A locust embryo as predictive developmental neurotoxicity testing system for pioneer axon pathway formation

Bode, Karsten; Bohn, Maja; Reitmeier, Jennifer; Betker, Philine; Stern, Michael; Bicker, Gerd

doi:10.1007/s00204-020-02929-6

“…(j) The inhibitory effect of 15 µM fipronil sulfone was alleviated by coapplication of the rho kinase inhibitor, Y-27632, in a dose-dependent manner (EC50 4.22 µM) differentiation reported by Pamies et al (2018) and Pistollato et al (2017). On the other hand, in an in vivo setting, rotenone specifically inhibited directed neurite growth in an intact insect embryo (Bergmann et al 2019;Bode et al 2020). A negative effect of rotenone on cell migration out of neurospheres has been reported by Ishido and Suzuki (2010) with an IC50 of ca.…”

Section: Discussionmentioning

confidence: 99%

“…These results are in line with the findings by Krug et al ( 2013 ) and Ryan et al ( 2016 ) with regard to neurite outgrowth and the effects on neuronal differentiation reported by Pamies et al ( 2018 ) and Pistollato et al ( 2017 ). On the other hand, in an in vivo setting, rotenone specifically inhibited directed neurite growth in an intact insect embryo (Bergmann et al 2019 ; Bode et al 2020 ). A negative effect of rotenone on cell migration out of neurospheres has been reported by Ishido and Suzuki ( 2010 ) with an IC50 of ca.…”

Section: Discussionmentioning

confidence: 99%

Developmental Neurotoxicity of Fipronil and Rotenone on a Human Neuronal In Vitro Test System

Schmitz¹,

Dempewolf²,

Tan³

et al. 2021

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Pesticide exposure during in utero and early postnatal development can cause a wide range of neurological defects. However, relatively few insecticides have been recognized as developmental neurotoxicants, so far. Recently, discovery of the insecticide, fipronil, in chicken eggs has raised public concern. The status of fipronil as a potential developmental neurotoxicant is still under debate. Whereas several in vivo and in vitro studies suggest specific toxicity, other in vitro studies could not confirm this concern. Here, we tested fipronil and its main metabolic product, fipronil sulfone both at concentrations between 1.98 and 62.5 µM, alongside with the established developmental neurotoxicant, rotenone (0.004–10 µM) in vitro on the human neuronal precursor cell line NT2. We found that rotenone impaired all three tested DNT endpoints, neurite outgrowth, neuronal differentiation, and precursor cell migration in a dose-dependent manner and clearly separable from general cytotoxicity in the nanomolar range. Fipronil and fipronil sulfone specifically inhibited cell migration and neuronal differentiation, but not neurite outgrowth in the micromolar range. The rho-kinase inhibitor Y-27632 counteracted inhibition of migration for all three compounds (EC50 between 12 and 50 µM). The antioxidant, n-acetyl cysteine, could ameliorate the inhibitory effects of fipronil on all three tested endpoints (EC 50 between 84 and 164 µM), indicating the involvement of oxidative stress. Fipronil sulfone had a stronger effect than fipronil, confirming the importance to test metabolic products alongside original pesticides. We conclude that in vitro fipronil and fipronil sulfone display specific developmental neurotoxicity on developing human model neurons.

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“…The genome of the migratory locust had been sequenced a few years ago 1 which, alongside advancements in genome editing like CRISPR-Cas9, rekindled the interest in research regarding the locust olfactory system 52 . Moreover, recent investigations introduced locusts as helpful organisms for research on axonal regeneration in the olfactory system 53 and as predictive test system for developmental neurotoxicity in humans 54 , 55 . Both lines of research will require information how locust neurons exposed to axonal injury or developmental-toxicants respond to these challenges in terms of Ca 2+ concentration changes.…”

Section: Discussionmentioning

confidence: 99%

Cholinergic calcium responses in cultured antennal lobe neurons of the migratory locust

Bergmann¹,

Bicker²

2021

Sci Rep

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Locusts are advantageous organisms to elucidate mechanisms of olfactory coding at the systems level. Sensory input is provided by the olfactory receptor neurons of the antenna, which send their axons into the antennal lobe. So far, cellular properties of neurons isolated from the circuitry of the olfactory system, such as transmitter-induced calcium responses, have not been studied. Biochemical and immunocytochemical investigations have provided evidence for acetylcholine as classical transmitter of olfactory receptor neurons. Here, we characterize cell cultured projection and local interneurons of the antennal lobe by cytosolic calcium imaging to cholinergic stimulation. We bulk loaded the indicator dye Cal-520 AM in dissociated culture and recorded calcium transients after applying cholinergic agonists and antagonists. The majority of projection and local neurons respond with increases in calcium levels to activation of both nicotinic and muscarinic receptors. In local interneurons, we reveal interactions lasting over minutes between intracellular signaling pathways, mediated by muscarinic and nicotinic receptor stimulation. The present investigation is pioneer in showing that Cal-520 AM readily loads Locusta migratoria neurons, making it a valuable tool for future research in locust neurophysiology, neuropharmacology, and neurodevelopment.

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“…The genome of the migratory locust had been sequenced a few years ago 1 which, alongside advancements in genome editing like CRISPR-Cas9, rekindled the interest in research regarding the locust olfactory system 51 . Moreover, recent investigations introduced locusts as helpful organisms for research on axonal regeneration in the olfactory system 52 and as predictive test system for developmental neurotoxicity in humans 53,54 . Both lines of research will require information how locust neurons exposed to axonal injury or developmental-toxicants respond to these challenges in terms of Ca 2+ concentration changes.…”

Section: Discussionmentioning

confidence: 99%

Cholinergic calcium responses in cultured antennal lobe neurons of the migratory locust

Bergmann

¹

2021

Preprint

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Locusts are advantageous organisms to elucidate mechanisms of olfactory coding at the systems level. Sensory input is provided by the olfactory receptor neurons of the antenna, which send their axons into the antennal lobe. So far, cellular properties of neurons isolated from the circuitry of the olfactory system, such as transmitter-induced calcium responses, have not been studied. Biochemical and immunocytochemical investigations have provided evidence for acetylcholine as classical transmitter of olfactory receptor neurons. Here, we characterize cell cultured projection and local interneurons of the antennal lobe by cytosolic calcium imaging to cholinergic stimulation. We bulk loaded the indicator dye Cal-520 AM in dissociated culture and recorded calcium transients after applying cholinergic agonists and antagonists. The majority of projection and local neurons respond with increases in calcium levels to activation of both nicotinic and muscarinic receptors. In local interneurons, we reveal interactions lasting over minutes between intracellular signaling pathways, mediated by muscarinic and nicotinic receptor stimulation. The present investigation is pioneer in showing that Cal-520 AM readily loads Locusta migratoria neurons, making it a valuable tool for future research in locust neurophysiology, neuropharmacology, and neurodevelopment.

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A locust embryo as predictive developmental neurotoxicity testing system for pioneer axon pathway formation

Cited by 6 publications

References 65 publications

Developmental Neurotoxicity of Fipronil and Rotenone on a Human Neuronal In Vitro Test System

Developmental Neurotoxicity of Fipronil and Rotenone on a Human Neuronal In Vitro Test System

Cholinergic calcium responses in cultured antennal lobe neurons of the migratory locust

Cholinergic calcium responses in cultured antennal lobe neurons of the migratory locust

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