Neonatal Rabbit Research Articles

Comparative Analysis of Angora Rabbit Colostrum and Mature Milk Using Quantitative Proteomics.

Colostrum intake is a crucial determinant of survival in newborn rabbits. Neonates rely entirely on passive immunity transfer from their mothers while suckling colostrum. The goal of this study was to explore the protein differences of rabbit milk during different lactation periods. Our findings showed that the daily milk yield exhibited an increasing trend from the 2nd to the 21st day of lactation. A data-independent acquisition proteomics approach identified a total of 2011 proteins. Significantly, different abundances were found for 525 proteins in the colostrum and the mature milk samples. Eleven differentially abundant proteins (DAPs) were examined using parallel reaction monitoring, which verified the reliability of the proteomic data. Gene Ontology analysis revealed that these DAPs were primarily associated with glycosyltransferase activity, macromolecule transmembrane transporter activity, and regulation of acute inflammatory response. The dominant metabolic pathways of the DAPs involve the complement and coagulation cascades. A protein-protein interaction analysis identified apolipoprotein B, apolipoprotein A1, triose phosphate isomerase 1, and albumin as the hub proteins responsible for distinguishing differences between biological properties in rabbit colostrum and mature milk. These findings enhance our comprehension of the rabbit milk proteome, particularly in expanding our knowledge regarding the requirements of neonatal rabbits.

Long-term pulmonary and neurodevelopmental impairment in a fetal growth restriction rabbit model.

Fetal growth restriction (FGR) remains one of the main obstetrical problems worldwide, with consequences beyond perinatal life. Animal models with developmental and structural similarities to the human are essential to understand FGR long-term consequences and design novel therapeutic strategies aimed at preventing or ameliorating them. Herein, we described the long-term consequences of FGR in pulmonary function, structure, and gene expression, and characterized neurodevelopmental sequelae up to preadolescence in a rabbit model. FGR was induced at gestational day 25 by surgically reducing placental blood supply in one uterine horn, leaving the contralateral horn as internal control. Neonatal rabbits born near term were assigned to foster care in mixed groups until postnatal day (PND) 21. At that time, one group underwent pulmonary biomechanical testing followed by lung morphometry and gene expression analysis. A second group underwent longitudinal neurobehavioral assessment until PND 60 followed by brain harvesting for multiregional oligodendrocyte and microglia quantification. FGR was associated with impaired pulmonary function and lung development at PND 21. FGR rabbits had higher respiratory resistance and altered parenchymal biomechanical properties in the lungs. FGR lungs presented thicker alveolar septal walls and reduced alveolar space. Furthermore, the airway smooth muscle content was increased, and the tunica media of the intra-acinar pulmonary arteries was thicker. In addition, FGR was associated with anxiety-like behavior, impaired memory and attention, and lower oligodendrocyte proportion in the frontal cortex and white matter. In conclusion, we documented and characterized the detrimental pulmonary function and structural changes after FGR, independent of prematurity, and beyond the neonatal period for the first time in the rabbit model, and describe the oligodendrocyte alteration in pre-adolescent rabbit brains. This characterization will allow researchers to develop and test therapies to treat FGR and prevent its sequelae.

Characterizing post-branching nephrogenesis in the neonatal rabbit

Human nephrogenesis ends prior to birth in term infants (34–36 week gestation), with most (60%) nephrons forming in late gestation in two post-branching nephrogenesis (PBN) periods: arcading and lateral branch nephrogenesis. Preterm infants, however, must execute PBN postnatally. Extreme prematurity is associated with low nephron counts. Identifying additional model(s) that undergo PBN postnatally will help support postnatal PBN in preterm infants. The rabbit exhibits longer postnatal nephrogenesis than the mouse but whether it forms nephrons through PBN has not been determined. We performed morphologic and immunohistological assessments of rabbit nephrogenesis from birth (post-conceptual day 31 or 32) to PC49 using H&E and antibodies against SIX1, SIX2, WT1, ZO-1, and JAG1 in the postnatal period. We performed 3D rendering of the nephrogenic niche to assess for PBN, and supplemented the staining with RNAScope to map the expression of Six1, Six2 (nephron progenitors, NPC), and Ret (ureteric bud tip) transcripts to determine the nephrogenic niche postnatal lifespan. Unlike the mouse, rabbit SIX2 disappeared from NPC before SIX1, resembling the human niche. Active nephrogenesis as defined by the presence of SIX1 + naïve NPC/tip population persisted only until PC35–36 (3–5 postnatal days). 3D morphologic assessments of the cortical nephrons identified an elongated tubule with attached glomeruli extending below the UB tip, consistent with PBN arcades, but not with lateral branch nephrogenesis. We conclude that the rabbit shows morphologic and molecular evidence of PBN arcades continuing postnatally for a shorter period than previously thought. The rabbit is the first non-primate expressing SIX1 in the progenitor population. Our findings suggest that studies of arcading in postnatal nephrogenic niche should be performed within the first 5 days of life in the rabbit.

Micro array patch assisted transdermal delivery of high dose, ibuprofen sodium using thermoresponsive sodium alginate/poly (vinylcaprolactam) in situ gels depot

Current study reports the combined technique of microneedle array patches and thermoresponsive gels. Microneedles array patch mediated insitu skin depots were evaluated for sustain drug delivery using sodium alginate/Poly (vinylcaprolactam) thermoresponsive gels. Their phase transition property from sol-gel state was monitored with AR2000 rheometer. Ibuprofen sodium was loaded in optimized formulations. The non-soluble cross-linked microneedle array patches (MAPs) were prepared from variable biocompatible polymers using silicone micromoulds. The fabricated MAPs were evaluated for mechanical stability, inskin dissolution, insertion forces and moisture contents. The penetration depth of MAPs in neonatal rabbit skin was tracked by optical coherence tomography. The optimized MAPs (GP10000) were used as microporation source in skin owing to their stable nature. Pores formation in skin samples after MAPs treatment was confirmed by optical coherence tomography, dye binding and skin integrity analysis. The invitro permeation of Ibuprofen sodium from formulations was studied using Franz cells across intact skin and MAPs applied skin. It was concluded from the results that Ibuprofen sodium permeation was observed for longer time through MAPs treated skin as compared to intact skin. Confocal study confirmed the diffusion of drug loaded formulations in deeper tissues with higher intensity.

Maternal Daidzein Supplementation during Lactation Promotes Growth Performance, Immunity, and Intestinal Health in Neonatal Rabbits

The main purpose of the present research was to evaluate the effect of varying levels of DA inclusion in maternal diet, in the form of powder, on the born-weaning growth performance (days 1–30) and intestinal health of neonatal rabbits. A total of 152 delivered maternal does (3.94 ± 0.05 kg) were allocated into four groups, with thirty-eight replicates of one doe each, and fed with a control diet (CON) supplemented with different levels of powdered DA (85 mg/kg (DA85), 170 mg/kg (DA170), and 340 mg/kg (DA340)) during lactation. The results show that dietary DA increased individual body weight at days 21 and 30 (p = 0.03 and p < 0.01) and showed a linear and quadratic effect on individual body weight and average daily gain (ADG) (p < 0.05), although ADG was not affected by DA. The serum progesterone (P) (p = 0.05) and glutathione peroxidase (GSH-Px) (p = 0.01) concentrations of maternal rabbits were increased in all maternal DA-supplemented groups and showed a linear and quadratic effect (p < 0.05), while a linear effect was observed in estrogen (E-2) (p < 0.05). Interestingly, the concentrations of the serum insulin-like growth factor-1 (IGF-I) (p < 0.01) and immunoglobulin (IgA and IgG) (p < 0.01 and p = 0.01) in neonatal rabbits were increased in all dietary DA groups, and both showed a linear and quadratic effect (p < 0.05). In addition, the serum interleukin (IL-1β) (p = 0.03) and catalase (CAT) (p = 0.04) concentrations were affected by DA supplements, but linear and quadratic effects were only observed in the catalase (CAT) of neonatal rabbits (p < 0.05). Importantly, the duodenal and jejunal villus heights (p < 0.04 and p = 0.01) and the ratio of villus height to crypt depth (p = 0.02 and p = 0.01) in neonatal rabbits were elevated in all DA-supplemented treatments, while a linear and quadratic effect was observed in jejunum, but a quadratic effect was observed in duodenum (p < 0.05). The ileal villus height (p < 0.01) was also elevated in all DA-supplemented treatments exhibiting both linear and quadratic effects (p < 0.05). Moreover, the expression levels of occludin (p = 0.04), zonula occludens-1 (ZO-1) (p < 0.01), IGF-1 (p = 0.03), and solute carrier family 5 member 9 (SCL5A9) (p < 0.01) in jejunum were markedly elevated in the DA175 and 340 mg/kg DA-supplemented groups and were showing linear and quadratic effects (p < 0.05). Finally, inflammation-related gene expression levels such as NF-κB, TLR4, and MYD88 in jejunal (p = 0.01, p = 0.04, and p < 0.01) and ileal (p = 0.04, p = 0.04, and p = 0.03) epithelium were decreased in the DA170 and DA340 groups (p < 0.05), while linear and quadratic effects were observed for TLR4 in jejunum and for NF-κB and TLR4 MYD88 in ileum (p < 0.05). In summary, as a functional additive, maternal DA supplementation with 170 and 340 mg/kg DA during lactation can promote the growth of neonatal rabbits, which is related to improved antioxidative capacity and immunity, as well as improved intestinal health in neonatal rabbits.

Spinal motoneurons respond aberrantly to serotonin in a rabbit model of cerebral palsy.

Cerebral palsy (CP) is caused by a variety of factors that damage the developing central nervous system. Impaired motor control, including muscle stiffness and spasticity, is the hallmark of spastic CP. Rabbits that experience hypoxic-ischaemic (HI) injury in utero (at 70%-83% gestation) are born with muscle stiffness, hyperreflexia and, as recently discovered, increased 5-HT in the spinal cord. To determine whether serotonergic modulation of spinal motoneurons (MNs) contributes to motor deficits, we performed ex vivo whole cell patch clamp in neonatal rabbit spinal cord slices at postnatal day (P) 0-5. HI MNs responded to the application of α-methyl 5-HT (a 5-HT1 /5-HT2 receptor agonist) and citalopram (a selective 5-HT reuptake inhibitor) with increased amplitude and hyperpolarization of persistent inward currents and hyperpolarized threshold voltage for action potentials, whereas control MNs did not exhibit any of these responses. Although 5-HT similarly modulated MN properties of HI motor-unaffected and motor-affected kits, it affected sag/hyperpolarization-activated cation current (Ih ) and spike frequency adaptation only in HI motor-affected MNs. To further explore the differential sensitivity of MNs to 5-HT, we performed immunostaining for inhibitory 5-HT1A receptors in lumbar spinal MNs at P5. Fewer HI MNs expressed the 5-HT1A receptor compared to age-matched control MNs. This suggests that HI MNs may lack a normal mechanism of central fatigue, mediated by 5-HT1A receptors. Altered expression of other 5-HT receptors (including 5-HT2 ) likely also contributes to the robust increase in HI MN excitability. In summary, by directly exciting MNs, the increased concentration of spinal 5-HT in HI-affected rabbits can cause MN hyperexcitability, muscle stiffness and spasticity characteristic of CP. Therapeutic strategies that target serotonergic neuromodulation may be beneficial to individuals with CP. KEY POINTS: We used whole cell patch clamp electrophysiology to test the responsivity of spinal motoneurons (MNs) from neonatal control and hypoxia-ischaemia (HI) rabbits to 5-HT, which is elevated in the spinal cord after prenatal HI injury. HI rabbit MNs showed a more robust excitatory response to 5-HT than control rabbit MNs, including hyperpolarization of the persistent inward current and threshold voltage for action potentials. Although most MN properties of HI motor-unaffected and motor-affected kits responded similarly to 5-HT, 5-HT caused larger sag/hyperpolarization-activated cation current (Ih ) and altered repetitive firing patterns only in HI motor-affected MNs. Immunostaining revealed that fewer lumbar MNs expressed inhibitory 5-HT1A receptors in HI rabbits compared to controls, which could account for the more robust excitatory response of HI MNs to 5-HT. These results suggest that elevated 5-HT after prenatal HI injury could trigger a cascade of events that lead to muscle stiffness and altered motor unit development.

Thermoregulation in periparturient rabbit does and their neonatal kits with different litter sizes during West African winter

The study evaluated the changes in body temperatures and hematological parameters in periparturient rabbit does and the dynamics of body surface temperature in neonatal rabbit kits during West African winter. The variables were measured in 21 rabbits, comprising periparturient (n = 12) and dry (n = 9) does, and after kindling, the kits were grouped based on litter size, into small (4 kits per litter; n = 24) and large (5–7 kits per litter; n = 34) litter. Results revealed that all body temperatures of the does were lower (p < 0.05) during gestation compared with lactation period or dry does in all hours of the day, and the PCV was significantly higher in lactating than dry does. At birth, small litter kits had higher (p < 0.05) body weight and morning body surface temperature than the large litter kits. The large litter kits showed distinct (p < 0.05) diurnal variation in body surface temperature from birth, while diurnal variation in the small litter kits was absent (P > 0.05) in the first three days after birth. It was concluded that pregnant rabbits and large litter kits may have poor thermoregulation during West African winter, and thus, the need for special protective housing to improve productivity.

Brown adipose tissue involution associated with progressive restriction in progenitor competence.

Human brown adipose tissue (BAT) undergoes progressive involution. This involution process is not recapitulated in rodents, and the underlying mechanisms are poorly understood. Here we show that the interscapular BAT (iBAT) of rabbits whitens rapidly during early adulthood. The transcriptomic remodeling and identity switch of mature adipocytes are accompanied by loss of brown adipogenic competence of progenitors. Single-cell RNA sequencing reveals that rabbit and human iBAT progenitors highly express the FSTL1 gene. When iBAT involutes in rabbits, adipocyte progenitors reduce FSTL1 expression and are refractory to brown adipogenic recruitment. Conversely, FSTL1 is constitutively expressed in mouse iBAT to sustain WNT signaling and prevent involution. Progenitor incompetence and iBAT paucity can be induced in mice by genetic deletion of the Fstl1 gene or ablation of Fstl1+ progenitors. Our results highlight the hierarchy and dynamics of the BAT progenitor compartment and implicate the functional incompetence of FSTL1-expressing progenitors in BAT involution.

Biological constraints on configural odour mixture perception.

Animals, including humans, detect odours and use this information to behave efficiently in the environment. Frequently, odours consist of complex mixtures of odorants rather than single odorants, and mixtures are often perceived as configural wholes, i.e. as odour objects (e.g. food, partners). The biological rules governing this 'configural perception' (as opposed to the elemental perception of mixtures through their components) remain weakly understood. Here, we first review examples of configural mixture processing in diverse species involving species-specific biological signals. Then, we present the original hypothesis that at least certain mixtures can be processed configurally across species. Indeed, experiments conducted in human adults, newborn rabbits and, more recently, in rodents and honeybees show that these species process some mixtures in a remarkably similar fashion. Strikingly, a mixture AB (A, ethyl isobutyrate; B, ethyl maltol) induces configural processing in humans, who perceive a mixture odour quality (pineapple) distinct from the component qualities (A, strawberry; B, caramel). The same mixture is weakly configurally processed in rabbit neonates, which perceive a particular odour for the mixture in addition to the component odours. Mice and honeybees also perceive the AB mixture configurally, as they respond differently to the mixture compared with its components. Based on these results and others, including neurophysiological approaches, we propose that certain mixtures are convergently perceived across various species of vertebrates/invertebrates, possibly as a result of a similar anatomical organization of their olfactory systems and the common necessity to simplify the environment's chemical complexity in order to display adaptive behaviours.

Comparison of the Effects of Citicoline and Piracetam on Hypoxic-ischemic Brain Damage in Neonatal Rabbits.

Objectives Perinatal hypoxic-ischemic brain injuries have been a major cause of mortality and neurodevelopmental morbidities in newborns. Citicoline and Piracetam have been used as nootropic agents in a number of studies. In this investigation, we aimed to determine the effects of these agents solely and in combination in hypoxic-ischemic brain damage in rabbit neonates.Materials & Methods Hypoxic-ischemic brain damage was induced by the occlusion of both uterine arteries of dams for eight minutes. The subjects were randomly divided into five groups as follows (n=6 per group): control group without hypoxia (C1), control group with hypoxic-ischemic damage (C2), the third group (P) received Piracetam (100 mg/kg), the fourth group (T) administered with Citicoline (250 mg/kg), and the fifth (PT) received both. The preventive effects of the two drugs on hypoxic-ischemic brain damage were microscopically investigated by the rates of damage to the hippocampus.ResultsNeuronal destruction rates in C1, C2, P, T, and PT were 4%, 45%, 37.5%, 12.5% (P=0.01 vs. C2), and 20% (P=0.03 vs. C2), respectively. The total means of hypoxic-ischemic damage, cell edema, neuronal degeneration, and eosinophilic degeneration were lower in the T group compared to C2 (P<0.05). Conclusion According to our results and previous findings, Citicoline as a treatment for hypoxic-ischemic brain injuries could be beneficial, and it has priority over neuroprotective agents like Piracetam. Moreover, the combination of Citicoline and Piracetam showed no superior effect in contrast with Citicoline alone. However, experimental studies on larger populations and clinical trials are highly suggested.

A new anesthesia protocol enabling longitudinal lung-function measurements in neonatal rabbits by micro-CT.

Micro-computed tomography (micro-CT) imaging is an emerging technology with many applications in small animals, for example, the study of pulmonary diseases, although clear guidelines and critical mass of evidence are still missing in the preclinical literature. The neonatal rabbit is a valuable model for studying pulmonary development. However, the longitudinal monitoring of lung function by micro-CT can be challenging. Distinctive datasets corresponding to the end-inspiration and end-expiration phases need to be generated and analyzed to derive lung-functional parameters. The quality of CT scans and the reliability of parameters obtained remain highly dependent on the anesthesia protocol used. Three different anesthetic protocols were tested. The combination of dexmedetomidine 0.25 mg/kg injected intraperitoneally followed by 1% isoflurane was found to facilitate CT imaging at 4 and 11 days after birth. Contrarily, isoflurane and ketamine-xylazine were found unsuitable and thus not investigated further. Total lung volumes significantly increased at day 11 compared with baseline in both respiratory phases, whereas lung tissue remained constant. As expected, functional residual capacity, air-to-tissue ratio, and minute ventilation were significantly increased at day 11 in each animal. Those parameters were correlated with inspiratory capacity, compliance, elastance, and resistance of both respiratory system and tissue component, as measured by flexiVent. Lung development was also evaluated by histomorphometric analyses. In conclusion, we have identified a safe and suitable anesthesia protocol for micro-CT imaging in neonatal rabbits. Moreover, the possibility to longitudinally measure lung function in the same subject dramatically reduced the intraexperimental variability.

Characterization of microglial phagocytosis and dendrimer nanoparticle uptake in a neonatal rabbit model of cerebral palsy

&lt;img src=” https://s3.amazonaws.com/production.scholastica/article/29729/large/prnano_732021ga.jpg?1635418524”&gt; Hydroxyl dendrimers target reactive microglia in multiple neuroinflammatory models, including cerebral palsy. Insights on the differential uptake of dendrimers between subpopulations of micro-glia will enable the better design of precise nanomedicines to treat neuroinflammation. We have pre-viously demonstrated that microglia of rabbits with cerebral palsy undergo change to a pro-inflammatory phenotype that selectively takes up dendrimer nanoparticles proportional to the sever-ity of the injury. However, the functional changes in microglia associated with this activated pheno-type and related mechanisms of dendrimer uptake are not well understood. Here, we established a method for isolating microglia from cerebral palsy and healthy neonatal rabbit brains and assessed microglial (1) phagocytic activity ex vivo using fluorescent E. coli bioparticles and (2) dendrimer uptake in vivo after intravenous administration of dendrimer conjugated with a fluorescent dye (D-Cy5). Flow cytometry studies showed that the surface marker CD11b is reliably expressed and could isolate microglia from rabbit brain tissue. Furthermore, a significantly higher proportion of microglia isolated from cerebral palsy rabbits demonstrated increased phagocytosis when compared to controls. To assess differences in dendrimer uptake, microglia were isolated from the brain 24 hours after D-Cy5 administration. D-Cy5 localized only in CD11b+ microglia with differential up-take in subpopulations of microglia and was significantly higher in microglia from cerebral palsy rabbits when compared to healthy controls. This study demonstrates that PAMAM hydroxyl den-drimers are preferentially taken up by microglia, especially in cerebral palsy rabbits, and that sub-populations of microglia demonstrate differential dendrimer uptake. Future work will continue to evaluate these subpopulations of microglia further to facilitate the design of precise nanotherapeu-tics for targeting specific profiles of microglial activation.

Fetal surgery has no additional effect to general anesthesia on brain development in neonatal rabbits

Fetal surgery is part of modern fetal medicine, and some procedures, such as fetal spina bifida repair, are performed under general anesthesia. Fetuses are operated on in a time window when the developing brain is extremely vulnerable to external, potentially harmful factors. To date, little is known about the effect of fetal surgery on fetal brain development. This study aimed to assess the effect of fetal surgery on the developing fetal brain in the rabbit model. This was a randomized, sham-controlled study in time-mated pregnant does at 28 days' gestation (term, 31 days), which corresponds to the start of the peak of brain development and end of the second trimester of pregnancy in humans. We included 4 different groups in this experiment: no-surgery, general anesthesia, general anesthesia+hysterotomy, and general anesthesia+fetal surgery. In 11 does, anesthesia was induced using propofol and maintained for 75 minutes with 3.6 vol% (4% is the equivalent of 1 minimum alveolar concentration) sevoflurane. Maternal blood pressure, heart rate, oxygen saturation, temperature, end-tidal CO2 were continuously monitored. For each operated doe, 6 fetuses were part of the experiment. Randomization determined which cornual sac and what opposing third sac were assigned to fetal surgery: hysterotomy, fetal injection (atropine, fentanyl, and cisatracurium), fetal skin incision, and suturing. Only hysterotomy was performed on the opposing cornual and third amniotic sacs of the does. The fetus in these experimental sacs was used as internal unmanipulated control (general anesthesia). All fetuses (n=38) from unmanipulated does (n=4) served as external controls (no-surgery). At term, the does were delivered by cesarean delivery under ketamine-medetomidine sedation and local anesthesia. The pups underwent standardized motoric and sensory neurologic testing on day 1 followed by euthanasia and brain harvesting for histologic assessment of neurons, synapses, proliferation, and glial cells. Maternal vital signs were stable during surgery. Survival was similar in the 4 groups (75%-94%), and brain-to-body weight ratio was comparable; only the no-surgery pups had a higher brain weight. On postnatal day 1, the pups in the 4 groups had a comparable neurobehavioral outcome on both motoric and sensory testing. In the prefrontal cortex, no-surgery pups had significantly higher neuron density than pups who underwent maternal surgery, but there was no difference among pups that underwent general anesthesia, hysterotomy, or fetal surgery. The measurements of proliferation had a similar outcome: a higher proliferation rate in the prefrontal cortex of no-surgery pups. Moreover, synaptic density values were higher in the no-surgery pups, but there was no difference observed among pups who underwent general anesthesia, hysterotomy, and fetal surgery. Lastly, there was no difference in gliosis among the 4 groups. In rabbits, fetal surgery through hysterotomy under maternal general anesthesia did not affect brain development, in addition to the effects of general anesthesia per se.

Prebiotics Inulin Metabolism by Lactic Acid Bacteria From Young Rabbits.

Inulin as a commercial prebiotic could selectively promote the growth of beneficial gut microbes such as lactic acid bacteria (LAB). Whether LAB in rabbit gut possesses the capability to metabolize and utilize inulin is little known. Therefore, this study recovered 94 LAB strains from neonate rabbits and found that only 29% (28/94) could metabolize inulin with both species- and strain-specificity. The most vigorous inulin-degrading strain, Lacticaseibacillus paracasei YT170, could efficiently utilize both short-chain and long-chain components through thin-layer chromatography analysis. From genomic analysis, a predicted fosRABCDXE operon encoding putative cell wall-anchored fructan β-fructosidase, five fructose-transporting proteins and a pts1BCA operon encoding putative β-fructofuranosidase and sucrose-specific IIBCA components were linked to long-chain and short-chain inulin utilization respectively. This study provides a mechanistic rationale for effect of inulin administration on rabbits and lays a foundation for synbiotic applications aimed at modulating the intestinal microbiota of young rabbits.

A neonatal leporine model of age-dependent natural heart regeneration after myocardial infarction

ObjectivesNeonatal rodents and piglets naturally regenerate the injured heart after myocardial infarction. We hypothesized that neonatal rabbits also exhibit natural heart regeneration after myocardial infarction. MethodsNew Zealand white rabbit kits underwent sham surgery or left coronary ligation on postnatal day 1 (n = 94), postnatal day 4 (n = 11), or postnatal day 7 (n = 52). Hearts were explanted 1 day postsurgery to confirm ischemic injury, at 1 week postsurgery to assess cardiomyocyte proliferation, and at 3 weeks postsurgery to assess left ventricular ejection fraction and scar size. Data are presented as mean ± standard deviation. ResultsSize of ischemic injury as a percentage of left ventricular area was similar after myocardial infarction on postnatal day 1 versus on postnatal day 7 (42.3% ± 5.4% vs 42.3% ± 4.7%, P = .9984). Echocardiography confirmed severely reduced ejection fraction at 1 day after postnatal day 1 myocardial infarction (33.7% ± 5.3% vs 65.2% ± 5.5% for postnatal day 1 sham, P = .0001), but no difference at 3 weeks after postnatal day 1 myocardial infarction (56.0% ± 4.0% vs 58.0% ± 3.3% for postnatal day 1 sham, P = .2198). Ejection fraction failed to recover after postnatal day 4 myocardial infarction (49.2% ± 1.8% vs 58.5% ± 5.8% for postnatal day 4 sham, P = .0109) and postnatal day 7 myocardial infarction (39.0% ± 7.8% vs 60.2% ± 5.0% for postnatal day 7 sham, P < .0001). At 3 weeks after infarction, fibrotic scar represented 5.3% ± 1.9%, 14.3% ± 4.9%, and 25.4% ± 13.3% of the left ventricle area in the postnatal day 1, postnatal day 4, and postnatal day 7 groups, respectively. An increased proportion of peri-infarct cardiomyocytes expressed Ki67 (15.9% ± 1.8% vs 10.2% ± 0.8%, P = .0039) and aurora B kinase (4.0% ± 0.9% vs 1.5% ± 0.6%, P = .0088) after postnatal day 1 myocardial infarction compared with sham, but no increase was observed after postnatal day 7 myocardial infarction. ConclusionsA neonatal leporine myocardial infarction model reveals that newborn rabbits are capable of age-dependent natural heart regeneration.

The effect of xenon on fetal neurodevelopment following maternal sevoflurane anesthesia and laparotomy in rabbits

BackgroundThere is concern that maternal anesthesia during pregnancy impairs brain development of the human fetus. Xenon is neuroprotective in pre-clinical models of anesthesia-induced neurotoxicity in neonates. It is not known if xenon also protects the developing fetal brain when administered in addition to maternal sevoflurane-anesthesia during pregnancy. ObjectiveTo investigate the effects of sevoflurane and xenon on neurobehaviour and neurodevelopment of the offspring in a pregnant rabbit model. MethodsPregnant rabbits on post-conception day 28 (term = 31d) underwent two hours of general anesthesia with 1 minimum alveolar concentration (MAC) of sevoflurane in 30% oxygen (n = 17) or 1 MAC sevoflurane plus 50-60 % xenon in 30% oxygen (n = 10) during a standardized laparotomy while receiving physiological monitoring. A sham-group (n = 11) underwent monitoring alone for two hours. At term, the rabbits were delivered by caesarean section. On the first postnatal day, neonatal rabbits underwent neurobehavioral assessment using a validated test battery. Following euthanasia, the brains were harvested for neurohistological analysis. A mixed effects-model was used for statistical analysis. ResultsMaternal cardiopulmonary parameters during anesthesia were within the reference range. Fetal survival rates were significantly higher in the sham-group as compared to the sevoflurane-group and the fetal brain/body weight ratio was significantly lower in the sevoflurane-group as compared with the sham- and xenon-group. Pups antenatally exposed to anesthesia had significantly lower motor and sensory neurobehavioral scores when compared to the sham-group (mean ± SD; sevo: 22.70 ± 3.50 vs. sevo+xenon: 22.74 ± 3.15 vs. sham: 24.37 ± 1.59; overall p = 0.003; sevo: 14.98 ± 3.00 vs. sevo+xenon: 14.80 ± 2.83 vs. sham: 16.43 ± 2.63; overall p = 0.006; respectively). Neuron density, neuronal proliferation and synaptic density were reduced in multiple brain regions of the exposed neonates. The co-administration of xenon had no measurable neuroprotective effects in this model. ConclusionsIn rabbits, sevoflurane anesthesia for a standardized laparotomy during pregnancy resulted in impaired neonatal neurobehavior and a decreased neuron count in several regions of the neonatal rabbit brain. Co-administration of xenon did not prevent this effect.

Food Entrainment, Arousal, and Motivation in the Neonatal Rabbit Pup.

In the newborn rabbit, the light entrainable circadian system is immature and once a day nursing provides the primary timing cue for entrainment. In advance of the mother’s arrival, pups display food anticipatory activity (FAA), and metabolic and physiological parameters are synchronized to this daily event. Central structures in the brain are also entrained as indicated by expression of Fos and Per1 proteins, GFAP, a glial marker, and cytochrome oxidase activity. Under fasting conditions, several of these rhythmic parameters persist in the periphery and brain, including rhythms in the olfactory bulb (OB). Here we provide an overview of these physiological and neurobiological changes and focus on three issues, just beginning to be examined in the rabbit. First, we review evidence supporting roles for the organum vasculosum of lamina terminalis (OVLT) and median preoptic nucleus (MnPO) in homeostasis of fluid ingestion and the neural basis of arousal, the latter which also includes the role of the orexigenic system. Second, since FAA in association with the daily visit of the mother is an example of conditioned learning, we review evidence for changes in the corticolimbic system and identified nuclei in the amygdala and extended amygdala as part of the neural substrate responsible for FAA. Third, we review recent evidence supporting the role of oxytocinergic cells of the paraventricular hypothalamic nucleus (PVN) as a link to the autonomic system that underlies physiological events, which occur in preparation for the upcoming next daily meal. We conclude that the rabbit model has contributed to an overall understanding of food entrainment.

Mechanistic Insights Into the Reduced Pacemaking Rate of the Rabbit Sinoatrial Node During Postnatal Development: A Simulation Study.

Marked age- and development- related differences have been observed in morphology and characteristics of action potentials (AP) of neonatal and adult sinoatrial node (SAN) cells. These may be attributable to a different set of ion channel interactions between the different ages. However, the underlying mechanism(s) have yet to be elucidated. The objective of this study was to determine the mechanisms underlying different spontaneous APs and heart rate between neonatal and adult SAN cells of the rabbit heart by biophysical modeling approaches. A mathematical model of neonatal rabbit SAN cells was developed by modifying the current densities and/or kinetics of ion channels and transporters in an adult cell model based on available experimental data obtained from neonatal SAN cells. The single cell models were then incorporated into a multi-cellular, two-dimensional model of the intact SAN-atrium to investigate the functional impact of altered ion channels during maturation on pacemaking electrical activities and their conduction at the tissue level. Effects of the neurotransmitter acetylcholine on the pacemaking activities in neonatal cells were also investigated and compared to those in the adult. Our results showed: (1) the differences in ion channel properties between neonatal and adult SAN cells are able to account for differences in their APs and the heart rate, providing mechanistic insight into understanding the reduced pacemaking rate of the rabbit sinoatrial node during postnatal development; (2) in the 2D model of the intact SAN-atria, it was shown that cellular changes during postnatal development impaired pacemaking activity through increasing the activation time and reducing the conduction velocity across the SAN; (3) the neonatal SAN model, with its faster beating rates, showed a greater sensitivity to parasympathetic modulation in response to acetylcholine than did the adult model. These results provide novel insights into the understanding of the cellular mechanisms underlying the differences in the cardiac pacemaking activities of the neonatal and adult SAN.

Glomerular developmental delay and proteinuria in the preterm neonatal rabbit.

Recent advances in neonatal care have improved the survival rate of those born premature. But prenatal conditions, premature birth and clinical interventions can lead to transient and permanent problems in these fragile patients. Premature birth (<36 gestational weeks) occurs during critical renal development and maturation. Some consequences have been observed but the exact pathophysiology is still not entirely known. This experimental animal study aims to investigate the effect of premature birth on postnatal nephrogenesis in premature neonatal rabbits compared to term rabbits of the same corrected age. We analyzed renal morphology, glomerular maturity and functional parameters (proteinuria and protein/creatinine ratio) in three cohorts of rabbit pups: preterm (G28), preterm at day 7 of life (G28+7) and term at day 4 of life (G31+4). We found no significant differences in kidney volume and weight, and relative kidney volume between the cohorts. Nephrogenic zone width increased significantly over time when comparing G31 + 4 to G28. The renal corpuscle surface area, in the inner cortex and outer cortex, tended to decrease significantly after birth in both preterm and term groups. With regard to glomerular maturity, we found that the kidneys in the preterm cohorts were still in an immature state (presence of vesicles and capillary loop stage). Importantly, significant differences in proteinuria and protein/creatinine ratio were found. G28 + 7 showed increased proteinuria (p = 0.019) and an increased protein/creatinine ratio (p = 0.023) in comparison to G31 +4. In conclusion, these results suggest that the preterm rabbit kidney tends to linger in the immature glomerular stages and shows signs of a reduced renal functionality compared to the kidney born at term, which could in time lead to short- and long-term health consequences.

Systemic dendrimer-drug nanomedicines for long-term treatment of mild-moderate cerebral palsy in a rabbit model

BackgroundNeuroinflammation mediated by microglia plays a central role in the pathogenesis of perinatal/neonatal brain injury, including cerebral palsy (CP). Therapeutics mitigating neuroinflammation potentially provide an effective strategy to slow the disease progression and rescue normal brain development. Building on our prior results which showed that a generation-4 hydroxyl poly(amidoamine) (PAMAM) dendrimer could deliver drugs specifically to activated glia from systemic circulation, we evaluated the sustained efficacy of a generation-6 (G6) hydroxyl-terminated PAMAM dendrimer that showed a longer blood circulation time and increased brain accumulation. N-acetyl-l-cysteine (NAC), an antioxidant and anti-inflammatory agent that has high plasma protein binding properties and poor brain penetration, was conjugated to G6-PAMAM dendrimer-NAC (G6D-NAC). The efficacy of microglia-targeted G6D-NAC conjugate was evaluated in a clinically relevant rabbit model of CP, with a mild/moderate CP phenotype to provide a longer survival of untreated CP kits, enabling the assessment of sustained efficacy over 15 days of life.MethodsG6D-NAC was conjugated and characterized. Cytotoxicity and anti-inflammatory assays were performed in BV-2 microglial cells. The efficacy of G6D-NAC was evaluated in a rabbit model of CP. CP kits were randomly divided into 5 groups on postnatal day 1 (PND1) and received an intravenous injection of a single dose of PBS, or G6D-NAC (2 or 5 mg/kg), or NAC (2 or 5 mg/kg). Neurobehavioral tests, microglia morphology, and neuroinflammation were evaluated at postnatal day 5 (PND5) and day 15 (PND15).ResultsA single dose of systemic ‘long circulating’ G6D-NAC showed a significant penetration across the impaired blood-brain-barrier (BBB), delivered NAC specifically to activated microglia, and significantly reduced microglia-mediated neuroinflammation in both the cortex and cerebellum white matter areas. Moreover, G6D-NAC treatment significantly improved neonatal rabbit survival rate and rescued motor function to nearly healthy control levels at least up to 15 days after birth (PND15), while CP kits treated with free NAC died before PND9.ConclusionsTargeted delivery of therapeutics to activated microglia in neonatal brain injury can ameliorate pro-inflammatory microglial responses to injury, promote survival rate, and improve neurological outcomes that can be sustained for a long period. Appropriate manipulation of activated microglia enabled by G6D-NAC can impact the injury significantly beyond inflammation.