Circadian disruption of hippocampus in an early senescence male mouse model.

Age-related cognitive decline and disruptions in circadian rhythms are growing problems as the average human life span increases. Multiple strains of the senescence-accelerated mouse (SAM) show reduced life span, and the SAMP8 strain in particular has been well documented to show cognitive deficits in behavior as well as a bimodal pattern of circadian locomotor activity. However, little is known about circadian regulation within the hippocampus of these strains of mice. Here we test the hypothesis that in this early senescence model, disruption of the molecular circadian clock in SAMP8 animals drives disrupted behavior and physiology. We found normal rhythms in PER2 protein expression in the SCN of SAMP8 animals at 4 months, despite the presence of disrupted wheel-running activity rhythms at this age. Interestingly, a significant rhythm in PER2 expression was not observed in the hippocampus of SAMP8 animals, despite a significant 24-h rhythm in SAMR1 controls. We also examined time-restricted feeding as a potential strategy to rescue disrupted hippocampal plasticity. Time-restricted feeding increased long-term potentiation at Schaffer collateral-CA1 synapses in SAMP8 mice (compared to SAMR1 controls). Overall, we confirm disrupted circadian locomotor rhythms in this early senescence model (as early as 4 months) and discovered that this disruption is not due to arrhythmic PER2 levels in the SCN; however, other extra-SCN circadian oscillators (i.e., hippocampus) are likely impaired with accelerated aging.

Dysregulated clock gene expression and abnormal diurnal regulation of hippocampal inhibitory transmission and spatial memory in amyloid precursor protein transgenic mice.

Patients with Alzheimer's disease (AD) often have fragmentation of sleep/wake cycles and disrupted 24-h (circadian) activity. Despite this, little work has investigated the potential underlying day/night disruptions in cognition and neuronal physiology in the hippocampus. The molecular clock, an intrinsic transcription-translation feedback loop that regulates circadian behavior, may also regulate hippocampal neurophysiological activity. We hypothesized that disrupted diurnal variation in clock gene expression in the hippocampus corresponds with loss of normal day/night differences in membrane excitability, synaptic physiology, and cognition. We previously reported disrupted circadian locomotor rhythms and neurophysiological output of the suprachiasmatic nucleus (the primary circadian clock) in Tg-SwDI mice with human amyloid-beta precursor protein mutations. Here, we report that Tg-SwDI mice failed to show day/night differences in a spatial working memory task, unlike wild-type controls that exhibited enhanced spatial working memory at night. Moreover, Tg-SwDI mice had lower levels of Per2, one of the core components of the molecular clock, at both mRNA and protein levels when compared to age-matched controls. Interestingly, we discovered neurophysiological impairments in area CA1 of the Tg-SwDI hippocampus. In controls, spontaneous inhibitory post-synaptic currents (sIPSCs) in pyramidal cells showed greater amplitude and lower inter-event interval during the day than the night. However, the normal day/night differences in sIPSCs were absent (amplitude) or reversed (inter-event interval) in pyramidal cells from Tg-SwDI mice. In control mice, current injection into CA1 pyramidal cells produced more firing during the night than during the day, but no day/night difference in excitability was observed in Tg-SwDI mice. The normal day/night difference in excitability in controls was blocked by GABA receptor inhibition. Together, these results demonstrate that the normal diurnal regulation of inhibitory transmission in the hippocampus is diminished in a mouse model of AD, leading to decreased daytime inhibition onto hippocampal CA1 pyramidal cells. Uncovering disrupted day/night differences in circadian gene regulation, hippocampal physiology, and memory in AD mouse models may provide insight into possible chronotherapeutic strategies to ameliorate Alzheimer's disease symptoms or delay pathological onset.

Time-restricted feeding rescues high-fat-diet-induced hippocampal impairment.

Feeding rodents a high-fat diet (HFD) disrupts normal behavioral rhythms, particularly meal timing. Within the brain, mistimed feeding shifts molecular rhythms in the hippocampus and impairs memory. We hypothesize that altered meal timing induced by an HFD leads to cognitive impairment and that restricting HFD access to the "active period" (i.e., night) rescues the normal hippocampal function. In male mice, ad-lib access to an HFD for 20 weeks increased body weight and fat mass, increased daytime meal consumption, reduced hippocampal long-term potentiation (LTP), and eliminated day/night differences in spatial working memory. Importantly, two weeks of time-restricted feeding (TRF) at the end of the chronic HFD protocol rescued spatial working memory and restored LTP magnitude, even though there was no change in body composition and total daily caloric intake. These findings suggest that short-term TRF is an effective mechanism for rescuing HFD-induced impaired cognition and hippocampal function.

High-Fat and High-Sucrose Diets Impair Time-of-Day Differences in Spatial Working Memory of Male Mice.

OBJECTIVE: This study aimed to investigate both the long-term and short-term impacts of high-fat diets (HFD) or high-sucrose diets (HSD) on the normal diurnal pattern of cognitive function, protein expression, and the molecular clock in mice. METHODS: This study used both 6-month and 4-week feeding strategies by providing male C57BL/6J mice access to either a standard chow, HFD, or HSD. Spatial working memory and synaptic plasticity were assessed both day and night, and hippocampal tissue was measured for changes in NMDA and AMPA receptor subunits (GluN2B, GluA1), as well as molecular clock gene expression. RESULTS: HFD and HSD both disrupted normal day/night fluctuations in spatial working memory and synaptic plasticity. Mice fed HFD altered their food intake to consume more calories during the day. Both diets disrupted normal hippocampal clock gene expression, and HFD reduced GluN2B levels in hippocampal tissue. CONCLUSIONS: Taken together, these results suggest that both HFD and HSD induce a loss of day/night performance in spatial working memory and synaptic plasticity as well as trigger a cascade of changes that include disruption to the hippocampal molecular clock.

Circadian regulation of membrane physiology in neural oscillators throughout the brain.

Twenty-four-hour rhythmicity in physiology and behavior are driven by changes in neurophysiological activity that vary across the light-dark and rest-activity cycle. Although this neural code is most prominent in neurons of the primary circadian pacemaker in the suprachiasmatic nucleus (SCN) of the hypothalamus, there are many other regions in the brain where region-specific function and behavioral rhythmicity may be encoded by changes in electrical properties of those neurons. In this review, we explore the existing evidence for molecular clocks and/or neurophysiological rhythms (i.e., 24 hr) in brain regions outside the SCN. In addition, we highlight the brain regions that are ripe for future investigation into the critical role of circadian rhythmicity for local oscillators. For example, the cerebellum expresses rhythmicity in over 2,000 gene transcripts, and yet we know very little about how circadian regulation drives 24-hr changes in the neural coding responsible for motor coordination. Finally, we conclude with a discussion of how our understanding of circadian regulation of electrical properties may yield insight into disease mechanisms which may lead to novel chronotherapeutic strategies in the future.

ETS transcription factor Etsrp / Etv2 is required for lymphangiogenesis and directly regulates vegfr3 / flt4 expression.

The molecular mechanisms initiating the formation of the lymphatic system, lymphangiogenesis, are still poorly understood. Here we have identified a novel role in lymphangiogenesis for an ETS transcription factor, Etv2/Etsrp, a known regulator of embryonic vascular development. Through the use of fully validated photoactivatable morpholinos we show that inducible Etv2 inhibition in zebrafish embryos at 1 day post-fertilization (dpf) results in significant inhibition of lymphangiogenesis, while development of blood vessels is unaffected. In Etv2-inhibited embryos and larvae, the number of lymphatic progenitors is greatly reduced, the major lymphatic vessel, the thoracic duct, is absent or severely fragmented, and lymphangiogenesis-associated marker expression, including lyve1b, prox1a, and vegfr3/flt4, is strongly downregulated. We also demonstrate that lymphatic progenitors in Etv2 deficient embryos fail to respond to Vegfc signaling. Chromatin immunoprecipitation and sequencing (ChIP-Seq) studies using differentiated mouse embryonic stem (ES) cells as well as luciferase reporter studies in the ES cells and in zebrafish embryos argue that Etv2 directly binds the promoter/enhancer regions of Vegfc receptor Vegfr3/Flt4 and lymphatic marker Lyve1, and promotes their expression. Together these data support a model where Etv2 initiates lymphangiogenesis by directly promoting the expression of flt4 within the posterior cardinal vein.

The implications of baseline bone-health assessment at initiation of androgen-deprivation therapy for prostate cancer.

OBJECTIVES: To assess bone-density testing (BDT) use amongst prostate cancer survivors receiving androgen-deprivation therapy (ADT), and downstream implications for osteoporosis and fracture diagnoses, as well as pharmacological osteoporosis treatment in a national integrated delivery system. PATIENTS AND METHODS: We identified 17 017 men with prostate cancer who received any ADT between 2005 and 2014 using the Veterans Health Administration cancer registry and administrative data. We identified claims for BDT within a 3-year period of ADT initiation. We then used multivariable regression to examine the association between BDT use and incident osteoporosis, fracture, and use of pharmacological treatment. RESULTS: We found that a minority of patients received BDT (n = 2 502, 15%); however, the rate of testing increased to >20% by the end of the study period. Men receiving BDT were older at diagnosis and had higher-risk prostate cancer (both P < 0.001). Osteoporosis and fracture diagnoses, use of vitamin D ± calcium, and bisphosphonates were all more common in men who received BDT. After adjustment, BDT, and to a lesser degree ≥2 years of ADT, were both independently associated with incident osteoporosis, fracture, and osteoporosis treatment. CONCLUSIONS: BDT is rare amongst patients with prostate cancer treated with ADT in this integrated delivery system. However, BDT was associated with substantially increased treatment of osteoporosis indicating an underappreciated burden of osteoporosis amongst prostate cancer survivors initiating ADT. Optimising BDT use and osteoporosis management in this at-risk population appears warranted.

Glycogen synthase kinase 3 regulates photic signaling in the suprachiasmatic nucleus.

Glycogen synthase kinase 3 (GSK3) is a serine-threonine kinase that regulates mammalian circadian rhythms at the behavioral, molecular and neurophysiological levels. In the central circadian pacemaker, the suprachiasmatic nucleus (SCN), inhibitory phosphorylation of GSK3 exhibits a rhythm across the 24 h day. We have recently shown that GSK3 is capable of influencing both the molecular clock and SCN neuronal activity rhythms. However, it is not known whether GSK3 regulates the response to environmental cues such as light. The goal of this study was to test the hypothesis that GSK3 activation mediates light-induced SCN excitability and photic entrainment. Immunofluorescence staining in the SCN of mice showed that late-night light exposure significantly increased GSK3 activity (decreased pGSK3ß levels) 30-60 min after the light-pulse. In addition, pharmacological inhibition of GSK3 blocked the expected light-induced excitability in SCN neurons; however, this effect was not associated with changes in resting membrane potential or input resistance. Behaviorally, mice with constitutively active GSK3 (GSK3-KI) re-entrained to a 6-h phase advance in the light-dark cycle in significantly fewer days than WT control animals. Furthermore, the behavioral and SCN neuronal activity of GSK3-KI mice was phase-advanced compared to WT, in both normal and light-exposed conditions. Finally, GSK3-KI mice exhibited normal negative-masking behavior and electroretinographic responses to light, suggesting that the enhanced photic entrainment is not due to an overall increased sensitivity to light in these animals. Taken together, these results provide strong evidence that GSK3 activation contributes to light-induced phase-resetting at both the neurophysiological and behavioral levels.

A tool set to allow rapid screening of dog families with PRA for association with candidate genes.

OBJECTIVE: To develop a method to rapidly screen candidate genes for association with recessively inherited progressive retinal atrophy (PRA) in pedigrees of dog in which a causative mutation has not been identified. ANIMAL STUDIED: Thirteen PRA-affected dogs were used in this study. PROCEDURES: Two microsatellite markers (MS) were designed flanking 45 candidate genes. MS markers were analyzed for heterozygosity and allelic richness. Two dog breeds, in which the causative mutation has been identified (Entlebucher Sennenhunds [ES] and PDE6A-mutant dogs [PDE6A]), were used to validate the MS marker panel. One breed in which the causative mutation is currently unknown (Old English Sheepdog [OES]) was investigated in this study utilizing the MS panel. RESULTS: Marker heterozygosity excluded 38 of 45 and 41 of 45 candidate genes (ES and PDE6A, respectively) with each true culprit gene remaining on the list of nonexcluded candidate genes. Additionally, 41 of 45 genes were excluded for OES. CONCLUSIONS: This tool set was used quickly and efficiently to narrow down 45 candidate genes for recessively inherited PRA in two types of dogs with known mutations and one type of dog with an unknown mutation.

Longitudinal Study of Transfusion Utilization in Hospitalized Veterans.

OBJECTIVE: Although transfusion guidelines have changed considerably over the past two decades, the adoption of patient blood management programs has not been fully realized across hospitals in the United States. PATIENTS AND METHODS: We evaluated trends in red blood cell (RBC), platelet and plasma transfusion at three Veterans Health Administration (VHA) hospitals from 2000 through 2010. RESULTS: There were 176,521 hospitalizations in 69,621 patients; of these, 13.6% of hospitalizations involved transfusion of blood products (12.7% RBCs, 1.4% platelets, 3.0% plasma). Transfusion occurred in 25.2% of surgical and 5.3% of medical hospitalizations. Transfusion use peaked in 2002 for surgical hospitalizations and declined afterwards (p<0.001). There was no significant change in transfusion use over time (p=0.126) for medical hospitalizations. In hospitalizations that involved transfusions, there was a 20.3% reduction in the proportion of hospitalizations in which ≥3 units of RBCs were given (from 51.7% to 41.1%; p<0.001) and a 73.6% increase when one RBC unit was given (from 8.0% to 13.8%; p<0.001) from 2000-2010. Of the hospitalizations with RBC transfusion, 9.6% involved the use of one unit over the entire study period. The most common principal diagnoses for medical patients receiving transfusion were anemia, malignancy, heart failure, pneumonia and renal failure. Over time, transfusion utilization increased in patients who were admitted for infection (p=0.009). CONCLUSION: Blood transfusions in three VHA hospitals have decreased over time for surgical patients but remained the same for medical patients. Further study examining appropriateness of blood products in medical patients appears necessary.

Mechanical Circulatory Support for the Failing Heart: Continuous-Flow Left Ventricular Assist Devices.

BACKGROUND: Heart transplantation remains the definitive therapy for patients with advanced heart failure; however, owing to limited donor organ availability and long wait times, continuous-flow left ventricular assist devices (LVADs) have become standard therapy. METHODS: This review summarizes the history, progression, function, and basic management of LVADs. Additionally, we provide some clinical pearls and important caveats for managing this unique patient population. RESULTS: Currently, the most common LVADs being implanted in the United States are second- and third-generation devices, the HeartMate II (Thoratec Corp., St. Jude Medical) and the HeartWare HVAD (HeartWare International, Inc.). A newer third-generation pump, the HeartMate III (Thoratec Corp., St. Jude Medical), is designed to create an artificial pulse and is currently under investigation in the United States. CONCLUSION: LVAD use is promising, will continue to grow, and has become standard therapy for advanced heart failure as a bridge to recovery, as destination therapy, and as a bridge to transplantation.

First records of Armigeres malayi and Armigeres milnensis in Timor-Leste.

Larval Armigeres malayi and larval Ar. milnensis were first collected from rainwater-filled broken coconut shells in the district of Manufahi, subdistrict Same, in southwest Timor-Leste in September 2010. In subsequent surveys, Ar. malayi and Ar. milnensis were frequently observed in water-filled coconut shells either as the sole culicid species, or coexisting with each other, or with larval Aedes albopictus or Culex spp. Although there have been a number of published surveys of Culicidae in Timor-Leste, these Armigeres species have not previously been recorded in this country. Little is known about the status of these species as potential vectors of human or animal disease; however, it has been suggested that Ar. milnensis is a potential vector of Dirofilaria immitis and other filariae, so they may merit further study from a human and veterinary health perspective, as well as for their role in local ecosystems, particularly their competitive impact on other mosquito species that oviposit in the same container habitats.

Influence of tumour necrosis factor-α polymorphism-308 and atopy on irritant contact dermatitis in healthcare workers.

BACKGROUND: Chronic irritant hand dermatitis is an issue for healthcare workers and may negatively impact infection control. OBJECTIVES: We examined the effects of a G to A transition at position -308 on the tumour necrosis factor-α (TNF-α) gene on chronically damaged skin of healthcare workers during exposure and recovery from repetitive hand hygiene, after intensive treatment, and on the irritant response in normal skin. PATIENTS/MATERIALS/METHODS: In 68 healthcare workers with irritant hand dermatitis, we genotyped TNF-α-308 and measured the epidermal response via quantitative digital imaging, erythema, dryness, and barrier integrity. RESULTS: Excess hand erythema decreased with hand hygiene exposure and increased during time off for AA/GA genotypes, but had opposite effects for GG. AA/GA had smaller reductions in dryness with lotion treatment and larger reductions in excess erythema than GG. The atopic diathesis and heightened neurosensory irritation resulting from water and lactic acid significantly influenced the responses. Repeated exposure to water and sodium lauryl sulfate (0.05, 0.1%) produced higher erythema in normal skin for AA/GA than for GG. CONCLUSIONS: This study provides evidence that the TNF-α polymorphism at -308 and an atopic history impact the severity of irritation and recovery from exposure and response to treatment for common hand skin products in both chronic irritant hand dermatitis and normal skin.

Hippocampal nAChRs mediate nicotine withdrawal-related learning deficits.

Nicotine modulation of learning may contribute to its abuse liability. The role of hippocampal nicotinic acetylcholine receptors (nAChRs) in the effects of acute, chronic and withdrawal from chronic nicotine on learning was assessed via intrahippocampal drug infusion in mice. Acute dorsal hippocampal nicotine infusion enhanced contextual fear conditioning. Conversely, chronic intrahippocampal infusion of a matched dose had no effect, and withdrawal from chronic infusion impaired learning. Thus, hippocampal functional adaptation, evidenced by learning deficits during abstinence, occurs with the transition from acute to chronic nicotine exposure. To investigate which hippocampal nAChRs mediate these adaptations, C57BL/6, beta2 nAChR subunit knockout (KO), and wildtype (WT) mice treated chronically with systemic nicotine received intrahippocampal dihydro-beta-erythroidine (a high affinity nAChR antagonist). Intrahippocampal dihydro-beta-erythroidine precipitated learning deficits in all but the KO mice. Therefore, the action of nicotine at hippocampal beta2 nAChRs mediates adaptations in hippocampal function that underlie withdrawal deficits in contextual fear conditioning.

Survey assessment of physical function following postmastectomy breast reconstruction.

BACKGROUND: Although isokinetic testing has demonstrated postoperative declines in trunk function following transverse rectus abdominis musculocutaneous (TRAM) flap reconstruction, there is little research assessing effects of these procedures on patient-reported measures of physical functioning. The authors used patient surveys to assess the affect of breast reconstruction on postoperative physical functioning. METHODS: Patients were recruited from 12 institutions and underwent expander/implant, pedicle TRAM flap, or free TRAM flap reconstruction. Preoperatively and 2 years postoperatively participants completed self-administered surveys, including the Medical Outcome Study Short Form-36 physical and role functioning subscales; the Functional Assessment of Cancer Therapy-Breast physical and functional well-being subscales; and new, condition-specific items. Multiple regressions evaluated the effects of procedure type on postoperative subscale scores. RESULTS: Of 295 patients recruited, 207 completed surveys at year 2, including 167 unilateral and 40 bilateral reconstructions. TRAM flaps were performed in 159 of 207 patients. There were 133 immediate and 74 delayed procedures. For unilateral reconstructions, no statistically significant procedure type differences were noted in the postoperative subscale scores. Reconstruction timing had a significant effect on function for only one of the subscales, with delayed reconstruction patients reporting significantly lower postoperative scores for Functional Assessment of Cancer Therapy-Breast functional well-being compared with women receiving immediate procedures. On most of the measures, high levels of preoperative functioning were significantly associated with better postoperative scores. CONCLUSION: Although deficits in trunk function have been demonstrated objectively following TRAM flap reconstruction, these changes have minimal impact on patients' daily physical functioning.

Associative learning, the hippocampus, and nicotine addiction.

The abuse liability of nicotine is comparable to or greater than that of a variety of addictive substances. However, the reinforcing and/or rewarding properties of addictive substances other than nicotine far outweigh the reinforcing and/or rewarding effects associated with nicotine use. These data suggest that, in addition to the intrinsic reinforcing effects of nicotine, other factors may contribute to nicotine addiction. One such factor is associative learning, or rather, the ability of nicotine to alter learning and memory processes that may underlie addiction. The present paper presents an overview of the role of learning in nicotine addiction. In addition, recent advances in the identification of behavioral processes, neural substrates, and cellular and molecular substrates that underlie nicotine-associated alterations in learning are reviewed. Particular attention has been paid to research that describes the role of the hippocampus and hippocampus-dependent learning processes in nicotine addiction.

Hippocampal alpha4beta2 nicotinic acetylcholine receptor involvement in the enhancing effect of acute nicotine on contextual fear conditioning.

Nicotine is known to enhance learning and memory in hippocampus-dependent tasks such as contextual fear conditioning. The present study was designed to directly examine whether the hippocampus plays a role in mediating this enhancement and which nicotinic acetylcholine receptor (nAChR) subtypes localized to the hippocampus are critical for enhanced learning. Contextual fear conditioning consisted of two white noise conditioned stimuli presentations, each coterminating with a 2 s, 0.57 mA footshock separated by a 120 s intertrial interval. Nicotine (0.09, 0.18, and 0.35 microg per side) was bilaterally infused into the dorsal hippocampus before training and testing. Infusions of nicotine into the dorsal hippocampus produced a dose-dependent enhancement of contextual fear conditioning. To determine which nAChRs are critical to the enhancing effect of nicotine, the preferential alpha4beta2 nAChR antagonist, dihydro-beta-erythroidine (DHbetaE) (6.00 and 18.00 microg per side), or the preferential alpha7 nAChR antagonist, methyllycaconitine (MLA) (13.50 and 27.00 microg per side), was bilaterally infused into the dorsal hippocampus before systemic injections of nicotine (0.09 mg/kg). DHbetaE infusions dose-dependently blocked the enhancement of contextual fear conditioning by nicotine, whereas MLA infusions yielded an intermediate effect. In addition, neither DHbetaE nor MLA had an effect on contextual fear conditioning in the absence of systemic nicotine. The present results suggest a critical role for alpha4beta2 nAChRs in the dorsal hippocampus for mediating the enhancing effect of nicotine on contextual fear conditioning.

Atomoxetine reverses nicotine withdrawal-associated deficits in contextual fear conditioning.

Recent evidence suggests that the cognitive symptoms of nicotine withdrawal and the cognitive symptoms of attention deficit hyperactivity disorder (ADHD) may share neural correlates. Thus, therapeutics that ameliorate ADHD symptoms may also ameliorate nicotine-withdrawal symptoms. The present research tested this hypothesis in an animal model of nicotine withdrawal-associated cognitive deficits using atomoxetine, a norepinephrine reuptake inhibitor that is approved by the FDA to treat the symptoms of ADHD. C57BL/6 mice were prepared with osmotic minipumps that administered 6.3 mg/kg/day of nicotine or saline, and the minipumps were removed after 12 days of continuous treatment. Twenty-four hours later, mice were trained in delay fear conditioning using two paired presentations of an auditory conditioned stimulus (CS) with a footshock unconditioned stimulus. Testing for freezing in response to the training context and for freezing in response to the CS occurred the next day. Nicotine-withdrawn mice and their saline-treated counterparts received either saline or atomoxetine before training and the context test. Consistent with previous research, the results indicate that mice withdrawn from chronic nicotine demonstrated lower levels of contextual fear conditioning than mice that were not withdrawn from chronic nicotine. Atomoxetine dose-dependently reversed the deficit, suggesting that nicotine withdrawal may be associated with changes in noradrenergic function, acetylcholinergic function, and/or with changes in cell signaling cascades that are activated by both nicotine and norepinephrine. These data suggest that atomoxetine may be efficacious for treating nicotine withdrawal-associated cognitive deficits that promote relapse in abstinent smokers.