Abstract HRV complexity metrics track the systemic cytokine response across mammalian inflammation models, but the dynamical mechanism translating molecular inflammation into HRV structure has remained unresolved, and no existing tool jointly extracts coordination-dynamics signatures from simultaneous vagus nerve electroneurogram (VENG) and ECG. We describe a modular open-source Python pipeline that (i) detects spectrally distinct latent states in 20 kHz whole-trunk VENG using a symmetric-Kullback–Leibler scan-statistic changepoint detector ported from earthquake seismology and verified against its R progenitor; (ii) places the inter-changepoint-interval series in the Haken–Kelso–Bunz coupled-oscillator framework, computes the Kuramoto order parameter R , fits the bistable potential V ( φ ) = − a cos φ − b cos 2 φ , and maps each timepoint into the Arnold-tongue (Ω, K ) plane; contrasts compound-action-potential band power between successive latent states; and (iii) couples the VENG analysis to a CIMVA HRV pipeline driven by a 5-algorithm Pro-MAC R-peak ensemble. Applied to a previously published two-animal neonatal piglet endotoxaemia cohort (Castel et al., 2020, 2024), the pipeline recovers known features of the (iv) preparation and surfaces three case-series observations that we frame as hypotheses for prospective replication: (1) a 6–8 s VENG state-switching period coincident with the HRV LF/HF spectral boundary; (2) qualitatively divergent coordination-dynamics trajectories under LPS versus LPS + VNS (Kuramoto R collapses from 0.69 to 0.12 without VNS, undergoes biphasic recovery to 0.99 with VNS, the two trajectories crossing at 60–75 min); (3) a regime-dependent empirical coupling between R and the HRV embedding scaling exponent eScalE ( R · eScalE ≈ 0.33 during moderate challenge, doubling to ≈0.80 during VNS overshoot). Pre-specified null comparisons (Poisson and bootstrap surrogates) demonstrate that R and the critical-slowing-down indicators carry information beyond their most obvious null alternatives. With N = 2 animals no inferential statistics are possible, so these findings are reported as hypothesis-generating; the manuscript closes with a pre-registered prospective protocol designed specifically to falsify them. Graphical Abstract Key Points We present an open-source Python pipeline that extracts coordination-dynamics invariants (Kuramoto phase coherence, Haken–Kelso–Bunz potential parameters, Arnold-tongue position, critical-slowing-down indicators) from simultaneously recorded vagus nerve electroneurogram (VENG) and ECG, starting from a spectral-KL scan-statistic changepoint detector ported from earthquake seismology. Applied to a previously published two-animal cohort (Castel et al., 2024), the pipeline reveals that VENG alternates between two spectrally distinct latent states at a baseline period of 6–8 s that coincides with the HRV low-frequency/high-frequency spectral boundary (∼0.15 Hz). We propose the VENG latent-state oscillator as a candidate mechanistic generator of vagal LF HRV; this is a hypothesis motivated by the timescale match and is not established by the present data. In this cohort, LPS endotoxaemia and vagus nerve stimulation drove qualitatively divergent trajectories in the coordination-dynamics phase space (Kuramoto R : 0.69 → 0.12 without VNS; biphasic to R → 0.99 with VNS at the IL-6/IL-8 cytokine peak). Because the design provides one animal per arm, these are case-series observations and not a between-arm statistical comparison. Inter-changepoint-interval variance rose >100-fold roughly 15 min before overt state-switching collapse in the unmitigated animal. Bootstrap nulls drawn from the same animal’s pre-collapse ICI pool reject the same-distribution alternative (§2.6), motivating a critical-slowing-down early-warning hypothesis that requires prospective replication before any clinical use is warranted. The pipeline surfaces a regime-dependent empirical coupling between VENG phase coherence ( R ) and HRV embedding scaling exponent (eScalE) in the VNS-treated animal: during moderate challenge R · eScalE ≈ 0.33 (CV ≈ 0.20), during VNS-enhanced overshoot (45, 90, 105 min) the product doubles to ≈0.80. The product is a post-hoc empirical construct, not a derived conservation law; we propose it as a falsification target for prospective cohorts (§6).

BACKGROUND. Glucosensing elements are widely distributed throughout the body and relay information about circulating glucose levels to the brain via the vagus nerve. However, while anatomical wiring has been established, little is known about the physiological role of the vagus nerve in glucosensing. The contribution of the vagus nerve to inflammation in the fetus is poorly understood. Increased glucose levels and inflammation act synergistically when causing organ injury, but their interplay remains incompletely understood. We hypothesized that vagotomy (Vx) will trigger a rise in systemic glucose levels and this will be enhanced during systemic and organ-specific inflammation. Efferent vagus nerve stimulation (VNS) should reverse this phenotype. METHODS. Near-term fetal sheep (n = 57) were surgically prepared using vascular catheters and ECG electrodes as the control and treatment groups (lipopolysaccharide (LPS), Vx + LPS, Vx + LPS + selective efferent VNS). The experiment was started 72 h postoperatively to allow for post-surgical recovery. Inflammation was induced with LPS bolus intravenously (LPS group, 400 ng/fetus/day for 2 days; n = 23). For the Vx + LPS group (n = 11), a bilateral cervical vagotomy was performed during surgery; of these n = 5 received double the LPS dose, LPS800. The Vx + LPS + efferent VNS group (n = 8) received cervical VNS probes bilaterally distal from Vx in eight animals. Efferent VNS was administered for 20 min on days 1 and 2 +/10 min around the LPS bolus. Fetal arterial blood samples were drawn on each postoperative day of recovery (-72 h, -48 h, and -24 h) as well as at the baseline and seven selected time points (3–54 h) to profile inflammation (ELISA IL-6, pg/mL), insulin (ELISA), blood gas, and metabolism (glucose). At 54 h post-LPS, a necropsy was performed, and the terminal ileum macrophages’ CD11c (M1 phenotype) immunofluorescence was quantified to detect inflammation. The results are reported for p < 0.05 and for Spearman R2 > 0.1. The results are presented as the median (IQR). RESULTS. Across the treatment groups, blood gas and cardiovascular changes indicated mild septicemia. At 3 h in the LPS group, IL-6 peaked. That peak was decreased in the Vx + LPS400 group and doubled in the Vx + LPS800 group. The efferent VNS sped up the reduction in the inflammatory response profile over 54 h. The M1 macrophage activity was increased in the LPS and Vx + LPS800 groups only. The glucose and insulin concentrations in the Vx + LPS group were, respectively, 1.3-fold (throughout the experiment) and 2.3-fold higher vs. control (at 3 h). The efferent VNS normalized the glucose concentrations. CONCLUSIONS. The complete withdrawal of vagal innervation resulted in a 72-h delayed onset of a sustained increase in glucose for at least 54 h and intermittent hyperinsulinemia. Under the conditions of moderate fetal inflammation, this was related to higher levels of gut inflammation. The efferent VNS reduced the systemic inflammatory response as well as restored both the concentrations of glucose and the degree of terminal ileum inflammation, but not the insulin concentrations. Supporting our hypothesis, these findings revealed a novel regulatory, hormetic, role of the vagus nerve in the immunometabolic response to endotoxin in near-term fetuses.

Background: Vagus nerve stimulation (VNS) reduced inflammation induced by lipopolysaccharide (LPS) in an adult rat sepsis model. A multi-dimensional heart rate variability (HRV) index reliably tracked the inflammatory profile in near-term sheep fetuses. The effects of VNS on neonates are not known. First, in a neonatal piglet model of sepsis, we present an approach to evaluate the effect of VNS on systemic inflammatory response induced by a high dose of LPS to mimic late-onset neonatal sepsis. Second, we present an analytical pipeline to validate our fetal-sheep-derived HRV inflammatory index in neonatal piglets to test its performance in different species, older developmental stages, and more robust septic responses. Methods: Three neonatal piglets of 7–14 days of age with 2.4–4 kg in body weight were used in this proof-of-principle study. Following anesthesia, electrodes were attached bilaterally to the cervical portion of the vagus nerve to allow for stimulation of the left vagus (VNS) and bilateral vagus electroneurogram (VENG). Electrocardiogram (ECG), blood pressure (BP), and VENG were recorded for the duration of the experiment. After baseline recording, the piglets were administered LPS at 2 mg/kg IV bolus. In the VNS-treated piglet, the vagus nerve was stimulated for 10 min prior to and 10 min after the injection of LPS. In both groups, each 15 min post LPS, an arterial blood sample was drawn for blood gas, lactate, and glucose as well as the inflammatory cytokines measured by a quantitative ELISA multiplex panel. At the end of the experiment, the piglets were euthanized. BP and ECG-derived HRV were calculated and VENG was analyzed. Results: The piglets developed a potent inflammatory response to the LPS injection with TNF-α, IL-1β, IL-6 and IL-8 peaking between 45 and 90-min post-injection. VNS diminished the LPS-induced systemic inflammatory response, with a decrease in measured cytokines levels ranging from two to ten-fold. We present a low-cost, easy-to-implement design of a VNS/VENG probe and a framework to analyze VENG in response to LPS. Furthermore, the HRV index accurately tracked cytokine temporal profiles’ which was reflected in the power-spectral and complex properties of VENG when applying the machine learning model derived from HRV. Discussion: We present a novel method to model, manipulate, and track neonatal sepsis using VNS/VENG. Our supportive findings suggest that (1) the HRV index of the systemic inflammatory response applies across species pre- and postnatally, (2) the HRV index performs well at different degrees of sepsis (i.e., nanogram and milligram doses of LPS), and (3) the present VNS paradigm effectively suppresses LPS-induced inflammation, even with high doses of LPS; an effect that is reflected by changes in the shared mathematical properties of both VENG and HRV. These findings suggest that the HRV inflammatory index reflects underlying changes in the VENG activity. The presented method lays a foundation for larger studies to investigate the mechanisms and therapeutic potential of early postnatal VNS intervention to counteract sepsis progression. Moreover, the presented experimental and analytical frameworks highlight the potential for HRV monitoring to serve as an early biomarker for tracking the systemic inflammatory response.

The efferent and afferent effects of the vagus nerve on the developing brain have remained enigmatic. Here we review the evidence of such effects on microglial plasticity in the sheep model of human fetal development, one of the most recognized and deployed models of human fetal physiology. We show that vagotomy alters microglial phenotype and that this effect is hormetic under conditions of mild systemic inflammation, as may occur antepartum with chorioamnionitis. We present the methodology to assess not only biomarker-based microglial activation but also the morphometric features of the microglia. Together, these assessments provide a more comprehensive toolbox of glial phenotypical characterizations, especially in the context of investigating the locoregional vagal control of glial function. The presented findings support the earlier discoveries in preclinical and clinical models of adult physiology whereby vagotomy appeared neuroprotective for Parkinson disease, explained, at least in part, by the effects on microglia. In addition, we present the approach to measure and the findings on regional cerebral blood flow changes in relation to vagus nerve manipulation. In summary, the body of evidence underscores the importance of both the efferent and the afferent vagal pathways, via the vagus nerve, in the programming of microglial phenotype in the developing brain. The significance of these relationships for developing and treating early susceptibility to neuroinflammatory and neurodegenerative disorders in later life requires further studies.

Background The chronically instrumented non-anesthetized fetal sheep (CINAFS) model has been a mainstay of human fetal development research for the past 60 years. As a large "two for one" animal model, involving the instrumentation of the ewe and her fetus, the model poses challenges to implement de novo and maintain overtime at the highest standards of operating procedures to ensure ongoing performance. A common yet conventionally underreported issue researchers face is a high rate of animal loss. Here, we investigate what determines the success of the CINAFS model of human development. Methods We conducted a retrospective cohort analysis consisting of 82 experiments spanning the course of six years. Our team identified 10 variables that we anticipated were likely to influence the experimental outcome, such as the time of year, animal size, and surgical complexity. To evaluate the role of each variable in contributing to the success of the model, a binary logit regression analysis with a Fisher scoring optimization was fit to the data (SAS, V9 engine, release 3.8, SAS Institute, Cary, NC, USA). A higher predictive probability indicates a larger impact by the given variable on the outcome of the experiment. A Wald chi-squared analysis was run on the data to control for confounders and determine significance. Results The single variable identified in this study as determining the success of experiment outcomes using the CINAFS model is the experience level of the team. Conclusion . The CINAFS model offers enormous potential to further our understanding of human fetal development and create interventional technologies related to fetal health. However, to improve experimental outcomes using the CINAFS model, stronger communication and training are needed. We discuss the implications of our findings for the successful implementation of this challenging yet scientifically advantageous animal model of human physiology.

The contribution of the vagus nerve to inflammation and glucosensing in the fetus is not understood. We hypothesized that vagotomy (Vx) will trigger a rise in systemic glucose levels and this will be enhanced during systemic and organ-specific inflammation. Efferent vagus nerve stimulation (VNS) should reverse this phenotype. Near-term fetal sheep (n=57) were surgically prepared with vascular catheters and ECG electrodes as control and treatment groups (lipopolysaccharide (LPS), Vx+LPS, Vx+LPS+selective efferent VNS). Fetal arterial blood samples were drawn for 7 days to profile inflammation (IL-6), insulin, blood gas and metabolism (glucose). At 54 h, a necropsy was performed; terminal ileum macrophages; CD11c (M1 phenotype) immunofluorescence was quantified to detect inflammation. Across the treatment groups, blood gas and cardiovascular changes indicated mild septicemia. At 3 h, in the LPS group IL-6 peaked; that peak was decreased in Vx+LPS400 and doubled in Vx+LPS800 group; the efferent VNS sped up the reduction of the inflammatory response profile over 54 h. M1 macrophage activity was increased in the LPS and Vx+LPS800 groups only. Glucose and insulin levels in the Vx+LPS group were respectively 1.3-fold and 2.3-fold higher vs. control at 3 h, and the efferent VNS normalized glucose levels. Complete withdrawal of vagal innervation results in a 72h delayed onset of sustained increase in glucose levels for at least 54h and intermittent hyperinsulinemia. Under conditions of moderate fetal inflammation, this is related to higher levels of gut inflammation; the efferent VNS reduces the systemic inflammatory response as well as restores both the levels of glucose and terminal ileum inflammation, but not the insulin levels. Our findings reveal a novel regulatory, hormetic, role of the vagus nerve in the immunometabolic response to endotoxin in near-term fetuses.

Little is known about the vagus nerve activity in near-term fetuses. The chronically instrumented unanesthetized fetal sheep model is used to study human fetal physiology, because it permits chronic instrumentation with catheters and electrodes, which allow repetitive blood sampling, substance injection and recording of bioelectrical activity. We describe the procedures required to manipulate the vagus nerve activity in this model. We refer the reader to the related published report in JoVE (doi: 10.3791/52581), available in full text via PubMed Canada and as video here.

An electrocardiogram (ECG)-derived heart rate variability (HRV) index reliably tracks the inflammatory response induced by low-dose lipopolysaccharide (LPS) in near-term sheep fetuses. We evaluated the effect of vagus nerve stimulation (VNS) on vagus nerve electroneurogram (VENG) and the systemic inflammatory response induced by a high dose of LPS in neonatal piglets to mimic late-onset neonatal sepsis. We tested if our HRV inflammatory index tracks inflammation in piglets and its relationship to VENG. Following anesthesia, electrodes were attached to the left vagal nerve; ECG and blood pressure (BP) were recorded throughout the experiment. Following baseline, the piglets were administered LPS as 2mg/kg IV bolus. In the VNS treated piglet, the vagus nerve was stimulated for 10 minutes prior to and 10 min after the injection of LPS. In both groups, every 15 min post LPS, the arterial blood sample was drawn for blood gas, metabolites, and inflammatory cytokines. At the end of the experiment, the piglets were euthanized. BP and HRV measures were calculated. The piglets developed a potent inflammatory response to the LPS injection with TNF-alpha, IL-1beta, IL-6 and IL-8 peaking between 45 and 90 min post-injection. VNS diminished the LPS-induced systemic inflammatory response varying across the measured cytokines from two to ten-fold. The HRV index tracked accurately the temporal profile of cytokines and VENG changes. This novel model allows manipulating and tracking neonatal sepsis: The HRV inflammatory index 1) applies across species pre- and postnatally and 2) performs well at different degrees of sepsis (i.e., nanogram and milligram doses of LPS); 3) the present VNS paradigm effectively suppresses LPS-induced inflammation, even at high doses of LPS. The potential of early postnatal VNS to counteract sepsis and of HRV monitoring to early detect and track it deserve further study.

BACKGROUND:The chronically instrumented pregnant sheep has been used as a model of human fetal development and responses to pathophysiologic stimuli. This is due to the unique amenability of the unanesthetized fetal sheep to the surgical placement and maintenance of catheters and electrodes, allowing repetitive blood sampling, substance injection, recording of bioelectrical activity, application of electric stimulation, and in vivo organ imaging. Recently, there has been growing interest in the pleiotropic effects of vagus nerve stimulation (VNS) on various organ systems such as innate immunity and inflammation, and metabolism. There is no approach to study this in utero and corresponding physiological understanding is scarce.NEW METHOD:Based on our previous presentation of a stable chronically instrumented unanesthetized fetal sheep model, here we describe the surgical instrumentation procedure allowing successful implantation of a cervical uni- or bilateral VNS probe with or without vagotomy.RESULTS:In a cohort of 68 animals, we present the changes in blood gas, metabolic, and inflammatory markers during the postoperative period. We detail the design of a VNS probe which also allows recording from the fetal nerve. We also present an example of fetal vagus electroneurogram (VENG) recorded from the VNS probe and an analytical approach to the data.COMPARISON WITH EXISTING METHODS:This method represents the first implementation of fetal VENG/VNS in a large pregnant mammalian organism.CONCLUSIONS:This study describes a new surgical procedure allowing to record and manipulate chronically fetal vagus nerve activity in an animal model of human pregnancy.

SummaryAccess to the respiratory tract of an anaesthetised animal is a vital line of life. An endotracheal tube ensures a secure airway that will allow the delivery of anaesthesia and facilitates mechanical ventilation. The case report by Miller and Auckburally (2020) described in this issue highlights the potential complications associated with endotracheal intubation. Intubation using a 30 mm ID endotracheal tube in the average sized horse (500 kg) has been documented to have a high rate of tracheal injury. The manufacturing specifications of endotracheal tubes may contribute to the incidence of tracheal injury. Further research is needed to help minimise the morbidity and potential mortality associated with this anaesthetic procedure.

This communication briefly describes the use of tricaine methanesulfonate (MS222) to induce chemical restraint/general anesthesia of a Mexican axolotl (Ambystoma mexicanum) for the endoscopic retrieval of a gastric foreign body. There is very little published scientific literature concerning the anesthesia of Mexican axolotls. The anesthesia used in this case was an immersion bath of tricaine methanesulfonate where the concentration of tricaine methanesulfonate was gradually increased to 500 mg/L (ppm) over a 15-min period. A loss of righting reflex was observed within 3 min of attaining the final concentration of the anesthetic bath. The first voluntary movements following the transfer to a freshwater bath occurred within 7 min. The recovery was uneventful. Tricaine methanesulfonate in this case proved to be an effective anesthetic agent for a short, minimally invasive procedure.

Fetal heart rate variability (fHRV) is an important indicator of health and disease, yet its physiological origins, neural contributions in particular, are not well understood. We aimed to develop novel experimental and data analytical approaches to identify fHRV measures reflecting the vagus nerve contributions to fHRV. In near-term ovine fetuses, a comprehensive set of 46 fHRV measures was computed from fetal pre-cordial electrocardiogram recorded during surgery and 72 hours later without (n=24) and with intra-surgical bilateral cervical vagotomy (n=15). The fetal heart rate did not change due to vagotomy. We identify fHRV measures specific to the vagal modulation of fHRV: Multiscale time irreversibility asymmetry index (AsymI), Detrended fluctuation analysis (DFA) alpha1, Kullback-Leibler permutation entropy (KLPE) and Scale dependent Lyapunov exponent slope (SDLE alpha). We provide a systematic delineation of vagal contributions to fHRV across signal-analytical domains which should be relevant for the emerging field of bioelectronic medicine and the deciphering of the vagus code. Our findings also have clinical significance for in utero monitoring of fetal health during surgery.

Neuroinflammation in utero may result in lifelong neurological disabilities. Astrocytes play a pivotal role, but the mechanisms are poorly understood. No early postnatal treatment strategies exist to enhance neuroprotective potential of astrocytes. We hypothesized that agonism on alpha7 nicotinic acetylcholine receptor (alpha7nAChR) in fetal astrocytes will augment their neuroprotective transcriptome profile, while the antagonistic stimulation of alpha7nAChR will achieve the opposite. Using an in vivo - in vitro model of developmental programming of neuroinflammation induced by lipopolysaccharide (LPS), we validated this hypothesis in primary fetal sheep astrocytes cultures re-exposed to LPS in presence of a selective alpha7nAChR agonist or antagonist. Our RNAseq findings show that a pro-inflammatory astrocyte transcriptome phenotype acquired in vitro by LPS stimulation is reversed with alpha7nAChR agonistic stimulation. Conversely, antagonistic alpha7nAChR stimulation potentiates the pro-inflammatory astrocytic transcriptome phenotype. Furthermore, we conduct a secondary transcriptome analysis against the identical alpha7nAChR experiments in fetal sheep primary microglia cultures and discuss the implications for neurodevelopment.

The non-neuronal, immunological effects of the cholinergic signaling are exerted on the system's scale of observation via the vagus nerve and on the cellular scale via α7 nicotinic acetylcholine receptor (nAChR) signaling in myeloid cells of the periphery or brain's microglia and astrocytes. The developmental effects of such multi-scale signaling can be conceived of as an example of psychoneuroimmunological (PNI) homeokinesis and, while reported in the literature, are not yet systematically well studied. To be better understood, the intricacy of the multi-scale interactions requires relevant preclinical animal models. Chronically instrumented non-anesthetized fetal sheep model comes with a strong track record of bench-to-bed translation and a large body of evidence for its strong resemblance to and relevance for human physiology on various scales of organization. Recently, there has been growing interest in pleiotropic effects of vagus nerve stimulation (VNS) on various organ systems such as innate immunity, metabolism, and emotion with implications for programming of PNI phenotype. Here we describe the procedures required to record and manipulate the vagus nerve activity in this large pregnant mammalian organism. Extending this in vivo model to in vitro, on the cellular scale, we present the method to manipulate the cholinergic signaling in ovine fetal microglia and astrocytes and analyze their responses on protein and RNA levels. Together these models can provide multi-scale-level mechanistic insights into the effects of cholinergic signaling on PNI phenotype.

Necrotic enteritis was studied in chickens using various in vivo infection models. Most of these use a combination of predisposing factors, such as coccidiosis and diet, with gavage or administration via the feed using Clostridium perfringens. In these models, the comparison of multiple C. perfringens strains for virulence studies requires a large number of hosts to obtain significant results. Mortality during the course of the study can be high depending on the experimental model, hence raising ethical concerns regarding animal welfare in research. The development of new infection models requiring fewer animals to study pathogenesis, yet providing statistically significant and valid results, is important in reducing animal use in research. Intestinal ligated loop models have been used to study clostridial infections in various species such as mice, rabbits and calves. Following surgical procedures to create ligated loop segments, C. perfringens strains are injected directly into the loops to establish a close contact between the bacteria and the intestinal mucosa. Samples of the small intestine and luminal contents are taken at the termination of the procedures after a few hours. Multiple bacterial strains can be inoculated in each animal, hence reducing the number of required subjects in the experiments. Also, procedures are performed under general anesthesia to reduce animal pain. In chickens, this model would be more appropriate than oral administration to compare C. perfringens strain pathogenicity because fewer animals are needed, no predisposing factors are required to induce the disease, and pain is controlled by analgesics. The intestinal ligated loop model is poorly described in chickens and standardization is essential for its optimal use. This manuscript provides all the necessary steps to create numerous intestinal ligated loops in chickens and brings information on the critical points to obtain valid results.

Walruses are a challenging species to anesthetize as a result of their large mass, limited access for drug delivery, unique physiology, and small number of reports describing anesthetic procedures. Three aquarium-housed walruses (Odobenus rosmarus) ranging in age from 3 to 11 yr old (344-1,000 kg) were anesthetized for dental or ophthalmic surgical procedures, with one animal anesthetized twice and one anesthetized three times. Preanesthetic medication was with intramuscular midazolam (0.1-0.2 mg/kg) and meperidine (2-3 mg/kg). A catheter was placed in the extradural intravertebral vein, and anesthesia was induced with propofol to effect. Orotracheal intubation was performed and anesthesia maintained with isoflurane in oxygen using a circle breathing system connected to a ventilator. Intermittent positive pressure ventilation was used in all procedures. For the ophthalmic surgery, the neuromuscular blocking agent, cisatracurium, was given intravenously to provide a central eye and optimal surgical conditions. The neuromuscular block was antagonized with edrophonium. Total anesthesia times ranged from 1.5 to 6 hr. Midazolam and meperidine were antagonized with flumazenil and naltrexone, respectively, in five of six cases. Nonsteroidal anti-inflammatory agents were provided for analgesia. Recoveries were calm and uneventful. The described anesthetic protocols and case management were successful under the conditions encountered.

Objective: The utility of fetal heart rate (FHR) monitoring can only be achieved with an acquisition sampling rate that preserves the underlying physiological information on the millisecond time scale (1000 Hz rather than 4 Hz). For such acquisition, fetal ECG (fECG) is required, rather than the ultrasound to derive FHR. We tested one recently developed algorithm, SAVER, and two widely applied algorithms to extract fECG from a single-channel maternal ECG signal recorded over the xyphoid process rather than the routine abdominal signal. Approach: At 126dG, ECG was attached to near-term ewe and fetal shoulders, manubrium and xyphoid processes (n = 12). fECG served as the ground-truth to which the fetal ECG signal extracted from the simultaneously-acquired maternal ECG was compared. All fetuses were in good health during surgery (pH 7.29 +/- 0.03, pO(2) 33.2 +/- 8.4, pCO(2) 56.0 +/- 7.8, O(2)Sat 78.3 +/- 7.6, lactate 2.8 +/- 0.6, BE -0.3 +/- 2.4). Main result: In all animals, single lead fECG extraction algorithm could not extract fECG from the maternal ECG signal over the xyphoid process with the F1 less than 50%. Significance: The applied fECG extraction algorithms might be unsuitable for the maternal ECG signal over the xyphoid process, or the latter does not contain strong enough fECG signal, although the lead is near the mother's abdomen. Fetal sheep model is widely used to mimic various fetal conditions, yet ECG recordings in a public data set form are not available to test the predictive ability of fECG and FHR. We are making this data set openly available to other researchers to foster non-invasive fECG acquisition in this animal model.

Neuroinflammation in utero may result in life-long neurological disabilities. Microglia play a pivotal role, but the mechanisms are poorly understood. No early postnatal treatment strategies exist to enhance neuroprotective potential of microglia. We hypothesized that agonism on α7 nicotinic acetylcholine receptor (α7nAChR) in fetal microglia will augment their neuroprotective transcriptome profile, while the antagonistic stimulation of α7nAChR will achieve the opposite. Using an in vivo - in vitro model of developmental programming of neuroinflammation induced by lipopolysaccharide (LPS), we validated this hypothesis in primary fetal sheep microglia cultures re-exposed to LPS in presence of a selective α7nAChR agonist or antagonist. Our RNAseq and protein level findings show that a pro-inflammatory microglial phenotype acquired in vitro by LPS stimulation is reversed with α7nAChR agonistic stimulation. Conversely, antagonistic α7nAChR stimulation potentiates the pro-inflammatory microglial phenotype. Surprisingly, under conditions of LPS double-hit an interference of a postulated α7nAChR - ferroportin signaling pathway may impede this mechanism. These results suggest a therapeutic potential of α7nAChR agonists in early re-programming of microglia in neonates exposed to in utero inflammation via an endogenous cerebral cholinergic anti-inflammatory pathway. Future studies will assess the role of interactions between inflammation-triggered microglial iron sequestering and α7nAChR signaling in neurodevelopment.

CASE DESCRIPTION A 4-hour-old 6.3-kg (13.9-lb) female alpaca cria was evaluated because of severe respiratory distress and difficulty nursing since birth. CLINICAL FINDINGS The cria had open-mouth breathing and cyanotic membranes, with no airflow evident from either nostril. Supplemental oxygen was delivered, and the patient was anesthetized and intubated orotracheally; a CT evaluation of the head confirmed bilateral membranous obstruction of the nasal cavities, consistent with complete bilateral choanal atresia. TREATMENT AND OUTCOME Choanal atresia was treated with an endoscopically assisted balloon-dilation technique, and temporary tracheostomy was performed. Stenosis recurred, requiring revision of the repair and intranasal stent placement 3 days after the first surgery. The tracheostomy tube was removed the next day. Complications during hospitalization included mucoid obstruction of the tracheostomy tube, granulation tissue development in the trachea near the tracheostomy site, mucoid stent obstruction, aspiration pneumonia, and presumed partial failure of passive transfer of immunity. The stents were removed 2 weeks after admission, and the cria was discharged 3 days later. The owner was advised that the animal should not be bred. At last follow-up 3 years later, the alpaca was doing well. CLINICAL RELEVANCE Surgical treatment with a balloon-dilation technique and placement of nasal stents with endoscopic guidance were curative in this neonatal alpaca with bilateral membranous choanal atresia. Computed tomography was useful to determine the nature of the atresia and aid surgical planning. Because a genetic component is likely, owners should be advised to prevent affected animals from breeding.

Department of Clinical Sciences Groupe de Recherche en Pharmacologic Animale du Quebec Faculte de Medecine Veterinaire Universite de Montreal, 3200 rue Sicotte