not different from concentrations in pigs fed diets without DDGS. The AID and ATTD of dry matter, gross energy, and crude protein were not affected by dietary Cu concentrations, but the AID and ATTD of acid hydrolyzed ether extract were greater (P < 0.05) in diets supplemented with Cu hydroxychloride compared with diets without Cu hydroxychloride. There was also a reduction (P < 0.05) in the concentration of microbial protein and a tendency for a reduction (P < 0.10) in the total concentration of VFA in feces when diets were supplemented with Cu hydroxychloride. In conclusion, supplementation of Cu hydroxychloride to diets improved AID and ATTD of acid hydrolyzed ether extract and reduced the concentration of microbial protein in the large intestine and this effect was observed in diets containing DDGS as well as in diets without DDGS.

Bovine Hemoglobin-Based Oxygen Carrier Treatment in a Severely Anemic Jehovah's Witness Patient After Cystoprostatectomy and Nephrectomy: A Case Report.

Patients who are Jehovah's Witnesses are known to the medical community for frequently declining blood products, even at times of life-threatening anemia. Alternatives to red blood cell transfusion are being developed, including hemoglobin (Hb)-based oxygen carriers. We present the case of a 77-year-old male Jehovah's Witness who underwent a cystoprostatectomy and radical nephrectomy with a postoperative Hb nadir of 4.5 g/dL. He received an Hb-based oxygen carrier, PEGylated carboxyhemoglobin bovine (Sanguinate), with gradual improvement in anemia symptoms and eventual discharge to a short-term rehabilitation facility.

Phylogenetic Analysis of the SNORD116 Locus.

The SNORD116 small nucleolar RNA locus (SNORD116@) is contained within the long noncoding RNA host gene SNHG14 on human chromosome 15q11-q13. The SNORD116 locus is a cluster of 28 or more small nucleolar (sno) RNAs; C/D box (SNORDs). Individual RNAs within the cluster are tandem, highly similar sequences, referred to as SNORD116-1, SNORD116-2, etc., with the entire set referred to as SNORD116@. There are also related SNORD116 loci on other chromosomes, and these additional loci are conserved among primates. Inherited chromosomal 15q11-q13 deletions, encompassing the SNORD116@ locus, are causative for the paternally-inherited/maternally-imprinted genetic condition, Prader-Willi syndrome (PWS). Using in silico tools, along with molecular-based and sequenced-based confirmation, phylogenetic analysis of the SNORD116@ locus was performed. The consensus sequence for the SNORD116@ snoRNAs from various species was determined both for all the SNORD116 snoRNAs, as well as those grouped using sequence and location according to a human grouping convention. The implications of these findings are put in perspective for studying SNORD116 in patients with inherited Prader-Willi syndrome, as well as model organisms.