Abstract
Objectives
The main function of myenteric neurons is the control of gut motility. As we recently showed that nitroxyl (HNO) induces intestinal smooth muscle relaxation, it was of interest to evaluate the effects of this signalling molecule on myenteric neurons in order to distinguish its properties in regard to myocytes.
Methods
Myenteric neurons isolated from the ileum of 4–10 days old rats were used. HNO-induced changes in intracellular concentration of Ca2+ or membrane potential and ion currents were measured using the Ca2+-sensitive fluorescent dye fura-2 AM or by electrophysiological whole-cell recordings, respectively. Changes in intracellular thiol groups pool were evaluated using thiol tracker violet. Angeli’s salt was used as HNO donor.
Results
The HNO donor Angeli’s salt induced a significant increase in the cytosolic Ca2+ concentration at the concentration 50 µM and a membrane hyperpolarization from a resting membrane potential of −56.1 ± 8.0 mV to −63.1 ± 8.7 mV (n=7). Although potassium channels primarily drive membrane potential changes in these cells, outwardly rectifying potassium currents were not significantly affected by 50 µM Angeli’s salt. Fast inward sodium currents were slightly but not significantly reduced by HNO. In more sensitive cells, HNO tended to reduce the pool of thiol groups.
Conclusions
As in the case of smooth muscle cells, HNO causes hyperpolarization of myenteric neurons, an effect also associated with an increase in intracellular Ca2+ concentration. Pathways other than activation of potassium currents appear to drive the hyperpolarization evoked by HNO.
Funding source: Deutsche Forschungsgemeinschaft
Award Identifier / Grant number: PO 2143/1-1
-
Research funding: This study was supported by a research fund from the German Research Foundation (PO 2143/1-1).
-
Author contributions: The author has accepted responsibility for the entire content of this manuscript and approved its submission.
-
Competing interests: Authors state no conflict of interest.
-
Informed consent: Not applicable.
-
Ethical approval: The local Institutional Review Board deemed the study exempt from review.
References
1. Gastreich-Seelig, M, Jimenez, M, Pouokam, E. Mechanisms associated to nitroxyl (HNO)-Induced relaxation in the intestinal smooth muscle. Front Physiol 2020;11:1–18. https://doi.org/10.3389/fphys.2020.00438.Search in Google Scholar PubMed PubMed Central
2. Pouokam, E, Vallejo, A, Martínez, E, Traserra, S, Jimenez, M. Complementary mechanisms of modulation of spontaneous phasic contractions by the gaseous signalling molecules NO, H2S, HNO and the polysulfide Na2S3 in the rat colon. J Basic Clin Physiol Pharmacol 2023;34:495–507. https://doi.org/10.1515/jbcpp-2021-0181.Search in Google Scholar PubMed
3. Haschke, G, Schafer, H, Diener, M. Effect of butyrate on membrane potential, ionic currents and intracellular Ca2+ concentration in cultured rat myenteric neurones. Neuro Gastroenterol Motil 2002;14:133–42. https://doi.org/10.1046/j.1365-2982.2002.00312.x.Search in Google Scholar PubMed
4. Pouokam, E, Rehn, M, Diener, M. Effects of H2O2 at rat myenteric neurones in culture. Eur J Pharmacol 2009;615:40–9. https://doi.org/10.1016/j.ejphar.2009.04.066.Search in Google Scholar PubMed
5. Rehn, M, Hübschle, T, Diener, M. TNF-α hyperpolarizes membrane potential and potentiates the response to nicotinic receptor stimulation in cultured rat myenteric neurones. Acta Physiol Scand 2004;181:13–22. https://doi.org/10.1111/j.1365-201x.2004.01269.x.Search in Google Scholar PubMed
6. Hamodeh, SA, Rehn, M, Haschke, G, Diener, M. Mechanism of butyrate-induced hyperpolarization of cultured rat myenteric neurones. Neuro Gastroenterol Motil 2004;16:597–604. https://doi.org/10.1111/j.1365-2982.2004.00545.x.Search in Google Scholar PubMed
7. Starodub, AM, Wood, JD. A-type potassium current in myenteric neurons from Guinea-pig small intestine. Neuroscience 2000;99:389–96. https://doi.org/10.1016/s0306-4522(00)00196-2.Search in Google Scholar PubMed
8. Vogalis, F, Hillsley, K, Smith, TK. Diverse ionic currents and electrical activity of cultured myenteric neurons from the Guinea pig proximal colon. J Neurophysiol 2000;83:1253–63. https://doi.org/10.1152/jn.2000.83.3.1253.Search in Google Scholar PubMed
9. Zholos, AV, Baidan, LV, Starodub, AM, Wood, JD. Potassium channels of myenteric neurons in Guinea-pig small intestine. Neuroscience 1999;89:603–18. https://doi.org/10.1016/s0306-4522(98)00337-6.Search in Google Scholar PubMed
10. Wood, JD. Physiology of the enteric nervous system. In: Physiology of the gastrointestinal tract. New York: Raven Press; 1994:423–82 pp.Search in Google Scholar
11. Evans, JR, Bielefeldt, K. Regulation of sodium currents through oxidation and reduction of thiol residues. Neuroscience 2000;101:229–36. https://doi.org/10.1016/s0306-4522(00)00367-5.Search in Google Scholar PubMed
12. Bianco, CL, Toscano, JP, Bartberger, MD, Fukuto, JM. The chemical biology of HNO signaling. Arch Biochem Biophys 2017;617:129–36. https://doi.org/10.1016/j.abb.2016.08.014.Search in Google Scholar PubMed PubMed Central
13. Cook, NM, Shinyashiki, M, Jackson, MI, Leal, FA, Fukuto, JM. Nitroxyl-mediated disruption of thiol proteins: inhibition of the yeast transcription factor Ace1. Arch Biochem Biophys 2003;410:89–95. https://doi.org/10.1016/s0003-9861(02)00656-2.Search in Google Scholar PubMed
14. Mandavilli, BS, Janes, MS. Detection of intracellular glutathione using ThiolTracker violet stain and fluorescence microscopy. In: Current protocols in cytometry; 2010, vol 53:9.35.1–8 pp.10.1002/0471142956.cy0935s53Search in Google Scholar PubMed
15. Jackson, MI, Fields, HF, Lujan, TS, Cantrell, MM, Lin, J, Fukuto, JM. The effects of nitroxyl (HNO) on H₂O₂ metabolism and possible mechanisms of HNO signaling. Arch Biochem Biophys 2013;538:120–9. https://doi.org/10.1016/j.abb.2013.08.008.Search in Google Scholar PubMed PubMed Central
16. Schindele, S, Pouokam, E, Diener, M. Hypoxia/reoxygenation effects on ion transport across rat colonic epithelium. Front Physiol 2016;7:1–10. https://doi.org/10.3389/fphys.2016.00247.Search in Google Scholar PubMed PubMed Central
© 2022 Walter de Gruyter GmbH, Berlin/Boston
