Neonatal Animals Research Articles

De novo methylation in male germ cells of the common marmoset monkey occurs during postnatal development and is maintained in vitro

ABSTRACTThe timing of de novo DNA methylation in male germ cells during human testicular development is yet unsolved. Apart from that, the stability of established imprinting patterns in vitro is controversially discussed. This study aimed at determining the timing of DNA de novo methylation and at assessing the stability of the methylation status in vitro. We employed the marmoset monkey (Callithrix jacchus) as it is considered the best non-human primate model for human testicular development. We selected neonatal, pre-pubertal, pubertal, and adult animals (n = 3, each) and assessed germ cell global DNA methylation levels by 5-methyl cytosine staining, and Alu elements and gene-specific methylation (H19, LIT1, SNRPN, MEST, OCT4, MAGE-A4, and DDX-4) by pyrosequencing. De novo methylation is progressively established during postnatal primate development and continues until adulthood, a process that is different in most other species. Importantly, once established, methylation patterns remained stable, as demonstrated using in vitro cultures. Thus, the marmoset monkey is a unique model for the study of postnatal DNA methylation mechanisms in germ cells and for the identification of epimutations and their causes.

Transcriptomic profiling reveals disordered regulation of surfactant homeostasis in neonatal cloned bovines with collapsed lungs and respiratory distress.

Respiratory distress is a major cause of mortality in cloned neonatal animals, but its pathogenesis remains poorly understood. Here, we used necropsy and histology procedures to evaluate the lungs of cloned neonatal bovines dying of respiratory distress, finding incomplete lung dilation, alveolar collapse, and thickened alveolar walls. Comparison of the transcriptomes between collapsed lungs of cloned bovines and their normal counterparts revealed 1373 differentially expressed genes in collapsed lungs (p < 0.05, fold change >1.5 or <1.5-1 ), many of which were associated with surfactant biosynthesis, secretion, transport, recycling, and degradation. ERK/MAPK and Notch signaling pathways were among the canonical pathways relevant to surfactant homeostasis. Expression of the genes encoding Surfactant protein B (SPB) and Surfactant protein C (SPC)-which control surfactant lipid packing, spreading, and stability-were significantly lower in collapsed lungs of cloned neonates at the transcript (p < 0.01) and protein levels (p < 0.05) relative to that in normal lungs. Thus, our results provide an initial view into the changes in gene expression in cloned newborns with lung collapse and respiratory distress, and present a valuable resource for developing novel preventive or therapeutic strategies to reduce the mortality rate of cloned animals and to improve the efficiency of somatic cell nuclear transfer technology.

Dendritic overgrowth and elevated ERK signaling during neonatal development in a mouse model of autism.

Autism spectrum disorder (hereafter referred to as “ASD”) is a heterogeneous neurodevelopmental condition characterized by impaired social communication and interactions, and restricted, repetitive activities or interests. Alterations in network connectivity and memory function are frequently observed in autism patients, often involving the hippocampus. However, specific changes during early brain development leading to disrupted functioning remain largely unclear. Here, we investigated the development of dendritic arbor of hippocampal CA1 pyramidal neurons in the BTBR T+tf/J (BTBR) mouse model of autism. BTBR mice display the defining behavioural features of autism, and also exhibit impaired learning and memory. We found that compared to control C57BL/6J (B6) animals, the lengths of both apical and basal dendrites were significantly greater in neonatal BTBR animals. Further, basal dendrites in the BTBR mice had higher branching complexity. In contrast, cross-sectional area of the soma was unchanged. In addition, we observed a similar density of CA1 pyramidal neurons and thickness of the neuronal layer between the two strains. Thus, there was a specific, compartmentalized overgrowth of dendrites during early development in the BTBR animals. Biochemical analysis further showed that the extracellular signal-regulated kinases (ERK) pathway was up-regulated in the hippocampus of neonatal BTBR animals. Since dendritic structure is critical for information integration and relay, our data suggest that altered development of dendrites could potentially contribute to impaired hippocampal function and behavior observed in the BTBR model, and that this might be related to increased activation of the ERK pathway.

Multilevel Analysis of Locomotion in Immature Preparations Suggests Innovative Strategies to Reactivate Stepping after Spinal Cord Injury.

Locomotion is one of the most complex motor behaviors. Locomotor patterns change during early life, reflecting development of numerous peripheral and hierarchically organized central structures. Among them, the spinal cord is of particular interest since it houses the central pattern generator (CPG) for locomotion. This main command center is capable of eliciting and coordinating complex series of rhythmic neural signals sent to motoneurons and to corresponding target-muscles for basic locomotor activity. For a long-time, the CPG has been considered a black box. In recent years, complementary insights from in vitro and in vivo animal models have contributed significantly to a better understanding of its constituents, properties and ways to recover locomotion after a spinal cord injury (SCI). This review discusses key findings made by comparing the results of in vitro isolated spinal cord preparations and spinal-transected in vivo models from neonatal animals. Pharmacological, electrical, and sensory stimulation approaches largely used to further understand CPG function may also soon become therapeutic tools for potent CPG reactivation and locomotor movement induction in persons with SCI or developmental neuromuscular disorder.

Long-term effects of enriched environment following neonatal hypoxia-ischemia on behavior, BDNF and synaptophysin levels in rat hippocampus: Effect of combined treatment with G-CSF

Increasing evidence shows that exposure to an enriched environment (EE) is neuroprotective in adult and neonatal animal models of brain ischemia. However, the mechanisms underlying this effect remain unclear. The aim of the current study was to investigate whether post-weaning EE would be effective in preventing functional deficits and brain damage by affecting markers of synaptic plasticity in a neonatal rat model of hypoxia-ischemia (HI). We also examined the possibility that granulocyte-colony stimulating factor (G-CSF), a growth factor with known neuroprotective effects in a variety of experimental brain injury models, combined with EE stimulation could enhance the potential beneficial effect of EE. Seven-day-old Wistar rats of either sex were subjected to permanent ligation of the left common carotid artery followed by 60min of hypoxia (8% O2) and immediately after weaning (postnatal day 21) were housed in enriched conditions for 4weeks. A group of enriched-housed rats had been treated with G-CSF immediately after HI for 5 consecutive days (50μg/kg/day). Behavioral examination took place approximately at three months of age and included assessments of learning and memory (Morris water maze) as well as motor coordination (Rota-Rod). Infarct size and hippocampal area were estimated following behavioral assessment. Synaptic plasticity was evaluated based on BDNF and synaptophysin expression in the dorsal hippocampus. EE resulted in recovery of post-HI motor deficits and partial improvement of memory impairments which was not accompanied by reduced brain damage. Increased synaptophysin expression was observed in the contralateral to carotid ligation hemisphere. Hypoxia-ischemia alone or followed by enriched conditions did not affect BDNF expression which was increased only in enriched-housed normal rats. The combined therapy of G-CSF and EE further enhanced cognitive function compared to EE provided as monotherapy and prevented HI-induced brain damage by altering synaptic plasticity as reflected by increased synaptophysin expression. The above findings demonstrate that combination of neuroprotective treatments may result in increased protection and it might be a more effective strategy for the treatment of neonatal hypoxic-ischemic brain injury.

Guidance on the risk assessment of substances present in food intended for infants below 16 weeks of age

Following a request from the European Commission to EFSA, the EFSA Scientific Committee (SC) prepared a guidance for the risk assessment of substances present in food intended for infants below 16 weeks of age. In its approach to develop this guidance, the EFSA SC took into account, among others, (i) an exposure assessment based on infant formula as the only source of nutrition; (ii) knowledge of organ development in human infants, including the development of the gut, metabolic and excretory capacities, the brain and brain barriers, the immune system, the endocrine and reproductive systems; (iii) the overall toxicological profile of the substance identified through the standard toxicological tests, including critical effects; (iv) the relevance for the human infant of the neonatal experimental animal models used. The EFSA SC notes that during the period from birth up to 16 weeks, infants are expected to be exclusively fed on breast milk and/or infant formula. The EFSA SC views this period as the time where health‐based guidance values for the general population do not apply without further considerations. High infant formula consumption per body weight is derived from 95th percentile consumption. The first weeks of life is the time of the highest relative consumption on a body weight basis. Therefore, when performing an exposure assessment, the EFSA SC proposes to use the high consumption value of 260 mL/kg bw per day. A decision tree approach is proposed that enables a risk assessment of substances present in food intended for infants below 16 weeks of age. The additional information needed when testing substances present in food for infants below 16 weeks of age and the approach to be taken for the risk assessment are on a case‐by‐case basis, depending on whether the substance is added intentionally to food and is systemically available.

Transfer of optogenetic vectors into the brain of neonatal animals to study neuron functions during subsequent periods of development

Optogenetics, that is, the control of cell activity using photosensitive ion channels, opsins, with the light of a specific wavelength, is increasingly being used to study the activities and functions of neurons. The expression of opsins in the cell membrane, making the cell sensitive to light, is achieved by means of the viral vectors, mainly constructed using lentiviruses or adeno-associated viruses (AAVs), with the inserted nucleotide sequences encoding the photochannel proteins. The specific promoter for the target cell type added to the transgene expression cassette provides opsin production only in these target cells. The aim of this work was to briefly describe the optogenetic method, as well as to analyze the possibility to use administration of viral vectors into the brain of neonatal animals to study the function of neurons in vivo during subsequent periods of development. In this analysis, 3-day-old rat pups are intracerebroventricularly injected with the optovector (pAAV-CAMKIIa-ChR2h134-YFP) coding for a neuron-activating photochannel and a marker, yellow fluorescent protein, under the CAMKIIa promoter, specific for glutamatergic neurons. The peak expression of the transferred gene is usually achieved 3–5 weeks after the vector transfer, which is also observed in our experiments. Stimulation of the hippocampal neurons with blue light in the 20-day-old animals injected with the optovector on 3rd day of their lives increases the discharge activity of these neurons, as well as the expression of c-Fos protein, a well-known marker of neuronal activation. The same experiments with older animals (60 days after the neonatal optochannel gene transfer) demonstrate neither any noticeable expression nor photoactivation of the target hippocampal neurons. Thus, the neonatal administration of a viral vector carrying an optochannel gene is suitable for the study of brain neurons in juvenile rats but requires additional assessment of the corresponding gene expression during subsequent periods of development.

Influence of central hemodynamics on VV ECMO oxygen delivery in neonatal animal model.

Recirculation of oxygenated blood in venovenous extracorporeal membrane oxygenation (VV ECMO) can decrease the oxygen delivery provided by the ECMO support. This study investigated the influence of central hemodynamics and catheter position on the amount of recirculation and oxygen delivery during VV ECMO. Recirculation was measured in seven newborn lambs (mean weight 4.7 kg) during VV ECMO using the ELSA Monitor (Transonic Systems, Inc., Ithaca, NY) and using the central venous line (CVL) method. The ECMO pump was set at the prescribed flow of 110-120 mL/kg/min for a targeted oxygen delivery rate of 6cc/kg/min without recirculation. Hemodynamic status before and during ECMO was also measured by the COstatus Monitor (Transonic Systems, Inc.,Ithaca, NY). Lambs with a higher cardiac index (>160 ml/min/kg), had a tendency to have higher percent oxygen delivery (65-94%, at prescribed flow) while lambs with lower cardiac index (<150 ml/min/kg), tended to have lower percent oxygen delivery (39-62%, at prescribed flow). ELSA recirculation measurements had a squared correlation coefficient R2 = 0.8 with the CVL method. The ELSA monitor provides an easy to use, non-invasive method to measure recirculation in VV ECMO. The data suggests that cardiac function may play an important prognostic role in achieving effective VV ECMO support.

Investigating the Potential Role of North American Animals as Hosts for Zika Virus

The recent emergence of the mosquito‐borne Zika virus (ZIKV) in the Americas has become a global public health concern. While little is known about potential animal reservoirs for ZIKV, nonhuman primates are thought to play a role during enzootic transmission cycles. We describe a series of experimental infections designed to investigate whether species within certain taxonomic groups in North America have the potential to serve as ZIKV amplifying hosts or reservoirs. The objectives of this study were to (1) identify potential animal reservoirs in North America with an emphasis on wildlife and livestock, (2) examine ZIKV pathogenicity in neonatal and adult animals, and (3) evaluate the ability of animal models to recapitulate neonatal birth defects as seen in human patients. Species investigated include armadillos, cottontail rabbits, goats, mink, chickens, pigeons, ground hogs, deer mice, cattle, raccoons, ducks, Syrian Golden hamsters, garter snakes, leopard frogs, house sparrows, and pigs. Animals ranging in age from neonates to adults were inoculated with 105 PFU of one of two ZIKV strains (PRVA BC59 and FSS13025) by subcutaneous and intradermal injection. All animals were monitored daily for clinical signs of disease. Blood samples and body temperatures were collected on days 0–5, 7, 14, 21 and 28 post inoculation. Plaque assays, polymerase chain reaction (PCR), and plaque reduction neutralization tests were performed to detect viral presence and antibody production. Infectious virus was isolated from seven frogs and one armadillo. In addition, neutralizing antibodies were detected in goats, rabbits, ducks, frogs, and pigs. This study indicates that the animals tested to date are unlikely to act as animal reservoirs for ZIKV. However, goats, rabbits, frogs, and pigs could potentially serve as a sentinel species. Future work includes examining the role of amphibians in the transmission dynamics of ZIKV. Understanding the transmission cycle and maintenance of ZIKV in animals will help in developing effective surveillance programs and preventative measures for future outbreaks.Support or Funding InformationThis material is based upon work supported by Colorado State University Animal Models Core and the U.S. Department of Homeland Security under Grant Award Number DHS‐2010‐ST‐061‐AG0001. The views and conclusions contained in this document are those of the authors and should not be interpreted as necessarily representing the official policies, either expressed or implied, of the U.S. Department of Homeland Security.

Rutin attenuates isoflurane-induced neuroapoptosis via modulating JNK and p38 MAPK pathways in the hippocampi of neonatal rats.

An increasing number of infants and children undergo surgery and are exposed to anesthesia as a part of medical care each year. Isoflurane is a commonly used anesthetic in the pediatric population. However, previous studies have reported widespread isoflurane-induced neuroapoptosis and cognitive impairments in neonatal animal models, raising concerns over the administration of isoflurane in the pediatric population. The current study investigated the effects of rutin, a flavonoid, on isoflurane-induced neuroapoptosis in a neonatal rodent model. Groups of neonatal rat pups were administered rutin at doses of 10, 20 or 40 mg/kg body weight from postnatal day 1 (P1) to P15. On P7, pups were exposed to 0.75% isoflurane for 6 h. Rat pups in the control groups did not receive rutin, and did not receive anesthesia in one group. Neuroapoptosis following isoflurane exposure was determined by TUNEL assay. The expression levels of cleaved caspase-3, apoptotic pathway proteins [Bcl2-associated agonist of cell death (Bad), phospho-Bad, Bax, B-cell lymphoma 2 (Bcl-2) and Bcl-xL and mitogen-activated protein kinases (MAPK)] signalling pathway proteins [c-Jun N-terminal kinase (JNK), phospho-JNK, extracellular-signal-regulated kinase 1/2 (ERK1/2), phosphoERK1/2, p38, phospho-p38 and phospho-c-Jun], were determined by western blot analysis. The Morris water maze test was used to assess the learning and memory of pups on P30 and P31. The present study found that rutin at the tested doses of 10, 20 and 40 mg significantly reduced (P<0.05) the isoflurane-induced elevation in apoptotic cell count. The expression levels of caspase-3, Bad, Bax and MAPK proteins, which were increased following isoflurane treatment, were rescued by rutin treatment. Furthermore, rutin prevented the increase in Bcl-xL, Bcl-2 and phospho-Bad expression following isoflurane treatment, and enhanced the memory of the rats. Rutin provided neuroprotection against isoflurane-induced neuronal apoptosis and improved the learning and memory of rats by effectively regulating the expression levels of proteins in the MAPK pathway.

Chicken Egg Yolk Antibodies (IgY) for Prophylaxis and Treatment of Rotavirus Diarrhea in Human and Animal Neonates: A Concise Review.

The rotavirus-induced diarrhea of human and animal neonates is a major public health concern worldwide. Until recently, no effective therapy is available to specifically inactivate the rotavirion particles within the gut. Passive immunotherapy by oral administration of chicken egg yolk antibody (IgY) has emerged of late as a fresh alternative strategy to control infectious diseases of the alimentary tract and has been applied in the treatment of diarrhea due to rotavirus infection. The purpose of this concise review is to evaluate evidence on the properties and performance of anti-rotavirus immunoglobulin Y (IgY) for prevention and treatment of rotavirus diarrhea in human and animal neonates. A survey of relevant anti-rotavirus IgY basic studies and clinical trials among neonatal animals (since 1994-2015) and humans (since 1982-2015) have been reviewed and briefly summarized. Our analysis of a number of rotavirus investigations involving animal and human clinical trials revealed that anti-rotavirus IgY significantly reduced the severity of clinical manifestation of diarrhea among IgY-treated subjects relative to a corresponding control or placebo group. The accumulated information as a whole depicts oral IgY to be a safe and efficacious option for treatment of rotavirus diarrhea in neonates. There is however a clear need for more randomized, placebo controlled and double-blind trials with bigger sample size to further solidify and confirm claims of efficacy and safety in controlling diarrhea caused by rotavirus infection especially among human infants with health issues such as low birth weights or compromised immunity in whom it is most needed.

Morphological and Morphometric Study of the Development of Leydig Cell population of Donkey (Equus asinus) Testis from Birth to Maturity

In this investigation, the postnatal morphologic and morphometric characteristics of Leydig cells and testicular interstitial tissue in 20 healthy donkey testes were studied. The volume percentage of the interstitial tissue was about 87.24% in neonates and decreased gradually with postnatal age reaching about 21.56% in mature animals. The Leydig cell population underwent evident morphologic and morphometric changes during the postnatal life. Morphologically, there were two discrete types of Leydig cells, which recognized side by side in the testis of neonatal and suckling animals. The first type (steroidogenic fetal Leydig cells), was large, contained partially shrunken nucleus together with vacuolated granular cytoplasm. This cellular type, were predominant in neonates, underwent degeneration and decreased gradually until being very few at two months of age. The second type (non-steroidogenic) was small, few at birth and contained large central nucleus. This type develops from undifferentiated fibroblastic elements and underwent gradual increase in number with age, where it dominated in the late suckling period. Some of these cells were degenerated and the remaining became transformed into the adult type of Leydig cells. The volume percentage and absolute number of Leydig cells displayed the same pattern of undulation during postnatal development, which being very high in late suckling animals. The volume percentage of the vascular components of the interstitial tissue displayed a slight decrease between neonatal and early suckling periods (from about 8% to about 7% of the interstitium); however, they increased steadily through premature and mature animals (about 12% and 20% respectively). The present study provides baseline information for further experimental or quantitative studies exploring the normal development of the Leydig cells in donkey testis and other related species.

Neonatal seizures and disruption to neurotransmitter systems

Neonatal seizures and disruption to neurotransmitter systems

Association of Anesthesia and Surgery During Childhood With Long-term Academic Performance

The results of preclinical studies suggest that anesthetic drugs administered to neonatal animals cause widespread neuronal apoptosis and later neurocognitive impairment. Adequately powered studies in the pediatric surgical population are scarce, and it is unclear whether such preclinical findings are relevant for the pediatric setting. To examine the association of anesthesia and surgery before age 4 years with long-term academic and cognitive performance indexed by school grades at age 16 years and IQ test scores at military conscription. This investigation was a cohort study among all children born in Sweden between January 1973 and December 1993. The dates of analysis were April 2013 to October 2015. Among all 2 174 073 Swedish children born between 1973 and 1993, we identified a primary study cohort of 33 514 children with 1 anesthesia and surgery exposure before age 4 years and no subsequent hospitalization and 159 619 matched unexposed control children. In addition, 3640 children with multiple surgical procedures before age 4 years were studied. Having at least 1 surgical procedure in the Swedish Patient Register before age 4 years. The mean school grades at age 16 years and IQ test scores at military conscription at age 18 years. The mean difference between the exposed cohort and unexposed cohort was estimated in a model that included sex, month of birth during the same year, gestational age at delivery, Apgar score at 5 minutes, maternal and paternal educational levels, annual taxable household income, cohabiting parents, and number of siblings. Among 33 514 exposed children (22 484 male and 11 030 female) and 159 619 unexposed children (105 812 male and 53 807 female) in the primary study cohort, 1 exposure before age 4 years was associated with a mean difference of 0.41% (95% CI, 0.12%-0.70%) lower school grades and 0.97% (95% CI, 0.15%-1.78%) lower IQ test scores. The magnitude of the difference was the same after multiple exposures. There was no difference in school grades with 1 exposure before ages 6 months, 7 to 12 months, 13 to 24 months, or 25 to 36 months. The overall difference was markedly less than the differences associated with sex, maternal educational level, or month of birth during the same year. Exposure to anesthesia and surgery before age 4 years has a small association with later academic performance or cognitive performance in adolescence on a population level. While more vulnerable subgroups of children may exist, the low overall difference in academic performance after childhood exposure to surgery is reassuring. These findings should be interpreted in light of potential adverse effects of postponing surgery.

Effects of Isolation on Stress Responses to Novel Stimuli in Subadult Chickens (Gallus gallus)

AbstractExtensive research has examined the effects of social isolation in neonatal and adult animal populations, but few studies have examined the effect of social isolation in early adulthood. Animals reaching reproductive age often experience extensive social changes as they leave their natal site, and a social stressor like isolation may uniquely affect this age group. Furthermore, adolescence is a time when sex differences in behavior become more pronounced. As such, the effects of social stressors are likely to vary by sex. In this study, we used noninvasive methods to evaluate stress responses to social change in male and female subadult chickens (Gallus gallus). Half of the birds experienced regular sessions of social isolation over the course of 2 wk, while the other half were never isolated. Subsequently, all of the animals were exposed to a suite of three novel probes, including an open‐field test. We monitored the birds’ behavioral (head movements) and physiological (fecal glucocorticoid metabolites, FGM) response to the tests. Our results indicate that, for subadult chickens, the effect of social isolation is sex dependent: Male FGM and behavioral responses did not change with subsequent experiences, in contrast to females. Females also exhibited more social reinstatement behavior compared to males. Our results are consistent with the expectations of differences between the sexes based on changes in the social environment due to sex‐biased dispersal patterns. For both sexes, the FGM and behavioral responses varied independently, which highlights the necessity for multiple measures of stress in animal populations.

Effect of the I/E ratio on CO2 removal during high-frequency oscillatory ventilation with volume guarantee in a neonatal animal model of RDS.

The objective of this study was to analyze the effect of I/E ratio on carbon dioxide (CO2) elimination during high-frequency oscillatory ventilation (HFOV) combined with volume guarantee (VG). Five 2-day-old piglets were studied before and after a bronchoalveolar lavage (BAL). The effect of an I/E ratio of 1:1 and 1:2 with (VG-ON) and without VG (VG-OFF) on PaCO2, as well as delta and mean airway pressures at the airway opening (∆Phf-ao, mPaw-ao) and at the tracheal level (∆Phf-t, mPaw-t) were evaluated at frequencies of 5, 8, 11, and 14Hz. With the VG-ON, PaCO2 was significant lower with the I/E ratio of 1:2 at 5Hz compared with the 1:1. mPaw-t was higher than mPaw-ao, with 1:1 I/E ratio, and on VG-ON, this difference was statistically significant. "In this animal study and with this ventilator, the I/E ratio of 1:1 compared to 1:2 in HFOV and VG-ON did not produce a higher CO2 lavage as when HFOV was used without the VG modality. Even more, a lower PaCO2 was found when using the lower frequency and 1:2 ratio compared to 1:1. So in contrast to non-VG HFOV mode, using a fixed tidal volume, no significant changes on CO2 elimination are observed during HFOV when the I/E ratios of 1:1 and 1:2 are compared at different frequencies." •The tidal volume on HFOV is determinant in CO 2 removal, and this is generated by delta pressure and the length of the inspiratory time. What is New: •HFOV combined with VG, an I/E ratio of 1:2 is more effective to remove CO 2 , and this is not related to the tidal volume.

Serotonin controls initiation of locomotion and afferent modulation of coordination via 5-HT7 receptors in adult rats.

Experiments on neonatal rodent spinal cord showed that serotonin (5-HT), acting via 5-HT7 receptors, is required for initiation of locomotion and for controlling the action of interneurons responsible for inter- and intralimb coordination, but the importance of the 5-HT system in adult locomotion is not clear. Blockade of spinal 5-HT7 receptors interfered with voluntary locomotion in adult rats and fictive locomotion in paralysed decerebrate rats with no afferent feedback, consistent with a requirement for activation of descending 5-HT neurons for production of locomotion. The direct control of coordinating interneurons by 5-HT7 receptors observed in neonatal animals was not found during fictive locomotion, revealing a developmental shift from direct control of locomotor interneurons in neonates to control of afferent input from the moving limb in adults. An understanding of the afferents controlled by 5-HT during locomotion is required for optimal use of rehabilitation therapies involving the use of serotonergic drugs. Serotonergic pathways to the spinal cord are implicated in the control of locomotion based on studies using serotonin type 7 (5-HT7 ) receptor agonists and antagonists and 5-HT7 receptor knockout mice. Blockade of these receptors is thought to interfere with the activity of coordinating interneurons, a conclusion derived primarily from in vitro studies on isolated spinal cord of neonatal rats and mice. Developmental changes in the effects of serotonin (5-HT) on spinal neurons have recently been described, and there is increasing data on control of sensory input by 5-HT7 receptors on dorsal root ganglion cells and/or dorsal horn neurons, leading us to determine the effects of 5-HT7 receptor blockade on voluntary overground locomotion and on locomotion without afferent input from the moving limb (fictive locomotion) in adult animals. Intrathecal injections of the selective 5-HT7 antagonist SB269970 in adult intact rats suppressed locomotion by partial paralysis of hindlimbs. This occurred without a direct effect on motoneurons as revealed by an investigation of reflex activity. The antagonist disrupted intra- and interlimb coordination during locomotion in all intact animals but not during fictive locomotion induced by stimulation of the mesencephalic locomotor region (MLR). MLR-evoked fictive locomotion was transiently blocked, then the amplitude and frequency of rhythmic activity were reduced by SB269970, consistent with the notion that the MLR activates 5-HT neurons, leading to excitation of central pattern generator neurons with 5-HT7 receptors. Effects on coordination in adults required the presence of afferent input, suggesting a switch to 5-HT7 receptor-mediated control of sensory pathways during development.

Linking Genomo- and Pathotype: Exploiting the Zebrafish Embryo Model to Investigate the Divergent Virulence Potential among Cronobacter spp.

Bacteria belonging to the genus Cronobacter have been recognized as causative agents of life-threatening systemic infections primarily in premature, low-birth weight and immune-compromised neonates. Apparently not all Cronobacter species are linked to infantile infections and it has been proposed that virulence varies among strains. Whole genome comparisons and in silico analysis have proven to be powerful tools in elucidating potential virulence determinants, the presence/absence of which may explain the differential virulence behaviour of strains. However, validation of these factors has in the past been hampered by the availability of a suitable neonatal animal model. In the present study we have used zebrafish embryos to model Cronobacter infections in vivo using wild type and genetically engineered strains. Our experiments confirmed the role of the RepF1B-like plasmids as “virulence plasmids” in Cronobacter and underpinned the importantce of two putative virulence factors—cpa and zpx—in in vivo pathogenesis. We propose that by using this model in vivo infection studies are now possible on a large scale level which will boost the understanding on the virulence strategies employed by these pathogens.

Сравнительная морфологическая характеристика первичных культур клеток надпочечников неонатальных животных

Сравнительная морфологическая характеристика первичных культур клеток надпочечников неонатальных животных

Rescue of the Functional Alterations of Motor Cortical Circuits in Arginase Deficiency by Neonatal Gene Therapy.

Arginase 1 deficiency is a urea cycle disorder associated with hyperargininemia, spastic diplegia, loss of ambulation, intellectual disability, and seizures. To gain insight on how loss of arginase expression affects the excitability and synaptic connectivity of the cortical neurons in the developing brain, we used anatomical, ultrastructural, and electrophysiological techniques to determine how single-copy and double-copy arginase deletion affects cortical circuits in mice. We find that the loss of arginase 1 expression results in decreased dendritic complexity, decreased excitatory and inhibitory synapse numbers, decreased intrinsic excitability, and altered synaptic transmission in layer 5 motor cortical neurons. Hepatic arginase 1 gene therapy using adeno-associated virus rescued nearly all these abnormalities when administered to neonatal homozygous knock-out animals. Therefore, gene therapeutic strategies can reverse physiological and anatomical markers of arginase 1 deficiency and therefore may be of therapeutic benefit for the neurological disabilities in this syndrome. These studies are one of the few investigations to try to understand the underlying neurological dysfunction that occurs in urea cycle disorders and the only to examine arginase deficiency. We have demonstrated by multiple modalities that, in murine layer 5 cortical neurons, a gradation of abnormalities exists based on the functional copy number of arginase: intrinsic excitability is altered, there is decreased density in asymmetrical and perisomatic synapses, and analysis of the dendritic complexity is lowest in the homozygous knock-out. With neonatal administration of adeno-associated virus expressing arginase, there is near-total recovery of the abnormalities in neurons and cortical circuits, supporting the concept that neonatal gene therapy may prevent the functional abnormalities that occur in arginase deficiency.