Peripheral Administration Research Articles

P36 Numeta G13% preterm neonatal parenteral nutrition solution – a licensed all-in-one triple chamber, ready to use and terminally sterilised parenteral nutrition for preterm newborn infants

AimsWe aim to evaluate the efficacy and safety of Numeta G13%E preterm neonatal parenteral nutrition (PN) in our neonatal population.In September 2017 a National Patient Safety Alert (NPSA) highlighted the risk of harm to babies when lipid was mistakenly run at the rate intended for the aqueous component resulting in significant lipid overdose.Although we have worked to implement many of the alert’s recommendations, we feel we can avoid this risk further by using an all-in-one PN solution.1Numeta meets current nutritional guidelines as per British Association of Perinatal Medicine (BAPM) but this project allows comparison of outcomes important to both patient and service between those achieved with our current regimen and those with the all-in-one regimen.2MethodsWe carried out a quality improvement project from April 2018 to April 2019. We collected data from 330 babies in our neonatal unit during six months before (154 babies) and after (176 babies) the adoption of the all-in-one solution.Our previous PN regimen consisted of a ‘menu’ of aqueous bags (starter, maintenance, ‘light’ and bespoke) and a separate lipid solution. All of them were suitable for peripheral or central administration. Numeta came with similar choices: starter, maintenance -for central administration only- and ‘lite’ and Numeta peripheral, suitable for peripheral administration. Bespoke bags were also available if clinically indicated. We set out our desired outcomes and measured parameters accordingly:Patient outcomesMetabolic stability: electrolyte, glucose, bilirubin and lipid measurements summarised by the need to change from standard PN regimen and/or requirement for insulin.Fluid balance summarised by the lowest weight during the first two weeks of life and time taken to regain birth weight.Growth summarised by change of standard deviation score of weight and head circumference between birth and discharge or transfer back to local hospital.Liver tolerance of lipid solutions summarised by incidence of cholestasis (>25 μmol/l conjugated fraction of serum bilirubin)Days and type of PNSepsisService outcomesNursing time taken to prepare PNCostWastageAccess to productResultsAlthough we finished collecting the data in April 2019, we are still in the process of analysing it and evaluating the final results. There have been no cases of lipid overdose and our neonates (including the preterm ones) have so far tolerated well the new parenteral nutrition solution. Average nursing time preparing Numeta went down from 18.5 minutes to 8 minutes and comparison of cost came in favour of Numeta. PN wastage was higher with Numeta (4.7% Maintenance, 10% ‘light’, peripheral 50%) especially in the first month during the transition phase. There was no significant increase of bespoke bags when Numeta was introduced.ConclusionsIn summary, so far we have not identified significant clinical differences between the first six months of the project -using our old standardised nutrition regimen- and the last months -on the new all-in-one solution. We have continued with Numeta preterm solution on the basis of assumed safety.ReferencesRisk of severe harm and death from infusing total parenteral nutrition too rapidly in babies. NHS improvement patient safety alert September 2017.The Provision of Parenteral Nutrition within Neonatal Services – A Framework for Practice. British Association of Perinatal Medicine (BAPM) April 2016 www.bapm.org

The pudendal nerve motor branch regenerates via a brain derived neurotrophic factor mediated mechanism

Peripheral nerve injuries can significantly reduce quality of life. While some recover, most do not recover fully, resulting in neuropathic pain and loss of sensation and motor function. Research on the mechanisms of peripheral nerve regeneration could elucidate poor patient outcomes and potential treatments. This study was designed to determine if brain derived neurotrophic factor (BDNF) is necessary for pudendal nerve regeneration and functional recovery. Peripheral administration of tyrosine kinase B functional chimera (TrkB) was used to inhibit the BDNF regenerative pathway. Female Sprague-Dawley rats received tyrosine kinase B functional chimera (TrkB) or saline after a pudendal nerve crush (PNC) or Sham PNC and were divided into three groups: Sham PNC, PNC + Saline, and PNC + TrkB. Seven days after injury, relative βII tubulin expression (1.0 ± 0.2) was significantly decreased after PNC + TrkB compared to PNC + saline (2.9 ± 1.0). Three weeks after injury, BDNF plasma concentration (1320.8 ± 278.1 pg/ml) was significantly reduced in PNC + TrkB compared to PNC + saline rats (2053.4 ± 211.0 pg/ml). Pudendal nerve motor branch firing rate (54.0 ± 9.5 Hz) was significantly decreased in the PNC + TrkB group compared to the PNC + saline group (120.4 ± 17.1 Hz); while nerve firing rate of the PNC + saline group was not significantly different from sham PNC rats (121.8 ± 26.6 Hz). This study demonstrated that peripheral administration of TrkB bound free BDNF and inhibited the regenerative response after PNC. BDNF is necessary for normal PN motor branch recovery after PNC.

Peripheral Latanoprost Administration Lowers Intraocular Pressure in the Wistar Rat.

PurposeInstillation of latanoprost eye drops into the conjunctival sac to lower intraocular pressure (IOP) is the most frequently used treatment for primary open-angle glaucoma. The aim of this study was to evaluate the influence of latanoprost on IOP in the rat when applied peripherally.MethodsA rodent-dedicated tonometer was used to measure IOP in conscious adult male normotensive Wistar rats habituated to the measurement procedure. Commercially available 0.005% latanoprost solutions were continuously delivered to the periphery of the eye over 7 days using mini-pumps inserted subcutaneously in the animal’s back, and IOP was measured daily. For comparison, a solution containing an equimolar concentration of latanoprost acid, an active compound of latanoprost, was similarly infused into the eyes of different Wistar rats.ResultsContinuous subcutaneous infusion of latanoprost gradually decreased the IOP; the stable nadir of IOP, which was 20% lower than that prior to the start of infusion, was reached on day 3. The effect was statistically significant and fully reversed 2 days after cessation of drug delivery. Continuous subcutaneous application of the solution containing an equimolar amount of latanoprost acid did not appreciably influence the IOP.ConclusionSubcutaneous continuous delivery of latanoprost decreased the IOP in the conscious normotensive Wistar rats in this study. If this effect is confirmed in humans, it may open the possibility of using peripheral systems of drug delivery, which could significantly improve patient compliance.

N-Acetylcysteine Inhibits Kynurenine Aminotransferase II

The tryptophan metabolite kynurenic acid (KYNA) may play an important role in normal and abnormal cognitive processes, most likely by interfering with α7 nicotinic and NMDA receptor function. KYNA is formed from its immediate precursor kynurenine either by non-enzymatic oxidation or through irreversible transamination by kynurenine aminotransferases. In the mammalian brain, kynurenine aminotransferase II (KAT II) is the principal enzyme responsible for the neosynthesis of rapidly mobilizable KYNA, and therefore constitutes an attractive target for pro-cognitive interventions. N-acetylcysteine (NAC), a brain-penetrant drug with pro-cognitive efficacy in humans, has been proposed to exert its actions by increasing the levels of the anti-oxidant glutathione (GSH) in the brain. We report here that NAC, but not GSH, inhibits KAT II activity in brain tissue homogenates from rats and humans with IC50 values in the high micromolar to low millimolar range. With similar potency, the drug interfered with the de novo formation of KYNA in rat brain slices, and NAC was a competitive inhibitor of recombinant human KAT II (Ki: 450 μM). Furthermore, GSH failed to S-glutathionylate recombinant human KAT II treated with the dithiocarbamate drug disulfiram. Shown by microdialysis in the prefrontal cortex of rats treated with kynurenine (50 mg/kg, i.p.), peripheral administration of NAC (500 mg/kg, i.p., 120 and 60 min before the application of kynurenine) reduced KYNA neosynthesis by ∼50%. Together, these results suggest that NAC exerts its neurobiological effects at least in part by reducing cerebral KYNA formation via KAT II inhibition.

In silico methods predict new blood-brain barrier permeable structure for the inhibition of kynurenine 3-monooxygenase

Kynurenine 3-monooxygenase (KMO) regulates the levels of bioactive substances in the kynurenine pathway of tryptophan catabolism and its activity is tied to so many diseases that finding an appropriate inhibitor for KMO has become an urgent task. This especially proved to be difficult for the central nervous system related diseases due to the requirement that the supposed inhibitor should be both blood brain barrier permeable and should not cause hydrogen peroxide as a harmful side product. In this in silico study, we present our step-wise approach, whose starting point is based on the important experimental observations. To tackle the problem, a library of 7561938 structures was obtained from Zinc15 database utilizing the tranche browser. From this library, a subset of 501777 structures was determined with the considerations of their functional groups that constrain their applicability. Then, the binding affinity ranking of this set of structures was determined via virtual screening. Starting from the structures whose affinities are the highest among this subset, the ADMET properties were checked through in silico methods and the binding properties of the selected inhibitor candidates were further investigated via molecular dynamics simulations and MM/GBSA calculations. According to the computational results of this study, ZINC_71915355 has passed all the evaluations and is a potentially BBB permeable structure that can inhibit KMO. Additionally, ZINC_19827377 was identified as a new potential KMO inhibitor which may be more suitable for peripheral administration. From the in silico study presented herein, ZINC_71915355 and ZINC_19827377 structures, which showed high binding affinity without harmful H2O2 production, along with the tailored properties can now serve as powerful candidates for KMO inhibition and these hits are worth of further experimental validation.

Brain Testosterone-CYP1B1 (Cytochrome P450 1B1) Generated Metabolite 6β-Hydroxytestosterone Promotes Neurogenic Hypertension and Inflammation.

Previously, we showed that peripheral administration of 6β-hydroxytestosterone, a CYP1B1 (cytochrome P450 1B1)-generated metabolite of testosterone, promotes angiotensin II-induced hypertension in male mice. However, the site of action and the underlying mechanism by which 6β-hydroxytestosterone contributes to angiotensin II-induced hypertension is not known. Angiotensin II increases blood pressure by its central action, and CYP1B1 is expressed in the brain. This study was conducted to determine whether testosterone-CYP1B1 generated metabolite 6β-hydroxytestosterone locally in the brain promotes the effect of systemic angiotensin II to produce hypertension in male mice. Central CYP1B1 knockdown in wild-type (Cyp1b1+/+) mice by intracerebroventricular-adenovirus-GFP (green fluorescence protein)-CYP1B1-short hairpin (sh)RNA attenuated, whereas reconstitution of CYP1B1 by adenovirus-GFP-CYP1B1-DNA in the paraventricular nucleus but not in subfornical organ in Cyp1b1-/- mice restored angiotensin II-induced increase in systolic blood pressure measured by tail-cuff. Intracerebroventricular-testosterone in orchidectomized (Orchi)-Cyp1b1+/+ but not in Orchi-Cyp1b1-/-, and intracerebroventricular-6β-hydroxytestosterone in the Orchi-Cyp1b1-/- mice restored the angiotensin II-induced: (1) increase in mean arterial pressure measured by radiotelemetry, and autonomic imbalance; (2) reactive oxygen species production in the subfornical organ and paraventricular nucleus; (3) activation of microglia and astrocyte, and neuroinflammation in the paraventricular nucleus. The effect of intracerebroventricular-6β-hydroxytestosterone to restore the angiotensin II-induced increase in mean arterial pressure and autonomic imbalance in Orchi-Cyp1b1-/- mice was inhibited by intracerebroventricular-small interfering (si)RNA-androgen receptor (AR) and GPRC6A (G protein-coupled receptor C6A). These data suggest that testosterone-CYP1B1-generated metabolite 6β-hydroxytestosterone, most likely in the paraventricular nucleus via AR and GPRC6A, contributes to angiotensin II-induced hypertension and neuroinflammation in male mice.

Peripheral soluble epoxide hydrolase inhibition reduces hypernociception and inflammation in albumin-induced arthritis in temporomandibular joint of rats

Rheumatoid arthritis (RA) is characterized by chronic inflammation of the synovial tissue, joint dysfunction, and damage. Epoxyeicosatrienoic acids (EETs) are endogenous anti-inflammatory compounds, which are quickly converted by the soluble epoxide hydrolase (sEH) enzyme into a less active form with decreased biological effects. The inhibition of the sEH enzyme has been used as a strategy to lower nociception and inflammation. The goal of this study was to investigate whether the peripheral treatment with the sEH enzyme inhibitor 1- trifluoromethoxyphenyl-3-(1-propionylpiperidin-4-yl) urea (TPPU) could prevent the hypernociception and inflammation in the albumin-induced arthritis model in rats’ temporomandibular joint (TMJ). After the induction of experimental arthritis, animals were assessed for nociceptive behavior test, leukocyte infiltration counts and histologic analysis, ELISA to quantify several cytokines and Western blotting. The peripheral pretreatment with TPPU inhibited the arthritis-induced TMJ hypernociception and leukocyte migration. Moreover, the local concentrations of proinflammatory cytokines were diminished by TPPU, while the anti-inflammatory cytokine interleukin-10 was up-regulated in the TMJ tissue. Finally, TPPU significantly decreased protein expression of iNOS, while did not alter the expression of MRC1. This study provides evidence that the peripheral administration of TPPU reduces hypernociception and inflammation in TMJ experimental arthritis.

Peripheral Administration of RF9 Does Not Affect Hypothalamic-PituitaryGonadal Axis in Normal Fed Adult Male Macaque

Stress represses hypothalamic-pituitary-gonadal axis (HPG-axis) but RF9, a synthetic peptide, rescues such repression. To assess the role of RF9 in regulating HPG-axis under normal physiological conditions in higher primates, RF9 was administered to intact adult male rhesus monkeys and response of the HPG-axis was examined by measuring plasma testosterone as an end parameter of the axis. Control group (n=4) received normal saline whereas the treated group (n=4) received RF9. On the first day of experiment, four bolus injections of normal saline (1ml/animal) were administered intravenously at 2-hr interval to the control monkeys. Similarly, on the second day of experiment, treated group received four iv bolus injections of RF9 (0.1mg/kg BW) at 2-hr interval. Serial blood samples were collected at 20 min interval during a 6-hr period which started just after first saline/RF9 injection. Plasma testosterone levels were measured by using a specific EIA. Overall means of plasma testosterone levels and plasma testosterone area under curve (AUC) and overall mean testosterone and mean testosterone AUC in short time windows following each injection of RF9 and saline were comparable between the groups. Our results demonstrate that RF9 has no role in regulating HPG-axis under normal physiological conditions in adult male monkeys.

Muramyl dipeptide-mediated immunomodulation on monocyte subsets exerts therapeutic effects in a mouse model of Alzheimer\u2019s disease

BackgroundMuramyl dipeptide (MDP) is a component derived from minimal peptidoglycan motif from bacteria, and it is a ligand for the NOD2 receptor. Peripheral administration of MDP converts Ly6Chigh into Ly6Clow monocytes. Previously, we have shown that Ly6Clow monocytes play crucial roles in the pathology of a mouse model of Alzheimer’s disease (AD). However, medications with mild immunomodulatory effects that solely target specific monocyte subsets, without triggering microglial activation, are rare.MethodsThree months old APPswe/PS1 transgenic male mice and age-matched C57BL/6 J mice were used for high frequency (2 times/week) over 6 months and low frequency (once a week) over 3 months of intraperitoneally MDP (10 mg/kg) administrations. Flow cytometry analysis of monocyte subsets in blood, and behavioral and postmortem analyses were performed.ResultsMemory tests showed mild to a strong improvement in memory function, increased expression levels of postsynaptic density protein 95 (PSD95), and low-density lipoprotein receptor-related protein 1 (LRP1), which are involved in synaptic plasticity and amyloid-beta (Aβ) elimination, respectively. In addition, we found monocyte chemoattractant protein-1(MCP-1) levels significantly increased, whereas intercellular adhesion molecule-1(ICAM-1) significantly decreased, and microglial marker (Iba1) did not change in the treatment group compared to the control. In parallel, we discovered elevated cyclooxygenase-2 (COX2) expression levels in the treated group, which might be a positive factor for synaptic activity.ConclusionsOur results demonstrate that MDP is beneficial in both the early phase and, to some extent, later phases of the pathology in the mouse model of AD. These data open the way for potential MDP-based medications for AD.

Oxytocin Ameliorates Impaired Behaviors of High Fat Diet-Induced Obese Mice.

Excessive intake of fat is a major risk factor for lifestyle-related diseases such as heart disease and also affects brain function such as object recognition memory, social recognition, anxiety behavior, and depression-like behavior. Although oxytocin (OXT) has been reported to improve object recognition, social recognition, anxiety behavior, and depression-like behavior in specific conditions, previous studies did not explore the impact of OXT in high-fat diet (HFD)-fed mice. Furthermore, it remains unclear whether intake of HFD affects OXT/oxytocin receptor (OXTR) in the brain. Here, we demonstrated that peripheral OXT administration improves not only social recognition but also object recognition and depressive-like behavior in HFD-fed mice. In contrast, peripheral OXT administration to HFD-fed male mice increased fear and anxiety-related behavior. In addition, we observed that intake of HFD decreased OXTR and c-fos mRNA expression in the hippocampus, specifically. Furthermore, peripheral OXT administration increased OXT mRNA expression in the hypothalamus. Altogether, these findings suggest that OXT has the potential to improve various recognition memory processes via peripheral administration but also has side effects that increase fear-related behavior in males.

On the role of the dopaminergic system in the memory deficits induced by maternal deprivation

Previous researches showed that maternal deprivation (MD) leads to memory deficits that persist until adulthood. The hippocampus, an important brain structure involved in memory processes, receives dopaminergic afferents from other brain areas that modulate memory. Here we demonstrated that MD results in object recognition memory deficits that are reverted by intra-hippocampal stimulation of D1-dopaminergic receptor and peripheral administration of a dopamine precursor. The D1-dopaminergic receptor and peripheral administration of a dopamine precursor also promoted memory persistence in control rats.

Loss of Protection by Antiepileptic Drugs in Lipopolysaccharide-primed Pilocarpine-induced Status Epilepticus is Mediated via Inflammatory Signalling

The evidences from various studies show the association of peripheral and neuronal inflammation with complex pathophysiology of status epilepticus (SE). In this view, the present work attempted to develop a model of neuronal inflammation mediated SE by combining both epileptic and inflammatory components of the disease and also to mimic SE co-morbid with systemic inflammation by peripheral administration of the lipopolysaccharide (LPS) 2 h prior to the pilocarpine (PILO) induction in C57BL/6 mice. We evaluated the anti-convulsant and neuroprotective effects of 7-day prophylactic treatment with three conventional anti-epileptic drugs (Sodium valproate, SVP 300 mg/kg p.o.; Carbamazepine CBZ 100 mg/kg p.o.; Levetiracetam; LEV 200 mg/kg p.o.) of widespread clinical use. Morris water maze and Rota rod tests were carried out 24-h post-exposure to evaluate the neurobehavioral co-morbidities associated with neuroinflammation-mediated status epilepticus. Upon priming with LPS, the loss of protection against PILO-induced seizures was observed by SVP and CBZ, however, LEV showed protection by delaying the seizures. Dramatic elevation in the seizure severity and neuronal loss demonstrated the possible pro-convulsant effect of LPS in the PILO model. Also, the decreased cytokine levels by the AEDs showed their association with NF-κB, IL-1β, IL-6, TNF-α and TGF-β pathways in PILO model. The loss of protective activities of SVP and CBZ in LPS+PILO model was due to increased cytokine levels associated with over-activation of neuroinflammatory pathways, however, partial efficacy of LEV is possibly due to association of other neuroinflammatory mechanisms. The current work provides direct evidence of the contribution of increased peripheral and neuronal inflammation in seizures via regulation of inflammatory pathways in the brain.

Effect of Peripheral Insulin Administration on Phosphorylation of Tau in the Brain.

Abnormally hyperphosphorylated tau is the major protein of neurofibrillary tangles in Alzheimer's disease. Insulin activates PI3K-AKT signaling and regulates tau phosphorylation. Impaired brain insulin signaling is involved in Alzheimer's disease pathogenesis. However, the effect of peripheral insulin on tau phosphorylation is controversial. In the present study, we determined the effect of peripheral insulin administration on tau phosphorylation in brain. We intraperitoneally injected a super physiological dose of insulin to mice and analyzed PI3K-AKT signaling and tau phosphorylation in brains by western blots. We found that peripherally administered insulin activated the PI3K-AKT signaling pathway immediately in the liver, but not in the brain. Tau phosphorylation in the mouse brain was found to be first decreased (15 min) and then increased (30 min and 60 min) after peripheral insulin administration and these changes correlated inversely with body temperature and the level of brain protein O-GlcNAcylation. Maintaining body temperature of mice post peripheral insulin administration prevented the insulin/hypoglycemia-induced tau hyperphosphorylation after peripheral insulin administration. These findings suggest that peripheral insulin can induce tau hyperphosphorylation through both hypothermia and downregulation of brain protein O-GlcNAcylation during hypoglycemia.

ErbB1-dependent signalling and vesicular trafficking in primary afferent nociceptors associated with hypersensitivity in neuropathic pain

Effective analgesic treatment for neuropathic pain remains an unmet need, so previous evidence that epidermal growth factor receptor inhibitors (EGFRIs) provide unexpected rapid pain relief in a clinical setting points to a novel therapeutic opportunity. The present study utilises rodent models to address the cellular and molecular basis for the findings, focusing on primary sensory neurons because clinical pain relief is provided not only by small molecule EGFRIs, but also by the anti-EGFR antibodies cetuximab and panitumumab, which are unlikely to access the central nervous system in therapeutic concentrations. We report robust, rapid and dose-dependent analgesic effects of EGFRIs in two neuropathic pain models, matched by evidence with highly selective antibodies that expression of the EGFR (ErbB1 protein) is limited to small nociceptive afferent neurons. As other ErbB family members can heterodimerise with ErbB1, we investigated their distribution, showing consistent co-expression of ErbB2 but not ErbB3 or ErbB4, with ErbB1 in cell bodies of nociceptors, as well as providing evidence for direct molecular interaction of ErbB1 with ErbB2 in situ. Co-administration of selective ErbB1 and ErbB2 inhibitors produced clear evidence of greater-than-additive, synergistic analgesia; highlighting the prospect of a unique new combination therapy in which enhanced efficacy could be accompanied by minimisation of side-effects. Peripheral (intraplantar) administration of EGF elicited hypersensitivity only following nerve injury and this was reversed by local co-administration of selective inhibitors of either ErbB1 or ErbB2. Investigating how ErbB1 is activated in neuropathic pain, we found evidence for a role of Src tyrosine kinase, which can be activated by signals from inflammatory mediators, chemokines and cytokines during neuroinflammation. Considering downstream consequences of ErbB1 activation in neuropathic pain, we found direct recruitment to ErbB1 of an adapter for PI 3-kinase and Akt signalling together with clear Akt activation and robust analgesia from selective Akt inhibitors. The known Akt target and regulator of vesicular trafficking, AS160 was strongly phosphorylated at a perinuclear location during neuropathic pain in an ErbB1-, ErbB2- and Akt-dependent manner, corresponding to clustering and translocation of an AS160-partner, the vesicular chaperone, LRP1. Exploring whether neuronal ion channels that could contribute to hyperexcitability might be transported by this vesicular trafficking pathway we were able to identify Nav1.9, (Nav1.8) and Cav1.2 moving towards the plasma membrane or into proximal axonal locations – a process prevented by ErbB1 or Akt inhibitors. Overall these findings newly reveal both upstream and downstream signals to explain how ErbB1 can act as a signalling hub in neuropathic pain models and identify the trafficking of key ion channels to neuronal subcellular locations likely to contribute to hyperexcitability. The new concept of combined treatment with ErbB1 plus ErbB2 blockers is mechanistically validated as a promising strategy for the relief of neuropathic pain.

The vagus nerve mediates the suppressing effects of peripherally administered oxytocin on methamphetamine self-administration and seeking in rats.

The neuropeptide oxytocin has emerged as a promising pharmacotherapy for methamphetamine (METH) addiction, and clinical trials of intranasal oxytocin are underway. However, there is debate as to how peripherally administered oxytocin alters brain signalling to modulate addiction processes. Interestingly, there is evidence for functional interactions between peripheral oxytocin administration and the vagus nerve. Therefore, this study investigated whether the effects of peripherally administered oxytocin require vagal signalling to reduce METH self-administration and reinstatement of METH-seeking behaviours. Male and female Sprague-Dawley rats underwent surgery for jugular catheterisation and either subdiaphragmatic vagotomy (SDV) or a sham operation. Rats were trained to self-administer METH, and the effect of peripherally administered oxytocin on METH intake was assessed. Rats then underwent extinction, and effects of oxytocin were assessed on cue- and METH-induced reinstatement of METH-seeking. Oxytocin treatment robustly attenuated METH intake in both sexes, and SDV entirely prevented the suppressant effect of oxytocin (0.3 mg/kg) on METH intake, and partially prevented the effects of 1 mg/kg oxytocin in both sexes. After extinction, SDV decreased the suppressing effects of oxytocin on cue- and METH-primed reinstatement in males, but not females. SDV was functionally confirmed by measuring food intake following administration of the vagal dependent peptide, cholecyostokin-8. Our data suggest that vagus nerve signalling is required for the inhibitory effects of peripherally administered oxytocin on METH self-administration and reinstatement, and that this vagal dependency is partially mediated by sex and drug withdrawal. This study has implications for the use of oxytocin as a therapy for METH use disorder for both sexes.

Recommendation of an ideal mixed fluid preparation to be used as partial parenteral nutrition in surgical Neonatal Intensive Care Unit

Background: This study was conducted to assess the role of Partial Parenteral Nutrition (PPN) and a specially prepared “mixed fluid formula” used as PPN in sick surgical neonates.Methods: This single institution based Randomised Control Trial was performed in Surgical NICU of a tertiary care Centre. Surgical neonates who required bowel rest for >3 days like Esophageal Atresia (22), Duodenal Atresia (38), Jejuno-Ileal Atresia (50), Necrotising Enterocolitis (29) and others (11), were chosen. As a CONTROL population half (75) neonates were allowed maintenance fluid only (Isolyte P) and rest (75) were given a combination of maintenance fluid (Isolyte P) and PPN (specially prepared mixed fluid). This mixed fluid each 100 ml was prepared with 65ml Isolyte P, 15 ml 25% Dextrose, 10 ml Normal Saline, 7 ml Astymin 3, 2 ml Multivitamin and 1 ml Injection KCl. Fluid was continued till enteral feeding was established.Results: Neonates according to their body weight were divided in three groups <1 kg (50), 1-2.5 kg (71) and >2.5 kg (29) and their post-operative outcome was assessed as Clinical Positive/Negative and Laboratory Positive/Negative. Results were separately assessed in 3 body weight groups under “Control” and “Study” which showed both Clinical and Biochemical improvement in “Study group”.Conclusions: The composition of mixed fluid, its nutritional value, calorie supplementation, compatible osmolarity for peripheral venous administration and most importantly its easy and cost-effective preparation were assessed thoroughly and we recommend this “mixed fluid” preparation as a very useful PPN for sick surgical neonates.

SAT-604 The Role of the Focal Adhesion Kinase Family in Leptin Receptor Signaling

Overweight and obesity are global concerns affecting nearly one third of the world population. These conditions are characterized by increased adiposity and are accompanied by a proportional increase in circulating leptin, an anorexigenic adipokine. Leptin is responsible for signaling peripheral energy status to the central nervous system to modulate food intake and energy expenditure. As such, neurons within the hypothalamus expressing the long isoform of leptin receptor (LepRb), a type I cytokine receptor, are primarily responsible for mediating the effects of leptin, which signal predominantly through the JAK2-STAT3 transduction mechanism. STAT3 is a latent transcription factor activated upon phosphorylation, which triggers its homodimerization and nuclear translocation. Evidence, however, for JAK2-independent, STAT3-dependent leptin receptor signaling mechanisms exist. FAK (focal adhesion kinase, Ptk2) and Pyk2 (protein tyrosine kinase 2b, Ptk2b) are a subset of nonreceptor protein tyrosine kinases and comprise the focal adhesion kinase family. FAK and Pyk2 are implicated in the regulation of cytokine receptor signaling. Furthermore, Pyk2 knockout mice have an obesity prone phenotype. Here, we studied the role of the focal adhesion kinases in leptin receptor signaling using genetic and pharmacological approaches. We found that overexpression of Pyk2 or FAK increased STAT3 phosphorylation (activation). Overexpression of a FAK or Pyk2 construct with impaired kinase activity, however, attenuated STAT3 phosphorylation, suggesting the increase in STAT3 phosphorylation is largely dependent upon kinase activity of FAK/Pyk2. Treatment of cells with a small molecule dual inhibitor of FAK and Pyk2 (PF431396) attenuated leptin-induced STAT3 phosphorylation in a mouse hypothalamic cell line. Importantly, this effect is independent of JAK2, as PF treatment of two independent JAK2-deficient cell lines exhibited similar attenuation of leptin-induced STAT3 phosphorylation. To assess the physiological relevance of FAK/Pyk2 in leptin receptor signaling in vivo, we administered PF compound to the lateral ventricle of 24-hour fasted lean wild-type mice followed by peripheral leptin administration. Intracerebroventricular (ICV) administration of PF suppressed the anorectic effect of leptin as evidenced by impaired inhibition of food intake upon refeeding. Accordingly, analysis of total hypothalamic lysates from these mice showed ICV PF impaired leptin-induced STAT3 phosphorylation. Taken together, these data suggest that Pyk2 and/or FAK play a role in leptin signal transduction.

SAT-603 Growth Hormone-Releasing Hormone (GHRH) Antagonists Stimulate Feeding in Mice

Growth hormone-releasing hormone (GHRH) is a hypothalamic neuropeptide which stimulates the synthesis and secretion of growth hormone (GH) in pituitary gland. GHRH was also found to modulate food intake in mammals. MIA-690 is a synthetic GHRH antagonist of the Miami (MIA) series with potent antitumor effects. To date, its role in hypothalamic feeding modulation has not been evaluated. In the present study, we aimed to investigate the effects of chronic MIA-690 administration on feeding behavior, locomotor activity and hypothalamic dopamine (DA), norepinephrine (NE), serotonin (5-hydroxytriptamine, 5-HT), orexigenic peptides [agouti-related peptide (AgRP) and neuropeptide Y (NPY)] and anorexigenic peptides [cocaine and amphetamine-regulated transcript (CART) and proopiomelanocortin (POMC)] activity.Adult C57/BL6 mice were treated daily for 4 weeks by subcutaneous administration of (5 µg) MIA-690 or vehicle solution. Food intake and body weight were recorded every 4 days throughout the study. Immediately after the last injection, locomotor activity in the home cage was recorded, and thereafter animals were sacrificed. Visceral, subcutaneous and brown fat depots were quickly excised and weighed. Hypothalamus was also dissected for evaluating gene expression of AgRP, NPY, CART and POMC by real-time reverse transcription polymerase chain reaction. In addition, hypothalamic DA, NE and 5-HT levels were measured by high performance liquid chromatography (HPLC) coupled to electrochemical detection.Our findings show that administration of MIA-690 increased food intake and body weight, without affecting locomotor activity. No difference was observed in visceral, subcutaneous and brown fat mass in animals treated with MIA-690 or vehicle. As for neuromodulatory effects, a significant increase of AgRP gene expression and NE levels, along with a reduction of 5-HT levels were found after MIA-690 treatment. On the other hand, we did not observe any alteration in NPY, POMC and CART gene expression, as well as DA levels, following MIA-690 administration.In conclusion, chronic peripheral administration of MIA-690 could play an orexigenic role paralleled by increased body weight. The stimulation of feeding could be mediated, at least in part, by increased AgRP gene expression and NE levels and decreased 5-HT levels, in the hypothalamus.

Emergency Care Provided by the Israeli Military Airborne Combat Evacuation Unit during Helicopter Winch Rescue Operations

Objectives The objective of this study was to evaluate the emergency care provided by the Israeli Military Airborne Combat Evacuation Unit (MACEU) during helicopter winching operations. Methods: A retrospective cohort study was performed of all patients rescued by winching by the MACEU between December 2011 and October 2018. Data were extracted from the electronic medical records of the unit registry. The data collected included helicopter type, scene times, demographics, mechanism of injury, interventions, medications, and survival. Results: During the study period, 208 civilians with a mean age of 36.8 ± 19.2 years were evacuated from inaccessible areas, 192 were from difficult terrain, 10 from sea vessels, and 5 from floods. All patients were winched up with a crewmember. No patient or crewmember was injured during winching. Overall, 156/208 (75%) had a traumatic injury, and 52/208 (25%) had a medical emergency. Sikorsky UH-60 “BlackHawk” helicopters and Sikorsky CH-53 “Sea Stallion” helicopters were used in 179 and 28 operations, respectively. Eighteen different procedures were performed by the medical personnel of the unit on scene and en route. The most performed procedures were peripheral vascular access establishment (60.6%), fluids administration (57.7%), oxygen supplementation (42.8%), analgesia (39.9%) and spine immobilization (37.5%). On scene, none of the patients was treated with a physician-only intervention. Thirty/208 (14.5%) patients were winched-up in darkness conditions. Eleven/208 (5.3%) apneic breathing patients were winched up ventilated by a crewmember. All the six patients who had oxygen saturation ≥89% after entrance into the cabin, survived. Conclusions: The reported MACEU experience provides useful information on the clinical characteristics, medical interventions, and outcomes of patients rescued using a winching operation. Study findings emphasize the importance of airway management and ventilation during winching.

Peripheral Reelin Administration Rescues Corticosterone-Induced Depression-Like Behaviour and Neurochemical Alterations in Male and Female Rats

Depression is characterized by anhedonia and despair and chronic stress is usually a predisposing factor. In our lab, we model depression by administering rats with the stress hormone, corticosterone, for several weeks. Corticosterone-treated rats consistently exhibit cognitive deficits, despair- and anhedonia-like behavior, along with neurochemical alterations associated with depression. These deficits are rescued by antidepressants, as well as intrahippocampally- and peripherally-administered reelin. Reelin regulates neuronal migration, synaptogenesis, and dendritic growth. To date, no studies have addressed sex differences in response to reelin treatment.