bluuebunny/crossref_metadata_2025 huggingface dataset:
Microglia Modulate Changes in Dorsal Motor Nucleus (DMNV) Neuron Excitability Following Sustained Hypoxia Exposure During a Critical Period of Neonatal Development
Authors: Mayer Catherine , Mac Farlane Peter
Published: June 2021

We showed previously that the respiratory neural control system exhibits a lethal vulnerability to sustained hypoxia (SH) exposure during the second postnatal week of life (SH=11% O2, postnatal (P) day 11–15). The vulnerability was associated with increased microglia expression and decreased serotonin (5‐HT) expression in the dorsal motor nucleus of vagus (DMNV), an area of the brainstem containing vagal preganglionic neurons which provide parasympathetic outflow to the heart (and viscera). We hypothesized that this increased microglia expression was associated with increased activity of DMNV neurons and that this could be reversed by treatment with minocycline, an inhibitor of microglia activation. We used whole‐cell patch clamp recordings to examine the excitatory properties of DMNV neurons in neonatal (P16) rats treated with SH from P11–P15. A subset of these animals received subQ injections of minocycline on P11, P13, and P15. SH exposure increased the frequency and amplitude of spontaneous EPSCs in DMNV neurons compared to normoxic (NX) rats; rise time, decay time, and area of the spontaneous EPSC's did not differ between groups. Minocycline treatment during SH prevented the increase in frequency and amplitude seen with SH exposure alone. In conclusion, neurons from the DMNV of SH exposed animals exhibit increased excitability suggesting increased vagal tone in vivo – an effect that appears modulated by microglia. These data may be important to understanding the potential effects of inadequate oxygenation on parasympathetic modulation of heart rate during particularly vulnerable periods of cardiorespiratory development.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
The Transcriptomic Profiles of Microglia from Respiratory Control Centers in Adult Offspring Exposed to Gestational Intermittent Hypoxia Differ by Sex and CNS Region
Authors: Ewald Andrea C , Kiernan Elizabeth A , Baker Tracy L , Watters Jyoti J
Published: June 2021

The in utero environment is well‐known to influence CNS development and microglial function, effects that can last into adulthood. However, fetal brain reprogramming has never been evaluated in the context of sleep‐disordered breathing (SDB) during pregnancy, despite its alarming increase in recent years and the detrimental consequences to the newborn. Here, we tested the hypothesis that maternal exposure to intermittent hypoxia during pregnancy would alter the microglial transcriptome of the adult offspring in CNS regions critical for respiratory neural control. To do this, we utilized a rat model of gestational intermittent hypoxia (GIH) in which dams received intermittent hypoxia (2 min alternating 10.5%/21% O2) or normoxia (2 min alternating 21% O2/21% O2; GNX) for 8 hrs/day from gestational days 10–21. Male and female offspring were grown to adulthood (8–12 wks old) and then challenged with vehicle (HBSS) or lipopolysaccharide (LPS; 1mg/kg, 3h) in order to assess transcriptional responses in both basal and inflammatory conditions. Microglia were immunomagnetically isolated from adult brainstem and cervical spinal cord and total RNA harvested. Sequencing libraries were generated using NEBNext Ultra RNA Library Prep Kit and Illumina paired‐end RNA‐sequencing was performed. The reference genome was built using Bowtie and clean reads were aligned to the reference genome using TopHat. FPKM quantification, differential gene expression levels, and gene ontology analyses in brainstem and spinal cord microglia from GNX and GIH male and female offspring were performed using the DESeq or GOSeq R packages, respectively. Preliminary bioinformatics analyses indicate strong differences in the normal transcriptomes of GNX brainstem and cervical spinal cord microglia. Further, we observed significant sex differences both in basal and LPS‐stimulated microglial gene expression in adult GNX microglia from both CNS regions. Interestingly, GIH potently up and down‐regulated more genes in female spinal cord microglia than in male, and effects of GIH were not detected in the brainstem in either sex. Together, these data suggest that the offspring microglial transcriptome can be changed by maternal exposure to intermittent hypoxia and that these effects can persist into adulthood. Given the importance of microglia‐neuron communication in the neural control of breathing, these microglial gene alterations may impact respiratory and brain immune function in the adult offspring.

Support or Funding Information

Supported by NS085226, HL105511 and Wisconsin Alumni Research Foundation UW2020 grant.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Increasing the excitability of V2a neurons restores diaphragm function following spinal cord injury
Authors: JENSEN VICTORIA N , Alilain Warren , Crone Steven A
Published: June 2021

We hypothesize that ipsilaterally projecting glutamatergic V2a neurons in the ventral spinal cord and brainstem contribute to recovery of diaphragm function following injury. To test our hypothesis, we used a transgenic mouse line that expresses DREADDs in V2a neurons in order to increase their excitability via activation of Gq signaling pathways following injection of the drug‐like molecule clozapine‐N‐oxide (CNO). We performed a high level C2 hemisection (C2Hx) spinal cord injury to paralyze the diaphragm ipsilateral to injury. Electromyography (EMG) recordings of the ipsilateral diaphragm confirmed paralysis. However, increasing V2a neuron excitability by intraperitoneal injection of CNO restored rhythmic burst activity to the paralyzed diaphragm within hours or days after injury. Moreover, the contralateral (uninjured side) diaphragm is able to maintain regular rhythmic breathing when V2a neuron activity is altered. These results indicate that targeting Gq signaling pathways in V2a neurons has the potential to restore function to respiratory muscles following spinal cord injury without significant adverse side effects on respiratory rhythm generation.

Support or Funding Information

UK Center for Clinical and Translational Science, Cincinnati Children's Hospital Medical Center Division of Neurosurgery, Albert J. Ryan Fellowship, University of Cincinnati Neuroscience Graduate Program

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
No Effect of Adenosine or Serotonin Receptor Blockade on the Biphasic Hypoxic Ventilatory Response of Neonatal Rats
Authors: Memishian Wendy , Turnage Mary Buford , Bavis Ryan W.
Published: June 2021

Newborn mammals generally exhibit a biphasic hypoxic ventilatory response (HVR) in which an initial increase in ventilation is followed by a progressive decline toward (or below) baseline. Previous studies have implicated a variety of inhibitory neurotransmitters and neuromodulators in the secondary ventilatory decline in one or more animal models. Here we investigated the contribution of adenosine receptors and serotonin receptors to the biphasic HVR in neonatal (P3‐5) rats. The HVR (10% or 12% O2) was measured by head‐body plethysmography after administration of an adenosine receptor antagonist (40 mg/kg caffeine citrate or 30 mg/kg aminophylline), a serotonin receptor antagonist (0.5 or 1.0 mg/kg SDZ SER 082 fumurate, 2.5 or 4.0 mg/kg methysergide maleate, or 0.5 mg/kg RS 127445 HCl), or the appropriate vehicle (n=5–22 per group). All drugs were administered intraperitoneally 30–45 minutes prior to recording the HVR. None of the drugs altered baseline ventilation, the magnitude of the HVR, or the shape of the biphasic HVR (Drug and Drug × Time, all P>0.05). Although the role of serotonin in the biphasic HVR is not well established, previous studies have supported a role for adenosine receptors in hypoxic ventilatory decline in neonatal rats (e.g., Bairam et al., Respir. Physiol. Neurobiol. 185: 57–66, 2013). The reason for these contrasting results is not known.

Support or Funding Information

Supported by NIH grant P20 GM‐103423 (Maine INBRE).

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
The Interaction of TRPV1 and EP3 Receptors in Cough Response of Guinea Pig
Authors: Gao Xiuping , Zhuang Jianguo , Zhao Lei , Wei Wan , Xu Fadi
Published: June 2021

Bronchopulmonary C‐fibers (PCFs) play an important role in cough generation. An interaction of TRPV1 and EP3 receptor in cough response has been reported, but the relevant mechanisms are not fully investigated. This study aims to identify: 1) the co‐localization of the two receptors in vagal pulmonary C‐neurons of the nodose/jugular ganglia; 2) the dependency of cough responses to citric acid (CA) on EP3 receptors and that to prostaglandin E2 (PGE2) on TRPV1 as well as the corresponding cough tachyphylaxis; and 3) the dependency of CA‐ or PGE2‐triggered currents on TRPV1 or EP3 receptors in the vagal pulmonary C‐neurons. Guinea pigs (GPs) were used. Immunohistochemical approach was applied to label TRPV1 and/or EP3 receptors in vagal pulmonary neurons retrogradely traced by DiI (Series I). Conscious GPs were exposed to aerosolized CA (150 mM) or PGE2 (0.43 mM) w/wo pretreatment of aerosolized antagonist (JNJ 17203212 or capsazepine for TRPV1 and L‐798106 for EP3 receptor) (Series II). In addition, some GPs were exposed to the same CA or PGE2 twice with a 3‐hour interval (CA‐CA and PGE2‐PGE2) and others to CA followed by PGE2 or vice versa (CA‐PGE2 and PGE2‐CA) (Series III). Whole‐cell patch clamp was employed to test the effects of above antagonists on the currents triggered by CAP or PGE2 (Series IV). The results were summarized as follows. 1) Almost all EP3‐labeled vagal pulmonary sensory neurons co‐expressed TRPV1 receptors with smaller cell sizes (< 20 μm) and the population of these neurons occupied 1/3 of the neurons single‐labeled by TRPV1. 2) TRPV1 antagonist significantly reduced the cough number evoked by PGE2, but EP3 antagonist did not affect that by CA. 3) Compared to the CA‐induced cough, the PGE2‐induced cough has extremely higher tachyphylaxis. In sharp contrast, no tachyphylaxis was observed in CA‐PGE2 and PGE2‐CA groups. 4) Both CA‐currents and PGE2‐currents were recorded in the same cells of some vagal pulmonary C‐neurons with the former significantly reduced by TRPV1 antagonist and the latter decreased by EP3 antagonist; PGE2 significantly strengthened the CA‐currents and this effect was abolished by EP3 antagonist; and most interestingly, the TRPV1 antagonist significantly diminished the PGE2‐currents. In conclusion, PGE2 inhalation evokes cough likely via acting on a subgroup of vagal pulmonary sensory TRPV1 neurons co‐expressing EP3 receptors. TRPV1 receptors of these neurons are able to facilitate PGE2‐evoked cough via interacting with EP3 receptors. Endogenous PGE2 seems to have a limited impact on CA‐evoked cough, while exogenous PGE2 is capable of amplifying TRPV1‐mediated respiratory and vagal pulmonary C‐neural responses. Moreover, their interaction fails to affect the cough tachyphylaxis.

Support or Funding Information

NIH HL119683

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Liquiritin Apioside (LA) Attenuates Apneic Responses to Stimulation of Laryngeal Chemoreceptors but not Mechanoreceptors in Rat Pups
Authors: Wei Wan , Gao Xiuping , Zhao Lei , Zhuang Jianguo , Jiao Yang , Xu Fadi
Published: June 2021

Laryngeal chemoreceptors (TRPV1; acid‐sensitive ion channels, ASIC; and osmotic receptors) and mechanoreceptors play an important role in triggering apnea, especially in infants, which has been related to the pathogenesis of sudden infant death syndrome (SIDS). LA, a main flavonoid component of licorice widely used as an antitussive herbal medicine in China even via taking lozenge, is reportedly able to suppress cough responses to inhalation of aerosolized capsaicin (a selective stimulant of TRPV1) in conscious guinea pigs. These results imply an inhibitory impact of LA on airway TRPV1; however, it remains unknown whether LA affects the apneic responses to selective stimulation of these laryngeal receptors, and if so whether its effect is achieved by directly acting on the superior laryngeal nerve (SLN). In anesthetized and spontaneously breathing rat pups, the following stimulations were applied after intralaryngeally loading 50 μl LA (10 μg/μl) and vehicle for 10 min. Intralaryngeal perfusion of capsaicin (CAP), hydrochloric acid solution (HCl), and distilled water (DW) for 10 s was employed to selectively stimulate local TRPV1, ASIC, and osmotic receptors respectively. Intralaryngeal delivery of air‐puff (AP for 2 s) with peak pressure at ~40 mmHg was utilized to stimulate laryngeal mechanoreceptors. The results showed that CAP, HCl, DW, and AP triggered an immediate apnea with TE prolonged by 24‐, 17‐, 18‐ and 3‐fold respectively. LA per se did not alter baseline ventilation but shortened the CAP‐, HCl‐, and DW‐evoked apneic durations by 75%, 68%, and 54% respectively (P < 0.01) with little effect on the apneic response to AP. To determine the direct effect of LA on the SLN, the latter was bilaterally isolated and pretreated by perilaryngeal LA treatment (10 μg/μl) or vehicle for 20 min. Perilaryngeal LA treatment produced a similar inhibition in CAP‐ and HCl‐induced apneic responses but failed to change DW‐induced apnea. Our results, for the first time, suggest that LA applied on the laryngeal mucosa depresses the stimulatory effects of CAP, HCl, and DW on laryngeal chemoreceptors in which CAP and HCl are capable of directly act on TRPV1 and ASIC of SLN. This finding may extend the therapeutic use of LA to treat a subgroup of SIDS victims with laryngeal disorders and explain, at least partially, the antitussive effect of LA due the critical role of laryngeal sensory afferents in cough initiation. The cellular mechanisms underlying this inhibition are warranted for further investigation.

Support or Funding Information

NIH HL119683

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Cigarette Smoke Disrupts Calcium Homeostasis in Airway Smooth Muscle Cells
Authors: Lin Jinheng , Taggart Michael , Tarran Robert , Gray Michael
Published: June 2021

Tobacco smoking is the largest risk factor for developing chronic obstructive pulmonary disease (COPD), and is associated with airway hyperresponsiveness. Hyperresponsiveness is linked to abnormal calcium metabolism, which regulates the contractility and hence the tone of airway smooth muscle (ASM). However, whether cigarette smoke (CS) directly alters or regulates calcium signaling in ASM cells (ASMC) is not fully known. In airway epithelial cells, acute CS exposure increases intracellular Ca2+ concentrations ([Ca2+]i). We hypothesise that CS impedes ASM function by disrupting calcium homeostasis in ASMC, and the present study aims to unravel the effects of CS on calcium handling in ASMC. Primary rat and human ASMC were isolated via enzymatic digestion, and maintained in culture up to 8 passages. ASMC were seeded onto glass coverslips, and deprived of serum for 24–48 hours prior to calcium imaging studies using fura‐2‐AM. ASMC were exposed to either whole CS or CS extract (CSE) from 3R4F reference cigarettes (University of Kentucky) with the particulate phase filtered out. We found that both CS and CSE elevated [Ca2+]i in rat and human ASMC, and this increase was not abolished following depletion of sarcoplasmic‐reticulum calcium stores with thapsigargin or cyclopiazonic acid. The increase in [Ca2+]i in response to CSE was concentration‐dependent. Removal of extracellular calcium attenuated the CSE‐induced elevation of [Ca2+]i in hASMC, suggesting that CSE may activate calcium influx pathways in cultured hASMC. We speculate that this elevation of [Ca2+]i would lead to exacerbated ASM contraction, especially under inflamed conditions in COPD, and hence narrowing of the airway lumen. Further investigations into the source of [Ca2+]i increase, identity of potential calcium influx channels activated, and the direct effects of CS/CSE on ASM contraction are on‐going.

Support or Funding Information

This work is supported by Newcastle University's JJ Hunter Bequest scholarship.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Sex differences in lung miRNA‐712 expression and inflammatory cytokines in a mouse model of asthma and air pollution exposure
Authors: Weaver Ashley Morgon , Fuentez Nathalie , Steckbeck Rachel , Riviera Lidys , Nicoleau Marvin , Silveyra Patricia
Published: June 2021

Background

MicroRNAs (miRNAs) are short RNAs (18–22 nucleotides) that play a critical role in post‐transcriptional gene regulation. Specifically miRNA 712‐5p has been shown to play a major role in the regulation of inflammatory cytokine expression, which is one of the hallmarks of asthma. Asthma has been shown to afflict more adult females than males. Asthmatic females are also more susceptible to air pollution exposure, which leads to higher rates of respiratory disease. The hypothesis of our work is that differential expression of specific miRNAs that regulate inflammatory cytokine expression in the male and female lung upon exposure to an air pollutant could contribute to the mechanisms responsible for the higher susceptibility of females to respiratory disease.

Methods

To test this hypothesis, we developed an asthma model in which male and female C57BL/6 mice were sensitized with house dust mite (HDM) or phosphate buffer saline (PBS) for five‐weeks, followed by a three‐hour ozone or filtered air exposure challenge to induce inflammation. Following exposure, lungs were harvested and total RNA was extracted and converted to cDNA. Relative miRNA expression linked to inflammatory cytokine production was measured by real‐time PCR and normalized with U6. Cytokine gene expression was also determined by real time PCR.

Results

Preliminary results show that there are sex‐specific differences in miR‐712 expression as well as inflammatory cytokine gene expression in asthmatic mice. The data shows that both males and females express higher amount of miR‐712 upon sensitization to HDM compared to PBS and that ozone also increases the levels of miR‐712 expressed in both sexes. Upon exposure to ozone, females sensitized with PBS express higher amounts of miR‐712 than males. However, males sensitized with HDM and exposed to ozone express more miR‐712 than females.

Conclusion

MiR‐712 is differentially expressed between asthmatic males and females following exposure to a pollutant. Since, miR‐712 is known to regulate inflammation, the differential expression of this miRNA could account for the increased susceptibility of asthmatic adult females to pollution. The results of this study could help us to further understand the sex‐specific mechanisms of asthma, and develop novel therapies.

Support or Funding Information

NIH (HL133520, HL141618)

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Microglial Depletion Abolishes Compensatory Plasticity Induced by Prolonged Reductions in Respiratory Neural Activity
Authors: Mickelson Carly Rose , Gumnit Maia , Agbeh Abiye , Ewald Andrea , Meza Armand , Ouellette Jonathan , Braegelmann Kendra , Watters Jyoti , Baker Tracy
Published: June 2021

Reduced respiratory neural activity elicits a compensatory enhancement in inspiratory motor output to the diaphragm, a form of plasticity known as inactivity‐induced inspiratory motor facilitation (iMF). Mechanisms that induce iMF differ depending on the pattern of reduced respiratory neural activity; spinal TNF‐α and subsequent activation of TNFR2 on phrenic motor neurons is necessary to induce iMF following prolonged reductions in respiratory neural activity, whereas spinal retinoic acid synthesis and subsequent activation of RARa on phrenic motor neurons is necessary to induce iMF following intermittent reductions in respiratory neural activity. Little is known regarding the cell subtypes required for iMF induced by either stimulus. Since microglia are an important source of TNF‐α in the CNS, we tested the hypothesis that microglia are necessary for iMF following prolonged, but not intermittent, reductions in respiratory neural activity. Male and female rats received vehicle or pexidartinib (PLX3397; 80 mg/kg, 7 days po), a colony stimulating factor 1 receptor (CSFR1) inhibitor that results in microglial apoptosis. PLX3397 significantly reduced Iba1+ and CD11b+ cells throughout the brain and spinal cord (−82±8%, n=4, p<0.05). The capacity for prolonged (30 min) and intermittent (5, ~1 min) reductions in respiratory neural activity to elicit iMF was then assessed in urethane‐anesthetized, mechanically ventilated rats. In vehicle‐treated rats, both intermittent and prolonged reductions in respiratory neural activity triggered significant increases in phrenic inspiratory output (75 ± 16% and 99 ± 32% baseline, n=6 each, p<0.05), indicating iMF. By contrast, in PLX‐treated rats, only intermittent (63 ± 13% baseline, n=6, p<0.05), but not prolonged (21 ± 6% baseline, n=6, p>0.05) reductions in respiratory neural activity increased phrenic inspiratory output, suggesting that microglial depletion selectively impairs iMF activated by prolonged, but not intermittent, reductions in respiratory neural activity. These data suggest that in response to prolonged reductions in inspiratory‐related synaptic inputs to phrenic motor neurons, microglia may release TNF‐α to trigger compensatory enhancement in phrenic inspiratory output. Studies are underway to investigate this idea further.

Support or Funding Information

NIH R01 HL105511 and NS085226

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Female Mate Choice Influences on Monogamous Pair Bond Formation in Prairie Voles
Authors: Francis Amie , Curtis J. Thomas
Published: June 2021

The prairie vole is an established model for social behavior, most notably, the monogamous pair bond formed between a mated pair. The formation of this bond hinges upon estrogen surges in the female shortly after being introduced to a potential mate. Our lab has correlated earlier estrogen surges with earlier achievement of pregnancy, which we have also found to be a determining factor in the formation of a monogamous pair bond for male prairie voles. Currently, our lab is examining how female mate choice influences the formation of a monogamous pair bond via both behavioral and endocrine measures.

This study has three objectives: 1) To establish whether endocrine measures provide supporting data for behavioral measures of pair bond formation; 2) To examine whether female prairie voles allowed to pair with a mate of their choice will show an earlier estrogen surge and a shorter latency pregnancy; and 3) To determine if pairing and pregnancy is a greater stressor for animals not paired with their mate of choice.

Sexually naïve female prairie voles were allowed to choose between two males and subsequently paired either with the male of their choice or the male they did not choose. An additional group of females was paired randomly with a male outside of the choice paradigm. Fecal samples were collected prior to and immediately after mate choice, and at select time points throughout the next 30 days or until the birth of a litter. Fecal samples were analyzed via ELISA for 17b‐estradiol and corticosterone content to examine sex hormone changes associated with pairing and pregnancy, as well as stress levels throughout pairing and pregnancy. These measures will be combined with partner preference testing on either the female or male subject to establish whether female mate choice can affect endocrine changes that may impact monogamous pair bond formation.

Regardless of sex, animals in pairs that include the non‐chosen male tend to display social affiliation patterns that differ from those pairs that include the chosen male or a randomly assigned male. Despite differences in affiliative behavior, latency to birth in all groups was similar. These data suggest that female mate choice may influence pair bond formation, despite the lack of differences in latency to pregnancy.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Effect of Creatine‐Monohydrate on Cognitive Function in Subjects Who Differ in Dietary Meat Consumption
Authors: Smith Kaitlyn , Weickel Caroline , Petersen Jahkeema , Skelton Michele
Published: June 2021

The purpose of this study was to examine the effects of 4 weeks of creatine supplementation on cognitive function using the ImPACT test in vegetarians, subjects that consumed 1–10 servings of beef, chicken, pork, or fish per week, and subjects that consumed greater than 10 servings of beef, chicken, pork or fish per week. In a randomized, double‐blind placebo‐controlled study, twenty‐five subjects were divided into two groups. One group received creatine while the other group received a placebo. Subjects completed a 5‐day loading phase (20 grams per day) followed by a 24‐day maintenance phase (5 grams per day). Cognitive function was assessed prior to supplementation and immediately after a 4‐week supplementation phase. The ImPACT test was used and provides 6 measures of cognitive function. There were no significant changes in any of the cognitive measures between groups. However, vegetarians in the creatine group scored significantly higher than subjects who consumed greater than 10 servings of beef, chicken, pork or fish per week on the visual memory test. Preliminary analysis suggests that acute supplementation of creatine may enhance cognition in vegetarians but have no effect on those who consume meat regularly in their diet.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
The Effect of Perinatal Blockade of Androgen Receptors on Adult Rat Behaviors and the Expression of Estrogen and Androgen Receptors in specific brain regions
Authors: Gopalan Chaya , Colley Zach , Niepoetter Paige , Waller Christopher , Cashikar Anil
Published: June 2021

The brain, by default, differentiates into a female brain unless testosterone (T) is present during a critical window of time. In rats, there is a surge of T around the time of birth, more specifically four days prior to and four days after birth, referred to as the perinatal period. The purpose of this study was to examine the effects of blockade of androgen receptors (AR) during the perinatal period using flutamide (F) on sexual motivation (SM) and an open field (OF) test as well as the expression of AR and estrogen receptors (ER)‐alpha and ER‐beta in the hypothalamus, hippocampus, and amygdala. Four timed‐pregnant rats were divided into two groups: one group (N=2) received F during perinatal period whereas the other group (N=2) served as the control group receiving vehicle (95% Olive oil‐5% ethanol). Once the pups reached adulthood, the male rats underwent SM testing where they were exposed to either an estrus or an ovariectomized rat. Both F and control groups of male rats spent more time with the estrus rat compared to the ovariectomized rat suggesting that SM was not affected by their AR blockade during perinatal period. Open field testing recorded animal movements between the two zones, the center and the periphery, within an open arena. Male rats moved between the two zones more than the female rats did. At the end of the behavioral study, animals were sacrificed and the brains were harvested to isolate the hippocampus, hypothalamus, and amygdala. Real time PCR testing was carried out to quantify the levels of AR, ER alpha, and ER beta using B‐Actin as an internal control. Perinatal blockade of AR had no effect on the expression of any of the receptors in the three regions tested in the adult rats. Thus, the increased movement by the male rats in the OF testing is not associated with changes in the expression of AR, ER‐alpha, or ER‐beta.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Sleep Restriction Worsens Daily Life Executive Functioning in Adolescents with ADHD
Authors: Fershtman Chaya E. M. , Kapadia Delna K. , Becker Stephen P.
Published: June 2021

Attention‐deficit/hyperactivity disorder (ADHD) is a persistent neurodevelopmental disorder characterized by inattention and/or hyperactivity‐impulsivity. Adolescents with ADHD commonly experience deficits in areas of executive function (EF). These deficits are associated with academic, social, and emotional impairments. Experimental research shows shortened sleep duration to cause poorer daily life EF in typically developing adolescents. However, no previous research has looked at the casual relationship between inadequate sleep and executive functioning deficits in teenagers with ADHD. The objective of this study was to investigate whether sleep duration impacted daily life executive functioning. The hypothesis was that restricted sleep would negatively impact daily life executive functioning in adolescents with ADHD. A three‐week at home sleep manipulation protocol was implemented, using an experimental crossover design. Participants completed a phase stabilization week, followed by a sleep restriction (SR) week (6.5 hours) and sleep extension (SE) week (9.5). SR and SE conditions were counterbalanced across participants. Each week, participants attended laboratory visits at the medical center where executive functioning was measured with a validated rating scale, the Behavior Rating Inventory of Executive Function, Second Edition (BRIEF2). The BRIEF2 measured different areas of EF domains such as working memory, inhibition, planning/organization, emotional control, and initiation. Sleep duration was monitored with actigraphy and sleep diaries. Analyses looked at 48 participants who successfully adhered to the study protocol and who had complete actigraphy data. The results of a paired sample t‐test showed significantly (ps <.05) more deficits in most daily life EF domains during the sleep restriction phase compared to the sleep extension phase. This study provides the first evidence for shortened sleep duration as a causal contributor to worsened executive functioning in adolescents with ADHD. Increased sleep may significantly impact academic, social, and emotional functioning in adolescents with ADHD, and sleep may be an important future target for future intervention.

Support or Funding Information

This research was supported by the National Institute of Mental Health (NIMH; grant R03MH109787; Dr. Becker). The content is solely the responsibility of the authors and does not necessarily represent the official views of the NIMH.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Optogenetic Stimulation of Cholinergic Neurons in the Brainstem Induces Splenic Nerve Activity and Attenuates Systemic Inflammation
Authors: Kressel Adam M , Tsaava Téa , Chang Eric H , Addorisio Meghan E , Chang Qing , Pavlov Valentin A , Chavan Sangeeta S , Tracey Kevin J
Published: June 2021

The inflammatory reflex is a well‐defined neural circuit composed of afferent and efferent fibers that travel via the vagus nerve to regulate peripheral tumor necrosis factor (TNF) production. Electrical stimulation of the efferent fibers reduces splenic TNF output in an animal model of endotoxemia. However, the exact origin of these vagus nerve fibers in the brainstem and the means by which they innervate the spleen to modulate TNF levels is not yet understood. Using optogenetics, we selectively stimulated cholinergic neurons in the dorsal motor nucleus of the vagus nerve (DMV), the autonomic brainstem nucleus from which the efferent vagus nerve fibers originate. A fiber‐optic cannula was inserted using stereotactic guidance into the DMV of transgenic mice expressing channelrhodopsin under the choline acetyltransferase promoter (ChAT‐ChR2‐EYFP). Mice were subjected to either optogenetic or sham stimulation (n=20 per group) for five minutes (473nm laser, 20Hz, 25% duty cycle). After 24 hours, animals were subjected to endotoxemia by intraperitoneal administration of lipopolysaccharide (0.25mg/kg), blood was collected after 90 minutes, and serum TNF was quantitated. Optogenetic stimulation of the cholinergic neurons in the DMV of ChAT‐ChR2‐EYFP mice significantly decreased endotoxin‐induced serum TNF levels compared to sham controls (p=0.0004). Splenic nerve activity, recorded during DMV stimulation using a cuffed two‐channel electrode, was significantly increased over baseline (p=0.0002). Blocking the signals transmitted in the vagus nerve by adding bupivacaine, a sodium channel antagonist, eliminated this effect (p=0.0028), implicating a functional synapse between the vagus and splenic nerves. These studies provide the first direct evidence for the central origin of the efferent vagus nerve fibers regulating TNF production during endotoxemia and suggest novel anti‐inflammatory approaches based on targeting DMV cholinergic signaling.

Support or Funding Information

This study was funded by NIH/NIGMS/NIAID.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Pharmacological baroreflex testing with fMRI reveals baroreflex mediated brainstem nuclei
Authors: Gerlach Darius Andreas , Manuel Jorge , Hoff Alex , Kronsbein Hendrik , Hoffmann Fabian , Heusser Karsten , Ehmke Heimo , Jordan Jens , Beissner Florian , Tank Jens
Published: June 2021

Background

Brainstem nuclei mediate baroreflex adjustments in efferent sympathetic and parasympathetic traffic. Yet, human brainstem physiology is poorly understood given the lack of suitable methodology. We developed a novel approach combining pharmacological testing, beat‐by‐beat cardiovascular monitoring, and high‐resolution functional magnetic resonance imaging (fMRI) to assess human baroreflex regulation at the level of the brainstem.

Method and materials

In 11 healthy men (30.7 ±6.6 years; 24.1 ±1.9 kg/m2), we monitored continuous finger arterial blood pressure and ECG using customized hardware during multiband fMRI brain acquisitions. We applied repeated intravenous phenylephrine (PHE, 25 and 75 μg, n=8) and nitroprusside (NTP 25 and 75 μg, n=8) boluses using a remote controlled injector. Brainstem and hypothalamus fMRI images were analyzed to identify brainstem nuclei involved in baroreflex‐mediated blood pressure control using with masked general linear model (GLM). BP changes were correlated with the time‐courses of blood‐oxygen‐level dependent (BOLD) signals by mixed‐effects general linear model.

Results

Pharmacological baroreflex testing yielded baroreflex sensitivity measurements with typical blood pressure and RR‐interval patterns. Correlation of the baroreflex‐mediated changes in systolic blood pressure with BOLD signals clearly revealed brainstem nuclei. The solitary tract (NTP: 7.3; PHE: 6.0 t‐value), the caudal ventrolateral medulla (PHE: 5.8 t‐value); the rostral ventrolateral medulla (NTP: 5.6; PHE: 7.1 t‐value), the nucleus ambiguus (PHE: 5.8 t‐value), the paraventricular nucleus (NTP:7.1; PHE: 4.5 t‐value), and raphe nuclei (NTP: 5.7; PHE: 6.0 t‐value), were identified with high sensitivity and corrected for multiple comparisons (p < 0.01).

Conclusion

We developed novel approach testing baroreflex regulation at the level of the brainstem in humans. The methodology identified baroreflex‐mediated activation and deactivation patterns consistent with previous investigations in animal models. The methodology can be applied to elucidate human physiology and mechanisms of autonomic cardiovascular disease.

Support or Funding Information

This study was funded by the DLR internal cost object 2475023.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Central Terminals of Trigeminal Nerves that Innervate the Nose and Nasal Passages Colocalize with Medullary Dorsal Horn Neurons Activated during Voluntary Diving in Rats
Authors: McCulloch Paul , Margosiak Malgorzata , DiNovo Karyn
Published: June 2021

Underwater submersion in mammals initiates the diving response, an autonomic cardiorespiratory reflex that includes apnea, a parasympathetically‐mediated bradycardia, and a sympathetically‐mediated increase in total peripheral resistance. The initial synapse of this reflex occurs within the ventral medullary dorsal horn (MDH) of the spinal trigeminal nucleus caudalis. The anterior ethmoidal and nasopalatine nerves, trigeminal nerves that innervate the internal nasal passages, also send their central terminations to the ventral MDH. The objective of this research was to detect colocalization of secondary MDH neurons activated by voluntary diving with the central terminations of the trigeminal nerves innervating the internal nasal passages. MDH activated neurons were identified by Fos protein production, while central terminals of the trigeminal nerves were located by the transganglionic tracer Wheat Germ Agglutinin (WGA). Sprague‐Dawley rats (N=12) were trained to voluntarily dive through a 5 m underwater Plexiglas maze. After training, 6 rats then received WGA injections into the anterior ethmoidal nerve, while 6 rats received WGA injections into the nasopalatine nerve. Four days post‐WGA injection, 3 rats from each nerve injection group performed 12 dive trials over 1 hour to activate brainstem neurons and initiate Fos production; 3 rats from each nerve injection group were non‐diving controls. Brainstem tissue from all rats was immunohistochemically processed to visualize both Fos and WGA, and was inspected using a Nikon C1 confocal microscope and Nikon Eclipse Ti2 inverted fluorescent microscope. Results indicate repetitive voluntary diving increases the number of Fos‐positive neurons within the ventral tip of the superficial MDH and adjacent paratrigeminal nucleus. WGA results show central termination sites from both the anterior ethmoidal and nasopalatine nerves within this same MDH location. Additionally, MDH Fos‐positive neurons from diving animals show extensive colocalization with WGA injected into both the anterior ethmoidal nerve and nasopalatine nerve compared to non‐diving control rats. We conclude both the anterior ethmoidal and nasopalatine nerves can provide afferent information necessary to activate MDH neurons during initiation of the mammalian diving response.

Support or Funding Information

Supported by MWU Intramural funding (PFM) and CGS MBS program (MM)

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Enhancement of Hypothalamic Corticotrophin Releasing Hormone Signal Contributes to Elevated Sympathetic Outflow in Spontaneous Hypertension Rat
Authors: Zhou Jing‐Jing , Shao Jian‐Ying , Ma HuiJie , Pan Hui‐Lin , Li De‐Pei
Published: June 2021

The paraventricular nucleus (PVN) of the hypothalamus is crucially involved in the regulation of blood pressure and sympathetic outflow in hypertension. The PVN contains neurons synthesizing corticotrophin releasing hormone (CRH), a neuropeptide that is critical in regulating neuroendocrine and cardiovascular functions in physiological and disease conditions. The mRNA and protein expression levels are significantly increased in the hypothalamus in patients with primary hypertension. However, the role of hypothalamic CRH signal in the elevated sympathetic vasomotor tone in hypertension remains unknown. In this study, we tested a hypothesis that enhanced hypothalamic CRH signals contributes to elevated sympathetic vasomotor tone in primary hypertension. Western blotting analysis by using PVN tissues revealed that CRH receptor (CRHR) 1 expression level was significantly higher in male adult spontaneously hypertensive rats (SHRs) than in Wistar‐Kyoto (WKY) rats. However, CRHR1 expression level in the PVN was not significantly affected by lowing ABP by celiac ganglionectomy in SHRs. In brain slice preparation, bath application of CRH induced a significantly greater membrane depolarization and an increase in the firing rate of spinally projecting PVN neurons in SHRs than in WKY rats. A CRHR1 antagonist NBI 35965, not CRHR2 antagonist antisauvagine‐30, decreased the basal firing rate of spinally projecting PVN neurons in SHRs but not in WKY rats. However, CRH did not change neither the miniature inhibitory post‐synaptic currents nor the miniature excitatory post‐synaptic currents of labeled PVN neurons in both SHRs and WKY rats. Furthermore, bilateral microinjection of CRH into the PVN induced dose‐dependently greater increases in arterial blood pressure (ABP), heart rate (HR), and renal sympathetic activity (RSNA) in anesthetized SHRs than in WKY rats. CRH‐induced sympathoexcitatory effect was eliminated by microinjection of NBI 35965 but not antisauvagine‐30. Microinjection of NBI 35965 into the PVN significantly reduced basal ABP, HR, and RSNA in SHRs. These data suggest that upregulation of CRFR1 receptor in the PVN is involved in hyperactivity of spinally projecting PVN neurons and contributes to the elevated sympathetic outflow in essential hypertension.

Support or Funding Information

Supported by grants from National Institutes of Health (HL139523 and HL142133).

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Group III Metabotropic Glutamate Receptors Regulate Excitability of Hypothalamic Presympathetic Neurons and Sympathetic Output in Hypertension
Authors: Zhou Jing‐Jing , Pachuau Judith , Li De‐Pei , Pan Hui‐Lin
Published: June 2021

Enhanced glutamatergic input to the hypothalamic paraventricular nucleus (PVN) contributes to the development of hypertension. Group III metabotropic glutamate receptors (mGluRs) are expressed in the PVN. In the present study, we determined the role of group III mGluRs in the control of the excitability of spinally projecting PVN neurons and sympathetic outflow in hypertension. We found that stimulation of group III mGluRs with L‐(+)‐2‐amino‐4‐phosphonobutyric acid (L‐AP4) consistently inhibited the firing activity of PVN neurons in normotensive WKY rats. However, L‐AP4 inhibited 37%, but excited 48%, of spinally projecting PVN neurons in spontaneously hypertensive rats (SHRs). Both the inhibitory and excitatory effects of L‐AP4 on the firing activity were abolished by the group III mGluR antagonist CPPG. Inclusion of GDP‐β‐S in the intracellular recording solution eliminated the excitatory effect of L‐AP4 on the firing activity of PVN neurons in SHRs, but it did not significantly change the inhibitory effect of L‐AP4 on PVN neurons in WKY rats or SHRs. L‐AP4 significantly reduced the frequency of spontaneous excitatory postsynaptic currents of PVN neurons in both WKY rats and SHRs. Interestingly, pretreatment of brain slices with the group I mGluR agonist DHPG converted the L‐AP4 effect on PVN neurons from inhibitory to excitatory in WKY rats. Also, L‐AP4 consistently inhibited PVN neurons in SHRs after treatment with the group I mGluR antagonist MPEP. In addition, microinjection of L‐AP4 into the PVN dose‐dependently decreased arterial Bp and lumbar sympathetic nerve activity (LSNA) in WKY rats and SHRs. Microinjection of L‐AP4 in the presence of MPEP produced a more significant reduction in arterial BP and LSNA than did with microinjection of L‐AP4 alone in SHRs. These findings suggest that group III mGluRs in the PVN regulate the synaptic plasticity and excitability of PVN presympathetic neurons and sympathetic vasomotor activity in hypertension. The signaling of postsynaptic group III mGluRs is influenced by increased group I mGluR activity in PVN presympathetic neurons in hypertension. Supported by NIH grants HL131161 and HL142133.

Support or Funding Information

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Phenotyping a Mouse with Vascular Smooth Muscle Specific Deletion of the Gamma Subunit of the Epithelial Sodium Channel (γENaC)
Authors: Hasan Mahpara , Mutchler Stephanie , Tan Roderick J , Kohan Donald E , Kleyman Thomas R
Published: June 2021

The epithelial sodium channel (ENaC) has an important role in mediating myogenic tone of blood vessels. Previous studies found that mice with whole body knockdown of bENaC had an increased blood pressure, thought to be due to the loss of myogenic tone in the renal vasculature leading to inflammation and glomerular damage. However, because these results were obtained in an animal with global suppression of bENaC expression, the contributions of vascular smooth muscle (VSM) ENaC cannot be separated from effects of suppressed ENaC expression at other sites. To address this concern, we generated a VSM ENaC knockout animal using cre‐lox technology. Contrary to the bENaC hypomorphs, these mice did not have increased baseline blood pressure, as measured by radiotelemetry. Our preliminary results do not show significant differences in the myogenic response of the mesenteric or thoracodorsal arteries as assessed by pressure myography, however, the vessels do have greater angiotensin II‐mediated vasoconstriction. Preliminary results reveal that VSM ENaC KO mice exhibited increased glomerular filtration rate (GFR) at baseline, and that the knockouts had more proteinuria following a 4‐week treatment with high salt diet and aldosterone. However, wild type and VSM ENaC KO mice had similar responses to renal ischemia reperfusion injury, as assessed by serum creatinine and message levels for kidney injury markers NGAL and Kim1. Taken together, these data reveal that VSM gENaC may be playing a distinct role from that of bENaC, potentially only contributing to the myogenic response in the microvasculature of the kidney under conditions of stress.

Support or Funding Information

NIH P30 2P30DK079307‐11, NIH T32 2T32DK061296‐16

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.