Ragnhild Elisabeth Heimtun Paulsen

Sigrid Bjørnstad, Lars Peter Engeset Austdal, Borghild Roald, Joel Clinton Glover, Ragnhild Elisabeth Paulsen: Cracking the Egg: Potential of the Developing Chicken as a Model System for Nonclinical Safety Studies of Pharmaceuticals. Journal of Pharmacology and Experimental Therapeutics, Dec 2015

L P E Austdal, S Bjørnstad, G H Mathisen, P K Aden, I Mikkola, R E Paulsen, K E Rakkestad: Glucocorticoid Effects on Cerebellar Development in a Chicken Embryo Model: Exploring Changes in PAX6 and Metalloproteinase-9 After Exposure to Dexamethasone. Journal of Neuroendocrinology, Oct 2016

Ajay Yadav, Steven Verhaegen, Panagiotis Filis, Diana Domanska, Robert Lyle, Arvind Y M Sundaram, Magnus Leithaug, Gunn Charlotte Østby, Mona Aleksandersen, Hanne Friis Berntsen, Karin Elisabeth Zimmer, Paul A Fowler, Ragnhild Elisabeth Paulsen, Erik Ropstad: Exposure to a human relevant mixture of persistent organic pollutants or to perfluorooctane sulfonic acid alone dysregulates the developing cerebellum of chicken embryo. Environment International, Aug 2022

Denis Zosen, Elena Kondratskaya, Oykum Kaplan-Arabaci, Fred Haugen, Ragnhild Elisabeth Paulsen: Antidepressants escitalopram and venlafaxine up-regulate BDNF promoter IV but down-regulate neurite outgrowth in differentiating SH-SY5Y neurons. Neurochemistry International, Oct 2023

• Arabaci, Öyküm Kaplan; Zosen, Denis; Domanska, Diana; Bjørnstad, Sigrid; Santini, Simone & Karaca, Umut [Show all 12 contributors for this article] (2025). The antidepressant venlafaxine induces upregulation of miR-92 and histological alterations in a chicken embryo model. Neurochemistry International. ISSN 0197-0186. 190, p. 1–11. doi: 10.1016/j.neuint.2025.106056. Full text in Research Archive Show summary

  Antidepressants are commonly prescribed for treating anxiety disorders and depression, including pregnant patients. These medications influence the absorption and metabolism of key neurotransmitters such as serotonin, norepinephrine, and dopamine. However, many aspects of antidepressant function remain unclear, especially regarding their impact on neurodevelopment when used during pregnancy. To explore this, we employed a 3R-friendly chicken embryo model to investigate the cellular and molecular implications of antidepressants use. We focused on commonly used antidepressants for the study: escitalopram, which is a selective serotonin reuptake inhibitor; and venlafaxine, a serotonin and noradrenaline reuptake inhibitor, both of which are frequently prescribed during pregnancy. Drug distribution analysis showed that both antidepressant drugs reached clinical concentrations in the brain of developing chicken embryos and persisted for several hours. High-throughput miRNA-sequencing showed that venlafaxine upregulated 13 different miRNAs in the chicken cerebellum. Specifically, miR-92 and its human ortholog, miR-363, were significantly upregulated in chicken cerebellum and neuronally differentiated human SH-SY5Y cells, respectively. Neurite metrics showed a significant reduction in neurite outgrowth with venlafaxine, but not with escitalopram. Furthermore, venlafaxine caused histological alterations, such as increased white matter and reduced thickness of the cortical and molecular layer in the cerebellum. Both drugs reduced the thickness of the external granular layer and the number of mitotic events, impacting this crucial germinal zone of the brain. These findings highlight distinct neurodevelopmental effects of escitalopram and venlafaxine and raise awareness for potential adverse neurological effects in offspring exposed to antidepressants in utero.

• Arabaci, Öyküm Kaplan; Dančišinová, Zuzana & Paulsen, Ragnhild Elisabeth Heimtun (2025). The Chicken Embryo: An Alternative Animal Model in Development, Disease and Pharmacological Treatment. Pharmacology Research & Perspectives. 13(2). doi: 10.1002/prp2.70086. Full text in Research Archive
• Rajalingam, Dhaksshaginy; Boix, Fernando; Khoder, Ala Zaim; Andersen, Jannike Mørch & Paulsen, Ragnhild Elisabeth Heimtun (2023). Distribution of morphine and methadone to the brain in a developmental chicken embryo model. Toxicology and Applied Pharmacology. ISSN 0041-008X. 479. doi: 10.1016/j.taap.2023.116731. Full text in Research Archive Show summary

  The use and/or misuse of opioids by pregnant women would expose the fetuses to these drugs during critical stages of development with serious effects for the newborn, like the neonatal abstinence syndrome (NAS). We have revisited an established chicken model for NAS to describe the distribution of morphine and methadone to the brain and explore its validity as a valuable alternative to rodent models. For this purpose, chicken eggs were injected with a single dose of 10 mg/kg or 20 mg/kg morphine or 20 mg/kg methadone onto the chorioallantoic membrane (CAM) on embryonal day 13. Whole brains and lungs were harvested and the concentrations of morphine, methadone and their subsequent metabolites (morphine-3-glucuronide and EDDP, respectively) determined in the brain and lungs at different time points using LC-MS/MS. Morphine and methadone, as well as their metabolites, were detected both in the brain and lungs, with significantly higher concentrations in the lungs. Pharmacokinetic modelling showed that the distribution of morphine to the brain followed a first-order absorption with transit compartments and linear elimination, with concentrations linearly dependent on dose. Moreover, methadone, but not morphine, reduced μ receptor (the main morphine receptor) binding, which can be of relevance for opioid tolerance. The present study is the first to report the brain distribution of morphine, which can be described by standard pharmacokinetic processes, and methadone in the developing chicken embryo. The present findings supplement the already established model and support the use of this chicken model to study NAS.

• Zosen, Denis; Kondratskaya, Elena; Arabaci, Öyküm Kaplan; Haugen, Fred & Paulsen, Ragnhild Elisabeth Heimtun (2023). Antidepressants escitalopram and venlafaxine up-regulate BDNF promoter IV but down-regulate neurite outgrowth in differentiating SH-SY5Y neurons. Neurochemistry International. ISSN 0197-0186. 169. doi: 10.1016/j.neuint.2023.105571. Full text in Research Archive Show summary

  Antidepressants are used to treat depression and some anxiety disorders, including use in pregnant patients. The pharmacological actions of these drugs generally determine the uptake and metabolism of a series of neurotransmitters, such as serotonin, norepinephrine, or dopamine, along with an increase in BDNF expression. However, many aspects of antidepressant action remain unknown, particularly whether antidepressants interfere with normal neurodevelopment when taken by pregnant women. In order to reveal cellular and molecular implications crucial to the functioning of pathways related to antidepressant effects, we performed an investigation on neuronally differentiating human SH-SY5Y cells. To our knowledge, this is the first time human SH-SY5Y cells in cultures of purely neuronal cells induced by controlled differentiation with retinoic acid are followed by short-term 48-h exposure to 0.1–10 μM escitalopram or venlafaxine. Treatment with antidepressants (1 μM) did not affect the electrophysiological properties of SH-SY5Y cells. However, the percentage of mature neurons exhibiting voltage-gated sodium currents was substantially higher in cultures pre-treated with either antidepressant. After exposure to escitalopram or venlafaxine, we observed a concentration-dependent increase in activity-dependent BDNF promoter IV activation. The assessment of neurite metrics showed significant down-regulation of neurite outgrowth upon exposure to venlafaxine. Identified changes may represent links to molecular processes of importance to depression and be involved in neurodevelopmental alterations observed in postpartum children exposed to antidepressants antenatally.

• Wæhler, Hallvard Austin; Labba, Nils-Anders Johannes; Paulsen, Ragnhild Elisabeth Heimtun; Sandve, Geir Kjetil Ferkingstad & Eskeland, Ragnhild (2023). ANDA: an open-source tool for automated image analysis of in vitro neuronal cells. BMC Neuroscience. 24(1), p. 1–11. doi: 10.1186/s12868-023-00826-z. Full text in Research Archive Show summary

  Background Imaging of in vitro neuronal differentiation and measurements of cell morphologies have led to novel insights into neuronal development. Live-cell imaging techniques and large datasets of images have increased the demand for automated pipelines for quantitative analysis of neuronal morphological metrics. Results ANDA is an analysis workflow that quantifies various aspects of neuronal morphology from high-throughput live-cell imaging screens of in vitro neuronal cell types. This tool automates the analysis of neuronal cell numbers, neurite lengths and neurite attachment points. We used chicken, rat, mouse, and human in vitro models for neuronal differentiation and have demonstrated the accuracy, versatility, and efficiency of the tool. Conclusions ANDA is an open-source tool that is easy to use and capable of automated processing from time-course measurements of neuronal cells. The strength of this pipeline is the capability to analyse high-throughput imaging screens.

• Yadav, Ajay; Verhaegen, Steven; Filis, Panagiotis; Domanska, Diana; Lyle, Robert & Sundaram, Arvind [Show all 14 contributors for this article] (2022). Exposure to a human relevant mixture of persistent organic pollutants or to perfluorooctane sulfonic acid alone dysregulates the developing cerebellum of chicken embryo. Environment International. ISSN 0160-4120. 166, p. 1–13. doi: 10.1016/j.envint.2022.107379. Full text in Research Archive Show summary

  Prenatal exposure to persistent organic pollutants (POPs) is associated with neurodevelopmental disorders. In the present study, we explored whether a human-relevant POP mixture affects the development of chicken embryo cerebellum. We used a defined mixture of 29 POPs, with chemical composition and concentrations based on blood levels in the Scandinavian population. We also evaluated exposure to a prominent compound in the mixture, perfluorooctane sulfonic acid (PFOS), alone. Embryos (n = 7–9 per exposure group) were exposed by injection directly into the allantois at embryonic day 13 (E13). Cerebella were isolated at E17 and subjected to morphological, RNA-seq and shot-gun proteomics analyses. There was a reduction in thickness of the molecular layer of cerebellar cortex in both exposure scenarios. Exposure to the POP mixture significantly affected expression of 65 of 13,800 transcripts, and 43 of 2,568 proteins, when compared to solvent control. PFOS alone affected expression of 80 of 13,859 transcripts, and 69 of 2,555 proteins. Twenty-five genes and 15 proteins were common for both exposure groups. These findings point to alterations in molecular events linked to retinoid X receptor (RXR) signalling, neuronal cell proliferation and migration, cellular stress responses including unfolded protein response, lipid metabolism, and myelination. Exposure to the POP mixture increased methionine oxidation, whereas PFOS decreased oxidation. Several of the altered genes and proteins are involved in a wide variety of neurological disorders. We conclude that POP exposure can interfere with fundamental aspects of neurodevelopment, altering molecular pathways that are associated with adverse neurocognitive and behavioural outcomes.

• Labba, Nils-Anders Johannes; Wæhler, Hallvard Austin; Houdaifi, Nora; Zosen, Denis; Haugen, Fred & Paulsen, Ragnhild Elisabeth Heimtun [Show all 8 contributors for this article] (2022). Paracetamol perturbs neuronal arborization and disrupts the cytoskeletal proteins SPTBN1 and TUBB3 in both human and chicken in vitro models. Toxicology and Applied Pharmacology. ISSN 0041-008X. 449. doi: 10.1016/j.taap.2022.116130. Full text in Research Archive Show summary

  Epidemiological studies have linked long-term/high-dose usage of paracetamol (N-acetyl-para-aminophenol, APAP) during pregnancy to adverse neuropsychiatric outcomes, primarily attention-deficit hyperactive disorder (ADHD), in the offspring. Though variable, ADHD has been associated with phenotypic alterations characterized by reductions in grey matter densities and aberrations in structural connectivity, effects which are thought to originate in neurodevelopment. We used embryonic chicken cerebellar granule neurons (CGNs) and neuronally differentiating human NTERA2 cells (NT2Ns) to investigate the in vitro effects of APAP on cell viability, migration, neuritogenesis, and the intracellular levels of various proteins involved in neurodevelopment as well as in the maintenance of the structure and function of neurites. Exposure to APAP ranging from 100 to 1600 μM yielded concentration- and time-dependent reductions in cell viability and levels of neurite arborization, as well as reductions in the levels of the cytoskeletal protein β2-spectrin, with the highest APAP concentration resulting in between 50 and 75% reductions in the aforementioned metrics over the course of 72 h. Exposure to APAP also reduced migration in the NT2Ns but not CGNs. Moreover, we found concentration- and time-dependent increases in punctate aggregation of the cytoskeletal protein β3-tubulin following exposure to APAP in both cell model systems, with the highest APAP concentration approximately doubling the number of aggregates over 72–120 h. Our findings demonstrate that APAP negatively perturbs neurite arborization degree, with concurrent reductions in the protein levels of β2-spectrin and disruption of the integrity of β3-tubulin, both proteins of which play important roles in neuronal structure and function.

• Lauvås, Anna Jacobsen; Lislien, Malene; Holme, Jørn Andreas; Dirven, Hubert; Paulsen, Ragnhild Elisabeth Heimtun & Alm, Inger Margit [Show all 13 contributors for this article] (2022). Developmental neurotoxicity of acrylamide and its metabolite glycidamide in a human mixed culture of neurons and astrocytes undergoing differentiation in concentrations relevant for human exposure. Neurotoxicology. ISSN 0161-813X. 92, p. 33–48. doi: 10.1016/j.neuro.2022.07.001. Full text in Research Archive Show summary

  Neural stem cells (NSCs) derived from human induced pluripotent stem cells were used to investigate effects of exposure to the food contaminant acrylamide (AA) and its main metabolite glycidamide (GA) on key neurodevelopmental processes. Diet is an important source of human AA exposure for pregnant women, and AA is known to pass the placenta and the newborn may also be exposed through breast feeding after birth. The NSCs were exposed to AA and GA (1 ×10-8 - 3 ×10-3 M) under 7 days of proliferation and up to 28 days of differentiation towards a mixed culture of neurons and astrocytes. Effects on cell viability was measured using Alamar Blue™ cell viability assay, alterations in gene expression were assessed using real time PCR and RNA sequencing, and protein levels were quantified using immunocytochemistry and high content imaging. Effects of AA and GA on neurodevelopmental processes were evaluated using endpoints linked to common key events identified in the existing developmental neurotoxicity adverse outcome pathways (AOPs). Our results suggest that AA and GA at low concentrations (1 ×10-7 - 1 ×10-8 M) increased cell viability and markers of proliferation both in proliferating NSCs (7 days) and in maturing neurons after 14-28 days of differentiation. IC50 for cell death of AA and GA was 5.2 × 10-3 M and 5.8 × 10-4 M, respectively, showing about ten times higher potency for GA. Increased expression of brain derived neurotrophic factor (BDNF) concomitant with decreased synaptogenesis were observed for GA exposure (10-7 M) only at later differentiation stages, and an increased number of astrocytes (up to 3-fold) at 14 and 21 days of differentiation. Also, AA exposure gave tendency towards decreased differentiation (increased percent Nestin positive cells). After 28 days, neurite branch points and number of neurites per neuron measured by microtubule-associated protein 2 (Map2) staining decreased, while the same neurite features measured by βIII-Tubulin increased, indicating perturbation of neuronal differentiation and maturation.

• Zosen, Denis; Austdal, Lars Peter Engeset; Bjørnstad, Sigrid; Lumor, Josephine Sena & Paulsen, Ragnhild Elisabeth Heimtun (2022). Antiepileptic drugs lamotrigine and valproate differentially affect neuronal maturation in the developing chick embryo, yet with PAX6 as a potential common mediator. Neurotoxicology and Teratology. ISSN 0892-0362. 90. doi: 10.1016/j.ntt.2021.107057. Full text in Research Archive
• Lindeman, Birgitte; Johansson, Ylva; Andreassen, Mathilda; Husøy, Trine; Dirven, Hubert & Hofer, Tim [Show all 13 contributors for this article] (2021). Does the food processing contaminant acrylamide cause developmental neurotoxicity? A review and identification of knowledge gaps. Reproductive Toxicology. ISSN 0890-6238. 101, p. 93–114. doi: 10.1016/j.reprotox.2021.02.006. Full text in Research Archive Show summary

  There is a worldwide concern on adverse health effects of dietary exposure to acrylamide (AA) due to its presence in commonly consumed foods. AA is formed when carbohydrate rich foods containing asparagine and reducing sugars are prepared at high temperatures and low moisture conditions. Upon oral intake, AA is rapidly absorbed and distributed to all organs. AA is a known human neurotoxicant that can reach the developing foetus via placental transfer and breast milk. Although adverse neurodevelopmental effects have been observed after prenatal AA exposure in rodents, adverse effects of AA on the developing brain has so far not been studied in humans. However, epidemiological studies indicate that gestational exposure to AA impair foetal growth and AA exposure has been associated with reduced head circumference of the neonate. Thus, there is an urgent need for further research to elucidate whether pre- and perinatal AA exposure in humans might impair neurodevelopment and adversely affect neuronal function postnatally. Here, we review the literature with emphasis on the iden-tification of critical knowledge gaps in relation to neurodevelopmental toxicity of AA and its mode of action and we suggest research strategies to close these gaps to better protect the unborn child.

• Zosen, Denis; Hadera, Mussie Ghezu; Lumor, Josephine Sena; Andersen, Jannike Mørch & Paulsen, Ragnhild Elisabeth Heimtun (2021). Chicken embryo as animal model to study drug distribution to the developing brain. Journal of pharmacological and toxicological methods. ISSN 1056-8719. 112, p. 1–5. doi: 10.1016/j.vascn.2021.107105. Full text in Research Archive
• Davidsen, Nichlas; Lauvås, Anna Jacobsen; Myhre, Oddvar; Ropstad, Erik; Carpi, Donatella & Gyves, Emilio Mendoza-de [Show all 11 contributors for this article] (2021). Exposure to human relevant mixtures of halogenated persistent organic pollutants (POPs) alters neurodevelopmental processes in human neural stem cells undergoing differentiation. Reproductive Toxicology. ISSN 0890-6238. 100, p. 17–34. doi: 10.1016/j.reprotox.2020.12.013. Full text in Research Archive Show summary

  Halogenated persistent organic pollutants (POPs) like perfluorinated alkylated substances (PFASs), brominated flame retardants (BFRs), organochlorine pesticides and polychlorinated biphenyls (PCBs) are known to cause cancer, immunotoxicity, neurotoxicity and interfere with reproduction and development. Concerns have been raised about the impact of POPs upon brain development and possibly neurodevelopmental disorders. The developing brain is a particularly vulnerable organ due to dynamic and complex neurodevelopmental processes occurring early in life. However, very few studies have reported on the effects of POP mixtures at human relevant exposures, and their impact on key neurodevelopmental processes using human in vitro test systems. Aiming to reduce this knowledge gap, we exposed mixed neuronal/glial cultures differentiated from neural stem cells (NSCs) derived from human induced pluripotent stem cells (hiPSCs) to reconstructed mixtures of 29 different POPs using concentrations comparable to Scandinavian human blood levels. Effects of the POP mixtures on neuronal proliferation, differentiation and synaptogenesis were evaluated using in vitro assays anchored to common key events identified in the existing developmental neurotoxicity (DNT) adverse outcome pathways (AOPs). The present study showed that mixtures of POPs (in particular brominated and chlorinated compounds) at human relevant concentrations increased proliferation of NSCs and decreased synapse number. Based on a mathematical modelling, synaptogenesis and neurite outgrowth seem to be the most sensitive DNT in vitro endpoints. Our results indicate that prenatal exposure to POPs may affect human brain development, potentially contributing to recently observed learning and memory deficits in children.

• Yadav, Ajay; Verhaegen, Steven; Hadera, Mussie Ghezu; Berntsen, Hanne Friis; Berg, Vidar & Lyche, Jan Ludvig [Show all 13 contributors for this article] (2021). Peripherally administered persistent organic pollutants distribute to the brain of developing chicken embryo in concentrations relevant for human exposure. Neurotoxicology. ISSN 0161-813X. 88, p. 79–87. doi: 10.1016/j.neuro.2021.10.013. Full text in Research Archive Show summary

  Persistent organic pollutants (POPs) can reach the fetal brain and contribute to developmental neurotoxicity. To explore the distribution of POPs to the fetal brain, we exposed chicken embryos to a POP mixture, containing 29 different compounds with concentrations based on blood levels measured in the Scandinavian human population. The mixture was injected into the allantois at embryonic day 13 (E13), aiming at a theoretical concentration of 10 times human blood levels. POPs concentrations in the brain were measured at 0.5, 1, 2, 4, 6, 24, 48, and 72 h after administration. Twenty-seven of the individual compounds were detected during at least one of the time-points analyzed. Generally, the concentrations of most of the measured compounds were within the order of magnitude of those reported in human brain samples. Differences in the speed of distribution to the brain were observed between the per- and polyfluoroalkyl substances (PFASs), which have protein binding potential, and the lipophilic polychlorinated biphenyls (PCBs), organochlorine pesticides (OCPs) and brominated flame retardants (BFRs). Based on pharmacokinetic modeling, PFASs were best described by a one compartment model. PFASs displayed relatively slow elimination (Kel) and persisted at high levels in the brain. Lipophilic OCPs and PCBs could be fitted to a 2-compartment model. These showed high levels in the brain relative to the dose administrated as calculated by area under the curve (AUC)/Dose. Altogether, our study showed that chicken is a suitable model to explore the distribution of POPs into the developing brain at concentrations which are relevant for humans.

• Yadav, Ajay; Verhaegen, Steven; Verbruggen, Evelien; Kerhoas, Marie; Huiberts, Eva Henriette Willemijn & Hadera, Mussie Ghezu [Show all 10 contributors for this article] (2021). A human relevant mixture of persistent organic pollutants (POPs) and perfluorooctane sulfonic acid (PFOS) differentially affect glutamate induced excitotoxic responses in chicken cerebellum granule neurons (CGNs) in vitro. Reproductive Toxicology. ISSN 0890-6238. 100, p. 109–119. doi: 10.1016/j.reprotox.2021.01.008. Full text in Research Archive Show summary

  Primary cultures of cerebellar granule neurons (CGNs) derived from chicken embryos were used to explore the effects on developmental neurotoxicity by a complex defined mixture of persistent organic pollutants (POPs). Its chemical composition and concentrations were based on blood levels in the Norwegian/Scandinavian popula-tion. Perfluorooctane sulfonic acid (PFOS) alone, its most abundant compound was also evaluated. Different stages of CGNs maturation, between day in vitro (DIV) 1, 3, and 5 were exposed to the POP mixture, or PFOS alone. Their combination with glutamate, an excitatory endogenous neurotransmitter important in neuro-development, also known to cause excitotoxicity was evaluated. Outcomes with the mixture at 500x blood levels were compared to PFOS at its corresponding concentration of 20 μM. The POP mixture reduced tetrazolium salt (MTT) conversion at earlier stages of maturation, compared to PFOS alone. Glutamate-induced excitotoxicity was enhanced above the level of that induced by glutamate alone, especially in mature CGNs at DIV5. Glutathione (GSH) concentrations seemed to set the level of sensitivity for the toxic insults from exposures to the pollutants. The role of N-methyl-D-aspartate receptor (NMDA-R) mediated calcium influx in pollutant exposures was investigated using the non-competitive and competitive receptor antagonists MK-801 and CGP 39551. Obser-vations indicate a calcium-independent, but still NMDA-R dependent mechanism in the absence of glutamate, and a calcium- and NMDA-R dependent one in the presence of glutamate. The outcomes for the POP mixture cannot be explained by PFOS alone, indicating that other chemicals in the mixture contribute its overall effect.

• Berntsen, Hanne Friis; Moldes-Anaya, Angel; Bjørklund, Cesilie Granum; Ragazzi, Lorenzo; Haug, Trude M & Strandabø, Rønnaug [Show all 10 contributors for this article] (2020). Perfluoroalkyl acids potentiate glutamate excitotoxicity in rat cerebellar granule neurons. Toxicology. ISSN 0300-483X. 445. doi: 10.1016/j.tox.2020.152610. Full text in Research Archive Show summary

  Perfluoroalkyl acids (PFAAs) are persistent man-made chemicals, ubiquitous in nature and present in human samples. Although restrictions are being introduced, they are still used in industrial processes as well as in consumer products. PFAAs cross the blood-brain-barrier and have been observed to induce adverse neurobehavioural effects in humans and animals as well as adverse effects in neuronal in vitro studies. The sulfonated PFAA perfluorooctane sulfonic acid (PFOS), has been shown to induce excitotoxicity via the N-methyl-D-aspartate receptor (NMDA-R) in cultures of rat cerebellar granule neurons (CGNs). In the present study the aim was to further characterise PFOS-induced toxicity (1–60 μM) in rat CGNs, by examining interactions between PFOS and elements of glutamatergic signalling and excitotoxicity. Effects of the carboxylated PFAA, perfluorooctanoic acid (PFOA, 300–500 μM) on the same endpoints were also examined. During experiments in immature cultures at days in vitro (DIV) 8, PFOS increased both the potency and efficacy of glutamate, whereas in mature cultures at DIV 14 only increased potency was observed. PFOA also increased potency at DIV 14. PFOS-enhanced glutamate toxicity was further antagonised by the competitive NMDA-R antagonist 3-((R)-2-Carboxypiperazin-4-yl)-propyl-1-phosphonic acid (CPP) at DIV 8. At DIV 8, PFOS also induced glutamate release (9–13 fold increase vs DMSO control) after 1−3 and 24 h exposure, whereas for PFOA a large (80 fold) increase was observed, but only after 24 h. PFOS and PFOA both also increased alanine and decreased serine levels after 24 h exposure. In conclusion, our results indicate that PFOS at concentrations relevant in an occupational setting, may be inducing excitotoxicity, and potentiation of glutamate signalling, via an allosteric action on the NMDA-R or by actions on other elements regulating glutamate release or NMDA-R function. Our results further support our previous findings that PFOS and PFOA at equipotent concentrations induce toxicity via different mechanisms of action.

• Evjen, Sigvart; Åstrand, Ove Alexander Høgmoen; Gaarder, Mona; Paulsen, Ragnhild Elisabeth; Fiksdahl, Anne & Knuutila, Hanna K (2020). Degradative behavior and toxicity of alkylated imidazoles. Industrial & Engineering Chemistry Research. ISSN 0888-5885. 59(2), p. 587–595. doi: 10.1021/acs.iecr.9b05100. Full text in Research Archive Show summary

  The thermal and oxidative degradation behavior and the corrosiveness of imidazole (Im), 2-methylimidazole (2MIm), 2-ethyl-4-methylimidazole (2E4MIm), 2,4,5-trimethylimidazole (2,4,5MIm), and 1,2,4,5-tetramethylimidazole (1,2,4,5MIm) were investigated in a CO2 rich environment. The imidazoles demonstrated high thermal stability in all solutions; however, Im and 2MIm were corrosive. Polyalkylated imidazoles were less stable toward oxidation compared to Im. Rat cell (PC-12) toxicology screening of Im and 45 alkylated imidazoles showed reduced toxicity for polyalkylated imidazoles compared to Im. Cell viability correlated negatively with cLogP predictions when cLogP > 3. The high rate of oxidative degradation and formation of potentially carcinogenic degradation products will prevent the use of polyalkylated imidazoles in industrial processes directed toward CO2.

• Berntsen, Hanne Friis; Duale, Nur; Bjørklund, Cesilie Granum; Rangel-Huerta, Oscar Daniel; Dyrberg, Kine & Hofer, Tim [Show all 13 contributors for this article] (2020). Effects of a human-based mixture of persistent organic pollutants on the in vivo exposed cerebellum and cerebellar neuronal cultures exposed in vitro. Environment International. ISSN 0160-4120. 146. doi: 10.1016/j.envint.2020.106240. Full text in Research Archive Show summary

  Exposure to persistent organic pollutants (POPs), encompassing chlorinated (Cl), brominated (Br) and perfluoroalkyl acid (PFAA) compounds is associated with adverse neurobehaviour in humans and animals, and is observed to cause adverse effects in nerve cell cultures. Most studies focus on single POPs, whereas studies on effects of complex mixtures are limited. We examined the effects of a mixture of 29 persistent compounds (Cl + Br + PFAA, named Total mixture), as well as 6 sub-mixtures on in vitro exposed rat cerebellar granule neurons (CGNs). Protein expression studies of cerebella from in vivo exposed mice offspring were also conducted. The selection of chemicals for the POP mixture was based on compounds being prominent in food, breast milk or blood from the Scandinavian human population. The Total mixture and sub-mixtures containing PFAAs caused greater toxicity in rat CGNs than the single or combined Cl/Br sub-mixtures, with significant impact on viability from 500x human blood levels. The potencies for these mixtures based on LC50 values were Br + PFAA mixture > Total mixture > Cl + PFAA mixture > PFAA mixture. These mixtures also accelerated induced lipid peroxidation. Protection by the competitive N-methyl-D-aspartate (NMDA) receptor antagonist 3-((R)-2-Carboxypiperazin-4-yl)-propyl-1-phosphonic acid (CPP) indicated involvement of the NMDA receptor in PFAA and Total mixture-, but not Cl mixture-induced toxicity. Gene-expression studies in rat CGNs using a sub-toxic and marginally toxic concentration ((0.4 nM-5.5 µM) 333x and (1 nM-8.2 µM) 500x human blood levels) of the mixtures, revealed differential expression of genes involved in apoptosis, oxidative stress, neurotransmission and cerebellar development, with more genes affected at the marginally toxic concentration. The two important neurodevelopmental markers Pax6 and Grin2b were downregulated at 500x human blood levels, accompanied by decreases in PAX6 and GluN2B protein levels, in cerebellum of offspring mice from mothers exposed to the Total mixture throughout pregnancy and lactation. In rat CGNs, the glutathione peroxidase gene Prdx6 and the regulatory transmembrane glycoprotein gene Sirpa were highly upregulated at both concentrations. In conclusion, our results support that early-life exposure to mixtures of POPs can cause adverse neurodevelopmental effects.

• Yadav, Ajay; Amber, Mazia; Zosen, Denis; Labba, Nils Anders; Huiberts, Eva Henriette Willemijn & Samulin-Erdem, Johanna Maria [Show all 13 contributors for this article] (2020). A human relevant mixture of persistent organic pollutants (POPs) and perfluorooctane sulfonic acid (PFOS) enhance nerve growth factor (NGF)-induced neurite outgrowth in PC12 cells. Toxicology Letters. ISSN 0378-4274. 338, p. 85–96. doi: 10.1016/j.toxlet.2020.12.007. Full text in Research Archive Show summary

  Disruption of neurite outgrowth is a marker for neurotoxicity. Persistent organic pollutants (POPs) are potential developmental neurotoxicants. We investigated their effect on neurite outgrowth in PC12 rat pheochromocytoma cells, in absence or presence of nerve growth factor (NGF), an inducer of neuronal differentiation. Cells were exposed for 72 h to a defined mixture of POPs with chemical composition and concentrations based on blood levels in the Scandinavian population. We also evaluated perfluorooctane sulfonic acid (PFOS) alone, the most abundant compound in the POP mixture. Only higher concentrations of POP mixture reduced tetrazolium salt (MTT) conversion. High-content analysis showed a decrease in cell number, but no changes for nuclear and mitochondrial cellular health parameters. Robust glutathione levels were observed in NGF-differentiated cells. Live imaging, using the IncuCyte ZOOM platform indicated ongoing cell proliferation over time, but slower in presence of NGF. The pollutants did not inhibit neuritogenesis, but rather increased NGF-induced neurite length. PFOS induced neurite outgrowth to about 50 % of the level seen with the POP mixture. Neither the POP mixture nor PFOS affected neurite length in the absence of NGF. Our observations indicate that realistic complex mixtures of environmental pollutants can affect neuronal connectivity via NGF-induced neurite outgrowth.

• Fjelldal, Marthe Fredheim; Hadera, Mussie Ghezu; Kongstorp, Mette; Austdal, Lars Peter Engeset; Sulovic, Ana & Andersen, Jannike Mørch [Show all 7 contributors for this article] (2019). Opioid receptor-mediated changes in the NMDA receptor in developing rat and chicken. International Journal of Developmental Neuroscience. ISSN 0736-5748. 78(C), p. 19–27. doi: 10.1016/j.ijdevneu.2019.07.009. Full text in Research Archive
• Fjelldal, Marthe Fredheim; Freyd, Thibaud; Evenseth, Linn; Sylte, Ingebrigt; Ring, Avi & Paulsen, Ragnhild Elisabeth (2019). Exploring the overlapping binding sites of ifenprodil and EVT‐101 in GluN2B‐containing NMDA receptors using novel chicken embryo forebrain cultures and molecular modeling. Pharmacology Research & Perspectives. doi: 10.1002/prp2.480. Full text in Research Archive Show summary

  N‐methyl‐d‐aspartate receptors (NMDAR) are widely expressed in the brain. GluN2B subunit‐containing NMDARs has recently attracted significant attention as potential pharmacological targets, with emphasis on the functional properties of allosteric antagonists. We used primary cultures from chicken embryo forebrain (E10), expressing native GluN2B‐containing NMDA receptors as a novel model system. Comparing the inhibition of calcium influx by well‐known GluN2B subunit‐specific allosteric antagonists, the following rank order of potency was found: EVT‐101 (EC50 22 ± 8 nmol/L) > Ro 25‐6981 (EC50 60 ± 30 nmol/L) > ifenprodil (EC50 100 ± 40 nmol/L) > eliprodil (EC50 1300 ± 700 nmol/L), similar to previous observations in rat cortical cultures and cell lines overexpressing chimeric receptors. The less explored Ro 04‐5595 had an EC50 of 186 ± 32 nmol/L. Venturing to explain the differences in potency, binding properties were further studied by in silico docking and molecular dynamics simulations using x‐ray crystal structures of GluN1/GluN2B amino terminal domain. We found that Ro 04‐5595 was predicted to bind the recently discovered EVT‐101 binding site, not the ifenprodil‐binding site. The EVT‐101 binding pocket appears to accommodate more structurally different ligands than the ifenprodil‐binding site, and contains residues essential in ligand interactions necessary for calcium influx inhibition. For the ifenprodil site, the less effective antagonist (eliprodil) fails to interact with key residues, while in the EVT‐101 pocket, difference in potency might be explained by differences in ligand‐receptor interaction patterns.

• Berntsen, Hanne Friis; Bjørklund, Cesilie Granum; Strandabø, Rønnaug; Haug, Trude; Moldes-Anaya, Angel & Fuentes-Lazaro, Judit [Show all 10 contributors for this article] (2018). PFOS-induced excitotoxicity is dependent on Ca2+ influx via NMDA receptors in rat cerebellar granule neurons. Toxicology and Applied Pharmacology. ISSN 0041-008X. 357, p. 19–32. doi: 10.1016/j.taap.2018.08.015. Full text in Research Archive Show summary

  Perfluoroalkyl acids (PFAAs) are persistent compounds used in many industrial as well as consumer products. Despite restrictions, these compounds are found at measurable concentrations in samples of human and animal origin. In the present study we examined whether the effects on cell viability of two sulfonated and four carboxylated PFAAs in cultures of cerebellar granule neurons (CGNs), could be associated with deleterious activation of the N-methyl-d-aspartate receptor (NMDA-R). PFAA-induced effects on viability in rat CGNs and unstimulated PC12 cells were examined using the MTT assay. Cells from the PC12 rat pheochromocytoma cell line lack the expression of functional NMDA-Rs and were used to verify lower toxicity of perfluorooctanesulfonic acid (PFOS) in cells not expressing NMDA-Rs. Protective effects of NMDA-R antagonists, and extracellular as well as intracellular Ca2+ chelators were investigated. Cytosolic Ca2+ ([Ca2+]i) was measured using Fura-2. In rat CGNs the effects of the NMDA-R antagonists MK-801, memantine and CPP indicated involvement of the NMDA-R in the decreased viability induced by PFOS and perfluorohexanesulfonic acid (PFHxS). No effects were associated with the four carboxylated PFAAs studied. Further, EGTA and CPP protected against PFOS-induced decreases in cell viability, whereas no protection was afforded by BAPTA-AM. [Ca2+]i significantly increased after exposure to PFOS, and this increase was completely blocked by MK-801. In PC12 cells a higher concentration of PFOS was required to induce equivalent levels of toxicity as compared to in rat CGNs. PFOS-induced toxicity in PC12 cells was not affected by CPP. In conclusion, PFOS at the tested concentrations induces excitotoxicity in rat CGNs, which likely involves influx of extracellular Ca2+ via the NMDA-R. This effect can be blocked by specific NMDA-R antagonists.

• Sørvik, Irene Beate; Solum, Eirik Johansson; Labba, Nils Anders; Hansen, Trond Vidar & Paulsen, Ragnhild Elisabeth (2018). Differential effects of some novel synthetic oestrogen analogs on oxidative PC12 cell death caused by serum deprivation. Free radical research. ISSN 1071-5762. 52(2), p. 273–287. doi: 10.1080/10715762.2018.1430363. Full text in Research Archive
• Myhre, Oddvar; Låg, Marit; Villanger, Gro Dehli; Oftedal, Bente Margaret; Øvrevik, Johan & Holme, Jørn Andreas [Show all 10 contributors for this article] (2018). Early life exposure to air pollution particulate matter (PM) as risk factor for attention deficit/hyperactivity disorder (ADHD): Need for novel strategies for mechanisms and causalities. Toxicology and Applied Pharmacology. ISSN 0041-008X. 354, p. 196–214. doi: 10.1016/j.taap.2018.03.015. Full text in Research Archive
• Stahl, Katja; Rahmani, Soulmaz; Prydz, Agnete; Skauli, Nadia; MacAulay, Nanna & Mylonakou, Maria-Niki [Show all 13 contributors for this article] (2018). Targeted deletion of the aquaglyceroporin AQP9 is protective in a mouse model of Parkinson's disease. PLOS ONE. 13(3). doi: 10.1371/journal.pone.0194896. Full text in Research Archive Show summary

  More than 90% of the cases of Parkinson’s disease have unknown etiology. Gradual loss of dopaminergic neurons of substantia nigra is the main cause of morbidity in this disease. External factors such as environmental toxins are believed to play a role in the cell loss, although the cause of the selective vulnerability of dopaminergic neurons remains unknown. We have previously shown that aquaglyceroporin AQP9 is expressed in dopaminergic neurons and astrocytes of rodent brain. AQP9 is permeable to a broad spectrum of substrates including purines, pyrimidines, and lactate, in addition to water and glycerol. Here we test our hypothesis that AQP9 serves as an influx route for exogenous toxins and, hence, may contribute to the selective vulnerability of nigral dopaminergic (tyrosine hydroxylase-positive) neurons. Using Xenopus oocytes injected with Aqp9 cRNA, we show that AQP9 is permeable to the parkinsonogenic toxin 1-methyl-4-phenylpyridinium (MPP+). Stable expression of AQP9 in HEK cells increases their vulnerability to MPP+ and to arsenite—another parkinsonogenic toxin. Conversely, targeted deletion of Aqp9 in mice protects nigral dopaminergic neurons against MPP+ toxicity. A protective effect of Aqp9 deletion was demonstrated in organotypic slice cultures of mouse midbrain exposed to MPP+ in vitro and in mice subjected to intrastriatal injections of MPP+ in vivo. Seven days after intrastriatal MPP+ injections, the population of tyrosine hydroxylase-positive cells in substantia nigra is reduced by 48% in Aqp9 knockout mice compared with 67% in WT littermates. Our results show that AQP9 –selectively expressed in catecholaminergic neurons—is permeable to MPP+ and suggest that this aquaglyceroporin contributes to the selective vulnerability of nigral dopaminergic neurons by providing an entry route for parkinsonogenic toxins. To our knowledge this is the first evidence implicating a toxin permeable membrane channel in the pathophysiology of Parkinson’s disease.

• Gervin, Kristina; Nordeng, Hedvig Marie Egeland; Eskeland, Ragnhild; Paulsen, Ragnhild Elisabeth & Lyle, Robert (2017). Farmakoepigenetikk: Samspillet mellom legemidler og epigenetikk. Norsk Farmaceutisk Tidsskrift (NFT). ISSN 0029-1935. 125(8), p. 30–34. Full text in Research Archive Show summary

  Epigenetikk er læren om arvelige mekanismer som regulerer aktiviteten til genene i ulike celler og vev som ikke involverer en endring i selve DNA-sekvensen. Om gener er skrudd av eller på er bestemt av epigenetiske merkelapper festet til det enkelte genet. Hensikten med denne artikkelen er å gi en kortfattet introduksjon til epigenetikk og farmakoepigenetikk. Epigenetiske faktorer antas å være viktige i utviklingen av en rekke sykdommer, som visse kreftformer og utviklingsforstyrrelser i nervesystemet. Det finnes allerede legemidler på markedet som virker via epigenetiske mekanismer ved å påvirke det epigenetiske maskineriet som fester de epigenetiske merkelappene til DNA-et, eller proteiner som DNA-et er kveilet rundt. Spesielt innen kreftmedisin satses det stort på utvikling av legemidler som kan normalisere avvikende genuttrykk via epigenetiske mekanismer. Lite er kjent om legemidler kan gi opphav til bivirkninger via epigenetiske mekanismer.

• Sørvik, Irene Beate & Paulsen, Ragnhild Elisabeth (2017). High and low concentration of 17?-estradiol protect cerebellar granule neurons in different time windows. Biochemical and Biophysical Research Communications - BBRC. ISSN 0006-291X. 490(3), p. 676–681. doi: 10.1016/j.bbrc.2017.06.100. Full text in Research Archive Show summary

  17α-estradiol is a hormonally inactive isomer of 17β-estradiol, but with similar potency as neuroprotector. However, we have previously reported that pretreatment with high concentration (10 μM) of both estrogens abolishes their neuroprotection in rat cerebellar granule neurons. Here, we have examined neuroprotective properties of 17α-estradiol against glutamate-induced excitotoxicity in chicken cerebellar granule neurons using low (1 nM) and high concentration.17α-Estradiol, 1 nM, was neuroprotective when glutamate was administered after a pretreatment period of 24 h, but not when coadministered with glutamate. In contrast, 10 μM was protective when coadministered with glutamate, but was not efficient when glutamate was administered after a pretreatment period. The difference in protection was linked to a stronger calcium response during glutamate exposure in the non-protective treatments. In addition, the pretreatment period with 10 μM was accompanied by increased protein level of the N-methyl-d-aspartate receptor subunit NR2B and reduced glutathione level. Thus, 17α-estradiol has a concentration and time dependent protective effect against glutamate-induced cell death.

• Bjørnstad, Sigrid; Samara, Athina; Erichsen, Aage; Paulsen, Ragnhild Elisabeth; Glover, Joel & Roald, Borghild (2016). Hampered lung maturation in methimazole-induced hypothyroidism in fetal chicken: morphological and molecular correlates to human fetal development. Neonatology. ISSN 1661-7800. 110(2), p. 83–92. doi: 10.1159/000444656. Full text in Research Archive
• Austdal, Lars Peter Engeset; Bjørnstad, Sigrid; Mathisen, Gro Haarklou; Aden, Petra; Mikkola, Ingvild & Paulsen, Ragnhild Elisabeth [Show all 7 contributors for this article] (2016). Glucocorticoid effects on cerebellar development in a chicken embryo model: exploring changes in PAX6 and Metalloproteinase-9 after exposure to dexamethasone. Journal of Neuroendocrinology (JNE). ISSN 0953-8194. 28(12). doi: 10.1111/jne.12438. Full text in Research Archive
• Austdal, Lars Peter Engeset; Mathisen, Gro Haarklou; Løberg, Else Marit & Paulsen, Ragnhild Elisabeth (2016). Calcium-induced apoptosis of developing cerebellar granule neurons depends causally on NGFI-B. International Journal of Developmental Neuroscience. ISSN 0736-5748. 55, p. 82–90. doi: 10.1016/j.ijdevneu.2016.10.003. Full text in Research Archive
• Bjørnstad, Sigrid; Austdal, Lars Peter Engeset; Roald, Borghild; Glover, Joel & Paulsen, Ragnhild Elisabeth (2015). Cracking the egg: potential of the developing chicken as a model system for nonclinical safety studies of pharmaceuticals. Journal of Pharmacology and Experimental Therapeutics. ISSN 0022-3565. 355, p. 386–396. doi: 10.1124/jpet.115.227025. Full text in Research Archive
• Yazdani, Mazyar; Paulsen, Ragnhild Elisabeth; Gjøen, Tor & Hylland, Ketil (2015). Reactive Oxygen Species and Cytotoxicity in Rainbow Trout Hepatocytes: Effects of Medium and Incubation Time. Bulletin of Environmental Contamination and Toxicology. ISSN 0007-4861. 94, p. 193–198. doi: 10.1007/s00128-014-1433-0. Full text in Research Archive
• Rakkestad, Kirsten Eline; Sørvik, Irene Beate; Øverby, Gunn Rebecca; Debernard, Karen A. Boldingh; Mathisen, Gro Haarklou & Paulsen, Ragnhild Elisabeth (2014). 17a-estradiol down-regulates glutathione synthesis in serum deprived PC-12 cells. Free radical research. ISSN 1071-5762. 48(10), p. 1170–1178. doi: 10.3109/10715762.2014.930455. Full text in Research Archive
• Solum, Eirik Johansson; Cheng, Jing-Jy; Sørvik, Irene Beate; Paulsen, Ragnhild Elisabeth; Vik, Anders & Hansen, Trond Vidar (2014). Synthesis and biological evaluations of new analogs of 2-methoxyestradiol: Inhibitors of tubulin and angiogenesis. European Journal of Medicinal Chemistry. ISSN 0223-5234. 85, p. 391–398. doi: 10.1016/j.ejmech.2014.08.002. Full text in Research Archive
• Bjørnstad, Sigrid; Paulsen, Ragnhild Elisabeth; Erichsen, Aage; Glover, Joel & Roald, Borghild (2014). Type I and II pneumocyte differentiation in the developing fetal chicken lung: Conservation of pivotal proteins from birds to human in the struggle for life at birth. Neonatology. ISSN 1661-7800. 105(2), p. 112–120. doi: 10.1159/000355346. Full text in Research Archive
• Aziz, Gulzeb; Odlo, Kristin; Hansen, Trond Vidar; Paulsen, Ragnhild Elisabeth & Mathisen, Gro Haarklou (2013). Combretastatin A-4 and structurally related triazole analogues induce caspase-3 and reactive oxygen species-dependent cell death in PC12 cells. European Journal of Pharmacology. ISSN 0014-2999. 703(1-3), p. 25–32. doi: 10.1016/j.ejphar.2013.01.043. Full text in Research Archive
• Åstrand, Ove Alexander; Aziz, Gul zeb; Ali, Sidra Farzand; Paulsen, Ragnhild Elisabeth; Hansen, Trond Vidar & Rongved, Pål (2013). Synthesis and initial in vitro biological evaluation of two new zinc-chelating compounds: Comparison with TPEN and PAC-1. Bioorganic & Medicinal Chemistry. ISSN 0968-0896. 21(17), p. 5175–5181. doi: 10.1016/j.bmc.2013.06.037. Full text in Research Archive
• Åstrand, Ove Alexander Høgmoen; Austdal, Lars Peter Engeset; Paulsen, Ragnhild Elisabeth; Hansen, Trond Vidar & Rongved, Pål (2013). Synthesis and characterization of new selective Zn2+ fluorescent probes for functionalization: in vitro Cell imaging applications. Tetrahedron. ISSN 0040-4020. 49(40), p. 8645–8654. doi: 10.1016/j.tet.2013.07.049. Full text in Research Archive
• Mathisen, Gro Haarklou; Yazdani, Mazyar; Rakkestad, Kirsten Eline; Aden, Petra; Bodin, Johanna & Samuelsen, Mari [Show all 13 contributors for this article] (2013). Prenatal exposure to bisphenol A interferes with the development of cerebellar granule neurons in mice and chicken. International Journal of Developmental Neuroscience. ISSN 0736-5748. 31(8), p. 762–769. doi: 10.1016/j.ijdevneu.2013.09.009. Full text in Research Archive
• Strøm, Bjørn Oddvar & Paulsen, Ragnhild Elisabeth (2012). Apoptosis inducer NGFI-B is degraded by the proteasome and stabilized by treatment with EGF. Biochemical and Biophysical Research Communications - BBRC. ISSN 0006-291X. 417(4), p. 1292–1297. doi: 10.1016/j.bbrc.2011.12.132. Full text in Research Archive
• Bølling, Anette Kocbach; Øvrevik, Johan; Samuelsen, Jan Tore; Holme, Jørn Andreas; Rakkestad, Kirsten Eline & Mathisen, Gro Haarklou [Show all 9 contributors for this article] (2012). Mono-2-ethylhexylphthalate (MEHP) induces TNF-alpha release and macrophage differentiation through different signalling pathways in RAW264.7 cells. Toxicology Letters. ISSN 0378-4274. 209(1), p. 43–50. doi: 10.1016/j.toxlet.2011.11.016. Full text in Research Archive
• Debernard, Karen A. Boldingh; Mathisen, Gro Haarklou & Paulsen, Ragnhild Elisabeth (2012). Differences in NGFI-B, Nurr1, and NOR-1 expression and nucleocytoplasmic translocation in glutamate-treated neurons. Neurochemistry International. ISSN 0197-0186. 61(1), p. 79–88. doi: 10.1016/j.neuint.2012.04.002. Full text in Research Archive
• Mathisen, Gro Haarklou; Fallgren, Åsa Birgitta; Strøm, Bjørn Oddvar; Debernard, Karen A. Boldingh; Mohebi, Beata Urbanczyk & Paulsen, Ragnhild Elisabeth (2011). Delayed translocation of NGFI-B/RXR in glutamate stimulated neurons allows late protection by 9-cis retinoic acid. Biochemical and Biophysical Research Communications - BBRC. ISSN 0006-291X. 414(1), p. 90–95. doi: 10.1016/j.bbrc.2011.09.028. Full text in Research Archive
• Debernard, Karen A. Boldingh; Aziz, Gul zeb; Gjesvik, Annine Thomassen & Paulsen, Ragnhild Elisabeth (2011). Cell death induced by novel procaspase-3 activators can be reduced by growth factors. Biochemical and Biophysical Research Communications - BBRC. ISSN 0006-291X. 413(2), p. 364–369. doi: 10.1016/j.bbrc.2011.08.107. Full text in Research Archive
• Aden, Petra; Paulsen, Ragnhild Elisabeth; Mæhlen, Jan; Løberg, Else Marit; Goverud, Ingeborg Løstegaard & Liestøl, Knut [Show all 7 contributors for this article] (2011). Glucocorticoids dexamethasone and hydrocortisone inhibit proliferation and accelerate maturation of chicken cerebellar granule neurons. Brain Research. ISSN 0006-8993. 1418, p. 32–41. doi: 10.1016/j.brainres.2011.08.053. Full text in Research Archive
• Aziz, Gulzeb; Akselsen, Øyvind Wilhelm; Hansen, Trond Vidar & Paulsen, Ragnhild Elisabeth (2010). Procaspase-activating compound 1 induces a caspase-3-dependent cell death in cerebellar granule neurons. Toxicology and Applied Pharmacology. ISSN 0041-008X. 247, p. 238–242. Full text in Research Archive
• Rakkestad, Kirsten Eline; Holme, Jørn Andreas; Paulsen, Ragnhild E; Schwarze, Per E & Becher, Rune (2010). Mono(2-ethylhexyl) phthalate induces both pro- and anti-inflammatory responses in rat alveolar macrophages through crosstalk between p38, the lipoxygenase pathway and PPAR alpha. Inhalation Toxicology. ISSN 0895-8378. 22(2), p. 140–150. doi: 10.3109/08958370903019885. Full text in Research Archive
• Strøm, Bjørn Oddvar; Aden, Petra; Mathisen, Gro Haarklou; Lømo, Jon; Davanger, Svend & Paulsen, Ragnhild Elisabeth (2010). Transfection of chicken cerebellar granule neurons used to study glucocorticoid receptor regulation by nuclear receptor 4A (NR4A). Journal of Neuroscience Methods. ISSN 0165-0270. 193(1), p. 39–46. doi: 10.1016/j.jneumeth.2010.08.012. Full text in Research Archive
• Aden, Petra; Goverud, Ingeborg Løstegaard; Liestøl, Knut; Løberg, Else Marit; Paulsen, Ragnhild Elisabeth & Maehlen, Jan [Show all 7 contributors for this article] (2008). Low-potency glucocorticoid hydrocortisone has similar neurotoxic effects as high-potency glucocorticoid dexamethasone on neurons in the immature chicken cerebellum. Brain Research. ISSN 0006-8993. 1236, p. 39–48. doi: 10.1016/j.brainres.2008.07.095. Full text in Research Archive
• Maiga, Ababacar Ibrahim; Malterud, Karl Egil; Mathisen, Gro Haarklou; Paulsen, Ragnhild Elisabeth; Thomas-Oates, Jane & Bergström, Ed [Show all 9 contributors for this article] (2007). Cell protective antioxidants from the root bark of Lannea velutina A. Rich., a Malian medicinal plant. Journal of Medicinal Plants Research. 1(4), p. 66–79. Full text in Research Archive
• Mathisen, Gro Anita; Thorkildsen, Inger Helene & Paulsen, Ragnhild Elisabeth (2007). Secretory PLA(2)-IIA and ROS generation in peripheral mitochondria are critical for neuronal death. Brain Research. ISSN 0006-8993. 1153, p. 43–51. doi: 10.1016/j.brainres.2007.03.067. Full text in Research Archive Show summary

  In this study the role of mitochondrial secretory PLA2-IIA in glutamate-induced cell death in cultured cerebellar granule neurons has been investigated. Inhibition of secretory PLA2-IIA blocked glutamate-induced cell death. Since PLA2 may generate reactive oxygen species (ROS), we have investigated ROS production, detected as dihydrorhodamine 123 oxidation and nitrotyrosine modifications of proteins, following glutamate-treatment in the absence or presence of an inhibitor of secretory PLA2-IIA. There was an increased generation of ROS in both glutamate- and buffer-treated neurons compared to untreated neurons. Scavenging with dihydrorhodamine 123 reduced glutamate-induced death (60 %), showing that ROS detected in glutamate-treated neurons were associated with cell death. However, ROS detected in buffer-treated neurons were not associated with toxicity. Glutamate-treatment led to ROS production predominantly in peripheral mitochondria, whereas buffer-treatment led to ROS production in somal mitochondria. Inhibition of secretory PLA2-IIA i) reduced the generation of ROS after glutamate- treatment, ii) reduced the ROS production in peripheral mitochondria in glutamate-treated neurons, consistent with the fact that calcium entry through glutamate (NMDA) receptors has a privileged access to peripheral mitochondria, and iii) did not reduce the generation of ROS after buffer-treatment. In conclusion, activation of NMDA receptors induces ROS which is critical for neuronal death, due to secretory PLA2-IIA associated with peripheral mitochondria.

• Fallgren, Åsa Birgitta; Mathisen, Gro Anita; Mæhlen, Jan; Blomhoff, Rune & Paulsen, Ragnhild Elisabeth (2007). Preconditioning with estradiol abolishes its neuroprotection in cerebellar neurons. Biochemical and Biophysical Research Communications - BBRC. ISSN 0006-291X. 352, p. 966–972. doi: 10.1016/j.bbrc.2006.11.139. Full text in Research Archive
• Jacobs, Chris; Aden, Petra Käte; Mathisen, Gro Anita; Khuong, Erica; Gaarder, Mona & Løberg, Else Marit [Show all 9 contributors for this article] (2006). Chicken cerebellar granule neurons rapidly develop excitotoxicity in culture. Journal of Neuroscience Methods. ISSN 0165-0270. 156, p. 129–135. Full text in Research Archive Show summary

  Rat cerebellar granule cell culture is widely used as a model to study factors that control neuronal differentiation and death (e.g. excitotoxicity). However, a main drawback of this model is its dependence on depolarizing culture condition (25 mM potassium). In addition, it is quite expensive to maintain and requires animal facilities. Here we report that cerebellar granule neuron cultures from chicken may be used as an alternative model to study excitotoxicity. Surprisingly, fetal chicken cells may be grown in a physiological potassium concentration (5 mM potassium). They develop excitotoxicity rapidly in culture (fully developed at 3 days in vitro), and respond to glutamate excitotoxicity similar to rat cultures (ROS production and activation of caspase-3).

• Jacobs, Chris; Trinh, Minh D.; Rootwelt, Terje; Lømo, Jon & Paulsen, Ragnhild Elisabeth (2006). Dexamethasone induces cell death which may be blocked by NMDA receptor antagonists but is insensitive to Mg2+ in cerebellar granule neurons. Brain Research. ISSN 0006-8993. 1070, p. 116–123. doi: 10.1016/j.brainres.2005.10.093. Full text in Research Archive Show summary

  Since dexamethasone may elevate the Ca2+ influx through NMDA receptors, we have investigated mechanisms of dexamethasone toxicity in rat cerebellar granule neurons. Dexamethasone concentrations over 0.1 μM induced cell death that reached about 20% of the death induced by glutamate. Dexamethasone-induced cell death was reduced by more than 80% by the mineralocorticoid antagonist RU 28318 or the NMDA receptor antagonists MK 801 and CGP 39551, whereas RU 28318 rescued only 30% of cells treated with glutamate, indicating that dexamethasone requires NMDA receptors to induce acute neuronal toxicity and that a fraction of the neurons showed this toxicity. Mg2+ reduced the cell death induced by glutamate at potassium concentrations of 1 mM and 5 mM, but not at 25 mM. In contrast, cell death induced by dexamethasone was not significantly reduced by Mg2+ in any of the potassium concentrations. Both glutamate and dexamethasone induced toxicity with translocation of the apoptosis inducer NGFI-B to the mitochondria seen after 30 min–2 h concomitant with activation of apoptosis inducing factor (AIF) and caspase-3. In conclusion, dexamethasone induces a rapid toxicity which is blocked by NMDA receptor antagonists other than Mg2+, and involves mitochondrial apoptosis inducer NGFI-B.

• Hankø, Erik; Hansen, Thor Willy; Almaas, Runar; Paulsen, Ragnhild Elisabeth & Rootwelt, Terje (2006). Synergistic Protection of a General Caspase Inhibitor and MK-801 in Bilirubin-Induced Cell Death in Human NT2-N Neurons. Pediatric Research. ISSN 0031-3998. 59(1), p. 72–77. doi: 10.1203/01.pdr.0000191135.6358. Full text in Research Archive Show summary

  Unconjugated bilirubin (UCB) induces both apoptosis and necrosis in neurons. To investigate the role of caspases and excitotoxicity in UCB-induced cell death, we exposed NT2-N neurons to 5 µM UCB (a concentration known to induce apoptosis) or 2 µM staurosporine (positive apoptosis control) and investigated the effects of treatments with the specific caspase-3 inhibitor, zDEVD.FMK (20 and 100 µM), or the general caspase inhibitor, zVAD.FMK (20 and 100 µM), and/or the N-methyl-D-aspartate (NMDA) receptor antagonist MK-801 (10 µM) during a 24- or 48-h exposure. UCB increased caspase-3 activity 2.3-fold after 6 h. Despite this, treatment with zDEVD.FMK did not prevent cell death. zVAD.FMK (100 µM only) reduced apoptotic morphologies induced by UCB after 48 h, however, this effect was not reflected in MTT assays. MK-801 attenuated UCB-induced cell death after 24 h, but not significantly after 48 h. Similar effects were seen in the MTT assay. Combined treatment with MK-801 and zVAD.FMK synergistically reduced apoptotic morphologies after 24 and 48 h. MTT assays showed comparable effects after 24 h but did not show significant differences after 48 h. We conclude that NMDA receptor-mediated pathways and caspase-mediated pathways are involved in UCB-induced cell death in human NT2-N neurons. Concomitant inhibition of both pathways results in synergistic protection

• Rosseland, Carola; Wierød, Lene; Oksvold, M.P.; Werner, H; Østvold, Anne C. & Thoresen, G. Hege [Show all 9 contributors for this article] (2005). Cytoplasmic retention of peroxide-activated ERK provides survival in primary cultures of rat hepatocytes. Hepatology. ISSN 0270-9139. 42, p. 200–2007. Full text in Research Archive
• Jacobs, Chris & Paulsen, Ragnhild Elisabeth (2005). Crosstalk between ERK2 and RXR regulates nuclear import of transcription factor NGFI-B. Biochemical and Biophysical Research Communications - BBRC. ISSN 0006-291X. 336, p. 646–652. doi: 10.1016/j.bbrc.2005.08.143. Full text in Research Archive
• Jacobs, Chris; Boldingh, Karen Astrid; Slagsvold, Hege Holte; Thoresen, Gunn Hege & Paulsen, Ragnhild Elisabeth (2004). ERK2 prohibits apoptosis-induced subcellular translocation of orphan nuclear receptor NGFI-B/TR3. Journal of Biological Chemistry. ISSN 0021-9258. 279(48), p. 50097–50101. Full text in Research Archive
• Slagsvold, Hege H.; Rosseland, Carola; Jacobs, Chris; Khuong, Erica; Gaarder, Mona & Fallgren, Åsa Birgitta [Show all 8 contributors for this article] (2003). High molecular weight DNA fragments are processed by caspase sensitive or caspase independent pathways in cultures of cerebellar granule neurons. Brain Research. ISSN 0006-8993. 984, p. 111–121. Full text in Research Archive
• Slagsvold, Hege H.; Østvold, Anne Carine; Fallgren, Åsa Birgitta & Paulsen, Ragnhild Elisabeth (2002). Nuclear receptor and apoptosis initiator NGFI-B is a substrate for kinase ERK2. Biochemical and Biophysical Research Communications - BBRC. ISSN 0006-291X. 291, p. 1146–1150. Full text in Research Archive
• Klungland, Arne; Paulsen, Ragnhild Elisabeth; Rolseth, Veslemøy; Yamada, Y.; Ueno, Y. & Wiik, Pål [Show all 9 contributors for this article] (2001). 5-Formyluracil and its nucleoside derivatives confer toxicity and mutagenicity to mammalian cells by interfering with normal RNA and DNA metabolism. Toxicology Letters. ISSN 0378-4274. 119, p. 71–78. Full text in Research Archive
• Oksvold, Morten Pedersen; Skarpen, Ellen; Wierød, Lene; Paulsen, Ragnhild Elisabeth & Huitfeldt, Henrik (2001). Re-localization of activated EGF receptor and its signal transducers to multivesicular compartments downstream of early endosomes in response to EGF. European Journal of Cell Biology. ISSN 0171-9335. 80(4), p. 285–294. Full text in Research Archive
• Paulsen, Ragnhild Elisabeth & Ottersen, Ole Petter (2000). Glutamate, apoptose og nekrose i sentralnervesystemet. Tidsskrift for Den norske legeforening (Tidsskriftet). ISSN 0029-2001. 120, p. 63–64. Full text in Research Archive
• Paulsen, Ragnhild Elisabeth & Ottersen, Ole Petter (2000). Glutamat, apoptose og nekrose i nervesystemet. Tidsskrift for Den norske legeforening (Tidsskriftet). ISSN 0029-2001. 120, p. 63–64. Full text in Research Archive
• Slagsvold, Hege H.; Marvik, Ole-Jørgen; Eidem, Guro; Kristoffersen, Nina & Paulsen, Ragnhild Elisabeth (2000). Detection of high molecular weight DNA fragments characteristic of early stage apoptosis in cerebellar granule cells exposed to glutamate. Experimental Brain Research. ISSN 0014-4819. 135, p. 173–178. Full text in Research Archive
• Olsen, Christel; Rustad, Anne; Fonnum, Frode; Paulsen, Ragnhild Elisabeth & Hassel, Bjørnar (1999). 3-nitropropionic acid: an astrocyte-sparing neurotoxin in vitro. Brain Research. ISSN 0006-8993. 850, p. 144–149. Full text in Research Archive
• Olsen, Christel Moræus; Rustad, Anne; Paulsen, Ragnhild Elisabeth Heimtun & Hassel, Bjørnar (1999). 3-Nitropropionic acid: an astrocyte-sparing neurotoxin in vitro. Brain Research. ISSN 0006-8993. doi: 10.1016/s0006-8993(99)02115-0. Full text in Research Archive
• Ciani, Elisabetta; Rizzi, Simona; Paulsen, Ragnhild Elisabeth & Contestabile, Antonio (1997). Chronic pre-explant blockade of the NMDA receptor affects survival of cerebellar granule cells explanted in vitro. Developmental Brain Research. ISSN 0165-3806. 99, p. 112–117. Full text in Research Archive
• Myhrer, Trond & Paulsen, Ragnhild Elisabeth (1997). Infusion of D-cycloserine into temporal-hippocampal areas and restoration of mnemonic function in rats with disrupted glutamatergic temporal systems. European Journal of Pharmacology. ISSN 0014-2999. 328, p. 1–7. Full text in Research Archive
• Paulsen, Ragnhild Elisabeth; Bardal, Kjersti Granås; Johnsen, Helge; Rolseth, Veslemøy & Sterri, Sigrun Hanne (1995). Three related brain nuclear receptors, NGFI-B, Nurr1, and NOR-1, as transcriptional activators. Journal of Molecular Neuroscience. ISSN 0895-8696. 6, p. 249–255. Full text in Research Archive
• Hassel, Bjørnar; Paulsen, Ragnhild Elisabeth Heimtun; Hamre, Ragnhild Paulsen; Johnsen, Arnt Magne & Fonnum, Frode (1992). Selective inhibition of glial cell metabolism in vivo by fluorocitrate. Brain Research. ISSN 0006-8993. doi: 10.1016/0006-8993(92)90616-h. Full text in Research Archive

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• Rakkestad, Kirsten Eline; Dahl, Jon Einar; Paulsen, Ragnhild Elisabeth; Låg, Marit & Samuelsen, Jan Tore (2017). Do silica nanoparticles induce oxidative stress by different mechanisms in different mammalian cell lines? Full text in Research Archive
• Treimo, Alexandra Isabel Sveinsen; Borgå, Katrine; Samuelsen, Jan Tore; Paulsen, Ragnhild Elisabeth & Rakkestad, Kirsten Eline (2017). Are nanoparticles used in dental materials neurotoxic? Full text in Research Archive
• Hadera, Mussie Ghezu; Andersen, Jannike Mørch & Paulsen, Ragnhild Elisabeth (2017). Effects of methadone in the developing chicken embryo as a model of neurodevelopmental toxicity. Reproductive Toxicology. ISSN 0890-6238. 72, p. 216–217. doi: 10.1016/j.reprotox.2017.06.030. Full text in Research Archive
• Yazdani, Mazyar; Paulsen, Ragnhild Elisabeth; Gjøen, Tor & Hylland, Ketil (2013). Reactive oxygen species and cytotoxicity in rainbow trout hepatocytes: effects of medium and incubation time. Full text in Research Archive
• Yazdani, Mazyar; Paulsen, Ragnhild Elisabeth; Gjøen, Tor & Hylland, Ketil (2011). Comparison of three fluorescence probes for ROS detection in primary cultures of rainbow trout hepatocytes. Full text in Research Archive
• Yazdani, Mazyar; Ellesat, Kathrin; Paulsen, Ragnhild Elisabeth & Hylland, Ketil (2010). Developing a fish hepatocyte model for oxidative stress studies. Full text in Research Archive
• Moen, Lars Fredrik; Movig, Trude; Jacobs, Chris; Debernard, Karen Boldingh; Mathisen, Gro Anita & Paulsen, Ragnhild Elisabeth (2007). Nuclear import and transcriptional activity of NGFI-B - effects of ATP, RXRa, RARa, 9cRA, ATRA, and TTNPB. Full text in Research Archive
• Debernard, Karen A. Boldingh; Strøm, Bjørn Oddvar; Jacobs, Chris & Paulsen, Ragnhild Elisabeth (2007). EGF Protects Neurons From Glutamate-Induced Cell Death by Erk2-Mediated Nuclear Targeting of Apoptosis-Inducer NGFI-B/TR3. Full text in Research Archive
• Debernard, Karen A. Boldingh; Strøm, Bjørn Oddvar; Jacobs, Chris & Paulsen, Ragnhild Elisabeth (2007). Vekstfaktoren EGF beskytter nerveceller mot død ved Erk2-mediert kjernelokalisering av apoptose-inducer NGFI-B. Full text in Research Archive
• Mathisen, Gro Anita; Fallgren, Åsa Birgitta; Gaarder, Mona; Mæhlen, Jan; Blomhoff, Rune & Paulsen, Ragnhild Elisabeth (2007). Preconditioning with estradiol abolishes its neuroprotection in cerebellar granule neurons. Full text in Research Archive
• Debernard, Karen A. Boldingh; Strøm, Bjørn Oddvar; Jacobs, Chris & Paulsen, Ragnhild Elisabeth (2007). EGF protects neurons from cell death by ERK2-mediated nuclear targeting of apoptosis-inducer NGFI-B/TR3. Full text in Research Archive
• Debernard, Karen Boldingh; Jacobs, Chris M & Paulsen, Ragnhild Elisabeth (2006). Activation of ERK2 leads to nuclear targeting of apoptosis inducer NGFI-B in cerebellar granule neurons. Full text in Research Archive
• Mathisen, Gro Anita & Paulsen, Ragnhild Elisabeth (2006). Different reactive oxygen species are critical for toxicity in two cell death models in cerebellar granule neurons. Free radical research. ISSN 1071-5762. 40, p. S91–S91. Full text in Research Archive
• Aden, Petra; Jacobs, Chris; Minh, D. Trinh; Rootwelt, Terje; Lømo, Jon & Paulsen, Ragnhild Elisabeth (2006). Dexamethasone induces apoptotic cell death through NMDA receptor activation in cultured cerebellar granule cells. Full text in Research Archive
• Debernard, Karen Boldingh; Jacobs, Chris; Thoresen, G. Hege & Paulsen, Ragnhild Elisabeth (2006). EGF protects cerebellar granule neurons from glutamate-induced cell death by ERK2-mediated inhibition of mitochondrial targeting of orphan nuclear receptor NGFI-B/TR3. Full text in Research Archive
• Mathisen, Gro Anita & Paulsen, Ragnhild Elisabeth (2006). Different reactive oxygen species are critical for toxicity in two cell death models in cerebellar granule neurons. Full text in Research Archive
• Aden, Petra; Jacobs, Chris; Khuong, Erica; Goverud, Ingeborg Løstegaard; Løberg, Else Marit & Paulsen, Ragnhild Elisabeth [Show all 8 contributors for this article] (2004). Neurotoxicological studies in fetal chicken: Effect of glucocorticoids on apoptosis in cerebellar granule cells in vivo and in vitro. Full text in Research Archive
• Lindgren, Carola; Holte, Hege Ruge; Solberg, Kristin & Paulsen, Ragnhild Elisabeth (2000). Caspase 3, enzyme activity and level of expression, in rat cerebellar granule cells after glutamate exposure. Full text in Research Archive Show summary

  Caspase 3, enzyme activity and level of expression, in rat cerebellar granule cells after glutamate exposure. C. Lindgren, H. Holte, K. Solberg, R. Paulsen. Institute of Pharmacy, University of Oslo. Norway. The enzyme activity of the apoptotic marker Caspase 3 was investigated in rat cerebellar granule neurons after sublethal glutamate exposure as compared to the Caspase 3 activity in two different cell deaths models: cell death induced by K+/serum deprivation, previously described to induce apoptosis, and calcium-ionophore, known to induce necrosis. The enzyme activity was analysed at different time-points after treatment, using the fluorogenic Caspase 3 substrate Ac-DEVD-AMC. The enzyme activity of Caspase 3 showed a many-fold and prolonged increase following K+/serum deprivation that preceded the morphological cell loss, whereas calcium-ionophore induced a rapid loss of viability but induced no increase in Caspase 3 activity. Highly toxic glutamate exposure has earlier been described to induce necrosis with no detectable increase in Caspase 3 activity (Armstrong et al. J.Neurosci.17(2):253-6, 1997), whereas a less severe, but still toxic, insult of glutamate induces apoptosis with highly increased Caspase 3 activity (Du et al. Proc. Natl. Acad. Sci. USA, 94:11657-62, 1997). Interestingly, a sublethal exposure of glutamate induced a significant decrease in Caspase 3 activity compared to control 6 hours after treatment. The mRNA-level of caspase 1 and caspase 3 in cerebellar granule neurons following lethal glutamate exposure was also analysed at different time-points, using RT-PCR. Caspase 3 mRNA was significantly increased following toxic glutamate exposure at 6 hours after treatment, whereas caspase 1 showed an attenuated mRNA level at all time-points.

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